Specimen processing/set ups
SWABS:
1) Nasopharyngeal (flu pcr/respiratory culture) (green top)
2) red top (throat culture/group a strep/group b strep/wound)
3) red top with notch (mrsa pcr/staph aureus (mssa)/mrsa pcr)
4) puritan ez kit/red top in viral media: for biofire (filmarray) respiratory panel pcr
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throats bench:
B-hemolytic streptococci belonging to Lancefield Group A, Streptococcus pyogenes, members are the main etiological agents of human respiratory infections, including tonsillitis, strep throat, bronchopneumonia, scarlet fever, and skin infections such as erysipelas, and cellulitis. These organisms are also responsible for the development of infections such as glomerulonephritis and rheumatic fever.
Members of Group C and G are B-hemolytic and have been implicated with erysipelas, puerperal fever, and pharyngitis.
Group D streptococci include the enterococci, such as E. faecalis.
Streptococcal virulence is due to its ability to produce a wide variety of virulence factors, including hemolysins, leukocidins, erythrogenic toxin, hyaluronidase, streptokinase, and nucleases.
Members of Group C and G are B-hemolytic and have been implicated with erysipelas, puerperal fever, and pharyngitis.
Group D streptococci include the enterococci, such as E. faecalis.
Streptococcal virulence is due to its ability to produce a wide variety of virulence factors, including hemolysins, leukocidins, erythrogenic toxin, hyaluronidase, streptokinase, and nucleases.
Rapid streptococcus tests that come out negative does not rule out strep throat or bacterial pharyngitis, therefore, specimens that are negative by rapid test should also be cultured. This takes anywhere from 18-24 hours or more. Bacteria may be plated on special blood agar that enhances the growth of Gram-positive cocci, such as SSA agar (Streptococcus Selective Agar), which contains antibiotics that prevent the growth of most Gram-negative organisms and promote or enhance the growth of Gram-positive ones. Plates are read at 24 and 48 hours and observed for beta-hemolysis. Beta-hemolytic organisms should be tested with the streptococcal panel, the PathoDX, to look for Group A, B, C, F, or G streptococcus, Arcanobacterium haemolyticum, Fusobacterium necrophorum, or Corynebacterium diphtheriae. View the images below to see what organisms to look for and their beta-hemolytic patterns on SSA blood agar:
streptococcus groups:
LANCEFIELD GROUP A (GAS): Streptococcus Pyogenes:
Causative agent of:
- Strep throat/pharyngitis/tonsillitis
- swollen glands, sore throat, fever, redness, white spots on tonsils, possible red "strawberry spots" on the back of the throat and tongue, difficulty swallowing/painful swallowing, nausea, vomiting, irritability, chills
- swollen glands, sore throat, fever, redness, white spots on tonsils, possible red "strawberry spots" on the back of the throat and tongue, difficulty swallowing/painful swallowing, nausea, vomiting, irritability, chills
- Skin and soft tissue infections (cellulitis)
- affects the deeper layers of skin, including the lower dermis and subdermal/subcutaneous layers
- affects the deeper layers of skin, including the lower dermis and subdermal/subcutaneous layers
- Erysipelas (bacterial infection of the upper dermis, the layer of skin just under the epidermis)
- skin is bright red, inflamed, rough, leather-like
- not as deep as cellulitis
- can be caused by scratching when the bacteria enters through breaks in the skin
- skin is bright red, inflamed, rough, leather-like
- Impetigo
- red crusty sores and itch and ooze and spread
- affects the superficial layers of skin (epidermis)
- red crusty sores and itch and ooze and spread
- Necrotizing fasciitis
- flesh-eating disease, death of skin and soft tissue, turns purple/blue, spreads rapidly, can be serious and result in disfiguration, amputation, surgery, or even death if not treated quickly
- flesh-eating disease, death of skin and soft tissue, turns purple/blue, spreads rapidly, can be serious and result in disfiguration, amputation, surgery, or even death if not treated quickly
- Toxic Shock Syndrome
- Glomerulonephritis
- post-streptococcal kidney infection
- blood in urine (hematuria)
- post-streptococcal kidney infection
- Rheumatic Fever
- post-streptococcal fever and joint pain
- can affect the heart as well
- fever and rash
- immune response
- may have permanent effects
- post-streptococcal fever and joint pain
- Scarlet Fever
- fever and rash with strep throat due to immune response to the bacterial infection
- if untreated, may lead to rheumatic fever and/or post-streptococcal glomerulonephritis
- fever and rash with strep throat due to immune response to the bacterial infection
- Pneumonia (rarely)
- Otitis (middle ear infection)
- common in children
- bacteria spreads from the throat to the middle ears
- common in children
- Bacteremia or sepsis
- bacteria has spread into the bloodstream
- bacteria has spread into the bloodstream
- Tonsillar abscess
- pocket of pus behind the tonsil(s)
- treated with antibiotics
- may need to be drained
- pocket of pus behind the tonsil(s)
- Joint, Kidney or Heart Damage
- Sometimes Fatal
This type of streptococcus can be very contagious, especially "Strep Throat". It can be spread through direct contact of mucous membranes or sores on the skin. It is beta-hemolytic, catalase-negative, CAMP negative, PYR positive, Hippurate negative, susceptible to Bacitracin, Gram-positive cocci in chains and pairs. It may cause either non-invasive (mild) or invasive (severe) infections. Most of the time it is still susceptible to Penicillin. If someone has an allergy to Penicillin, then Erythromycin, Macrolides, or Cephalosporins may be prescribed instead. It causes many infection per year across the globe, at a rate of >500,000 worldwide. Virulence and variance of severity of the infection is linked to both epigenetic, phenotypic host and environmental factors. In fact, immunocompromised individuals are at higher risk for infection.
Humans may carry streptococci in their nose or on their skin without displaying any symptoms, but can still carry it and spread it to others who may develop an infection. However, carriers are not as contagious as those with an active infection. Non-invasive infections are more common and less severe than invasive ones. When the bacteria colonizes the throat, it has a preference for epithelial cells. Non-invasive infections include Strep Throat (pharyngitis) (15-30% of childhood infections and 10% of adult cases), Scarlet Fever, impetigo (a skin infection), cellulitis, abscesses and erysipelas. Because these types of infections can be spread via person-to-person contact, cleanliness and washing of the hands frequently is important. Strep Throat is contagious about a week before actual onset of symptoms.
Invasive infections are less common and more severe. When the bacteria gets into the bloodstream and infects organs or deeper tissues, this is considered to be an invasive infection. These infections include bacteremia, sepsis, Toxic Shock Syndrome (TSS), and necrotizing fasciitis (NF). It may also include pneumonia (pulmonary alveolus). Complications may include ear infections, glomerulonephritis, acute rheumatic fever, joint (septic arthritis), kidney and/or heart damage. Rarely, the bacteria may cause a bone infection called osteomyelitis, meningitis or sinusitis. These can all lead to shock, organ failure, or even death, but this is rare. This is why prevention, early recognition, proper diagnosis and treatment are extremely important. Blood counts, blood cultures and urine cultures are important in the detection of bloodstream or more invasive infections.
Virulence Factors:
Acute Rheumatic Fever:
Humans may carry streptococci in their nose or on their skin without displaying any symptoms, but can still carry it and spread it to others who may develop an infection. However, carriers are not as contagious as those with an active infection. Non-invasive infections are more common and less severe than invasive ones. When the bacteria colonizes the throat, it has a preference for epithelial cells. Non-invasive infections include Strep Throat (pharyngitis) (15-30% of childhood infections and 10% of adult cases), Scarlet Fever, impetigo (a skin infection), cellulitis, abscesses and erysipelas. Because these types of infections can be spread via person-to-person contact, cleanliness and washing of the hands frequently is important. Strep Throat is contagious about a week before actual onset of symptoms.
Invasive infections are less common and more severe. When the bacteria gets into the bloodstream and infects organs or deeper tissues, this is considered to be an invasive infection. These infections include bacteremia, sepsis, Toxic Shock Syndrome (TSS), and necrotizing fasciitis (NF). It may also include pneumonia (pulmonary alveolus). Complications may include ear infections, glomerulonephritis, acute rheumatic fever, joint (septic arthritis), kidney and/or heart damage. Rarely, the bacteria may cause a bone infection called osteomyelitis, meningitis or sinusitis. These can all lead to shock, organ failure, or even death, but this is rare. This is why prevention, early recognition, proper diagnosis and treatment are extremely important. Blood counts, blood cultures and urine cultures are important in the detection of bloodstream or more invasive infections.
Virulence Factors:
- M protein: causes antibodies to be secreted, which cross-react with autoantigens on cells or connective tissue, especially of the endocardium of the heart and synovial layer of the joints, which can lead to significant illness and damage
- Hemolysins: these cause hemolysis and rupture of red blood cells, resulting in the rashes that sometimes accompany streptococcal infections
- Streptoloysin O (exotoxin): lyses red blood cells, which is linked to formation of petechiae or rashes in humans, lysis of red blood cells in agar on bacterial culture, and attacks the cholesterol in the cell membranes of eukaryotic cells in humans, including immune system white blood cells, enabling the bacteria to evade phagocytosis
- Extracellular Enzymes: break down and penetrate blood cells
- Deep oropharyngeal abscess: Ampicillin/Sulbactam, Amoxicillin/Clavulanic Acid, Clindamycin, Aspiration and Drainage
- Toxic Shock Syndrome: Penicillin and Clindamycin, IgG
- Necrotizing Fasciitis: High-Dose Penicillin and Clindamycin, Surgery
Acute Rheumatic Fever:
- Usually caused by a complication of a respiratory infection linked to GAS
- Most commonly affects children (ages 5-15)
- Onset is 1-3 weeks post-streptococcal infection
- Signs and symptoms: pancarditis, endocarditis, joint pain and inflammation and arthritis, subcutaneous nodules, erythema (red rash), chorea (movement)
- Tests to confirm: DNAse, antibody (ASO) serology testing, ESR (will be elevated)
- Uncommon complication of Strep Throat or skin infection
- Type III Hypersensitivity Reaction (Delayed)
- Occurs around 10 days post-streptococcal throat infection or around 3 weeks post-streptococcal skin infection
- Inflammation of the kidney with blood in the urine (hematuria)
- Symptoms: pale skin, fatigue and lethargy, headache, loss of appetite, dull back pain, swelling (edema), elevated blood pressure, blood in urine
- Extremely rare complication that causes an acute, rapid onset of chorea (purposeless movement), neurological changes, OCD, personality changes, behavioral problems, aggression, slurred speech, tic disorders, feeling hot/skin feels hot, fever in children as a result of an autoimmune response to a GAS infection
Streptococcus pyogenes (Group A) on Sheep Blood Agar. S. pyogenes produces Beta-hemolysis on Sheep Blood Agar. Streptococcus pyogenes (Group A) is the causative agent of streptococcal pharyngitis, or Strep Throat. SSA agar is also helpful and is a selective and differential agar utilized for isolation of streptococcus. Placing 2 small stabs in the 1st quadrant can aid in creating a slightly anaerobic environment to encourage the growth and hemolysis of Group A strep. Always be aware that there are some strains that may not produce hemolysis yet be positive for Group A strep. Colonies are small, cream-colored, round and glistening/shiny and most are beta-hemolytic with medium zones of beta hemolysis surrounding the colonies.
LANCEFIELD GROUP B (GBS): Streptococcus agalactiae:
Group B streptococcal infections are caused by Streptococcus agalactiae. This can result in either serious illness such as meningitis or sepsis or even death in newborns, elderly people, or those who are immunocompromised. GBS is normally part of the human microflora, colonizing the gastrointestinal and genitourinary tract of about 30% of healthy adults. Those who are colonized with the bacterium are referred to as "carriers", and carry it harmlessly without displaying any symptoms of illness or disease. The bacterium is Gram-positive cocci in chains, beta-hemolytic (thin layer), catalase-negative, CAMP positive, hippurate positive, and is a facultative anaerobe. It is surrounded by a capsule of exopolysaccharides, and there are 10 serotypes based on the reactivity of the capsule. NOTE: There are also non-hemolytic strains of Group B streptococcus out there, so be mindful of this during your testing and identification on the bench.
Virulence Factors:
Samples That Can Be Collected at 35-37 Weeks of Gestation:
Virulence Factors:
- Capsular polysaccharide (sialic acid): evades the immune system and phagocytosis by phagocytes (neutrophils, macrophages)
- Porforin (pore-forming toxin)
- B-haemolysin
Samples That Can Be Collected at 35-37 Weeks of Gestation:
- Vaginal Swab
- Rectal Swab
- Rectovaginal Swab
- Swabs need to be inoculated in a selective enrichment broth such as LIM broth or Todd Hewitt broth with antibiotics
- After 18-24 hours, inoculate the broth onto blood agar plates or CHROMAgar
- CAMP positive test
- Latex agglutination positive with GBS antiserum
- Granada agar growth as orange or pink-red colonies or blue colonies
- Chromogenic growth as colored colonies (protect from light)
A loopful of isolate is placed in broth containing hippuric acid and incubated for 4 hours. Ninhydrin reagent is added. After 4 hours, read the broth and if it is purple, it indicates that it is hippurate positive. This is another test that aids in the identification of GBS. Hippuric acid is a benzene ring, which is a 6-carbon ring. It contains one amino acid, serine, off one of its carbon rings. Hippuricase enzyme snips the benzene ring from serine and separates them, and this test detects the amino acid serine, resulting in a purple color if it is present.
LANCEFIELD GROUP C: Streptococcus equi, S. dysgalactiae subspp dysgalactiae or equisimilis, S. zooepidemicus
Group C streptococci produce huge zones of beta-hemolysis on blood agar. S. dysgalactiae are significant for humans because this bacterium can cause tonsillitis, skin infections, severe necrotizing fasciitis, cellulitis, endocarditis, bacteremia, pneumonia, meningitis, post-streptococcal glomerulonephritis, acute rheumatic fever, or significant respiratory infections. It also causes pharyngitis in humans, as well as other pyrogenic infections similar to GAS. Individuals with underlying illnesses, such as cancer or diabetes, are at higher risk for opportunistic infection.
S. dysgalactiae is an opportunistic pathogen that may cause pharyngitis with symptoms similar to that of Group A streptococci. There is also an emerging invasive strain of this organism that is categorized with Group A strep since it is so similar to Group A strep. S. zooepidemicus is an opportunistic pathogen of humans. S. equi causes respiratory infections in horses. S. dysgalactiae subspp equisimilis is a mammalian pathogen, but is rarely associate with human infections such as meningitis, bacteremia, skin and soft tissue infections. Because it is part of the normal oral flora of many animals, it may also be involved in bite infections. It produces the virulence factor hyaluronidase, which digests hyaluronic acid and chondroitin. This organism is also part of Group G streptococci as well. This subspp is mostly beta-hemolytic and it is PYR negative and resistant to bacitracin and hyaluronidase negative. S. dysgalactiae subspp dysgalactiae tends to be predominately alpha-hemolytic as opposed to the predominately beta-hemolytic strains.
LANCEFIELD GROUP D: Streptococcus equinus, S. bovis, S. gallolyticus, Enterococci (E. faecalis, E. raffinosus, etc...)
Streptococcus equinus is rarely found in humans but can very rarely cause bacteremia, sepsis, peritonitis and infective endocarditis in humans. Streptococcus bovis is a group consisting of S. bovis, S. caprinus, and S. gallolyticus. It is associated with UTI, endocarditis, peritonitis and colorectal cancer in humans. It has also been associated with neonatal septicemia and meningitis, as well as bacterial peritonitis in patients with underlying liver cirrhosis. S. gallolyticus has been linked to colon adenomas and carcinomas. Diets high in starch and sugar may increase risk of infection and malignancy. S. bovis is catalase and oxidase negative. Most are non (gamma)-hemolytic and some are alpha-hemolytic.
E. faecalis is part of the family of Lancefield Group D Streptococci. It is linked to significant nosocomial (hospital-acquired) infections. Some of these infections are life-threatening. Many strains have become antibiotic-resistant. It is nonmotile, it utilizes/ferments glucose, it does not produce gas, it is catalase-negative or weakly positive, and it grows consistently in nutrient broth and is a facultative anaerobe. It utilizes glycerol, lactate, malate, citrate, arginine, agmatine, and keto acids. It is PYR and Hippurate +. It is beta-lactamase negative. Infections linked to this organism include:
E. faecalis is part of the family of Lancefield Group D Streptococci. It is linked to significant nosocomial (hospital-acquired) infections. Some of these infections are life-threatening. Many strains have become antibiotic-resistant. It is nonmotile, it utilizes/ferments glucose, it does not produce gas, it is catalase-negative or weakly positive, and it grows consistently in nutrient broth and is a facultative anaerobe. It utilizes glycerol, lactate, malate, citrate, arginine, agmatine, and keto acids. It is PYR and Hippurate +. It is beta-lactamase negative. Infections linked to this organism include:
- UTI
- Septicemia
- Bacteremia
- Endocarditis
- Meningitis
- Others
LANCEFIELD GROUP F: Streptococcus anginosus group, S. milleri group (Includes S. intermedius, S. anginosus, S. constellatus)
The Streptococcus anginosus group consists of S. anginosus, S. intermedius, and S. constellatus. It is part of the commensal flora of the mouth, throat, vaginal tract, GI tract. This bacterium has been known to cause bacteremia and sepsis, typically seeding from abscesses, especially liver and brain abscesses, and occasionally spleen abscesses. S. anginosus has a sweet, buttery caramel or butterscotch scent on SBA. Some strains are gamma-hemolytic (about 1/2), some strains are alpha (about 36%) and a few are beta-hemolytic. Colonies tend to be tiny and either raised or flat. S. constellatus is associated with upper respiratory tract and upper body abscesses. Cystic Fibrosis patients are prone to exacerbations with this organism. S. intermedius is associated with purulent infections, periodontitis, liver and brain abscesses. These are members of the S. milleri group.
This group of bacteria ferments glucose, sucrose, lactose and trehalose. They hydrolyze esculin and arginine as well. Most are resistant to Bacitracin and Nitrofurazone but susceptible to Penicillin, Ampicillin, Erythromycin, and Tetracycline.
This group of bacteria ferments glucose, sucrose, lactose and trehalose. They hydrolyze esculin and arginine as well. Most are resistant to Bacitracin and Nitrofurazone but susceptible to Penicillin, Ampicillin, Erythromycin, and Tetracycline.
LANCEFIELD GROUP G STREPTOCOCCI: Streptococcus canis, S. dysgalactiae subspp equisimilis
Streptococcus canis is part of the normal respiratory flora of dogs and cats, and is occasionally isolated from a dog or cat bite in humans. It is also part of the normal skin and oral flora of humans. It occasionally causes pharyngitis with symptoms similar to that of Group A streptococci, as well as skin infections. It has been associated with rare cases of meningitis and sepsis in humans. It is a clinically significant opportunistic pathogen and nosocomial pathogen. It is often seen in polymicrobial infections, particularly those including S. aureus. Skin and soft tissue infections with this organism have begun emerging in recent years. Malignancy, alcoholism and diabetes predispose individuals to a higher risk of infection with this organism. Endocarditis, septic arthritis, pulmonary infections, bone and joint infections, eye infections, peritonitis, neonatal infections, bacteremia, and infections involving prosthetic devices have been seen with this organism.
Most of the time, these bacteria are beta-hemolytic and produce large zones of beta-hemolysis. The colonies tend to be small, yellowish, convex. S. dysgalactiae is the most significant for humans in terms of disease. It can cause pharyngitis, tonsillitis, skin infections, severe necrotizing fasciitis, cellulitis, bone and joint infection, pneumonia, and bacteremia. Invasive disease is most common in individuals with underlying health conditions such as cancer or diabetes. Subspecies equisimilus has also been linked to the above infections plus post-streptococcal glomerulonephritis and rheumatic fever and rheumatoid arthritis.
Virulence Factors:
Most of the time, these bacteria are beta-hemolytic and produce large zones of beta-hemolysis. The colonies tend to be small, yellowish, convex. S. dysgalactiae is the most significant for humans in terms of disease. It can cause pharyngitis, tonsillitis, skin infections, severe necrotizing fasciitis, cellulitis, bone and joint infection, pneumonia, and bacteremia. Invasive disease is most common in individuals with underlying health conditions such as cancer or diabetes. Subspecies equisimilus has also been linked to the above infections plus post-streptococcal glomerulonephritis and rheumatic fever and rheumatoid arthritis.
Virulence Factors:
- M protein (helps evade the immune system and phagocytosis by inactivating complement)
- Protein G (binds immunoglobulins and inhibits antibody response)
- DrsG (a protein that inhibits the effects of antimicrobial peptides secreted by immune cells)
- Toxins and hemolysis, including streptolysin O and streptolysin S
- Streptokinase enzyme (enables fibrinolysis so that the bacteria can spread in the bloodstream)
- Ability to form biofilm
If you see the beta hemolytic patterns above on the throat cultures, the following steps should be performed:
1) Perform the PathoDX and test with Group A latex reagent first. NOTE: there are other kits other laboratories may use, including Streptex kit. These steps are for the PathoDX kit.
-2 drops of reagent 1 in glass tube
-2 drops of reagent 2 in glass tube
-Use wooden applicator to touch 5 colonies and mix with reagents
-4 drops of reagent 3 in glass tube
-Pipette 1-2 drops onto card into each circle being tested
-1 drop of latex reagent in same circle for each circle being tested
-Mix with wooden applicator stick and rotate card for 15-60 seconds to mix the specimen and latex, being careful to keep the solution inside the circle so you don't mix with the next circle (could produce a false-positive in the next circle if the one next to it is positive)
-Positive samples will show visible clumping/agglutination (blue clumps with a light blue or clear background, uniformly throughout)
OR
1) Perform the Streptex test panel beginning with Group A latex reagent first.
-3 drops of reagent 1 in glass tube (blue-green sodium nitrite solution)
-3 drops of reagent 2 in glass tube (yellow acetic acid solution)
-use a wooden applicator to touch 5 colonies and mix with reagents
-incubate at room temp. for 1 minute
-3 drops of reagent 3 in glass tube (Tris buffer neutralizing solution)
-mix well; solution will turn purple
-pipette 1 drop of solution onto the card in each circle
-shake the latex reagents really well and place a drop next to the solution in the circle
-mix with wooden applicator stick and rotate the card for up to 60 seconds, being careful to keep the solution inside the circle
-Positive samples will show visible clumping/agglutination (blue clumps with a light blue or clear background, uniformly throughout)
2) If the isolate is negative for Group A, perform the Panel Test (Groups B, C, F, G). If positive, report out which one is positive.
3) If all streptococcal latex reagents are negative, perform a Gram stain and look at the images below to rule out the following organisms:
1) Perform the PathoDX and test with Group A latex reagent first. NOTE: there are other kits other laboratories may use, including Streptex kit. These steps are for the PathoDX kit.
-2 drops of reagent 1 in glass tube
-2 drops of reagent 2 in glass tube
-Use wooden applicator to touch 5 colonies and mix with reagents
-4 drops of reagent 3 in glass tube
-Pipette 1-2 drops onto card into each circle being tested
-1 drop of latex reagent in same circle for each circle being tested
-Mix with wooden applicator stick and rotate card for 15-60 seconds to mix the specimen and latex, being careful to keep the solution inside the circle so you don't mix with the next circle (could produce a false-positive in the next circle if the one next to it is positive)
-Positive samples will show visible clumping/agglutination (blue clumps with a light blue or clear background, uniformly throughout)
OR
1) Perform the Streptex test panel beginning with Group A latex reagent first.
-3 drops of reagent 1 in glass tube (blue-green sodium nitrite solution)
-3 drops of reagent 2 in glass tube (yellow acetic acid solution)
-use a wooden applicator to touch 5 colonies and mix with reagents
-incubate at room temp. for 1 minute
-3 drops of reagent 3 in glass tube (Tris buffer neutralizing solution)
-mix well; solution will turn purple
-pipette 1 drop of solution onto the card in each circle
-shake the latex reagents really well and place a drop next to the solution in the circle
-mix with wooden applicator stick and rotate the card for up to 60 seconds, being careful to keep the solution inside the circle
-Positive samples will show visible clumping/agglutination (blue clumps with a light blue or clear background, uniformly throughout)
2) If the isolate is negative for Group A, perform the Panel Test (Groups B, C, F, G). If positive, report out which one is positive.
3) If all streptococcal latex reagents are negative, perform a Gram stain and look at the images below to rule out the following organisms:
Other causes of non-streptococcal pharyngitis:
Arcanobacterium haemolyticum:
Arcanobacterium haemolyticum can cause pharyngitis. It is also associated with tonsillar abscess. If you see anything resembling this, perform a Vitek or Mass Spec ID and susceptibility test, and the organism may need to be sent out to the state reference laboratory for confirmation. This organism may also be involved in peritonsillar abscesses. This bacterium is Gram-positive-positive, non-motile, and is a facultative anaerobe. The name of this bacterium means "hidden bacterium".
Fusobacterium necrophorum:
Fusobacterium necrophorum is an anaerobe and it is a causative agent of pharyngitis, peritonsillar abscesses and the potentially fatal Lemierre's syndrome/disease, usually causing sepsis and being recovered in blood specimens (anaerobic). If this organism is suspected, perform a Vitek ID and susceptibility test. This organism should be reported to the physician ASAP.
Corynebacterium:
Corynebacterium diphtheriae or haemolyticum may cause pharyngitis. C. diphtheriae is extremely rare these days because the DTaP vaccination provides immunity against this pathogenic microbe. Other types of Corynebacterium or normal colonizers of the throat and respiratory tracts.
Actinomyces:
Actinomyces/Arcanobacterium pyogenes is a rare causative agent of pharyngitis and is very similar to A. haemolyticum.
Bordetella pertussis and parapertussis:
Neisseria spp:
Haemophilus spp:
Spirochetes:
Candida albicans:
Staphylococcus:
Strep throat
Scarlet Fever
rheumatic fever
glomerulonephritis
peritonsillar abscess
Vincent's Angina ulcer
otitis media: middle/inner ear infection
mastoiditis: infection of the bones of the jaw or the bones of the mastoid process behind the ear
Candidiasis: oral thrush
If you see yeast on the throat culture, and it is pure or predominant, perform a Gram stain and a germ tube test to rule out Candida albicans, the causative agent of thrush.
If your germ tube is negative, perform a Vitek yeast card ID or an API ID test and subculture the organism to a Cornmeal agar with 4 stabs in the first quadrant covered with a coverslip. Correlate your Vitek or API results with your Cornmeal agar results.
the pathophysiology of the conditions seen in the images above show the seriousness of why it is important to correctly and quickly identify the bacteria which cause these conditions and treat infections so that they do not spread.
Streptococcus pyogenes virulence factors:
Streptococcus pyogenes is the causative agent of Strep Throat, Scarlet Fever, Rheumatic Fever, Glomerulonephritis, and can cause skin infections. Its virulence is due to a number of factors:
1) Lancefield antigen A
2) M protein (there are >80 types), which is a major virulence factor for Group A streptococcus because it
-blocks complement
-protects the bacterium from phagocytosis
Plasma (B-Cells) secrete antibodies against M protein, opsonizing it and aiding in its destruction by macrophages and neutrophils.
3) Enzymes, which are major factors in its pathogenicity
-Streptolysin O (oxygen labile, inactivated by oxygen)
-destroys red blood cells and white blood cells
-responsible for the Beta-hemolysis on the Sheep Blood Agar
-is antigenic, leading to the formation of anti-streptolysin O ASO antibodies, which can be measured by ASO titers (immunoserology)
-Streptolysin S (oxygen stable, not inactivated by oxygen)
-also responsible for Beta-hemolysis on the Sheep Blood Agar
-Pyrogenic Exotoxin/Erythrogenic Toxin
-can cause Scarlet Fever (pyrogens released by the bacterium are responsible for the symptoms of fever, redness, heat, pain)
-some are superantigens that superstimulate T-cells to pour out cytokines, causing inflammation (swelling, pain)
-can cause streptococcal Toxic Shock Syndrome
-Streptokinase
-activates proteolytic plasmin
-breaks up fibrin blood clots
-Hyaluronidase
-increases tissue permeability
-plays a role in angiogenesis
-to use hyaluronon as a carbon source
-destroys polysaccharides that hold cell walls together in human cells
-makes it easier and quicker for the bacteria to spread
-DNAases
-Anti-C5a Peptidase
-minimizes the influx of neutrophils
-Polysaccharide capsule
-hides from the immune system
-Pili
-attachment to mucosal surfaces
Infections:
A. Local Invasion/Exotoxin Release:
1. Streptococcus Pharyngitis ("Strep Throat"), a self-limiting illness that tends to resolve within 5 days
2. Skin Infections
a. Folliculitis
b. Cellulitis
c. Impetigo
d. Necrotizing fascitis
3. Scarlet Fever, due to the release of pyrogenic toxin and erythrogenic toxin
4. Streptococcal Toxic Shock Syndrome
B. Delayed Antibody-Mediated Diseases (Hypersensitivity Reactions):
1. Rheumatic Fever (fever, rash, nodules, myocarditis, joint pain, and rarely, formation of chorea, or uncontrolled musculoskeletal movements)
2. Glomerulonephritis (hematuria, or blood in the urine, with damage to the glomeruli of the kidneys)
Incubation period is about 1-3 days
>700 million infections worldwide each year
Treatment:
Treatment includes Penicillin G or Clindamycin. Penicillin G is the original penicillin discovered by Alexander Fleming. The oral form of this medicine is acid stable in the stomach and commonly given for streptococcus pharyngitis caused by Group A beta-hemolytic streptococci. Other alternatives include the Cephalosporins or Macrolides.
STREPTOCOCCAL PATHODX PANEL:
If growth on the Sheep Blood Agar is heavy (in quadrant 4), a PathoDx panel is run.
Group B streptococci (GBS), S. agalactiae, is rarely found in the throat and rarely a cause of a pharyngeal infection. However, serious group B strep infections in adults can be fatal should they occur, especially in patients who are immunosuppressed or immunocompromised. On average, approximately 8% of adults with invasive group B strep infections (infections where the bacteria have entered a part of the body that is normally not exposed to bacteria) die. The risk of death is lower among younger adults, and adults who do not have other medical conditions.
Group C streptococci (GCS) and group G streptococci (GGS) are part of the normal commensal flora of the human upper respiratory tract and frequently colonize the skin, gastrointestinal tract and female genital tract without causing any symptoms. Occasionally, they are implicated in pharyngitis and in mono- and polymicrobial infections of the skin and soft tissues, bacteremia and endocarditis, septic arthritis and osteomyelitis, puerperal infections, and meningitis. These include S. equi, S. dysagalactiae, and S. canis.
Group D streptococci (E. faecalis, E. faecium)
Group F streptococci (GFS) are part of the normal oropharyngeal commensal flora. Rarely, these organisms (S. anginosus, S. milleri) can cause pharyngitis and may be associated with peritonsillar abscesses and bacteremia.
Group G streptococci (GGS) (S. canis) (See description above)
Other Pathogens That Can Cause Pharyngitis, Colonize the Oropharyngeal Passages, or Cause Other Types of Upper Respiratory Infections (SOP: Non-streptococcal Pharyngitis):
Fusobacterium necrophorum (GNR, fusiform, long, rounded or pointed ends, sometimes curved or variable in size; Associated with peritonsillar abscesses; May cause pharyngeal ulcers with thin pseudomembrane formation, and usually seen with spirochetes; This is very rare)
Arcanobacterium haemolyticum (GPR, Corynebacterium-like in appearance, rarely associated with pharyngitis)
Actinomyces pyogenes (GPR, branching, rarely associated with pharyngitis)
Corynebacterium diphtheriae, Corynebacterium ulcerans (C. diphtheriae is the causative agent of diphtheria, which forms a pseudomembrane over the tonsils/pharyngeal area; These are GPRs shaped like "drumsticks" or "Chinese Letters", and may be side-by-side in pallasides form)
Bordetella pertussis, Bordetella parapertussis (B. pertussis causes "Whooping Cough" and B. parapertussis has been associated with epiglottitis)
Neisseria gonorrhoeae (GNDC; pharyngitis)
Neisseria meningitidis (GNDC; meningitis; This has never officially been defined as a throat pathogen, per se, but throat swabs may be screened for this organism for epidemiological purposes)
Candida albicans (Yeast; Associated with thrush, particularly in infants, diabetics, immunocompromised, and those on broad spectrum or long-term antibiotics)
Haemophilus influenzae, Haemophilus parainfluenzae (GN coccobacilli/short rods; Associated with acute epiglottitis, which can be life-threatening)
Spirochetes (Spiral, GN; Associated with oropharyngeal abscesses or dental abscesses, usually mixed; Also associated with pharyngeal ulcers covered with a thin pseudomembrane and may spread to the heart, resulting in Vincent's angina; Usually seen with F. necrophorum, and this is very rare)
Streptococcus pneumoniae (GPC's in pairs and short chains, often "lancet"-shaped; May travel from the throat through the eustachian tube, resulting in ear infections or more seriously, meningitis; May travel from the upper respiratory tract to the lower respiratory tract, resulting in bacterial pneumonia)
Staphylococcus aureus (GPC; May be present in the throat culture without causing infection)
Coagulase Negative Staphylococci (Neonates) (GPC, coagulase negative, non-hemolytic; May colonize neonates and should be noted)
Pseudomonas aeruginosa (GNR; May colonize the throat and should be noted)
1) Lancefield antigen A
2) M protein (there are >80 types), which is a major virulence factor for Group A streptococcus because it
-blocks complement
-protects the bacterium from phagocytosis
Plasma (B-Cells) secrete antibodies against M protein, opsonizing it and aiding in its destruction by macrophages and neutrophils.
3) Enzymes, which are major factors in its pathogenicity
-Streptolysin O (oxygen labile, inactivated by oxygen)
-destroys red blood cells and white blood cells
-responsible for the Beta-hemolysis on the Sheep Blood Agar
-is antigenic, leading to the formation of anti-streptolysin O ASO antibodies, which can be measured by ASO titers (immunoserology)
-Streptolysin S (oxygen stable, not inactivated by oxygen)
-also responsible for Beta-hemolysis on the Sheep Blood Agar
-Pyrogenic Exotoxin/Erythrogenic Toxin
-can cause Scarlet Fever (pyrogens released by the bacterium are responsible for the symptoms of fever, redness, heat, pain)
-some are superantigens that superstimulate T-cells to pour out cytokines, causing inflammation (swelling, pain)
-can cause streptococcal Toxic Shock Syndrome
-Streptokinase
-activates proteolytic plasmin
-breaks up fibrin blood clots
-Hyaluronidase
-increases tissue permeability
-plays a role in angiogenesis
-to use hyaluronon as a carbon source
-destroys polysaccharides that hold cell walls together in human cells
-makes it easier and quicker for the bacteria to spread
-DNAases
-Anti-C5a Peptidase
-minimizes the influx of neutrophils
-Polysaccharide capsule
-hides from the immune system
-Pili
-attachment to mucosal surfaces
Infections:
A. Local Invasion/Exotoxin Release:
1. Streptococcus Pharyngitis ("Strep Throat"), a self-limiting illness that tends to resolve within 5 days
2. Skin Infections
a. Folliculitis
b. Cellulitis
c. Impetigo
d. Necrotizing fascitis
3. Scarlet Fever, due to the release of pyrogenic toxin and erythrogenic toxin
4. Streptococcal Toxic Shock Syndrome
B. Delayed Antibody-Mediated Diseases (Hypersensitivity Reactions):
1. Rheumatic Fever (fever, rash, nodules, myocarditis, joint pain, and rarely, formation of chorea, or uncontrolled musculoskeletal movements)
2. Glomerulonephritis (hematuria, or blood in the urine, with damage to the glomeruli of the kidneys)
Incubation period is about 1-3 days
>700 million infections worldwide each year
Treatment:
Treatment includes Penicillin G or Clindamycin. Penicillin G is the original penicillin discovered by Alexander Fleming. The oral form of this medicine is acid stable in the stomach and commonly given for streptococcus pharyngitis caused by Group A beta-hemolytic streptococci. Other alternatives include the Cephalosporins or Macrolides.
STREPTOCOCCAL PATHODX PANEL:
If growth on the Sheep Blood Agar is heavy (in quadrant 4), a PathoDx panel is run.
Group B streptococci (GBS), S. agalactiae, is rarely found in the throat and rarely a cause of a pharyngeal infection. However, serious group B strep infections in adults can be fatal should they occur, especially in patients who are immunosuppressed or immunocompromised. On average, approximately 8% of adults with invasive group B strep infections (infections where the bacteria have entered a part of the body that is normally not exposed to bacteria) die. The risk of death is lower among younger adults, and adults who do not have other medical conditions.
Group C streptococci (GCS) and group G streptococci (GGS) are part of the normal commensal flora of the human upper respiratory tract and frequently colonize the skin, gastrointestinal tract and female genital tract without causing any symptoms. Occasionally, they are implicated in pharyngitis and in mono- and polymicrobial infections of the skin and soft tissues, bacteremia and endocarditis, septic arthritis and osteomyelitis, puerperal infections, and meningitis. These include S. equi, S. dysagalactiae, and S. canis.
Group D streptococci (E. faecalis, E. faecium)
Group F streptococci (GFS) are part of the normal oropharyngeal commensal flora. Rarely, these organisms (S. anginosus, S. milleri) can cause pharyngitis and may be associated with peritonsillar abscesses and bacteremia.
Group G streptococci (GGS) (S. canis) (See description above)
Other Pathogens That Can Cause Pharyngitis, Colonize the Oropharyngeal Passages, or Cause Other Types of Upper Respiratory Infections (SOP: Non-streptococcal Pharyngitis):
Fusobacterium necrophorum (GNR, fusiform, long, rounded or pointed ends, sometimes curved or variable in size; Associated with peritonsillar abscesses; May cause pharyngeal ulcers with thin pseudomembrane formation, and usually seen with spirochetes; This is very rare)
Arcanobacterium haemolyticum (GPR, Corynebacterium-like in appearance, rarely associated with pharyngitis)
Actinomyces pyogenes (GPR, branching, rarely associated with pharyngitis)
Corynebacterium diphtheriae, Corynebacterium ulcerans (C. diphtheriae is the causative agent of diphtheria, which forms a pseudomembrane over the tonsils/pharyngeal area; These are GPRs shaped like "drumsticks" or "Chinese Letters", and may be side-by-side in pallasides form)
Bordetella pertussis, Bordetella parapertussis (B. pertussis causes "Whooping Cough" and B. parapertussis has been associated with epiglottitis)
Neisseria gonorrhoeae (GNDC; pharyngitis)
Neisseria meningitidis (GNDC; meningitis; This has never officially been defined as a throat pathogen, per se, but throat swabs may be screened for this organism for epidemiological purposes)
Candida albicans (Yeast; Associated with thrush, particularly in infants, diabetics, immunocompromised, and those on broad spectrum or long-term antibiotics)
Haemophilus influenzae, Haemophilus parainfluenzae (GN coccobacilli/short rods; Associated with acute epiglottitis, which can be life-threatening)
Spirochetes (Spiral, GN; Associated with oropharyngeal abscesses or dental abscesses, usually mixed; Also associated with pharyngeal ulcers covered with a thin pseudomembrane and may spread to the heart, resulting in Vincent's angina; Usually seen with F. necrophorum, and this is very rare)
Streptococcus pneumoniae (GPC's in pairs and short chains, often "lancet"-shaped; May travel from the throat through the eustachian tube, resulting in ear infections or more seriously, meningitis; May travel from the upper respiratory tract to the lower respiratory tract, resulting in bacterial pneumonia)
Staphylococcus aureus (GPC; May be present in the throat culture without causing infection)
Coagulase Negative Staphylococci (Neonates) (GPC, coagulase negative, non-hemolytic; May colonize neonates and should be noted)
Pseudomonas aeruginosa (GNR; May colonize the throat and should be noted)
stool bench
The Enterobacteriaceae are the facultative anaerobic, Gram-negative bacilli bacteria that live in the gut and are part of the natural/normal gastrointestinal flora. Most of these types of bacteria actually help prevent disease by breaking down foods along with enzymes, competing with pathogens for nutrients, and providing vitamins, such as vitamin K, for our diet. However, because we are colonized with this bacteria, they can become opportunistic pathogens if they are in the right place at the right time and the conditions are just right. Sometimes it may be an overgrowth of one type of organism when other flora are wiped out or damaged from strong antimicrobials given to treat other conditions (examples: yeast overgrowth or C. difficile overgrowth or Pseudomonas aeruginosa overcolonization). Examples of our normal gut flora and Enterobacteriaceae include the following:
Non-enterobacteriaceae are not or are never or rarely part of the normal gut flora, are considered pathogenic, and may cause severe gastrointestinal illness. These include organisms such as Salmonella, Shigella, Aeromonas, Plesiomonas, Yersinia enterocolitica, Bacillus cereus, Campylobacter spp, E. coli 0157:H7. These organisms cause illness via the fecal-oral route, through ingestion of contaminated food or water. Undercooked meats, poultry, and eggs, improperly washed lettuce and other vegetables, and contaminated peanut butter can transmit Salmonella. Some of these organisms cause bloody diarrhea, whereas others or other strains cause watery diarrhea only or a combination of diarrhea and constipation. Some cause vomiting and nausea as well. It is important to identify and treat the organisms that cause gastroenteritis.
- Klebsiella spp
- Escherichia coli
- Enterobacter spp
- Citrobacter spp
- Serratia spp
- Proteus spp
- Providencia spp
- Bacteroides spp
- Pseudomonas spp
- Candida albicans
- Clostridium spp
- Fusobacterium spp
- Pantoea spp
- Edwardsiella tarda
Non-enterobacteriaceae are not or are never or rarely part of the normal gut flora, are considered pathogenic, and may cause severe gastrointestinal illness. These include organisms such as Salmonella, Shigella, Aeromonas, Plesiomonas, Yersinia enterocolitica, Bacillus cereus, Campylobacter spp, E. coli 0157:H7. These organisms cause illness via the fecal-oral route, through ingestion of contaminated food or water. Undercooked meats, poultry, and eggs, improperly washed lettuce and other vegetables, and contaminated peanut butter can transmit Salmonella. Some of these organisms cause bloody diarrhea, whereas others or other strains cause watery diarrhea only or a combination of diarrhea and constipation. Some cause vomiting and nausea as well. It is important to identify and treat the organisms that cause gastroenteritis.
Aeromonas hydrophila on Blood Agar is seen here. Aeromonas hydrophila is found in aquatic environments throughout the globe (fresh water, polluted water, chlorinated water, brackish water, marine salt water). It may either colonize the GI tract, or may cause infection via ingestion of contaminated food or water (meat, dairy, produce). Exposed wounded skin or mucosal surfaces may also be infected via traumatic implantation by fish hooks or fish fins, contaminated water or soil. GI infection results in watery diarrhea. Other rare infections that may be caused by this organism include endocarditis, septicemia, meningitis, pneumonia, osteomyelitis, and conjunctivitis.
Aeromonas hydrophila on trypticase sheep agar 5% is seen here. It produces large, round, raised, mucoid, opaque colonies that are beta-hemolytic and buff-to-tan-to-slightly orange in color. Aeromonas hydrophila is oxidase positive, glucose and sucrose positive, LDC positive. This spp may express resistance to various antimicrobials.
If you have an oxidase + organism that looks like this (tan-gray, mucoid, possibly beta-hemolytic), perform TULM test (TSI, Urea, LIA and MIO slants), a Vitek ID and susceptibility test to rule out Aeromonas hydrophilia, Aeromonas veronii, or Aeromonas caviae (Of the 14 species of Aeromonas, these are the most clinically significant for humans).
Aeromonas veronii:
Info: CDC: Dr. Vila, professor of Microbiology, School of Medicine, University of Barcelona, and Consultant to the Clinical Microbiology Laboratory of the Hospital Clinic, Barcelona
The WHO: http://www.who.int/water_sanitation_health/dwq/en/admicrob2.pdf
Aeromonas veronii:
- Oxidase +
- Glucose-fermenter, Sucrose-fermenter
- Facultative anaerobe
- Decarboxylases Lysine and Ornithine
- Capable of producing beta-lactamases and resistance to ampicillin, mezlocillin and piperacillin
- Gram-negative bacilli, straight rods
- Beta-hemolytic, round, smooth, large, raised, opaque colonies (A. caviae are usually gamma)
- NLF and LF strains
- Primary habitat is fresh water, polluted or chlorinated water, brackish water, occasionally marine water
- Transmission to humans is via ingestion of contaminated water or seafood or by exposure of disrupted skin and mucosal surfaces to contaminated water or soil or via traumatic inoculation by fish fins or fishing hooks
- May transiently colonize the GI tract
- Ingestion of contaminated foods, such as dairy, meat or produce, or ingestion of contaminated water
- Produces various toxins and virulence factors, but the specific role of these is still under research (2 types of pili, flagella, S-layer, alt and last genes, which encode enterotoxins associated with watery diarrhea, attachment mechanisms, lectins, adhesins, haemolysins, aerolysin, pore-forming cytolysin, proteases; influenced by temperature)
- Spectrum of disease includes: Gastroenteritis, wound infections, bacteremia, sepsis, endocarditis, meningitis, pneumonia, conjunctivitis, and osteomyelitis and UTI
- The ability of most commercial ID systems to accurately ID Aeromonas spp from Vibrio spp is limited, and should be confirmed using conventional biochemicals or serotyping (string test)
- Found in mosquitos and leeches (symbiosis)
- May be a connection with outbreaks of diarrhea and both public/municipal drinking-water and from private, unchlorinated wells
- Associated with biofilm development
- Has been a rare cause of traveler’s diarrhea, with leukocyte increase and blood and mucous
- Occurence of 3 or more episodes of watery stools within a 24-hour period, with or without symptoms
- The occurence of unformed stools with vomiting, nausea, abdominal cramps, fever, chills, prostration, tenesmus
- Persistent diarrhea of >14 days duration
- Diarrhea is of 4 types: secretory (acute, water, and with vomiting; very common; dysenteric, acute with blood and mucus, common; chronic diarrhea lasting >10 days, common; or choleraic, or “rice water” stools, rare)
- A. veronii has a positive reaction to arginine dihydrolase and a negative response to bile-esculin hydrolysis and a production of ornithine decarboxylase
- A. caviae has been linked to chronic diarrhea lasting more than 1 year (enteritis) and a quinolone is the drug of choice
- Risk factors include age, immunocompetence, infection dose, underlying illness, and expression of virulence factors by the organism itself
Info: CDC: Dr. Vila, professor of Microbiology, School of Medicine, University of Barcelona, and Consultant to the Clinical Microbiology Laboratory of the Hospital Clinic, Barcelona
The WHO: http://www.who.int/water_sanitation_health/dwq/en/admicrob2.pdf
Plesiomonas spp, as seen in this image, produces shiny, opaque, matte, smooth, nonhemolytic, grayish-white colonies on TSA agar. On MacConkey agar, there are both non-lactose-fermenting and lactose-fermenting strains. Plesiomonas spp is oxidase +, LDC +, Ornithine decarboxylase +. This spp may express resistance to some antimicrobials.
Plesiomonas shigelloides is found in aquatic environments around the world. Infection via ingestion of contaminated water or food products may result in gastroenteritis and even septicemia, particularly in elderly patients who are immunocompromised and infants who have experienced a difficult delivery. If you have an oxidase + organism that looks like this, perform a TULM test (TSI, Urea, LIA and MIO slants), Vitek ID and susceptibility test to rule out Plesiomonas shigelloides.
MacConkey agar with normal lactose-fermenting enteric flora Escherichia coli on the left side, and non-lactose-fermenting colonies on the right side. Further testing must be performed for identification and susceptibility of the non-lactose-fermenting colonies on the right side. There are different strains of E. coli, the most potent being E. coli 0157:H7, an enterohemorrhagic (EHEC) strain of E. coli that produces watery, bloody diarrhea and can be deadly. It is also linked to hemolytic uremic syndrome (HUS), in which the kidneys go into failure and the individual may need dialysis. About 5-6% of infections with E. coli 0157:H7 develop into HUS. Other strains of E. coli with the ability to produce diarrhea include enterotoxogenic, enteroinvasive, enteropathogenic, and enteroaggregative strains.
If you have a NLF on the SMAC plate, perform the TULM test (TSI slant, Urea slant, LIA slant, MIO slant), indole test (+), EHEC test (+), Prolex test (+) and Vitek ID and susceptibility. You may need to send out to state reference laboratory for confirmation. E. coli 0157:H7 is a non-sorbitol-fermenter on SMAC agar. On the TSI, it produces an A/A reaction with gas, but no H2S. This means the entire slant will be yellow, and there will be cracks or bubbles in the agar, but no black precipitate. The urea test is negative for E. coli, which means it will stay yellow and not turn pink/fuschia. On MIO agar, E. coli is positive for motility, positive for indole, and positive for ornithine decarboxylation.
E. coli 0157:H7 can produce a severe cholera-like syndrome, traveler's diarrhea, and can grow in the small bowel and cause diarrhea in children because of its production of enterotoxins.
E. coli 0157:H7 can produce a severe cholera-like syndrome, traveler's diarrhea, and can grow in the small bowel and cause diarrhea in children because of its production of enterotoxins.
If you see this, perform a TULM test (TSI, Urea, LIA, and MIO slants), and perform a Vitek ID and susceptibility test (GNI/GNS). TSI reaction produces an alkaline slant (red) over an acid butt (yellow) with production of H2S gas, blackening of the agar, and production of gas (bubbles/cracks).
Salmonella typhi can be confirmed by blood culture during the first two days of illness.
Salmonella typhi can be confirmed by blood culture during the first two days of illness.
If you see blue-green colonies on Hektoen agar, perform a TULM test (TSI, Urea, LIA, and MIO slants), and perform a Vitek ID and susceptibility test (GNI/GNS). Shigella gives a TSI reaction of alkaline (red) over acid (yellow), no gas production and no H2S. It is also nonmotile.
During biochemical testing (ex: Wellcollex Shigella Kit), if a suspension is tested in polyvalent antiserum but does not result in agglutination, you can boil the serum for 15-20 minutes and then retest. If positive, agglutination will occur in polyvalent and group D antisera. This indicates that the organism possesses capsular antigens.
During biochemical testing (ex: Wellcollex Shigella Kit), if a suspension is tested in polyvalent antiserum but does not result in agglutination, you can boil the serum for 15-20 minutes and then retest. If positive, agglutination will occur in polyvalent and group D antisera. This indicates that the organism possesses capsular antigens.
Orange or yellow colonies or yellow with olive-green centered colonies are normal and OK on Hektoen agar. No further testing is required. This is normal gut flora fermenting the sugars in the medium.
Sorbitol-fermenting Escherichia coli on the SMAC plate, indicative of normal fecal flora is seen in this image. Non-sorbitol-fermenters on the SMAC plate should be further tested for the presence of enterohemorrhagic E. coli via biochemical testing, such as the ImmunoStat EHEC test, which tests for E. coli O157:H7, and other strains that produce shiga-like toxins, and tests such as the Prolex Latex Agglutination test, which test for just the E. coli O157 strain.
If you have growth on the Campylobacter plate that is oxidase + and catalase +, perform a Gram stain. If the Gram stain shows small, curved, "S" or "gull-wing" shaped GNRs, perform a Hippurate test. If the Hippurate test is positive (purple), the ID is C. jejuni, the only Campylobacter spp that is Hippurate +. If it is negative, it is another type of Campylobacter spp. Perform a Vitek GNI/GNS. You may need to send the isolate to the state reference laboratory for confirmation. Campylobacter spp grows well on the Campy BAP agar, as well as Skirrow's medium. Campylobacter spp require a moist, microaerophilic environment at 42 degrees Celsius, in 6% O2, 10-15% CO2, and 85-90% nitrogen for optimal recovery of this organism. It is susceptible to nalidixic acid and resistant to cephalothin. Campylobacter fetus is the only species of Campylobacter that has an optimal growth at 37 degrees Celsius versus 42 degrees Celsius.
Campylobacter coli is:
Campylobacter jejuni is
Campylobacter upsaliensis is
Liquid fecal specimens submitted for culture on babies should be routinely examined for Campylobacter, in addition to Salmonella and Shigella spp.
Liquid preserved samples in Cary-Blair should be stored in the refrigerator to preserve Campy until it is ready to be sent out to a reference lab or public health laboratory.
Campylobacter coli is:
- microaerophilic and more aerotolerant than the other strains, and has increased survival and transmission over other strains
- spread via raw or undercooked contaminated meats and vegetation, including soil and water and unpasteurized milk and infected animals
- incubation time: 24-72 hours
- causes campylobacteriosis: high fever, headache, nausea, cramps, diarrhea that may be bloody (self-limiting; treat with electrolytes and rehydration)
- diarrhea
- the most frequently reported foodborne illness in Europe
- is a slow-grower
- likes 42 degrees C temp.
- becomes spherical or coccoidal in shape if it's exposed to air for too long
- treatment: azithromycin or ciprofloxacin
- resistance: fluoroquinolones and the macrolides
- linked to Guillain-Barre' Syndrome (rare autoimmune reaction resulting in muscle weakness)
- diarrhea
Campylobacter jejuni is
- one of the most common causes of food-poisoning
- this type is associated with consumption of contaminated poultry
- linked to reactive arthritis and Guillain-Barre' Syndrome
- this one has more of the characteristic "seagull" shape than the other strains
- oxidase +
- microaerophilic
- catalase +
- hippurate +
- motility +
Campylobacter upsaliensis is
- typically spread by infected animals, especially dogs, consumption of raw or undercooked contaminated meat, or contaminated water
- catalase negative, unlike other strains that are catalase positive
- thermotolerant and microaerophilic
- oxidase positive, like the other strains
- carried by 66% of cats and 75% of dogs (asymptomatic)
- this is the second most common cause of campylobacteriosis in humans
- sometimes causes septicemia
- linked to Guillain-Barre' Syndroe
Liquid fecal specimens submitted for culture on babies should be routinely examined for Campylobacter, in addition to Salmonella and Shigella spp.
Liquid preserved samples in Cary-Blair should be stored in the refrigerator to preserve Campy until it is ready to be sent out to a reference lab or public health laboratory.
Heavy predominate or pure isolates of S. aureus are worked up because this may indicate staphylococcal food poisoning. Blood cultures may also come up positive if this is the case.
Heavy predominate or pure isolates of Pseudomonas aeruginosa represent a colonization that should be reported to the physician to be made aware of should the patient become ill or have surgery for preventative care.
tsi reactions (triple sugar iron agar):
Another TSI Reaction:
TSI (Triple sugar iron) agar slant results. From left: uninoculated (as control) P. aeruginosa: Glc (-), Lac/Suc (-), H2S (-), Gas (-) E. coli:Glc (+), Lac/Suc (+), H2S (-), Gas (+) Salmonella Typhimurium:Glc (+), Lac/Suc (-), H2S (+), Gas (+) Shigella flexneri:Glc (+), Lac/Suc (-), H2S (-), Gas (-) Abbreviations and Interpretation of results Glc:Glucose fermantation, (+) butter colored yellow, (-) red. Lac/Suc:Lactose and/or Sucrose fermentation, (+) slant colored yellow, (-) red. H2S:production of H2S, (+) a black precipitate formed in slant, (-) not formed. Gas:production of Gas(CO2, H2S, etc), (+) agar may be lifted off the bottom or break apart, (-) not formed.
urea agar slant results:
lysine iron agar (lia) slant results:
motility indole ornithine (mio) agar results:
gn broth: inoculate with 1 mL of runny stool, 2 ml of stool preserved in carey-blair medium, or 1-2 grams of formed stool
Clinical Significance of Stool Pathogens:
1) Aeromonas hydrophila-This is an oxidase (+), catalase (+), motility (+), indole (+), sucrose (+), mannitol (+), lactose (-), facultative anaerobe, Gram-negative, straight bacilli that is found in aquatic environments, such as in brackish water, fresh or seawater, polluted water, chlorinated water, sink traps, tap water faucets, drain pipes distilled water supplies and causes watery diarrhea (gastroenteritis). It is transmitted via the fecal-oral route, usually through contaminated seafood or contaminated dairy products, meats or produce. Individuals may be transiently colonized with it (gastrointestinal tract). Additionally, individuals may become infected by accidental exposure to contaminated fishing hooks or fish fins. On blood agar, it tends to be Beta-hemolytic and produces large, tan-yellow, mucoid colonies. It can also cause wound infections and bacteremia. It can be isolated on TSA, CIN agar, Enteric agars like MAC. NLF and/or LF on MAC. This is always a pathogen and is NEVER part of the normal flora in humans.
2) Plesiomonas spp-This species is also oxidase (+), catalase (+), motility (+), indole (+), sucrose (-), mannitol (-), lactose (+), and is also transmitted via ingestion of contaminated water, food or seafood. It is found in fresh surface waters and soil, particularly in warmer climates. This straight, rounded Gram-negative bacilli found in pairs, short chains, singular, long or short, filamentous and motile, also causes mild gastroenteritis with mild, watery diarrhea, can cause Traveler's Diarrhea, and rarely, septicemia. Asplenic patients are more at risk for infection, as well as those with domestic amphibians and reptiles as pets, because it can be transmitted via that route as well. It can be recovered on TSA and most enteric agars. It is non-hemolytic on TSA. Colonies average 1.5 mm in diameter and are shiny and gray, smooth, opaque, and may be slightly raised in the center. NLF and/or LF or MAC. This is always a pathogen and is never part of the normal flora in humans.
3) Salmonella spp-This Gram-negative bacilli with flagella causes mild to severe gastroenteritis with watery, bloody diarrhea. Salmonella is motile, it produces gas and H2S, and is glucose (+). On MacConkey and MacConkey with Sorbitol agars, it is a non-lactose fermenter (NLF). On Hektoen agar, it produces colonies with black centers due to its production of H2S gas. It is always a pathogen and is NEVER part of the normal fecal flora in humans. Salmonella is especially virulent due to its production of Vi antigen, a polysaccharide capsule that surrounds O antigen, protecting the organism from antibody attack and from phagocytosis.
A. Groups:
1) S. typhi
2) S. choleraesuis
3) S. enteritidis
Salmonella lives in the gastrointestinal tracts of animals. Humans become hosts when they are infected via the fecal-oral route during contamination of food or water with animal feces. Domestic pets such as turtles, birds, hamsters, gerbils and others can transmit the bacterium to humans. Eating undercooked, contaminated beef, chicken, and eggs can also cause illness.
B. Four Disease States:
1) Typhoid Fever (Enteric Fever)
-S. typhi invades epithelial cells and lymph nodes and becomes a facultative intracellular parasite
-Treatment: Ciprofloxacin or Cefriaxone
2) Carrier State (Asymptomatic)
-Chronic
-Carrier has a colonization of the organism in the gallbladder following recovery from Typhoid Fever and excretes the bacteria constantly
-Example: Typhoid Mary (1868)
-Treatment: removal of gallbladder
3) Sepsis
-Bloodstream invasion
-Invasion of lungs, brain, bone
-At risk: asplenic patients and sickle cell patients
-Treatment: long-term antibiotic therapy
4) Gastroenteritis (Diarrhea)
-Self-limiting (usually within 1 week)
-Hundreds of serotypes
-Nausea, abdominal pain, watery diarrhea with blood and mucous
-50% have a fever
-Release of a cholera-like toxin causes ileal inflammation and production of mucous
-Treatment: fluid and electrolyte replacement
4) Shigella spp-This Gram-negative, nonmotile bacillus causes dysentery, with acute abdominal pain and bloody diarrhea, with infection occurring with an infectious dose of as little as just 10 organisms. It, too, is spread via the fecal-oral route, usually through the ingestion of contaminated food. On MacConkey and MacConkey with Sorbitol agars, it is a non-lactose fermenter (NLF). On Hektoen agar, the colonies are green-to-blue. The organism is non-motile, does not produce gas or H2S. It is always a pathogen, and is NEVER part of the normal fecal flora in humans.
A. Four Species:
1) S. sonnei
2). S. flexneri
3) S. boydii
4) S. dysenteriae
Preschool-age children and infants and toddlers at daycare are at higher risk for infection with this organism, as are elderly persons in nursing homes and care centers. Shigella invades the intestinal and colon epithelial cells and the microvilli. It causes the following:
-the release of Shiga toxin, which destroys cells
-inflammation, due to the arrival of white blood cells
-fever
-ulcers in the colon
-sloughing off of cells
-pus and mucous in the stool
-flecks of blood in the stool
-diarrhea
-cramping and abdominal pain
-possible dehydration, as the colon becomes unable to reabsorb fluids and electrolytes
-the 60S ribosome is inactivated, protein synthesis is halted, so epithelial cells die and slough off
5) Campylobacter jejuni and other Campylobacter spp. This is a curved, Gram-negative rod with a single flagellum therefore, it is motile. It resembles "seagull wings" or "S" shapes because it is spiral-shaped. It is microaerophilic, growing best in an environment of a little amount of oxygen, 42 degrees Celsius, a moist environment, and CO2. It is hippurate hydrolysis (+), Catalase (+), Oxidase (+) (the only strain of Campylobacter that is oxidase +), nonfermentive, TSI H2S negative, and colonies grow as gray-to-pinkish, flat, mucoid, convex, spreading on the Campy CVA plate. It is sensitive to Cephalothin and resistant to Nalidixic Acid. The plates are read at 24 and 48 hours.
This organism causes gastroenteritis with diarrhea. There are >2 million cases per year in the USA alone. It is spread through contaminated water or milk and children are most commonly affected. C. jejuni is the most common species of Campylobacter implicated in bacterial foodborne disease in humans. Symptoms of Campylobacteriosis include:
-fever
-headache
-abdominal cramps and pain
-myalgia (muscle pain)
-bloody, loose, inflammatory diarrhea (dysentery)
-invasion of the small intestinal lining with spread of the organism
-release of LT toxin and cytotoxin that destroy the mucosal cells (cytolethal distending toxin)
-blocks cell division
-evades the immune system
-spread via the fecal-oral route by the ingestion of contaminated food or water, particularly raw meat (mostly chicken/poultry)
-self-limiting illness that lasts about 5-7 days
-affects the jejunum, the ileum, and the colon
-complications include an underlying cause of Guillain-Barre' Syndrome or perforation of the gastrointestinal contents in ileal infections, toxic megacolon, or dehydration
-occasionally associated with periodontitis
-contagious
Anywhere from 20-75% of raw chicken and raw ground chicken tested have been contaminated with Campylobacter spp, therefore, it is very important to wash your hands thoroughly after touching raw meat before touching your mouth or before eating. Some of the packaging tested has also been found to be contaminated, so throw it away right away and then wash/clean your surroundings and surfaces immediately. If you have cut chicken on a cutting board, it is also important not to reuse it to cut up other things, such as vegetables, because you may contaminate them with Campylobacter.
Other Campylobacter spp (about 12 species are associated with human disease):
6) Vibrio cholerae, V. hemolyticus, V. parahemolyticus, V. vulnificus and other Vibrio spp-Vibrio are motile, curved Gram-negative rods. Vibrio cholerae causes severe "rice water" (watery) diarrhea due to the production of the virulence factor toxin choleragen. Diarrhea may result in the loss of up to 1 liter of fluid per hour! Dehydration, shock, decreased pulse, sunken eyes, poor skin turgor, and even death are associated with this illness. Travelers to endemic areas are at risk for infection. V. hemolyticus, V. parahemolyticus, Vibrio vulnificus and others are associated with eating undercooked, contaminated seafood and shellfish or by accidentally ingesting contaminated water (brackish or salt). G-protein is inactivated, cAMP is produced, resulting in secretion of Na+ and Cl-, but they are not reabsorbed, leading to fluid, bicarbonate and potassium loss and an increase in intestinal motility. Vibrio are non-lactose-fermenting and grow as transparent clear-green colonies on TCBS agar where laboratories use this agar. The organism is transmitted through contaminated water. Treatment consists of fluid and electrolyte replacement, along with doxycycline to lessen the severity of the symptoms.
7) Yersinia enterocolitica-This small, motile, Gram-negative bacilli grows on Yersinia CIN agar at room temperature as "bull's eye" colonies with a distinct red center. This causes acute gastroenteritis. It is transmitted via the fecal-oral route through contaminated water or unpasteurized milk. Symptoms include:
-fever
-diarrhea
-abdominal pain and tenderness, particularly in the right lower quadrant
-mucosal ulceration of the ileum
-increased WBC count (leukocytosis) with increased segs (neutrophilia)
The organism produces virulence factors that enable it to invade the intestinal epithelial cells and the lymph nodes, leading to bloodstream infection and lymphadenopathy. Sepsis is also possible. Because of the enterotoxins it produces, it can survive and grow in the cold and in adverse conditions.
8) E. coli serotypes and E. coli 0157:H7-This non-lactose-fermenting (NLF), Gram-negative rod with peritrichous flagella (all over) is a very motile bacterium. This particular strain of E. coli is extremely virulent. Not only is it responsible for the very uncomfortable bloody Traveler's Diarrhea, but it can also lead to Hemolytic Uremic Syndrome (HUS), which can be deadly or which can result in kidney damage requiring permanent and lifelong dialysis. It is referred to as Enterohemorrhagic E. coli. Virulence factors include:
-pili, which enable it to hook into and "stick" to the intestinal mucosa
-shiga-like toxin (verotoxins)
1) STX1: Lowest risk of developing Hemolytic Uremic Syndrome
2) STX2: Highest risk of developing Hemolytic Uremic Syndrome
Together: Intermediate risk of developing Hemolytic Uremic Syndrome
Symptoms:
-bloody diarrhea
-abdominal cramps
-Hemolytic Uremic Syndrome (HUS)
-Kidney problems/failure
-Strokes in the brain
-Hemolytic anemia due to destruction of red blood cells and clogging up of the veins and arteries
-Edema and swelling
-Fluid around the lungs and heart and increased blood pressure
The STAT EHEC test (Enterohemorrhagic E. coli) test cartridge is performed to test for this (a variety of E. coli serotypes that produce STX1, STX2, or both).
Other Pathogenic Strains of E. coli Include:
A. Enterotoxogenic (ETEC)
-Traveler's Diarrhea
-Heat-labile toxin
-Heat-stable toxin
-Inhibits the reabsorption of Na+ and Cl- and stimulates the secretion of Cl- and HCO3 into the intestinal lumen
-Water and electrolyte loss may be as much as 20 liters a day
-Also causes "rice water" diarrhea
-Associated with outbreaks (contaminated vegetables like lettuce, tomatoes, etc...)
-Can occur along with a C. difficile infection at the same time
-You can perform the shiga toxin test to see which toxin or toxins are present
B. Enteroinvasive (EIEC)
-Fever
-Mucous and and bloody diarrhea
-Dehydration
-This is encoded on a bacterial plasmid
C. Enteropathogenic (EPEC)
-Also called uropathogenic
-One of the main causes of UTIs
-Hook-like pili and adhesives
D. Enteroaggregative (EAEC)
-Pathotype of E. coli associated with diarrhea, particularly in children
-"Stacked Brick" pattern of adhesion to endothelial cells, creating an aggregation or colonization of the bacterium to the mucosa (due to secretion of adhesins and fimbriae)
-Enterotoxins and Cytotoxins
-Damages host cells and results in inflammation
-An emerging enteric pathogen
-Traveler's Diarrhea
-Sporadic food-poisoning outbreaks with deaths
-May acquire a shiga toxin or shiga-like verotoxin
PROLEX:
The Prolex latex agglutination test is a test that is used to test for E. coli 0157 and other strains of E. coli, not just enterohemorrhagic.
http://www.pro-lab.com/products-ecoli.php?country=IE (Go to this link to watch a demonstration of how to perform this test)
EHEC:
It tests for enterotoxogenic (ETEC) E. coli, and tests for other serotypes including 026, 045, 0103, 0111, 0121, 0145, some causing Traveller's diarrhea and some linked also to shiga-toxin production and cases of hemolytic uremic syndrome (HUS), in which red blood cells are destroyed and the kidneys fail (mostly seen in kids <5 years of age and elderly with infections of the serotypes 0157 and 0157:H7.
http://www.meridianbioscience.com/Content/Assets/Files/2.2%20Foodborne%20Products/Procedure-Card-ImmunoCard-STAT-EHEC.pdf
http://www.meridianbioscience.com/Content/Assets/PackInsert/8.5x11_CLEAN_SN11169_REV_05-13_751630_ICS_EHEC_PI_8.14.13.pdf
https://vimeo.com/12298121 (EHEC Product Demonstration) (Go to this link to watch a demonstration of how to perform this test)
9) Coagulase Negative Staphylococcus (CNS) on Neonates
10) Pseudomonas overgrowth-Pseudomonas is an obligate aerobe, Gram-negative bacilli that is NLF on MacConkey agar, is oxidase + and is motile (not all strains will show motility in deep agar slants used for motility, however, even though all are motile). It produces a green fluorescent pigment called fluorescein and a blue pigment called pyocyanin.
A. Overgrowth can seed to other areas
B. Colonizes and infects sick, immunocompromised, hospital patients
C. Many strains are multi-drug-resistant
D. It produces exotoxin A, which stops protein synthesis
E. It produces an antiphagocytic and adhesive capsule, making treatment difficult
F. It can spread and cause pneumonia, osteomyelitis, burn infections, sepsis, UTI, pyelonephritis, endocarditis, outer ear infections, corneal infections, and infections in patients with Cystic Fibrosis (CF).
G. Treatment includes Penicillin and an Aminoglycoside.
11) Candida albicans yeast overgrowth-May occur in patients who are on broad-spectrum antibiotics that have wiped out the normal fecal flora, allowing overgrowth, or may occur in patients on medications such as antacids and anti-reflux to treat other conditions, which changes the pH of the intestines, leading to loss of normal flora, and allowing overgrowth of yeast. This is the only species of Candida that is germ-tube positive and produces terminal chlamydospores on cornmeal agar.
12) Staphylococcus aureus-If this Beta-hemolytic, Gram-positive staphylococci in clusters is found as a pure culture from a stool culture, this is indicative of bacterial food poisoning or gastroenteritis, usually from food that has sat out for too long without proper preservation (mayonnaise, eggs, rice, etc...)
-Releases preformed exotoxin, an enterotoxin that causes the symptoms of food poisoning
-Stimulates the increase of intestinal peristalsis (motility), resulting in an acute illness that comes on rapidly, but also ends quickly
-Associated with rice, mayonnaise, and/or egg dishes, meats that have sat out for >2 hours unrefrigerated
-Symptoms: nausea, vomiting, diarrhea, abdominal pain and cramping, fever, occasional dehydration
-Self-limiting illness that is over in about 12-24 hours
13) Bacillus cereus-food poisoning, blood infections (sepsis, bacteremia)
-Associated with rice or rice dishes that have been sitting out for too long unrefrigerated (>2 hours)
-Catalase +, Large, thick, boxcar-like Gram-positive rods with terminal endospores
-3 toxins: necrotic toxin (tissue death), cereolysin (hemolysis), phospholipase C (lecithinase)
-2 disease states:
-emetic (nausea and vomiting within 1-6 hours of ingestion; self limiting and resolves in <24 hours); Associated with rice/rice dishes)
-diarrheal (abdominal cramps and diarrhea about 9 hours post-ingestion; self-limiting and resolves in 24-36 hours; Associated with meat, veggie, or sauce dishes that have sat out unrefrigerated for >2 hours)
14) Clostridium botulinum-food poisoning
-Nausea, vomiting within 1 hour of ingestion
-Large, thick, boxcar-like Gram-positive rods
15) Clostridium difficile overgrowth:
-Antibiotic-associated pseudomembranous colitis
-Associated with broad spectrum antibiotics (ampicillin, clindamycin, cephalosporin)
-Exotoxins A and B
-Severe diarrhea, cramps, fever, red/inflamed mucosa, exudate, necrosis
-Treat with Vancomycin or Metronidazole
-GPR with spores, big and boxy and may look like "spoons"
Refer to the Bacteriology Page, Media Agar Section for further information about the agars used here.
Normal Fecal Flora/Enterics (No Significance in Stool Culture, Though They May Cause Infections in Other Areas of the Body if In the Wrong Place):
Bacteriodaceae (about 99% of the fecal flora), including B. fragilis, P. melaninogenicus, Fusobacterium, Peptostreptococcus, Peptococcus, Serratia, Enterobacter, Citrobacter, Providencia, Edwardsiella tarda, Proteus mirabilis, Proteus vulgaris, Klebsiella, E. coli, Veillonella spp, Eubacterium, Bifidobacterium
Fecal Flora that Occasionally Cause Infection or Colonization of the Gut if They Overgrow:
Pseudomonas, Clostridium difficile, E. coli, Candida albicans
CNS on Neonates
1) Candida albicans overgrowth-
a)Tells the doctor that the patient's normal fecal flora is decreased or wiped out due to strong antibiotics, antacids, anti-reflux medications, immunocompromisation or diabetes
b)Tells the doctor that the patient may have a disseminated, systemic yeast infection (check the blood cultures)
c)The patient may also have UTI and vaginitis (check other cultures)
d)An infant with Candida overgrowth may also develop a diaper rash
e)Treatment includes Amphotericin B or Fluconazole and replacement of the flora with yogurt, kefir, or supplements
2) Clostridium difficile overgrowth-
a)Antibiotic-associated pseudomembranous colitis associated with the use of broad-spectrum antibiotics (Ampicillin, Clindamycin, Cephalosporin)
b)Produces exotoxins
-Toxin A (diarrhea)
-Toxin B (cytotoxic to the colon)
c)Symptoms: severe diarrhea, cramps, fever, red/inflamed colon mucosa, exudate formation, necrosis
d)Treatment: Metronidazole or Vancomycin by mouth
Helicobacter pylori-
Helicobacter pylori is a Gram-negative, corkscrew-shaped, motile bacterium (spirilla) that has the ability to bore into the duodenal wall, creating ulcerations and is linked to stomach cancer. It is a causative agent of chronic gastritis, gastric and duodenal ulcers. Approximately 80% of individuals infected with this microbe are actually asymptomatic, and it is estimated that around half of the world's population may be colonized with H. pylori in their upper gastrointestinal tracts. It produces oxidase, catalase and urease.
Symptoms:
Those who are infected that do develop symptoms may experience the following:
-acute gastritis
-abdominal pain
-nausea
-dyspepsia
-bloating
-belching
-vomiting
-black stool (due to blood)
-10-20% lifetime chance of developing a peptic ulcer
-1-2% lifetime chance of developing stomach cancer
-chronic inflammation of areas of the stomach
-chance of developing colorectal polyps and colorectal cancer
The urease test and clotest (a modified urease test), as well as a rapid identification test are good tools to aid in the identification and treatment for infections with this organism.
MEDIA: TULMS
Triple Sugar Iron Agar (TSI):
*Stab straight down and streak the surface after touching the top of a well-isolated colony with a straight inoculation needle
*Keep the cap on loosely
*Incubate in ambient air at 35 degrees Celsius for 18-24 hours
1) Alkaline Slant/No Change In the Butt (K/NC, or K/K)=glucose, lactose, and sucrose non-utilizer
2) Alkaline Slant/Acid Butt (K/A)=glucose fermentation only ("yellow"=acid production due to glucose fermentation)
3) Acid Slant/Acid Butt (A/A)=glucose, sucrose and/or lactose fermenter
4) Black precipitate in the butt indicates the production of ferrous sulfide and H2S gas
5) Bubbles or cracks in the agar indicate gas production (CO2 or H2)
-Shigella (K/A, no gas, no H2S production)
-Salmonella (K/A, H2S production, no gas production)
-E. coli, Klebsiella spp, Enterobacter spp (A/A, gas production, no H2S production)
-Pseudomonas aeruginosa (K/K, no gas, no H2S)
Urea Hydrolysis:
*Streak surface of agar
*Loosen cap
*Incubate in ambient air at 35 degrees Celsius for 18-24 hours (sometimes it takes 48 hours to 7 days)
1)Positive: Color of slant changes from light yellow-orange to bright fuschia/magenta (Proteus vulgaris, P. mirabilis, Enterobacter cloacae, Klebsiella, Brucella, Bordetella, Yersinia enterocolitica, Morganella spp. H. pylori)
2)Negative: No color change (agar remains light yellow-orange) (E. coli, Salmonella, Shigella, Enterobacter aerogenes)
This test is used to see if an organism has the ability to produce urease, an enzyme that hydrolyzes urea, producing ammonia and CO2, which alkalizes the medium and shifts the pH from 6.8 to 8.1. The pH change is detected by a color indicator (phenol red).
Lysine Iron Agar (LIA):
*Stab straight down and streak the surface after touching the top of a well-isolated colony with a straight inoculation needle
*Keep the cap on loosely
*Incubate in ambient air at 35 degrees Celsius for 18-24 hours
1) LDC + (yellow slant/purple butt) (Salmonella)
2) LDC - (yellow slant/yellow butt)
3) H2S (yellow slant/black butt)
4) LDA + (yellow slant/burgandy butt) (Proteus)
5) LDA- (yellow slant/purple butt) (Salmonella, Shigella)
Motility Agar:
1) Motility + if there is movement away from/spreading out from the stab in the agar
2) Indole + if liquid on top turns pink-red after 1-2 drops of indole are added and mixed well
3) Ornithine + if yellow, - if purple
Motile:
-Yersinia at 22 degrees Celsius
-E.coli
-Listeria at 22 degrees Celsius
Non-motile:
-Shigella
-Klebsiella
-Yersinia at 35 degrees Celcius
Stool For Reducing Substances (Clinitest on Stool):
In this test, Benedict's solution (a blue, alkaline solution) is used to test for the presence of a reducing sugar in the stool. This includes monosaccharides and disaccharides, such as lactose and maltose. It also detects aldehydes and ketoses, such as fructose. It also tests for glucose and galactose, which may indicate diabetes (former) or hereditary galactosemia (latter). When reducing sugars are heated in the presence of an alkali over the Bunsen Burner, they convert to powerful reducing compounds (enediols). Reducing sugars reduce copper II ions (cupric) in the solution to copper I ions (cuprous), which form a copper oxide precipitate. This test is semi-quantitative. "A greenish precipitate indicates about 0.5 g% concentration (trace); yellow precipitate indicates 1 g% concentration; orange indicates 1.5 g% and red indicates 2 g% or higher concentration." ( http://en.wikipedia.org/wiki/Benedict%27s_reagent). Any amount from trace up to high concentration indicates a positive test.
A false positive reaction may occur if other reducing substances are present, such as ascorbic acid (vitamin C supplement). Contrasts used in radiological studies, Levodopa drugs administered to treat Parkinson's disease, and homogentisic acid may also cause false positive results.
Fecal Occult Blood Test:
This test is used to detect "hidden" blood in the stool that may not be visible to the naked eye. The presence of occult blood is often the first sign of colon or rectal cancer, and early detection is key for treatment and prolonged life. The rapid guaiac-based hemoccult stool blood tests detect heme, a component of blood, in the stool. It detects blood loss anywhere along the gastrointestinal tract, from the mouth to the colon. It can be helpful in detecting and diagnosing upper or lower GI bleeds, peptic ulcers, anemia, and even malignancies. It is a screening tool for these purposes, along with other tests.
Upper or lower GI bleeds may be due to gastric or colorectal cancers, polyps, adenoma, peptic ulcer, angiodysplasia of the colon, or sickle cell anemia. If positive, this test is followed up with a sigmoidoscopy and/or colonoscopy or endoscopy, bloodwork, and an M2-PK test.
Fecal Leukocytes Test (WBCs):
White blood cells, or leukocytes, are not normally seen in the stool unless inflammation or infection is present or suspected, therefore, this test is a useful screening tool for the detection of bacterial or parasite infections, or inflammatory conditions caused by autoimmune diseases or viruses. Salmonella, Shigella, Campylobacter, Amoebiasis, ulcerative colitis, Crohn's Disease are all conditions associated with the presence of fecal leukocytes (lactoferrin).
Stool pH:
A fecal pH test is a screening tool to indicate the level of pH of the stool. Because the stool is normally alkaline, an acidic stool may indicate a virus (ex: Rotavirus) infection, a bacterial infection (such as E. coli), or lactose intolerance. This is indicated by a stool pH of less than 5.5.
Fecal Elastase:
This test is used to detect and diagnose pancreatitis.
Stool Fats: Fecal Fat Test
This test is used to detect and diagnose the malabsorption of fats, or steatorrhea. Fatty stools may float, be a light color, have a greasy/oily appearance, and may be very foul-smelling. Causes of steatorrhea may be caused by a lack of bile salts (ex: gallbladder removal), liver damage, drugs (hypolipidemic drugs or medicines that block lipid absorption or treat acromegaly), defective pancreatic enzymes or mucosal cells, or if there is too much undigestible oil or fat in the diet that spills over into the stool.
This test is a screening tool that may aid the physician in detecting and diagnosing the following conditions:
1) Aeromonas hydrophila-This is an oxidase (+), catalase (+), motility (+), indole (+), sucrose (+), mannitol (+), lactose (-), facultative anaerobe, Gram-negative, straight bacilli that is found in aquatic environments, such as in brackish water, fresh or seawater, polluted water, chlorinated water, sink traps, tap water faucets, drain pipes distilled water supplies and causes watery diarrhea (gastroenteritis). It is transmitted via the fecal-oral route, usually through contaminated seafood or contaminated dairy products, meats or produce. Individuals may be transiently colonized with it (gastrointestinal tract). Additionally, individuals may become infected by accidental exposure to contaminated fishing hooks or fish fins. On blood agar, it tends to be Beta-hemolytic and produces large, tan-yellow, mucoid colonies. It can also cause wound infections and bacteremia. It can be isolated on TSA, CIN agar, Enteric agars like MAC. NLF and/or LF on MAC. This is always a pathogen and is NEVER part of the normal flora in humans.
2) Plesiomonas spp-This species is also oxidase (+), catalase (+), motility (+), indole (+), sucrose (-), mannitol (-), lactose (+), and is also transmitted via ingestion of contaminated water, food or seafood. It is found in fresh surface waters and soil, particularly in warmer climates. This straight, rounded Gram-negative bacilli found in pairs, short chains, singular, long or short, filamentous and motile, also causes mild gastroenteritis with mild, watery diarrhea, can cause Traveler's Diarrhea, and rarely, septicemia. Asplenic patients are more at risk for infection, as well as those with domestic amphibians and reptiles as pets, because it can be transmitted via that route as well. It can be recovered on TSA and most enteric agars. It is non-hemolytic on TSA. Colonies average 1.5 mm in diameter and are shiny and gray, smooth, opaque, and may be slightly raised in the center. NLF and/or LF or MAC. This is always a pathogen and is never part of the normal flora in humans.
3) Salmonella spp-This Gram-negative bacilli with flagella causes mild to severe gastroenteritis with watery, bloody diarrhea. Salmonella is motile, it produces gas and H2S, and is glucose (+). On MacConkey and MacConkey with Sorbitol agars, it is a non-lactose fermenter (NLF). On Hektoen agar, it produces colonies with black centers due to its production of H2S gas. It is always a pathogen and is NEVER part of the normal fecal flora in humans. Salmonella is especially virulent due to its production of Vi antigen, a polysaccharide capsule that surrounds O antigen, protecting the organism from antibody attack and from phagocytosis.
A. Groups:
1) S. typhi
2) S. choleraesuis
3) S. enteritidis
Salmonella lives in the gastrointestinal tracts of animals. Humans become hosts when they are infected via the fecal-oral route during contamination of food or water with animal feces. Domestic pets such as turtles, birds, hamsters, gerbils and others can transmit the bacterium to humans. Eating undercooked, contaminated beef, chicken, and eggs can also cause illness.
B. Four Disease States:
1) Typhoid Fever (Enteric Fever)
-S. typhi invades epithelial cells and lymph nodes and becomes a facultative intracellular parasite
-Treatment: Ciprofloxacin or Cefriaxone
2) Carrier State (Asymptomatic)
-Chronic
-Carrier has a colonization of the organism in the gallbladder following recovery from Typhoid Fever and excretes the bacteria constantly
-Example: Typhoid Mary (1868)
-Treatment: removal of gallbladder
3) Sepsis
-Bloodstream invasion
-Invasion of lungs, brain, bone
-At risk: asplenic patients and sickle cell patients
-Treatment: long-term antibiotic therapy
4) Gastroenteritis (Diarrhea)
-Self-limiting (usually within 1 week)
-Hundreds of serotypes
-Nausea, abdominal pain, watery diarrhea with blood and mucous
-50% have a fever
-Release of a cholera-like toxin causes ileal inflammation and production of mucous
-Treatment: fluid and electrolyte replacement
4) Shigella spp-This Gram-negative, nonmotile bacillus causes dysentery, with acute abdominal pain and bloody diarrhea, with infection occurring with an infectious dose of as little as just 10 organisms. It, too, is spread via the fecal-oral route, usually through the ingestion of contaminated food. On MacConkey and MacConkey with Sorbitol agars, it is a non-lactose fermenter (NLF). On Hektoen agar, the colonies are green-to-blue. The organism is non-motile, does not produce gas or H2S. It is always a pathogen, and is NEVER part of the normal fecal flora in humans.
A. Four Species:
1) S. sonnei
2). S. flexneri
3) S. boydii
4) S. dysenteriae
Preschool-age children and infants and toddlers at daycare are at higher risk for infection with this organism, as are elderly persons in nursing homes and care centers. Shigella invades the intestinal and colon epithelial cells and the microvilli. It causes the following:
-the release of Shiga toxin, which destroys cells
-inflammation, due to the arrival of white blood cells
-fever
-ulcers in the colon
-sloughing off of cells
-pus and mucous in the stool
-flecks of blood in the stool
-diarrhea
-cramping and abdominal pain
-possible dehydration, as the colon becomes unable to reabsorb fluids and electrolytes
-the 60S ribosome is inactivated, protein synthesis is halted, so epithelial cells die and slough off
5) Campylobacter jejuni and other Campylobacter spp. This is a curved, Gram-negative rod with a single flagellum therefore, it is motile. It resembles "seagull wings" or "S" shapes because it is spiral-shaped. It is microaerophilic, growing best in an environment of a little amount of oxygen, 42 degrees Celsius, a moist environment, and CO2. It is hippurate hydrolysis (+), Catalase (+), Oxidase (+) (the only strain of Campylobacter that is oxidase +), nonfermentive, TSI H2S negative, and colonies grow as gray-to-pinkish, flat, mucoid, convex, spreading on the Campy CVA plate. It is sensitive to Cephalothin and resistant to Nalidixic Acid. The plates are read at 24 and 48 hours.
This organism causes gastroenteritis with diarrhea. There are >2 million cases per year in the USA alone. It is spread through contaminated water or milk and children are most commonly affected. C. jejuni is the most common species of Campylobacter implicated in bacterial foodborne disease in humans. Symptoms of Campylobacteriosis include:
-fever
-headache
-abdominal cramps and pain
-myalgia (muscle pain)
-bloody, loose, inflammatory diarrhea (dysentery)
-invasion of the small intestinal lining with spread of the organism
-release of LT toxin and cytotoxin that destroy the mucosal cells (cytolethal distending toxin)
-blocks cell division
-evades the immune system
-spread via the fecal-oral route by the ingestion of contaminated food or water, particularly raw meat (mostly chicken/poultry)
-self-limiting illness that lasts about 5-7 days
-affects the jejunum, the ileum, and the colon
-complications include an underlying cause of Guillain-Barre' Syndrome or perforation of the gastrointestinal contents in ileal infections, toxic megacolon, or dehydration
-occasionally associated with periodontitis
-contagious
Anywhere from 20-75% of raw chicken and raw ground chicken tested have been contaminated with Campylobacter spp, therefore, it is very important to wash your hands thoroughly after touching raw meat before touching your mouth or before eating. Some of the packaging tested has also been found to be contaminated, so throw it away right away and then wash/clean your surroundings and surfaces immediately. If you have cut chicken on a cutting board, it is also important not to reuse it to cut up other things, such as vegetables, because you may contaminate them with Campylobacter.
Other Campylobacter spp (about 12 species are associated with human disease):
- C. coli (2nd most common after C. jejuni)-Causes enteritis, abdominal pain, diarrhea, bloody stool, fever
- C. fetus (opportunistic pathogen in humans)-Causes bacteremia and thrombophlebitis, and only species of Campylobacter that can cause septicemia, especially in the immunocompromised and in newborns; Can cause meningitis, pleurisy, respiratory infections, joint infections, and pericarditis
- C. upsaliensis
6) Vibrio cholerae, V. hemolyticus, V. parahemolyticus, V. vulnificus and other Vibrio spp-Vibrio are motile, curved Gram-negative rods. Vibrio cholerae causes severe "rice water" (watery) diarrhea due to the production of the virulence factor toxin choleragen. Diarrhea may result in the loss of up to 1 liter of fluid per hour! Dehydration, shock, decreased pulse, sunken eyes, poor skin turgor, and even death are associated with this illness. Travelers to endemic areas are at risk for infection. V. hemolyticus, V. parahemolyticus, Vibrio vulnificus and others are associated with eating undercooked, contaminated seafood and shellfish or by accidentally ingesting contaminated water (brackish or salt). G-protein is inactivated, cAMP is produced, resulting in secretion of Na+ and Cl-, but they are not reabsorbed, leading to fluid, bicarbonate and potassium loss and an increase in intestinal motility. Vibrio are non-lactose-fermenting and grow as transparent clear-green colonies on TCBS agar where laboratories use this agar. The organism is transmitted through contaminated water. Treatment consists of fluid and electrolyte replacement, along with doxycycline to lessen the severity of the symptoms.
7) Yersinia enterocolitica-This small, motile, Gram-negative bacilli grows on Yersinia CIN agar at room temperature as "bull's eye" colonies with a distinct red center. This causes acute gastroenteritis. It is transmitted via the fecal-oral route through contaminated water or unpasteurized milk. Symptoms include:
-fever
-diarrhea
-abdominal pain and tenderness, particularly in the right lower quadrant
-mucosal ulceration of the ileum
-increased WBC count (leukocytosis) with increased segs (neutrophilia)
The organism produces virulence factors that enable it to invade the intestinal epithelial cells and the lymph nodes, leading to bloodstream infection and lymphadenopathy. Sepsis is also possible. Because of the enterotoxins it produces, it can survive and grow in the cold and in adverse conditions.
8) E. coli serotypes and E. coli 0157:H7-This non-lactose-fermenting (NLF), Gram-negative rod with peritrichous flagella (all over) is a very motile bacterium. This particular strain of E. coli is extremely virulent. Not only is it responsible for the very uncomfortable bloody Traveler's Diarrhea, but it can also lead to Hemolytic Uremic Syndrome (HUS), which can be deadly or which can result in kidney damage requiring permanent and lifelong dialysis. It is referred to as Enterohemorrhagic E. coli. Virulence factors include:
-pili, which enable it to hook into and "stick" to the intestinal mucosa
-shiga-like toxin (verotoxins)
1) STX1: Lowest risk of developing Hemolytic Uremic Syndrome
2) STX2: Highest risk of developing Hemolytic Uremic Syndrome
Together: Intermediate risk of developing Hemolytic Uremic Syndrome
Symptoms:
-bloody diarrhea
-abdominal cramps
-Hemolytic Uremic Syndrome (HUS)
-Kidney problems/failure
-Strokes in the brain
-Hemolytic anemia due to destruction of red blood cells and clogging up of the veins and arteries
-Edema and swelling
-Fluid around the lungs and heart and increased blood pressure
The STAT EHEC test (Enterohemorrhagic E. coli) test cartridge is performed to test for this (a variety of E. coli serotypes that produce STX1, STX2, or both).
Other Pathogenic Strains of E. coli Include:
A. Enterotoxogenic (ETEC)
-Traveler's Diarrhea
-Heat-labile toxin
-Heat-stable toxin
-Inhibits the reabsorption of Na+ and Cl- and stimulates the secretion of Cl- and HCO3 into the intestinal lumen
-Water and electrolyte loss may be as much as 20 liters a day
-Also causes "rice water" diarrhea
-Associated with outbreaks (contaminated vegetables like lettuce, tomatoes, etc...)
-Can occur along with a C. difficile infection at the same time
-You can perform the shiga toxin test to see which toxin or toxins are present
B. Enteroinvasive (EIEC)
-Fever
-Mucous and and bloody diarrhea
-Dehydration
-This is encoded on a bacterial plasmid
C. Enteropathogenic (EPEC)
-Also called uropathogenic
-One of the main causes of UTIs
-Hook-like pili and adhesives
D. Enteroaggregative (EAEC)
-Pathotype of E. coli associated with diarrhea, particularly in children
-"Stacked Brick" pattern of adhesion to endothelial cells, creating an aggregation or colonization of the bacterium to the mucosa (due to secretion of adhesins and fimbriae)
-Enterotoxins and Cytotoxins
-Damages host cells and results in inflammation
-An emerging enteric pathogen
-Traveler's Diarrhea
-Sporadic food-poisoning outbreaks with deaths
-May acquire a shiga toxin or shiga-like verotoxin
PROLEX:
The Prolex latex agglutination test is a test that is used to test for E. coli 0157 and other strains of E. coli, not just enterohemorrhagic.
http://www.pro-lab.com/products-ecoli.php?country=IE (Go to this link to watch a demonstration of how to perform this test)
EHEC:
It tests for enterotoxogenic (ETEC) E. coli, and tests for other serotypes including 026, 045, 0103, 0111, 0121, 0145, some causing Traveller's diarrhea and some linked also to shiga-toxin production and cases of hemolytic uremic syndrome (HUS), in which red blood cells are destroyed and the kidneys fail (mostly seen in kids <5 years of age and elderly with infections of the serotypes 0157 and 0157:H7.
http://www.meridianbioscience.com/Content/Assets/Files/2.2%20Foodborne%20Products/Procedure-Card-ImmunoCard-STAT-EHEC.pdf
http://www.meridianbioscience.com/Content/Assets/PackInsert/8.5x11_CLEAN_SN11169_REV_05-13_751630_ICS_EHEC_PI_8.14.13.pdf
https://vimeo.com/12298121 (EHEC Product Demonstration) (Go to this link to watch a demonstration of how to perform this test)
9) Coagulase Negative Staphylococcus (CNS) on Neonates
10) Pseudomonas overgrowth-Pseudomonas is an obligate aerobe, Gram-negative bacilli that is NLF on MacConkey agar, is oxidase + and is motile (not all strains will show motility in deep agar slants used for motility, however, even though all are motile). It produces a green fluorescent pigment called fluorescein and a blue pigment called pyocyanin.
A. Overgrowth can seed to other areas
B. Colonizes and infects sick, immunocompromised, hospital patients
C. Many strains are multi-drug-resistant
D. It produces exotoxin A, which stops protein synthesis
E. It produces an antiphagocytic and adhesive capsule, making treatment difficult
F. It can spread and cause pneumonia, osteomyelitis, burn infections, sepsis, UTI, pyelonephritis, endocarditis, outer ear infections, corneal infections, and infections in patients with Cystic Fibrosis (CF).
G. Treatment includes Penicillin and an Aminoglycoside.
11) Candida albicans yeast overgrowth-May occur in patients who are on broad-spectrum antibiotics that have wiped out the normal fecal flora, allowing overgrowth, or may occur in patients on medications such as antacids and anti-reflux to treat other conditions, which changes the pH of the intestines, leading to loss of normal flora, and allowing overgrowth of yeast. This is the only species of Candida that is germ-tube positive and produces terminal chlamydospores on cornmeal agar.
12) Staphylococcus aureus-If this Beta-hemolytic, Gram-positive staphylococci in clusters is found as a pure culture from a stool culture, this is indicative of bacterial food poisoning or gastroenteritis, usually from food that has sat out for too long without proper preservation (mayonnaise, eggs, rice, etc...)
-Releases preformed exotoxin, an enterotoxin that causes the symptoms of food poisoning
-Stimulates the increase of intestinal peristalsis (motility), resulting in an acute illness that comes on rapidly, but also ends quickly
-Associated with rice, mayonnaise, and/or egg dishes, meats that have sat out for >2 hours unrefrigerated
-Symptoms: nausea, vomiting, diarrhea, abdominal pain and cramping, fever, occasional dehydration
-Self-limiting illness that is over in about 12-24 hours
13) Bacillus cereus-food poisoning, blood infections (sepsis, bacteremia)
-Associated with rice or rice dishes that have been sitting out for too long unrefrigerated (>2 hours)
-Catalase +, Large, thick, boxcar-like Gram-positive rods with terminal endospores
-3 toxins: necrotic toxin (tissue death), cereolysin (hemolysis), phospholipase C (lecithinase)
-2 disease states:
-emetic (nausea and vomiting within 1-6 hours of ingestion; self limiting and resolves in <24 hours); Associated with rice/rice dishes)
-diarrheal (abdominal cramps and diarrhea about 9 hours post-ingestion; self-limiting and resolves in 24-36 hours; Associated with meat, veggie, or sauce dishes that have sat out unrefrigerated for >2 hours)
14) Clostridium botulinum-food poisoning
-Nausea, vomiting within 1 hour of ingestion
-Large, thick, boxcar-like Gram-positive rods
15) Clostridium difficile overgrowth:
-Antibiotic-associated pseudomembranous colitis
-Associated with broad spectrum antibiotics (ampicillin, clindamycin, cephalosporin)
-Exotoxins A and B
-Severe diarrhea, cramps, fever, red/inflamed mucosa, exudate, necrosis
-Treat with Vancomycin or Metronidazole
-GPR with spores, big and boxy and may look like "spoons"
Refer to the Bacteriology Page, Media Agar Section for further information about the agars used here.
Normal Fecal Flora/Enterics (No Significance in Stool Culture, Though They May Cause Infections in Other Areas of the Body if In the Wrong Place):
Bacteriodaceae (about 99% of the fecal flora), including B. fragilis, P. melaninogenicus, Fusobacterium, Peptostreptococcus, Peptococcus, Serratia, Enterobacter, Citrobacter, Providencia, Edwardsiella tarda, Proteus mirabilis, Proteus vulgaris, Klebsiella, E. coli, Veillonella spp, Eubacterium, Bifidobacterium
Fecal Flora that Occasionally Cause Infection or Colonization of the Gut if They Overgrow:
Pseudomonas, Clostridium difficile, E. coli, Candida albicans
CNS on Neonates
1) Candida albicans overgrowth-
a)Tells the doctor that the patient's normal fecal flora is decreased or wiped out due to strong antibiotics, antacids, anti-reflux medications, immunocompromisation or diabetes
b)Tells the doctor that the patient may have a disseminated, systemic yeast infection (check the blood cultures)
c)The patient may also have UTI and vaginitis (check other cultures)
d)An infant with Candida overgrowth may also develop a diaper rash
e)Treatment includes Amphotericin B or Fluconazole and replacement of the flora with yogurt, kefir, or supplements
2) Clostridium difficile overgrowth-
a)Antibiotic-associated pseudomembranous colitis associated with the use of broad-spectrum antibiotics (Ampicillin, Clindamycin, Cephalosporin)
b)Produces exotoxins
-Toxin A (diarrhea)
-Toxin B (cytotoxic to the colon)
c)Symptoms: severe diarrhea, cramps, fever, red/inflamed colon mucosa, exudate formation, necrosis
d)Treatment: Metronidazole or Vancomycin by mouth
Helicobacter pylori-
Helicobacter pylori is a Gram-negative, corkscrew-shaped, motile bacterium (spirilla) that has the ability to bore into the duodenal wall, creating ulcerations and is linked to stomach cancer. It is a causative agent of chronic gastritis, gastric and duodenal ulcers. Approximately 80% of individuals infected with this microbe are actually asymptomatic, and it is estimated that around half of the world's population may be colonized with H. pylori in their upper gastrointestinal tracts. It produces oxidase, catalase and urease.
Symptoms:
Those who are infected that do develop symptoms may experience the following:
-acute gastritis
-abdominal pain
-nausea
-dyspepsia
-bloating
-belching
-vomiting
-black stool (due to blood)
-10-20% lifetime chance of developing a peptic ulcer
-1-2% lifetime chance of developing stomach cancer
-chronic inflammation of areas of the stomach
-chance of developing colorectal polyps and colorectal cancer
The urease test and clotest (a modified urease test), as well as a rapid identification test are good tools to aid in the identification and treatment for infections with this organism.
MEDIA: TULMS
Triple Sugar Iron Agar (TSI):
*Stab straight down and streak the surface after touching the top of a well-isolated colony with a straight inoculation needle
*Keep the cap on loosely
*Incubate in ambient air at 35 degrees Celsius for 18-24 hours
1) Alkaline Slant/No Change In the Butt (K/NC, or K/K)=glucose, lactose, and sucrose non-utilizer
2) Alkaline Slant/Acid Butt (K/A)=glucose fermentation only ("yellow"=acid production due to glucose fermentation)
3) Acid Slant/Acid Butt (A/A)=glucose, sucrose and/or lactose fermenter
4) Black precipitate in the butt indicates the production of ferrous sulfide and H2S gas
5) Bubbles or cracks in the agar indicate gas production (CO2 or H2)
-Shigella (K/A, no gas, no H2S production)
-Salmonella (K/A, H2S production, no gas production)
-E. coli, Klebsiella spp, Enterobacter spp (A/A, gas production, no H2S production)
-Pseudomonas aeruginosa (K/K, no gas, no H2S)
Urea Hydrolysis:
*Streak surface of agar
*Loosen cap
*Incubate in ambient air at 35 degrees Celsius for 18-24 hours (sometimes it takes 48 hours to 7 days)
1)Positive: Color of slant changes from light yellow-orange to bright fuschia/magenta (Proteus vulgaris, P. mirabilis, Enterobacter cloacae, Klebsiella, Brucella, Bordetella, Yersinia enterocolitica, Morganella spp. H. pylori)
2)Negative: No color change (agar remains light yellow-orange) (E. coli, Salmonella, Shigella, Enterobacter aerogenes)
This test is used to see if an organism has the ability to produce urease, an enzyme that hydrolyzes urea, producing ammonia and CO2, which alkalizes the medium and shifts the pH from 6.8 to 8.1. The pH change is detected by a color indicator (phenol red).
Lysine Iron Agar (LIA):
*Stab straight down and streak the surface after touching the top of a well-isolated colony with a straight inoculation needle
*Keep the cap on loosely
*Incubate in ambient air at 35 degrees Celsius for 18-24 hours
1) LDC + (yellow slant/purple butt) (Salmonella)
2) LDC - (yellow slant/yellow butt)
3) H2S (yellow slant/black butt)
4) LDA + (yellow slant/burgandy butt) (Proteus)
5) LDA- (yellow slant/purple butt) (Salmonella, Shigella)
Motility Agar:
1) Motility + if there is movement away from/spreading out from the stab in the agar
2) Indole + if liquid on top turns pink-red after 1-2 drops of indole are added and mixed well
3) Ornithine + if yellow, - if purple
Motile:
-Yersinia at 22 degrees Celsius
-E.coli
-Listeria at 22 degrees Celsius
Non-motile:
-Shigella
-Klebsiella
-Yersinia at 35 degrees Celcius
Stool For Reducing Substances (Clinitest on Stool):
In this test, Benedict's solution (a blue, alkaline solution) is used to test for the presence of a reducing sugar in the stool. This includes monosaccharides and disaccharides, such as lactose and maltose. It also detects aldehydes and ketoses, such as fructose. It also tests for glucose and galactose, which may indicate diabetes (former) or hereditary galactosemia (latter). When reducing sugars are heated in the presence of an alkali over the Bunsen Burner, they convert to powerful reducing compounds (enediols). Reducing sugars reduce copper II ions (cupric) in the solution to copper I ions (cuprous), which form a copper oxide precipitate. This test is semi-quantitative. "A greenish precipitate indicates about 0.5 g% concentration (trace); yellow precipitate indicates 1 g% concentration; orange indicates 1.5 g% and red indicates 2 g% or higher concentration." ( http://en.wikipedia.org/wiki/Benedict%27s_reagent). Any amount from trace up to high concentration indicates a positive test.
A false positive reaction may occur if other reducing substances are present, such as ascorbic acid (vitamin C supplement). Contrasts used in radiological studies, Levodopa drugs administered to treat Parkinson's disease, and homogentisic acid may also cause false positive results.
Fecal Occult Blood Test:
This test is used to detect "hidden" blood in the stool that may not be visible to the naked eye. The presence of occult blood is often the first sign of colon or rectal cancer, and early detection is key for treatment and prolonged life. The rapid guaiac-based hemoccult stool blood tests detect heme, a component of blood, in the stool. It detects blood loss anywhere along the gastrointestinal tract, from the mouth to the colon. It can be helpful in detecting and diagnosing upper or lower GI bleeds, peptic ulcers, anemia, and even malignancies. It is a screening tool for these purposes, along with other tests.
Upper or lower GI bleeds may be due to gastric or colorectal cancers, polyps, adenoma, peptic ulcer, angiodysplasia of the colon, or sickle cell anemia. If positive, this test is followed up with a sigmoidoscopy and/or colonoscopy or endoscopy, bloodwork, and an M2-PK test.
Fecal Leukocytes Test (WBCs):
White blood cells, or leukocytes, are not normally seen in the stool unless inflammation or infection is present or suspected, therefore, this test is a useful screening tool for the detection of bacterial or parasite infections, or inflammatory conditions caused by autoimmune diseases or viruses. Salmonella, Shigella, Campylobacter, Amoebiasis, ulcerative colitis, Crohn's Disease are all conditions associated with the presence of fecal leukocytes (lactoferrin).
Stool pH:
A fecal pH test is a screening tool to indicate the level of pH of the stool. Because the stool is normally alkaline, an acidic stool may indicate a virus (ex: Rotavirus) infection, a bacterial infection (such as E. coli), or lactose intolerance. This is indicated by a stool pH of less than 5.5.
Fecal Elastase:
This test is used to detect and diagnose pancreatitis.
Stool Fats: Fecal Fat Test
This test is used to detect and diagnose the malabsorption of fats, or steatorrhea. Fatty stools may float, be a light color, have a greasy/oily appearance, and may be very foul-smelling. Causes of steatorrhea may be caused by a lack of bile salts (ex: gallbladder removal), liver damage, drugs (hypolipidemic drugs or medicines that block lipid absorption or treat acromegaly), defective pancreatic enzymes or mucosal cells, or if there is too much undigestible oil or fat in the diet that spills over into the stool.
This test is a screening tool that may aid the physician in detecting and diagnosing the following conditions:
- Malabsorption
- Inflammatory Bowel Disease
- Celiac Disease
- Gluten Intolerance
- Problems with the Pancreas
- Gallstones or Other Bile Duct Blockage (may be due to cancer, tumor, narrowing as well)
- Tropical Sprue
- Recent viral infection
- Current parasite infection, such as Giardia
- Overgrowth of bacteria
- Cystic Fibrosis
- Cholangitis
- Short Bowel Syndrome
- Grave's Disease
- Addiction to certain diet pills
- Zollinger-Ellison Syndrome (gastrin-secreting tumor of the pancreas)
stool culture media:
tsa 5% sheep's blood agar: colony types to look for and hemolysis patterns to look for
Aeromonas hydrophila, A. veronii, A. caviae:
Aeromonas hydrophila:
- Facultative anaerobe
- Gram-negative rod (typical, straight, medium)
- Motile
- Beta-hemolytic on blood agar
- Ferments both sucrose and mannitol
- Indole +, Oxidase +
- Lysine decarboxylase +
- Esculin hydrolysis +
- Ornithine decarboxylase -
- Found in aquatic habitats around the world
- Causes infections by colonizing open trauma wounds or by ingesting contaminated water or food
- Produces an enterotoxin
- Symptoms include acute, watery diarrhea and/or UTI
- Can cause bacteremia and sepsis
- Mortality rates are high amongst immunocompromised patients following sepsis
Plesiomonas shigelloides:
bacillus cereus:
Bacillus cereus is an organism that commonly causes food poisoning from consumption of foods containing excessive populations of organisms or preformed enterotoxin. It is associated with cooked rice that has been sitting at room temperature for >2 hours without refrigeration. It can cause diarrhea and/or vomiting.
Colonies are large, golden tan, rough, spreading, smelly, umbonate and strongly beta-hemolytic with large zones of hemolysis surrounding them on sheeps' blood agar.
Colonies are large, golden tan, rough, spreading, smelly, umbonate and strongly beta-hemolytic with large zones of hemolysis surrounding them on sheeps' blood agar.
Bacillus cereus produces large, spreading, rough, tan-gray colonies with large zones of beta-hemolysis. It is oxidase variable, motility positive, catalase +, lipase + organism. Organisms suspected of being B. cereus should be tested for slide motility, catalase, oxidase, and lipase on the egg yolk agar. A Vitek GPI and GPS should be performed for confirmation.
fecal occult blood
Some fecal occult blood tests will detect blood along any part of the intestinal tract, including upper GI bleeds from ulcers or aspirin sensitivities, cancers, or lower GI bleeds such as colorectal cancers and hemorrhoids and fissures.
Reducing substances (clinitest):
Benedict's solution, a copper solution used to perform the reducing substances test in stool, comes initially as a blue liquid that is heated up with the stool over a bunsen burner (left). Once heated to boiling, stools negative for reducing substances will remain a dark blue-green (teal) color (middle) and positives will turn any shade of green to yellow to orange (right), as seen in the strongly positive tube to the right.
fecal fats
Oil red O or Sudan Red are used to dye lipids red so they can be observed if present in the stool. This test is useful for the dianosis of steatorrhea and malabsorption caused by a variety of things. An equal part of water and stool is first mixed to create a solution in one test tube. Another test tube is prepared with 95% ethanol alcohol. Another test tube is prepared with Oil red O. 2 drops each of the stool solution, alcohol and Oil red O are placed onto a clean slide and covered with a long coverslip. Observe under the microscope under low light and turn the condenser down since lipids are transparent. >60 lipids per high power field is considered abnormal.
Things that can interfere with the test and cause false positive results include the following taken within 1 week of the test:
Things that can interfere with the test and cause false positive results include the following taken within 1 week of the test:
- Metamucil
- Barium
- Oily salad dressing
- Bismuth
- Castor oil
- Mineral oil
- Olive oil
- Suppository
zollinger-ellison syndrome
malabsorption/steatorrhea (celiac disease, whipple disease, and damaged microvilli and intestinal crypts, preventing absorption)
normal intestinal microvilli (brush border; increases surface area for absorption and reabsorption of sugars, lipids, nutrients)
crohn's disease and ulcerative colitis: inflammation, friability, ulcerations, polyps, damaged microvilli and intestinal crypts
colorectal cancers
gallstones
Stool pathogens media growth patterns:
aeromonas
Aeromonas spp are oxidase (+), facultative anaerobic Gram-negative bacilli. They resemble members of the Enterobacteriaceae. The most clinically significant species include:
- A. hydrophila
- A. caviae
- A. veronii sobria
- Gastroenteritis/Diarrhea (chronic in adults, acute/severe in children) with blood and leukocytes in the stool
- Soft tissue and wound infections
- Bacteremia
- Endotoxins
- Hemolysins
- Enterotoxins
- Adherence Factors
Plesiomonas shigelloides are oxidase (+), Gram-negative bacilli that resemble the Enterobacteriaceae.
Diseases caused by this organism include:
vibrio spp
|
Vibrio spp are oxidase (+), motile, curved Gram-negative bacilli. Clinically significant species include:
- V. cholerae
- V. parahemolyticus
- V. vulnificus
- V. alginolyticus
- Cholera
- Gastroenteritis, usually from eating undercooked seafood/shellfish
- Wound infections
- Septicemia
- Otitis (ear infection)
- Cellulitis
Campylobacter spp:
Campylobacter spp are oxidase (+), microaerophilic, motile, spiral-shaped Gram-negative bacilli ("S" or "seagull wings") that are nonfermentive. Clinically significant species include:
- C. jejuni (only spp that is HIPPURATE +)
- C. coli
- C. fetus
- Campylobacteriosis
- Dysentery
- Bloody diarrhea
- Cramps, fever, pain
- Hemolytic Uremic Syndrome (rare complication)
- Thrombotic thrombocytopenic purpura (TTP) (rare complication)
- Guillain-Barre' Syndrome (rare complication)
- Gastrointestinal perforation (rare complication)
- Periodontitis
Salmonella spp
|
Salmonella spp is a Gram-negative, facultative anaerobic, motile member of the Enterobacteriaceae. They include:
- S. enterica
- S. typhi
- S. typhimurium
- S. bongori
- Salmonellosis (food poisoning) with back pain, GI tract infection, diarrhea, weakness, myalgia, enteric fever, bacteremia, local infection, and chronic reservoir state (survives in REC cells) ; It can also cause arthritis, UTI, CNS infection, bone infection, soft tissue infection
- Typhoid fever
- Paratyphoid fever
shigella spp
Shigella spp are nonmotile, facultative anaerobic, Gram-negative bacilli that are similar to Salmonella spp. There are 4 serogroups which include:
- S. flexneri
- S. boydii
- S. dysenteriae
- S. sonnei
- Shigellosis (diarrhea, gas, abdominal cramps, fever, headache, nausea, vomiting, reactive arthritis, bloody diarrhea with mucus and pus, sometimes alternating with constipation)
- Dysentery
- Hemolytic Uremic Syndrome (rare complication)
Escherichia coli 0157:H7
This is a reportable organism to Public Health.
yersinia enterocolitica
Yersinia enterocolitica forms colorless, nonfermenting colonies on MacConkey agar, yellow-orange colonies on HEK agar, produces an acid slant/acid butt (yellow/yellow) with no gas and no H2S or blackening of the agar on the TSI slant, and is positive for the urease test. This Gram-negative bacilli ferments carbohydrates. Symptoms of infection and illness with this organism include fever, abdominal pain, lower right quadrant pain and tenderness resembling appendicitis, diarrhea, and vomiting. It is oxidase -, indole -, urease +, ornithine decarboxylase +, sucrose +, motility + at 22 degrees Celsius and H2S negative. Other laboratory tests indicate an increased WBC count and increased PMNs. Lymph nodes may be enlarged. The organism is recovered on plates left at room temperature, and can be identified with the Yersinia CIN agar plate left at room temperature, producing colonies with pink centers resembling "bulls' eyes" or "targets". This is a reportable organism to Public Health.
staphylococcus aureus
clostridium botulinum
bacillus cereus
Yeast
pseudomonas aeruginosa
coagulase-negative staphylococcus
urines bench:
The MacConkey plate is used for the detection of lactose-fermenting and non-lactose-fermenting bacteria. It contains crystal violet and bile salts to prevent the growth of Gram-positive organisms and it contains a pH indicator (phenol red) that changes colors when lactose within the medium is fermented.
FACTS:
-The most common urine pathogen overall is E. coli (80-85% of cases)
-E. coli is also the #1 cause of nosomial (hospital-acquired) infections
-Other Enterobacteriaceae and nonenterobacteriaceae are the main cause of UTI (see all of the stool fecal flora colonizers as well as stool pathogens)
-Urines are streaked with the semi-quantitative streak for colony count
-If you see a gamma-hemolytic, gray-white colony on the BAP that is weakly catalase +, perform a PYR test and a bile esculin test to rule out Enterococcus spp
-If you have an Enterococcus spp with a pure purity plate and growth on a VSA plate, or the Vitek says that the organism is Vancomycin resistant, perform a Vancomycin E-test
-If you see a beta-hemolytic organism on the BAP and it is catalase -, and the culture is on a woman of child-bearing age, perform a Group B strep agglutination test to rule out GBS, because S. agalactiae is associated with birth defects, stillbirth, and meningitis in neonates with mothers who are carriers colonized with the organism.
-The Enterobacteriaceae are very common Gram-negative pathogens recovered in UTIs (ex: Klebsiella spp, Enterobacter spp, Proteus spp, Pseudomonas spp, E. coli, Citrobacter spp, Serratia spp, Etc...)
-If you suspect Corynebacterium, perform a urease test and, if it is positive, an API Coryne Strip to rule out C. urealyticum
-If you isolate a yeast, perform a germ tube test to rule out C. albicans, since it is germ tube +. If your yeast is germ tube -, set up an API Yeast or a Vitek Yeast card and a cornmeal agar with 4 stabs in the 1st quadrant and a coverslip over the stabs to promote pseudohyphae and chlamydospore production. Place in the fungal incubator at 37 degrees C.
-A common Gram-positive organism that causes urinary tract infections in sexually active young women is Staphylococcus saprophyticus (5-10% of cases). It is resistant to the Novobiocin disk.
-UTI's are one of the most common bacterial infections
-Approximately 6 million outpatient visits and around 300,000 hospital visits each year are due to UTIs
-About 10% of individuals will have a UTI at some time during their life time.
-A parasite that can cause a UTI is: Schistosoma haemotobium
UTI Complications:
-Occur as a result of diabetes, kidney disease, transplants, structural anomalies such as ureter strictures or shortened urethras, neurogenic bladders, spina bifida, MS, neurologic abnormalities, kidney infection, sickle cell anemia, kidney stones, a tipped bladder, an indwelling urinary catheter
-UTIs are the leading cause of Gram-negative sepsis in hospitalized patients (bacteremia, sepsis, bacteria travels to bloodstream)
-UTIs are the starting point for about 1/2 of all nosocomial (hospital-acquired) infections, and many are associated with urinary catheters. In fact, as much as 20% of patients who have been catheterized, even short-term, in the hospital may develop a UTI due to the nature of the material the catheter is made of and the nature of the ability of bacterial to produce a sticky biofilm and/or be displaced from the skin into the urethra and travel to the bladder and/or kidneys.
-Proteus, Pseudomonas, Klebsiella, Enterobacter spp are associated with more complicated UTI's and recurrent infections
Specimen Collection:
-Transport urine to the laboratory as soon as possible after collection
-Culture specimens within 30 minutes if possible, or refrigerate within 2 hours and culture them within 24 hours
-Urines in sterile cups are unpreserved
-Urines in gray-top tubes are preserved
-Request a repeat urine specimen collection if there is no evidence of refrigeration and the specimen is >2 hours old
-If the urine has been improperly collected, unlabeled or mislabeled, improperly transported or handled, and cannot be replaced, document this in the final report that the specimen quality may have been compromised
-Perform a semi-quantitative streak with the small, 0.001 uL loop. If it is a pediatric catherized urine, performed the same streak with the larger 0.01 uL loop.
-0.001 uL Loop=1,000 CFU/mL
-0.01 uL Loop=100 CFU/mL
-Urine Gram Stain: easy, cost-effective, sensitive, reliable; Drop 1 drop of well-mixed urine on a slide and allow to air dry; heat or methanol fix the smear, Gram-stain and observe under oil-immersion (100x) objective for the presence of at least 1 organism per oif
Urinalysis Tests That Aid in the Screening Process:
1) Nitrate Reductase (Greiss) Test-This test observes for the presence of urine nitrite. Nitrite present in the urine is an indicator of UTI. Most of the common urinary tract pathogens reduce nitrate to nitrite, producing this byproduct in the urine and leading to a positive test.
2) Leucocyte Esterase (LE) Test-This test tests for leukocyte esterase, an enzyme produced by polymorphonuclear neutrophils (PMNs) in the urine, a sign of inflammation.
3) WBC-A high WBC count in the urine is an indicator of inflammation and possible infection
4) RBC-A high RBC count in the urine may indicate possible infection in collaboration with other tests
5) Protein-A high protein count in the urine in conjunction with other positive tests may indicate possible infection
6) Specific Gravity-A high specific gravity in the urine in conjunction with other positive tests may indicate possible infection
Organisms that can Cause UTI:
Many UTI's are caused by the Enterobacteriaceae (normal gut flora) or the normal commensal skin flora (yeast, staphylococci) getting into a normally sterile body site by being at the wrong place at the wrong time in the right conditions. Some patients who are colonized with Group B streptococci (GBS, S. agalactiae) may also have GBS in their urine, and do not necessarily have an infection, but may be colonized with the organism. Common tests to rule out the causative agent for UTI include catalase, coagulase, indole, oxidase, PathoDx Strep Panel, Gram stain, urease, bile esculin, PYR, beta-lactamase, PBP2, motility, germ tube test, Cornmeal agar test, API strips, Vitek ID/Susceptibility, KB disk testing, and Etests.
-E. coli (#1 cause; community and nosocomial-acquired; Indole postive)
-S. saprophyticus (leading cause, 2nd to E. coli, of UTI in sexually active young women; community-acquired; Coagulase negative)
-S. epidermidis
-S. aureus
-Proteus mirabilis or vulgaris (pure)
-Pseudomonas aeruginosa or other spp
-Citrobacter spp
-Enterobacter spp
-Klebsiella oxytoca or pneumoniae
-Mycoplasma
-Ureaplasma urealyticum
-Salmonella spp
-Serratia marscecens
-Lactobacillus (heavy)
-Corynebacterium urealyticum
-Enterococcus faecalis
-Enterococcus gallinum
-Enterococcus casseiflavus
-Enterococcus raffinosus
-Enterococcus faecium
-S. agalactiae (GBS)
-MRSA
-Stenotrophomonas maltophilia
-Sphingomonas paucimobilis
-Edwardsiella tarda
-Yeast (pure)
-Hafnia spp
-Acinetobacter spp
-Alcaligenes spp
-Gardnerella vaginalis
-Mycobacterium spp
-Campylobacter spp
-H. influenzae
-Leptospira spp
-Schistosoma haemotobium
-Eikenella corrodens
-Edwardsiella tarda
-Morganella morganii
-Raoultella pantoea
-Providencia spp
-Streptococcus pneumoniae
-Rarely, CNS
-Listeria
-Bacillus spp
-The most common urine pathogen overall is E. coli (80-85% of cases)
-E. coli is also the #1 cause of nosomial (hospital-acquired) infections
-Other Enterobacteriaceae and nonenterobacteriaceae are the main cause of UTI (see all of the stool fecal flora colonizers as well as stool pathogens)
-Urines are streaked with the semi-quantitative streak for colony count
-If you see a gamma-hemolytic, gray-white colony on the BAP that is weakly catalase +, perform a PYR test and a bile esculin test to rule out Enterococcus spp
-If you have an Enterococcus spp with a pure purity plate and growth on a VSA plate, or the Vitek says that the organism is Vancomycin resistant, perform a Vancomycin E-test
-If you see a beta-hemolytic organism on the BAP and it is catalase -, and the culture is on a woman of child-bearing age, perform a Group B strep agglutination test to rule out GBS, because S. agalactiae is associated with birth defects, stillbirth, and meningitis in neonates with mothers who are carriers colonized with the organism.
-The Enterobacteriaceae are very common Gram-negative pathogens recovered in UTIs (ex: Klebsiella spp, Enterobacter spp, Proteus spp, Pseudomonas spp, E. coli, Citrobacter spp, Serratia spp, Etc...)
-If you suspect Corynebacterium, perform a urease test and, if it is positive, an API Coryne Strip to rule out C. urealyticum
-If you isolate a yeast, perform a germ tube test to rule out C. albicans, since it is germ tube +. If your yeast is germ tube -, set up an API Yeast or a Vitek Yeast card and a cornmeal agar with 4 stabs in the 1st quadrant and a coverslip over the stabs to promote pseudohyphae and chlamydospore production. Place in the fungal incubator at 37 degrees C.
-A common Gram-positive organism that causes urinary tract infections in sexually active young women is Staphylococcus saprophyticus (5-10% of cases). It is resistant to the Novobiocin disk.
-UTI's are one of the most common bacterial infections
-Approximately 6 million outpatient visits and around 300,000 hospital visits each year are due to UTIs
-About 10% of individuals will have a UTI at some time during their life time.
-A parasite that can cause a UTI is: Schistosoma haemotobium
UTI Complications:
-Occur as a result of diabetes, kidney disease, transplants, structural anomalies such as ureter strictures or shortened urethras, neurogenic bladders, spina bifida, MS, neurologic abnormalities, kidney infection, sickle cell anemia, kidney stones, a tipped bladder, an indwelling urinary catheter
-UTIs are the leading cause of Gram-negative sepsis in hospitalized patients (bacteremia, sepsis, bacteria travels to bloodstream)
-UTIs are the starting point for about 1/2 of all nosocomial (hospital-acquired) infections, and many are associated with urinary catheters. In fact, as much as 20% of patients who have been catheterized, even short-term, in the hospital may develop a UTI due to the nature of the material the catheter is made of and the nature of the ability of bacterial to produce a sticky biofilm and/or be displaced from the skin into the urethra and travel to the bladder and/or kidneys.
-Proteus, Pseudomonas, Klebsiella, Enterobacter spp are associated with more complicated UTI's and recurrent infections
Specimen Collection:
- Specimens should be obtained from early morning collection when possible, because of the increased bacterial concentration after overnight incubation in the bladder
- If patients are asymptomatic, three consecutive early morning specimens should be collected
- Specimens for urine culture should be clean catch midstream in a sterile container MIDSTREAM, CLEAN CATCH)
- If urine is catheterized, a closed system should be used, or a straight catheter should be used by a physician to obtain urine directly from the bladder. Foley catheters should be avoided since they increased the risk of UTI.
- Suprapubic Bladder Aspiration (urine is withdrawn into a syringe through a needle that has been inserted percutaneously)
-Transport urine to the laboratory as soon as possible after collection
-Culture specimens within 30 minutes if possible, or refrigerate within 2 hours and culture them within 24 hours
-Urines in sterile cups are unpreserved
-Urines in gray-top tubes are preserved
-Request a repeat urine specimen collection if there is no evidence of refrigeration and the specimen is >2 hours old
-If the urine has been improperly collected, unlabeled or mislabeled, improperly transported or handled, and cannot be replaced, document this in the final report that the specimen quality may have been compromised
-Perform a semi-quantitative streak with the small, 0.001 uL loop. If it is a pediatric catherized urine, performed the same streak with the larger 0.01 uL loop.
-0.001 uL Loop=1,000 CFU/mL
-0.01 uL Loop=100 CFU/mL
-Urine Gram Stain: easy, cost-effective, sensitive, reliable; Drop 1 drop of well-mixed urine on a slide and allow to air dry; heat or methanol fix the smear, Gram-stain and observe under oil-immersion (100x) objective for the presence of at least 1 organism per oif
Urinalysis Tests That Aid in the Screening Process:
1) Nitrate Reductase (Greiss) Test-This test observes for the presence of urine nitrite. Nitrite present in the urine is an indicator of UTI. Most of the common urinary tract pathogens reduce nitrate to nitrite, producing this byproduct in the urine and leading to a positive test.
2) Leucocyte Esterase (LE) Test-This test tests for leukocyte esterase, an enzyme produced by polymorphonuclear neutrophils (PMNs) in the urine, a sign of inflammation.
3) WBC-A high WBC count in the urine is an indicator of inflammation and possible infection
4) RBC-A high RBC count in the urine may indicate possible infection in collaboration with other tests
5) Protein-A high protein count in the urine in conjunction with other positive tests may indicate possible infection
6) Specific Gravity-A high specific gravity in the urine in conjunction with other positive tests may indicate possible infection
Organisms that can Cause UTI:
Many UTI's are caused by the Enterobacteriaceae (normal gut flora) or the normal commensal skin flora (yeast, staphylococci) getting into a normally sterile body site by being at the wrong place at the wrong time in the right conditions. Some patients who are colonized with Group B streptococci (GBS, S. agalactiae) may also have GBS in their urine, and do not necessarily have an infection, but may be colonized with the organism. Common tests to rule out the causative agent for UTI include catalase, coagulase, indole, oxidase, PathoDx Strep Panel, Gram stain, urease, bile esculin, PYR, beta-lactamase, PBP2, motility, germ tube test, Cornmeal agar test, API strips, Vitek ID/Susceptibility, KB disk testing, and Etests.
-E. coli (#1 cause; community and nosocomial-acquired; Indole postive)
-S. saprophyticus (leading cause, 2nd to E. coli, of UTI in sexually active young women; community-acquired; Coagulase negative)
-S. epidermidis
-S. aureus
-Proteus mirabilis or vulgaris (pure)
-Pseudomonas aeruginosa or other spp
-Citrobacter spp
-Enterobacter spp
-Klebsiella oxytoca or pneumoniae
-Mycoplasma
-Ureaplasma urealyticum
-Salmonella spp
-Serratia marscecens
-Lactobacillus (heavy)
-Corynebacterium urealyticum
-Enterococcus faecalis
-Enterococcus gallinum
-Enterococcus casseiflavus
-Enterococcus raffinosus
-Enterococcus faecium
-S. agalactiae (GBS)
-MRSA
-Stenotrophomonas maltophilia
-Sphingomonas paucimobilis
-Edwardsiella tarda
-Yeast (pure)
-Hafnia spp
-Acinetobacter spp
-Alcaligenes spp
-Gardnerella vaginalis
-Mycobacterium spp
-Campylobacter spp
-H. influenzae
-Leptospira spp
-Schistosoma haemotobium
-Eikenella corrodens
-Edwardsiella tarda
-Morganella morganii
-Raoultella pantoea
-Providencia spp
-Streptococcus pneumoniae
-Rarely, CNS
-Listeria
-Bacillus spp
The plates used for urine culture include 5% TSA Sheep's Blood Agar and MacConkey agar. Growth on MacConkey agar indicates Gram-negative rods that are either lactose-fermenters or non-lactose-fermenters or late-lactose-fermenters. Growth on the blood side only may indicate a Gram-positive organism or yeast. Hemolysis patterns are observed to aide in identification. Beta-hemolytic GP organisms include staphylococci and streptococci. Gamma or non-hemolytic GP organisms include things like Enterococcus faecalis or faecium, S. saprophyticus, S. epidermidis, or other CNS. Alpha hemolytic organisms may include Viridans streptococci, Lactobacillus, Streptococcus pnuemoniae, Gardnerella vaginalis, Gemella, etc...For this section, the Gram-stains tutorials and indiviual Gram stain types, as well as the biochemical section are helpful tools on this website for ID.
Urinalysis results should be read to see what was seen on the urine during that process, including bacteria, RBCs, WBCs, to see if the nitrate test was positive and to see if the leukocyte esterase test was positive. Other helpful indicators include a high protein and high specific gravity along with these tests. These are pretty good indicators that the patient has a UTI and may have already begun treatment.
Urinalysis results should be read to see what was seen on the urine during that process, including bacteria, RBCs, WBCs, to see if the nitrate test was positive and to see if the leukocyte esterase test was positive. Other helpful indicators include a high protein and high specific gravity along with these tests. These are pretty good indicators that the patient has a UTI and may have already begun treatment.
four major types of urinary tract infections:
1) URETHRITIS:
Urethritis is infection of the urethra, often associated with dysuria, or painful and difficult urination and frequency. This is also called a lower UTI and it is a common infection. Sometimes, this is caused by an STD, such as Neisseria gonorrhoeae, Chlamydia trachomatis, or Trichomonas vaginalis.
2) CYSTITIS:
Cystitis is a bladder infection, also a lower UTI. Symptoms include dysuria, a need to urinate frequently, and a sense of urgency. This is due to inflammation in the bladder and multiplication of bacteria in the urethra and the urine with tenderness and pain over the bladder. The urine may be cloudy, bloody, and/or have a putrid/foul odor to it. Green-yellow, thick, cloudy or milky urine is due to the presence of pus, a combination of bacteria, high white blood cell count, bacteria and cellular debris. This is called pyuria. Pyuria is indicated by the presence of 8+ leukocytes (WBCs) per cubic mm on microscopic examination of uncentrifuged urine.
3) ACUTE URETHRAL SYNDROME:
Patients who experience acute urethral syndrome are typically young females of child-bearing age who are sexually active. The symptoms include dysuria, frequency and urgency. Staphylococcus saprophyticus is a common cause of this type of UTI, which is introduced into the skin of the perineum through the urethra and into the bladder. Females with shorter urethras are at higher risk for this type of infection, including recurrent infections.
4) PYELONEPHRITIS:
Pyelonephritis is a kidney infection with inflammation of the kidney, the parenchyma, the calices, and the renal pelvis, caused by the travel of bacteria from the bladder to the kidneys. This is an upper UTI and includes fever, chills, general feeling of unwellness, malaise, flank pain, frequency, urgency, dysuria, and occasionally vomiting, diarrhea, tachycardia and lower abdominal pain as well. As many as 40% of patients with pyelonephritis have bacteremia as well, so blood cultures may become positive, too.
Urethritis is infection of the urethra, often associated with dysuria, or painful and difficult urination and frequency. This is also called a lower UTI and it is a common infection. Sometimes, this is caused by an STD, such as Neisseria gonorrhoeae, Chlamydia trachomatis, or Trichomonas vaginalis.
2) CYSTITIS:
Cystitis is a bladder infection, also a lower UTI. Symptoms include dysuria, a need to urinate frequently, and a sense of urgency. This is due to inflammation in the bladder and multiplication of bacteria in the urethra and the urine with tenderness and pain over the bladder. The urine may be cloudy, bloody, and/or have a putrid/foul odor to it. Green-yellow, thick, cloudy or milky urine is due to the presence of pus, a combination of bacteria, high white blood cell count, bacteria and cellular debris. This is called pyuria. Pyuria is indicated by the presence of 8+ leukocytes (WBCs) per cubic mm on microscopic examination of uncentrifuged urine.
3) ACUTE URETHRAL SYNDROME:
Patients who experience acute urethral syndrome are typically young females of child-bearing age who are sexually active. The symptoms include dysuria, frequency and urgency. Staphylococcus saprophyticus is a common cause of this type of UTI, which is introduced into the skin of the perineum through the urethra and into the bladder. Females with shorter urethras are at higher risk for this type of infection, including recurrent infections.
4) PYELONEPHRITIS:
Pyelonephritis is a kidney infection with inflammation of the kidney, the parenchyma, the calices, and the renal pelvis, caused by the travel of bacteria from the bladder to the kidneys. This is an upper UTI and includes fever, chills, general feeling of unwellness, malaise, flank pain, frequency, urgency, dysuria, and occasionally vomiting, diarrhea, tachycardia and lower abdominal pain as well. As many as 40% of patients with pyelonephritis have bacteremia as well, so blood cultures may become positive, too.
Lactobacillus spp are part of the normal urogenital/vaginal flora of females. They reside the the urinary, digestive and gential tracts of humans. They are rarely a cause of infection, but their colonization may be noted for physicians to be aware of. They convert lactose and sugars to lactic acid, and are a part of the lactic acid bacteria family. They help to maintain the normal pH of the vaginal tract. These bacilli may be long, curved, spiral, or just regular, straight Gram-positive rods. Lactobacillus are alpha-hemolytic and catalase -.
Viridans Group Streptococci are part of the commensal skin and oral flora. In culture, they may be either alpha-hemolytic or non-hemolytic. The term "viridans" refers the the greening of the agar that occurs by the alpha-hemolytic strains, and the halo that forms around colonies that are non-hemolytic. Because this group possesses no Lancefield antigens, pathogenicity tends to be low in normal healthy individuals. This streptococci group is resistant to the Optochin (Taxo P) disk test, and is negative for the bile esculin hydrolysis test. Some of the species Included in this group are: S. mutans, S. mitis, S. sanguinis, S. oralis, S. salivarious, S. gordonii, and S. anginosus (butterscotch or caramel scent). Because they are part of the normal oral flora, this group is associated with bacterial plaque formation, dental cavities, mouth and gingival infections, and endocarditis in patients with damaged heart valves or prosthetic valves. S. viridans group is the most common cause of subacute bacterial endocarditis because of its ability to adhere to fibrin-platelet aggregates. Patients with underlying structural disorders, immunocompromisation and immunosuppression are at higher risk of infections, and leukemia patients at at risk for severe sepsis with shock. Very rarely, it can cause bacteremia and sepsis in susceptible patients, catheter infections, and transient bacteremia in catheterized pediatric patients. Patients who are neutropenic, on prophylactic antibiotics, or who have a heavy colonization of the group are at higher risk of infection.
The 3 main types of infections to look for with this group include:
-Subacute Endocarditis (due to production of extracellular dextran that allows it to bind to cardiac valves)
-Slow growth and buildup of bacteria on the heart valve
-Symptoms: slow development of low-grade fevers, heart murmurs, fatigue, anemia
-Abscesses (S. anginosus, S. intermedius, S. constellatus)
-Microaerophilic
-Part of the normal GI flora
-Brain abscesses or abdominal abscesses found alone or in mixed cultures with anaerobes like B. fragilis
-If S. intermedius is found in a blood culture, suspect an abscess hiding in an organ somewhere
-Dental Infections (S. mitis or S. mutans)
S. viridans is also referred to as "nutrient-deficient streptococci". They grow as nonhemolytic or alpha-hemolytic, pinpoint, domed gray colonies on blood agar. S. anginosus has a butterscotch or caramel scent. S. lugdunensis smells like the scent of fresh hay, somewhat sweet. S. gordonii is a mouth colonizer and forms biofilm related to dental plaque. Rarely, it can cause acute bacterial endocarditis if introduced into the bloodstream.
The 3 main types of infections to look for with this group include:
-Subacute Endocarditis (due to production of extracellular dextran that allows it to bind to cardiac valves)
-Slow growth and buildup of bacteria on the heart valve
-Symptoms: slow development of low-grade fevers, heart murmurs, fatigue, anemia
-Abscesses (S. anginosus, S. intermedius, S. constellatus)
-Microaerophilic
-Part of the normal GI flora
-Brain abscesses or abdominal abscesses found alone or in mixed cultures with anaerobes like B. fragilis
-If S. intermedius is found in a blood culture, suspect an abscess hiding in an organ somewhere
-Dental Infections (S. mitis or S. mutans)
S. viridans is also referred to as "nutrient-deficient streptococci". They grow as nonhemolytic or alpha-hemolytic, pinpoint, domed gray colonies on blood agar. S. anginosus has a butterscotch or caramel scent. S. lugdunensis smells like the scent of fresh hay, somewhat sweet. S. gordonii is a mouth colonizer and forms biofilm related to dental plaque. Rarely, it can cause acute bacterial endocarditis if introduced into the bloodstream.
The term "diphtheroids" is a term that is used frequently to describe the non-pathogenic Corynebacterium spp. They are Gram + and Catalase + bacilli whose morphology is "Coryneform", meaning that the rods appear side-by-side like picket fences (palisades) or as "V" shapes, like drumsticks, or like Chinese letters. They are not motile, and they appear straight or slightly curved. Because they contain vacoles for storage of nutrients such as phosphate, they may appear to stain "dotted" because of the presence of metachromic granules. They may also display thickenings at each end. They are part of the normal skin flora.
The only pathogenic Corynebacterium species that may cause UTI is C. urealyticum, which is urease + and lipophilic (will display a + lipophilic test). It produces the urease enzyme, causing the urine to become alkaline. Therefore, because of this, it is linked to the formation of renal (kidney) stones or struvite calculi. Those who are immunosuppressed, on antibiotics, or who have an underlying urogenitary disorder or disease are at higher risk of infection.
The only pathogenic Corynebacterium species that may cause UTI is C. urealyticum, which is urease + and lipophilic (will display a + lipophilic test). It produces the urease enzyme, causing the urine to become alkaline. Therefore, because of this, it is linked to the formation of renal (kidney) stones or struvite calculi. Those who are immunosuppressed, on antibiotics, or who have an underlying urogenitary disorder or disease are at higher risk of infection.
Gardnerella vaginalis is the causative agent of bacterial vaginosis, which is typically due to a disruption of vaginal pH and disruption of the normal vaginal microflora. Even though it is usually isolated in vaginal cultures or wet mounts, it is occasionally isolated in urine. Microscopically, GV appears as small, pleomorphic, Gram-variable or Gram-negative coccobacilli and short rods, some curved. On blood agar, colonies are pinpoint, small, gray, alpha- or gamma-hemolytic, opaque colonies. GV is catalase -. It is also hippurate +, oxidase -, and maltose, fructose, and dextrose +.
Enterococcus spp are associated with UTI's. If this species is identified, a catalase test (-), a PYR test (+), Bile Esculin Agar test (+), and ID/Susceptibility should be performed. Once identified, the organism should be checked for Vancomycin susceptibility. If resistant, a confirmation E-test should be performed to verify true resistance. VRE is on the rise, and is associated with nosocomial (hospital-acquired) infections. Cephalosporin antibiotic therapy is associated with colonization and infection with VRE. E. faecalis and E. faecium are more commonly associated with UTI, but rarely, E. casseliflavus, E. gallinarum, and E. raffinosus, among other species, cause infection as well. E. casseliflavus and E. gallinarum show inherent resistance to Vancomycin. Nitrofurantoin is usually a good antibiotic of choice for treatment of UTI with Enterococcus spp. It is relatively resistant to penicillin. Enterococcus grows in the presence of 6.5% NaCl broth.
Streptococcus agalactiae is a beta-hemolytic, small cream-colored, glossy colony on SBA and is also known as Group B Streptococcus as part of the Lancefield grouping. If >10,000 CFU are present in the urine of a pregnant woman or woman of child-bearing age, this may represent of colonization of the bacterium, which should be noted. Though the female may be asymptomatic, should she become or currently be pregnant, GBS is associated with chorioamnioitis, stillbirth, premature delivery, post-partum infections, and bacterial septicemia of the newborn, which may lead to death or to long-term health problems. Additionally, pneumonia in the neonate, as well as meningitis, are also possible and linked to exposure of GBS from the colonization of the vaginal canal, vertical transmission from mother-to-infant, or during birth following the rupture of the membranes. Infection can occur during the birthing process.
Streptococcus pyogenes (Group A streptococcus) may be associated with an autoimmune response following strep throat, leading to glomerulonephritis, or it may be associated with a UTI. Streptococcus Groups C, F, and G are also occasionally associated with UTI or colonization.
S. aureus UTI's are usually secondary to infections that have seeded from elsewhere in the body. A pure culture of S. aureus is considered to be significant, no matter how many the number of CFU/mL. S. aureus is catalase + and coagulase +.
S. saprophyticus is a common cause of community-acquired UTI amongst women who are young, sexually active and of child-bearing age. It is part of the normal urogenital flora of females and of the perineum. However, it causes from 5-20% of UTI's in females of 14-27 years of age, 2nd only to infections by E. coli. This cause of cystitis typically occurs rapidly, as early as 24 hours after sexual activity, because the bacteria are displaced from the skin and enter the urethra into the bladder, leading to bacterial multiplication and infection. Pili and adhesives enable this bacterial to adhere strongly to the urinary epithelium and mucosa. It is part of the coagulase negative staphylococci and is resistant to Novobiocin. Colonies are smooth, glistening, opaque, white, occasionally yellowish with age, non-hemolytic. It smells strong and rather pungent. Sensitivities are not performed on this CNS.
Staphylococcus epidermidis is part of the family of coagulase negative staphylococci. It is susceptible to Novobiocin. It is part of the normal skin flora. It rarely causes UTI, and if it does, it is usually associated with a catheter. Colonies are circular, smooth, pale, matte, translucent, white, and non-hemolytic. It is catalase + and coagulase -. Sensitivities are performed on CNS other than S. saprophyticus. It is the main cause of acute infective endocarditis often secondary to IV drug abuse or immunocompromisation, and characterized by a rapid onset of shaking chills, high fevers and rapid destruction of the heart valves.
Members of Coagulase Negative Staphylococci include: S. epidermidis, S. haemolyticus, S. lugdunensis, S. capitis, S. caprae, S. warneri, S. schleiferi, S. hominis, S. auricularis, S. cohnii, S. xylosus, S. simulans, S. saprophyticus, S. kooslii. CNS members are catalase +, coagulase -, usually nonhemolytic, though some strains are beta-hemolytic, and colonies may be cream, white, tan or gray. Sensitivity testing is performed on CNS other than S. saprophyticus.
Members of Coagulase Negative Staphylococci include: S. epidermidis, S. haemolyticus, S. lugdunensis, S. capitis, S. caprae, S. warneri, S. schleiferi, S. hominis, S. auricularis, S. cohnii, S. xylosus, S. simulans, S. saprophyticus, S. kooslii. CNS members are catalase +, coagulase -, usually nonhemolytic, though some strains are beta-hemolytic, and colonies may be cream, white, tan or gray. Sensitivity testing is performed on CNS other than S. saprophyticus.
E. coli is a Gram-negative bacilli that is the number one cause of community-acquired UTI and nosocomial infection. It is usually a lactose-fermenter, but there are also some non-lactose fermenting strains on MacConkey agar. It is indole + and oxidase - and PYR-. E. coli may be gamma-hemolytic or beta-hemolytic on Blood Agar, and colonies are typically either gray or tan, may be spreading or circular and entire, and some strains are mucoid.
Citrobacter spp is a Gram-negative bacilli associated with UTI. It is a late lactose-fermenter, meaning that at 24 hours, it may appear to be a NLF, and at 48 hours, will appear LF. Some of the most common Citrobacter spp include C. koseri, C. freundii, C. braakii, C. youngae.
Enterobacter spp are lactose-fermenting Gram-negative bacilli that are associated with UTI. This organism is both oxidase - and indole -. At 24 hours, some strains appear as NLF even though they are LF, because they may be late LF. Enterobacter spp are indole - and motility +. Colonies are thick, white, raised, moist, smooth and entire on Blood Agar, and non-hemolytic. Many times they are mucoid.
Some of the most common Enterobacter species include E. cloacae, E. cancerogenous, and E. aerogenes, now known as Klebsiella aerogenes.
Some of the most common Enterobacter species include E. cloacae, E. cancerogenous, and E. aerogenes, now known as Klebsiella aerogenes.
Klebsiella spp is a Gram-negative bacilli and a lactose-fermenter that is oxidase -. Its mucoid appearance is due to the production of a slime capsule. Some strains produce ESBL (extended spectrum Beta-lactamase) and are MDRO's (multi-drug-resistant organisms). Klebsiella spp are indole - and motility -. On Blood Agar, colonies are white, shiny, mucoid, convex, smooth, glistening, and entire.
Some of the most common Klebsiella spp include K. pneumoniae, K. oxytoca, K. aerogenes, and K. variicola.
Some of the most common Klebsiella spp include K. pneumoniae, K. oxytoca, K. aerogenes, and K. variicola.
Proteus spp is associated with nosocomial UTI and this Gram-negative urease producer is a swarmer associated with production of struvite kidney stones. Proteus spp have a putrefactive odor like rotten chicken soup, though to some it smells like chocolate. P. mirabilis, P. vulgaris and P. penneri.
Serratia spp is associated with nosocomial UTI and is responsible for only about 2% of all UTI. After 48 hours, some strains produce a red pigmentation. S. marcescens is the most common.
Pseudomonas spp is an oxidase +, indole -, catalase +, Gram-negative bacilli that produces the pigment pyocyanin, which is a blue-green pigment that produces a metallic sheen on blood agar and gives this bacterium a fruity, grape-like scent. It is a fairly common cause of UTI. P. aeruginosa, P. fluorescens, P. alcaligenes, P. flavescens and P. stutzeri are the most common.
Acinetobacter spp is a non-lactose-fermenting, oxidase-, Gram-negative bacillus that is a rare cause of UTI. It is often a MDRO. It is nosocomial-acquired and usually associated with catheters. On Blood Agar, Acinetobacter spp produce colonies that are white, glistening, smooth, moist and viscid with an odor like dirty gym socks. A. baumannii is the most common strain.
This NLF Gram-negative bacilli is a rare cause of UTI.
This NLF Gram-negative bacilli is a rare cause of UTI. P. stuartii and P. rettgeri are the most common.
This NLF Gram-negative bacilli is a rare cause of UTI. Hafnia alvei is the most common strain.
This NLF Gram-negative bacilli is a rare cause of UTI. It produces large, smooth, glistening colonies with uneven edges, and a lavender-green to light purple pigment with a greenish discoloration underneath. Colonies produce an ammonia-like odor. They are oxidease -, maltose +, glucose/dextrose +, and multi-drug resistant. It is an NLF on MAC agar, bile esculin variable, lysine decarboxylase +, and intrinsically resistant to most beta-lactams and aminoglycosides and quinolones. It is a frequent colonizer of the skin and respiratory tracts of nosocomial patients. Infections typicaly involve debilitated patients who are hospitalized, in burn units, in the ICU, have undergone medical procedures or medical instrumentation, or those that have received multple antimicrobial agents. It is found in moist hospital environments. Most infections are nosocomial and may include catheter-related UTI, other IV, bacteremia, wound infections, pneumonia, and miscellaneous infections in other parts of the body. S. maltophilia is the most common strain.
This NLF Gram-negative bacilli is a rare cause of UTI.
Raoultella (Klebsiella) planticola is a NLF, Gram-negative bacilli that is oxidase +, nonmotile, a facultative anaerobe, mucoid, encapsulated, and a member of the Enterobacteriaceae family. This bacterium is found in the environment in soil, water, and on plant material. It smells like AMMONIA. There are 3 species of Raoultella:
1) R. planticola-Associated with UTI, pancreatitis
2) R. ornitholytica
3) R. terrigena-Does not produce bacterial disease in humans, however, it is linked to melamine toxicity and converts it to cyanuric acid, which may cause kidney damage
1) R. planticola-Associated with UTI, pancreatitis
2) R. ornitholytica
3) R. terrigena-Does not produce bacterial disease in humans, however, it is linked to melamine toxicity and converts it to cyanuric acid, which may cause kidney damage
Pantoea agglomerans is an Enterobacteriaceae that can cause UTI. It is LF on MacConkey agar and is a GNR. On blood agar, this microbe is orange-to-yellow-pigmented, mucoid, and it is motile. Infections are associated with catheters and trauma. It can cause opportunistic infections, particularly in immunocompromised individuals. It has caused bloodstream infections, endocarditis, UTI, periostitis, osteomyelitis, endophthalmitis in patients with interstitial lung disease, septic arthritis, and widespread infection. It was previously known as Erwinia and as Enterobacter agglomerans.
Aerococcus urinae is a Gram-positive microbe that shows alpha-hemolysis on blood agar and microscopically is often seen in tetrads and pairs and clusters. It is associated with UTI's and has also caused other infections, including bloodstream and infective endocarditis. It is catalase-negative. It is found in the air, in dust, in water and can be an environmental contaminant or it can cause invasive infection because it has the ability to form biofilms, thus causing opportunistic infections in susceptible individuals.
Pseudomonas luteola and stutzeri are GNR soil and water bacteria and rarely cause opportunistic but serious infections in humans. They are catalase and oxidase positive.
If a germ tube test is negative, an API test or Vitek ID should be performed to identify the type of yeast. The yeast should also be plated on a cornmeal agar with 4 small loop stabs in the first quadrant where the 2nd quadrant overlaps, and a coverslip placed over it, to enhance the production of pseudohyphae to aid in the identification process. To read the cornmeal agar, remove the stage clip and lower the stage. Take the lid off the cornmeal agar and set it on the flat stage. Adjust the 10x objective and bring it into focus. Lower the 20x objective over the coverslip and focus to observe for pseudohyphae and chlamydospores and other structures. Look around the coverslip edges and around the stab marks. To focus closer, you may look under the 40x objective lens as well. Make sure that the API or Vitek results and cornmeal agar results correlate.
Types of Infections and Clinical Manifestations:
-Asymptomatic:
-Urethritis: Infection of the urethra with dysuria (painful urination, difficulty, frequency); A common infection
-Cystitis: Infection of the bladder with dysuria, frequency, urgency, inflammation, bacteria, tenderness, pain, urine that may be grossly bloody, cloudy, or filled with pus and has a bad odor; localized infection with fever and sometimes chills
-Pyelonephritis: Kidney infection usually caused by bacterial infection with fever, flank pain, low back pain, frequency, urgency, dysuria, vomiting, diarrhea, chills, tachycardia, lower abdominal pain, possible bacteremia
-Glomerulonephritis: The same as pyelonephritis with blood in the urine and history of recent sore throat or Group A Streptococcal (S. pyogenes) infection
-Acute Urethral Syndrome: Young, sexually active women who are experiencing dysuria, pyuria, frequency, and urgency, but have <100,000K CFU/mL of bacteria on the urinary cultures
-Kidney Stones: Renal calculi or nephrolith; Formed in the kidneys by minerals found in the urine
-May cause a blockage/obstruction in the ureter
-Cause lower back/side pain (flank pain) that may radiate to the groin
-Renal colic
-Symptoms: pain, nausea, vomiting, fever, painful urination, chills, blood and pus in the urine
-May cause kidney dysfunction if severe enough
-May form in the bladder
-Calcium, struvite, uric acid, staghorn composition types
-Dehydration may play a role
-May need cytoscopy, shock wave ultrasonic therapy, medication or a stent
-Asymptomatic:
-Urethritis: Infection of the urethra with dysuria (painful urination, difficulty, frequency); A common infection
-Cystitis: Infection of the bladder with dysuria, frequency, urgency, inflammation, bacteria, tenderness, pain, urine that may be grossly bloody, cloudy, or filled with pus and has a bad odor; localized infection with fever and sometimes chills
-Pyelonephritis: Kidney infection usually caused by bacterial infection with fever, flank pain, low back pain, frequency, urgency, dysuria, vomiting, diarrhea, chills, tachycardia, lower abdominal pain, possible bacteremia
-Glomerulonephritis: The same as pyelonephritis with blood in the urine and history of recent sore throat or Group A Streptococcal (S. pyogenes) infection
-Acute Urethral Syndrome: Young, sexually active women who are experiencing dysuria, pyuria, frequency, and urgency, but have <100,000K CFU/mL of bacteria on the urinary cultures
-Kidney Stones: Renal calculi or nephrolith; Formed in the kidneys by minerals found in the urine
-May cause a blockage/obstruction in the ureter
-Cause lower back/side pain (flank pain) that may radiate to the groin
-Renal colic
-Symptoms: pain, nausea, vomiting, fever, painful urination, chills, blood and pus in the urine
-May cause kidney dysfunction if severe enough
-May form in the bladder
-Calcium, struvite, uric acid, staghorn composition types
-Dehydration may play a role
-May need cytoscopy, shock wave ultrasonic therapy, medication or a stent
Specimen Collection:
-Clean-catch, Midstream Urine
-Random Urine
-Catheterized Urine
-Suprapubic Bladder Aspiration
-Indwelling Catheter
-Kidney/Nephrostomy Tube Secretions (Percutaneous nephorostomy is a surgical procedure and radiological procedure performed in order to detect calcifications in the renal pelvis. The renal pelvis is purposefully punctured at the same time radiological imaging is performed with dye injection (contrast). An antegrade pyelogram is taken following injection of contrast dye into the renal pelvis with a fine needle. The nephrostomy tube is then placed in order to allow for drainage. Nephrostomy tubes are also placed when there is a blockage or obstruction between the kidneys and the urethra, such as when kidney stones are present. This allows for draining of the urine and to prevent stagnancy of the urine or kidney infection. Other than kidney stones, the most common cause of blockages and the need for placement of a nephrostomy tube is cancer, such as ovarian or colon cancer. Without the placement of the tube, pressure would otherwise build up and cause damage to the kidneys. Surgeons and/or radiologists pierce the skin and insert this special catheter, which stays in place in the urinary tract and empties into a bag on the outside, which is emptied whenever needed.)
Tests That May Indicate Bacterial UTI:
-Nitrate Reductase Test +
-Leukocyte Esterase Test +
-Urinalysis Showed Bacteria, Blood, Increased Protein, and/or Increased Specific Gravity
-Clean-catch, Midstream Urine
-Random Urine
-Catheterized Urine
-Suprapubic Bladder Aspiration
-Indwelling Catheter
-Kidney/Nephrostomy Tube Secretions (Percutaneous nephorostomy is a surgical procedure and radiological procedure performed in order to detect calcifications in the renal pelvis. The renal pelvis is purposefully punctured at the same time radiological imaging is performed with dye injection (contrast). An antegrade pyelogram is taken following injection of contrast dye into the renal pelvis with a fine needle. The nephrostomy tube is then placed in order to allow for drainage. Nephrostomy tubes are also placed when there is a blockage or obstruction between the kidneys and the urethra, such as when kidney stones are present. This allows for draining of the urine and to prevent stagnancy of the urine or kidney infection. Other than kidney stones, the most common cause of blockages and the need for placement of a nephrostomy tube is cancer, such as ovarian or colon cancer. Without the placement of the tube, pressure would otherwise build up and cause damage to the kidneys. Surgeons and/or radiologists pierce the skin and insert this special catheter, which stays in place in the urinary tract and empties into a bag on the outside, which is emptied whenever needed.)
Tests That May Indicate Bacterial UTI:
-Nitrate Reductase Test +
-Leukocyte Esterase Test +
-Urinalysis Showed Bacteria, Blood, Increased Protein, and/or Increased Specific Gravity
The lower urinary tract
the upper urinary tract
wounds bench, Including some anaerobes:
Skin flora and opportunistic pathogens:
Staphylococcus aureus and MRSA are part of the normal skin flora, but cause one of the most common opportunistic wound infections.
Coagulase negative staphylococci are also part of the normal skin flora are rarely cause opportunistic wound infections. These include:
Coagulase negative staphylococci are also part of the normal skin flora are rarely cause opportunistic wound infections. These include:
- S. epidermidis (associated with opportunistic infections, including catheter, surgical implants, pacemakers and artificial heart valves, nephrostomy tubes, dialysis equipment, plastic devices)
- S. saprophyticus
- S. lugdunensis
- S. simiae
- S. auricularis
- S. hominis (contributes to body odor)
- S. carnosus group: S. carnosus, S. condimenti, S. simulans, S. massiliensis, S. piscifermentans
- S. capitis, S. caprae, S. saccharolyticus
- S. haemolyticus
- S. warneri
- S. mutans
- S. mitis
- S. sanguinis
- S. sobrinus
clostridium perfringens:
staphylococcus aureus and mrsa:
Other types of Staphylococcus seen in wound cultures, mostly as part of the normal flora (Coagulase-negative staphylococci), include S. hominis, S. lugdunensis, S. haemolyticus, S. aeruginosa, S. werneri, S. capitis, S. epidermidis, S. saprophyticus, and others
Boils are small abscesses with superficial pustules filled with fluid pus. Furuncles are boils formed by deep folliculitis (hair follicle infections) resulting in pus-filled nodules that are tender, warm and painful to the touch. Recurring furuncles are referred to as furunculosis.
A carbuncle is an abscess that is larger than a furuncle or boil and usually contains at least one, if not more, openings to the skin to drain pus. This infection is caused by S. aureus, MRSA, or S. pyogenes. It is contagious and spreads easily on the body. A carbuncle is basically several skin boils combined and is most commonly found on the nape of the neck, ranging from pea-size to golf ball-size, as seen in the image above. Patients who are immunocompromised, diabetic, IV drug users, or anemic are at higher risk for developing this type of infection.
A stye is basically a bacterial boil either on the outside or on the inside of the eye (mucosa). S. aureus is the bacteria that most often causes this condition.
An intraoral dental sinus, or gum boil, is a boil on the gum caused by bacteria such as S. aureus or S. pyogenes.
Cellulitis is a bacterial infection of the skin spreading to the dermis and subcutaneous fat. It is usually caused by S. aureus or S. pyogenes.
Impetigo is a highly contagious bacterial skin infection caused by S. aureus, MRSA or S. pyogenes. It may present as a non-bullous form with honey-colored scabbing as sebum dries and heals over, a bullous form with larger sores and scabbing, or a blistering form, particularly in infants and more often caused by the bacteria S. pyogenes. It is most commonly seen in children in daycare, preschool, or of elementary school age.
Sebaceous cysts are fluid-filled pockets or infections of the sebaceous (oil-producing) glands. Sebum is found in these types of cysts, resulting in sebum and pus underneath the skin that is often movable. Cysts may be opened and drained or removed if they cause too much discomfort and trouble. They are commonly found on the nape of the neck.
Abscesses are superficial or deep skin localized pustules filled with fluid pus, resulting in redness, swelling, heat, pain and loss of function. Sometimes this includes fever and a general feeling of unwellness. An abscess is basically a defense mechanism of the immune system intended to wall-off the bacterial infection so that it does not spread any further. Bacteria that slip through this barricade into the bloodstream may result in bacteremia, spread of infection, seeding to other areas of the body, and even septicemia/sepsis. Common abscesses include complications due to folliculitis or boils, incisional abscesses (post-surgical), or perianal abscesses (anal fistulas). Pus is a collection of dead bateria, cellular debris, fluid and white blood cells of the immune system.
A JP Drain is a closed suction device used as a post-operative drain to drain fluids and/or abscesses following various surgeries.
Staphylococcus epidermidis:
staphylococcus saprophyticus:
staphylococcus lugdunensis:
Staphylococcus lugdunensis produces colonies that are typically beta-hemolytic, stick to the agar, and produce a yellow-to-tan pigment. Colonies are 2–4 mm in diameter after a 48-hour incubation period. They also can have a characteristic sweet odor that smells like fresh hay and is "earthy". S. lugdunensis is slide coagulase negative most of the time, however, it may be tube coagulase + and is PYR +. It is associated with a variety of infections, but as far as wound infections, has been known to cause furuncles, abscesses and even cellulitis, skin and soft tissue infections, and its most common infection is of the toe(s). It can progress to joint and bone infections as well. This is part of the commensal flora of the perianal region and has been associated with genital abscesses. In kids, this organism has been associated with otitis media. In middle aged individuals, it has been associated with infections of the groin, armpits, buttocks, and incisional sites, and has been associated with mastitis of the breast. In elderly persons and diabetic patients, the most common infections associated with this organism are wound infections of the toes, fingers, and leg ulcers. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2668335/.
enterococcus faecalis and enterococcus faecium:
Enterococcus faecalis and faecium are part of the normal gastrointestinal flora. They are nonhemolytic and unique because they grow well in 40% bile and 6.5% NaCl. They are associated with UTI, biliary tract infections, bacteremia, and subacute bacterial endocarditis. They are the second most common cause of nosocomial infections in the US. Most are resistant to Penicillin, Ampicillin and some are resistant to Vancomycin as well. It is very important to identify individual members of enterococci of the group D streptococci because enterococci often show more antibiotic resistance than other group D streptococci. They are catalase negative, PYR + and BEA +. They appear as gray-white and shiny/flat colonies with a narrow zone of alpha hemolysis and blood agar.
Pasteurella multocida and spp:
Pasteurella multocida is the most common cause of Pasteurella infection in humans, however, other species of Pasteurella can cause skin and soft tissue infections. For example, P. canis is also linked to serious disease in immunocompromised patients following a domestic animal bite by a cat, dog or rabbit. P. multocida and P. canis are carried in the mouths of dogs and cats and rabbits and are indole +, oxidase +, catalase +, urease -, and ornithine decarboxylase +. The Pasteurellae spp are Gram-negative coccobacilli that exhibit bipolar staining. On blood agar, Pasteurella grows as smooth, circular, gray, small non-hemolytic colonies. Pasteurella spp are nonmotile, and P. multocida ferments dextrose/glucose, maltose and sucrose in the sugars test. It is nonmotile and it does not grow on the MacConkey agar. P. pneumotropica is urease +, a differentiating factor between it and P. multocida, which is urease -. Some stains of Pasteurella take 48 hours to show up oxidase +. For this reason, when testing for oxidase activity, it is recommended that a 5% aqueous solution of tetramethyl-phenylenediamine dihydrochloride be used rather than commercially available strips, such as the dryslide.
This bacteria is never part of the normal human flora and is transmitted to humans through domestic pet bites, scratches, or licking of wounds by the pet. Immunocompromised patients are at higher risk of infection, particularly those with underlying illnesses and conditions such as diabetes. Wound, skin, soft tissue infections, peritonitis, conjunctivitis, and even arthritis and joint and bone infections may be caused by this bacteria. Other Pasteurella spp that cause these infections include P. anatis, P. dagmatis, P. langaa, P. pneumotropica (urease +), and P. stomatis.
Pasteurellosis typically exhibits itself fairly quickly, even within 1-2 hours after a bite or scratch. Symptoms are cutaneous and include inflammation, swelling, edema, redness and tenderness with severe pain and even a serosanguineous exudate discharge. This is typically indicated by leukocytosis and an elevated neutrophil count. Illness may occur with headache, fever, vomiting, diarrhea and swollen lymph nodes. If the bacteria spreads, it can lead to bacteremia and septic arthritis. Rarely, fulminant sepsis may manifest, with high fever and chills, vomiting and shock. Some patients also have bilateral pneumonia. Rarely, endocarditis or meningitis may be complications of the disease. Infection may be polymicrobial.
OTHER BACTERIA CARRIED IN THE MOUTHS OF CATS (NOTE: MOST CAT BITE INFECTIONS ARE POLYMICROBIAL):
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3122494/
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3122494/
This bacteria is never part of the normal human flora and is transmitted to humans through domestic pet bites, scratches, or licking of wounds by the pet. Immunocompromised patients are at higher risk of infection, particularly those with underlying illnesses and conditions such as diabetes. Wound, skin, soft tissue infections, peritonitis, conjunctivitis, and even arthritis and joint and bone infections may be caused by this bacteria. Other Pasteurella spp that cause these infections include P. anatis, P. dagmatis, P. langaa, P. pneumotropica (urease +), and P. stomatis.
Pasteurellosis typically exhibits itself fairly quickly, even within 1-2 hours after a bite or scratch. Symptoms are cutaneous and include inflammation, swelling, edema, redness and tenderness with severe pain and even a serosanguineous exudate discharge. This is typically indicated by leukocytosis and an elevated neutrophil count. Illness may occur with headache, fever, vomiting, diarrhea and swollen lymph nodes. If the bacteria spreads, it can lead to bacteremia and septic arthritis. Rarely, fulminant sepsis may manifest, with high fever and chills, vomiting and shock. Some patients also have bilateral pneumonia. Rarely, endocarditis or meningitis may be complications of the disease. Infection may be polymicrobial.
OTHER BACTERIA CARRIED IN THE MOUTHS OF CATS (NOTE: MOST CAT BITE INFECTIONS ARE POLYMICROBIAL):
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3122494/
- Pasteurella multocida (#1 cause)
- Other Pasteurella spp
- Streptococcus spp, particularly S. mitis (#2 cause), S. sanguis, S. intermedius, S. constellatus, S. agalactiae
- Group F streptococci
- Staphylococci (#3 cause), particularly S. epidermidis and S. warneri
- Neisseria spp (#4 cause), particularly N. weaveri and N. zoodegmatis
- Moraxella catarrhalis (#5 cause)
- Corynebacterium
- Enterococcus durans and faecalis
- Bacillus firmus and B. circulans
- Bergeyella zoohelcum
- Capnocytophaga
- Acinetobacter
- Pseudomonas
- Actinomyces
- Brevibacterium
- Gemella morbillorum
- Actinobacillus
- Alcaligenes
- Enterobacter cloacae
- Erysipelothrix rhusopathiae
- Rothia
- Riemerella
- Klebsiella oxytoca
- Lactobacillus spp
- Eikenella corrodens
- Flavimonas
- Aeromonas hydrophila
- Pantoea
- Rhodococcus
- Streptomyces
- Haemophilus felis
- Bartonella henselae
- Fusobacterium nucleatum (#1 anaerobe cause)
- Porphyromonas
- Bacteroides
- Prevotella
- Proprionibacterium
- Peptostreptococcus
- Filifactor
- Eubacterium
- Clostridium
- Veillonella
- Francisella tularensis (RARE in wild cats)
- Yersinia pestis (RARE in wild cats)
- Sporothrix schenckii
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3122494/
- Bacteroides
- Porphyromonas
- Pasteurella spp (#1 cause)
- Streptococcus (#2 cause)
- Staphylococcus (#3 cause)
- Neisseria (#4 cause)
- Corynebacterium
- Moraxella
- Enterococcus
- Bacillus
- Pseudomonas
- Actinomyces
- Brevibacterium
- Gemella
- E. coli
- Weeksella
- Klebsiella
- Lactobacillus
- Citrobacter
- Flavobacterium
- Micrococcus
- Proteus
- Stenotrophomonas
- Capnocytophaga
- Eikenella corrodens
- Flavomonas
- Dermabacter
- Oerskovia
- Pediococcus
- Stomatococcus
- Fusobacterium nucleatum (#1 anaerobe cause)
- Porphyromonas
- Bacteroides
- Prevotella
- Propionibacterium
- Peptostreptococcus
- Blastomyces
- Bordetella bronchiseptica
staphylococcus intermedius:
bordetella bronchiseptica:
Bordetella bronchiseptica is associated on occasion with infected dog bite wounds in humans or other infections, such as rare respiratory infections in patients who are immunocompromised. This organism is a small, Gram-negative coccobacillus that produces small, raised, beta-hemolytic colonies on blood agar and colorless, non-lactose-fermenting colonies on MacConkey agar. It is motility + (unlike B. parapertussis, which is nonmotile), catalase +, oxidase +, citrate +, nitrate +, and rapid urease +, often within 4 hours. It does not ferment any carbohydrates. Because this organism is part of the normal flora of dogs, and produces Kennel Cough, a history of close contact with a domestic pet or working closely with animals may predispose an individual to a higher risk of becoming infected with the organism.
Bartonella henselae:
Bartonella henslae and Bartonella clarridgeiae are short, Gram-negative bacilli capable of causing "Cat Scratch Fever". These bacterium are able to grow on CHOC agar, but not on blood agar, and they are fastidious organisms. Cat Scratch Fever is caused by cat scratches, bites, or flea bites that transmit the bacteria to humans. These oxidase negative bacilli are also causative agents of bacterema, endocarditis, bacillary angiomatosis (immunocompromised patients), and peliosis hepatitis (immunocompromised patients). There are approximately 24,000 cases per year in the USA alone, and 80% of these cases are children. Illness begins with a painless papule or pustule and the incubation period is 1-7 weeks. Typically, Cat Scratch Fever is a chronic but self-limiting disease resulting in adenopathy to severe, systemic illness affecting multiple organs and may include eye infection and severe muscle pain. It usually takes awhile to properly diagnose. Symptoms range from gastrointestinal illness with on-and-off recurring fevers, chills, bronchitis, and/or enlarged liver/spleen.
Bartonella quintana causes trench fever, relapsing fever, bacteremia, endocarditis, and chronic lymphadenopathy in humans. It is carried by the body louse and causes flu-like symptoms with headache, bone pain, rash, and splenomegaly.
Bartonella bacilliformis is the causative agent of Carrion's Disease, and is carried by sandflies. It manifests as recurring fevers with headaches, severe muscle pain, abdominal pain and severe anemia. Later, nodular lesions appear on the skin (red-purple) and may bleed or ulcerate.
Bartonella elizabethae causes endocarditis.
Afipia felis is an oxidase +, motility +, nonfermentive, fastidious, Gram-negative bacilli.
Bartonella quintana causes trench fever, relapsing fever, bacteremia, endocarditis, and chronic lymphadenopathy in humans. It is carried by the body louse and causes flu-like symptoms with headache, bone pain, rash, and splenomegaly.
Bartonella bacilliformis is the causative agent of Carrion's Disease, and is carried by sandflies. It manifests as recurring fevers with headaches, severe muscle pain, abdominal pain and severe anemia. Later, nodular lesions appear on the skin (red-purple) and may bleed or ulcerate.
Bartonella elizabethae causes endocarditis.
Afipia felis is an oxidase +, motility +, nonfermentive, fastidious, Gram-negative bacilli.
streptobacillus moniliformis:
Streptobacillus moniliformis is a small, twisted Gram-negative bacilli that may chain and is the causative agent of "rat bite fever", or Haverhill Fever. It is microaerophilic.
Spirillum minus is also a spiral-shaped bacterium that can cause "rat bite fever". Having a pet rat increases one's chance of becoming infected with these two organisms, and scratches, even minor ones, by pet rats carrying this organism may result in human infection if the conditions are just right. This organism causes a disease called sodoku, a form of rat-bite fever, seen mainly in Asia. It is a zoonotic pathogen. Bites or scratches by infected rats or mice result in inflammation and ulcerations. Symptoms include fever that continues to recur, swollen lymph nodes, fatigue, headache, and may progress to severe complications such as endocarditis, myocarditis, meningitis, hepatosplenomegaly, or even death (anywhere from 6-10%).
capnocytophaga:
Capnocytophaga canimorsus is a Gram-negative bacillus found in the mouths of animals, such as dogs, and is associated with infection from dog bites.
pantoea agglomerans:
Streptococcus pyogenes:
Necrotizing fasciitis is a flesh-eating bacteria syndrome resulting from infection by certain toxic strains of S. pyogenes, S. aureus, MRSA, C. perfringens, B. fragilis, V. vulnificus, A. hydrophila). This is caused mostly by S. pyogenes, or Group A Streptococci, a "flesh-eating" strain that has M proteins that block phagocytosis. This allows the bacteria to quickly move through tissues, causing damage and tissue death (necrosis). The streptococci enter through a break in the skin (trauma, a cut, a pimple, etc...) and follow the fascia (the protective covering overlying muscles but underneaththe subcutaneous layers of skin and fat). Very quickly, within just one day, a patient will exhibit swelling, fever, heat, pain, redness that moves quickly from the initial infection site. Eating away at the tissues, the skin color changes from a healthy pink to red to purple to blue to black, and large blisters form, called "bullae". Upon initial skin death, the muscles may also become infected as well (myositis). The damaged tissues have to be removed surgically and possible amputation may need to be performed. Quick diagnosis and antibiotic therapy is crucial. The mortality rate with this disease is up to 50% or more, even with treatment.
M protein is a major virulence factor for GAS because it blocks the activation of immune system complement and protects the bacteria from being destroyed or removed by phagocytosis. The bacterium also possesses streptolysins O and S. Streptolysin O is inactivated by oxygen, and it destroys red blood cells, white blood cells, and is responsible for the beta-hemolysis on blood agar, and it is antigenic. ASO titers are an immunological test that can be performed in the laboratory to detect or confirm recent infection. Streptolysin S is oxygen stable, is not antigenic, and also contributes to beta-hemolysis on the plate and destruction of red and white blood cells in the body. Streptokinase enzyme breaks up fibrin clots, enabling the flesh-eating bacteria to quickly travel. Other enzymes that bore through the tissues include hyaluronidsease, DNAase, anti-C5a peptidase.
M protein is a major virulence factor for GAS because it blocks the activation of immune system complement and protects the bacteria from being destroyed or removed by phagocytosis. The bacterium also possesses streptolysins O and S. Streptolysin O is inactivated by oxygen, and it destroys red blood cells, white blood cells, and is responsible for the beta-hemolysis on blood agar, and it is antigenic. ASO titers are an immunological test that can be performed in the laboratory to detect or confirm recent infection. Streptolysin S is oxygen stable, is not antigenic, and also contributes to beta-hemolysis on the plate and destruction of red and white blood cells in the body. Streptokinase enzyme breaks up fibrin clots, enabling the flesh-eating bacteria to quickly travel. Other enzymes that bore through the tissues include hyaluronidsease, DNAase, anti-C5a peptidase.
eikenella corrodens:
Human bite wounds: The human oropharyngea consists of a variety of bacteria that can cause opportunistic wound infections if the skin is broken during a bite by the teeth. In fact, it is one of the most commonly encountered bacterial infections encountered from bites or clenched fist wounds. Typically, these infections are polymicrobial. Bacteria that can cause human bite wound infections include the following: Eikenella corrodens. It is a fastidious organism (Gram-negative coccobacillus) that smells like bleach and characteristically "pits" the agar about half of the time. Small gray colonies with a greenish discoloration of the agar are characteristic of this organism. It is oxidase +, indole -, catalase -, and urease -. It does hydrolyze both ornithine and lysine. Immunocompromised patients are more at risk for infection, and blood cultures may come up positive. It is one of the HACEK bacteria occasionally associated with subacute bacterial endocarditis. This organism Gram-stains as a medium-length, straight Gram-negative rod that may be vacuolated and pale-staining, and resist decolorization. It does not grow very well on either sheep's blood or chocolate agars.
A certain percentage of cats and dogs are also known to carry this bacterium in their mouths, and cat or dog bites may also lead to polymicrobial infection with this organism and others. This organism is usually susceptible to the penicillins and the beta-lactam antimicrobials.
A certain percentage of cats and dogs are also known to carry this bacterium in their mouths, and cat or dog bites may also lead to polymicrobial infection with this organism and others. This organism is usually susceptible to the penicillins and the beta-lactam antimicrobials.
bergeyella spp:
Bergeyella zoohelcum is a rare zoonotic pathogen that is linked to dog or cat bites or scratches and can cause infections in humans following a bite. In fact, it has been linked to acute cellulitis. It is carried in the mouths and saliva of a large percentage of dogs and a moderate percentage of cats and other animals. This Gram-negative rod stains as short, straigh rods that may form "II-forms" like the Sphingobacterium, and it is fastidious and it is difficult to ID it and perform susceptibility by traditional methods, so PCR is a great definitive method for ID and susceptibility. Infections have been underreported. It grows well on blood agar, but not on MacConkey agar. It is oxidase +, indole +, catalase +, urease +, and nonmotile and asaccharolytic. It is an aerobic bacterium that has been categorized as part of the Flavobacterium. Colonies are tan-to-yellow in color and sticky, and typically slow-growers. This organism is biochemically similar to Weeksella. It also grows slowly in broth media. In order to detect growth, plates must be incubated at 35-37 degrees Celsius in CO2 for a minimum of 48 hours.
weeksella virosa:
Weeksella virosa is biochemically similar to Bergeyella zoohelcum. It is asaccharolytic, oxidase +, catalase +, and does not grow on MacConkey agar. It is a short, straight Gram-negative rod that may form "II-forms" like Sphingobacterium. It grows slowly on blood and CHOC agars, and in broth medium. It is an environmental bacteriumno not part of the normal human flora, and the mode of transmission is uncertain, as it is rarely found in clinical material. Its role in human disease is uncertain, but it may be linked to rare cases of asymptomatic bacteruria and is rarely isolated from the female genital tract. At 24 hours, colonies are small and once mature, they are mucoid and adherent with a tan-to-brown pigmentation. It is urease - and indole + and will grow on selective media for N. gonorrhoeae (MTM, ML agars). It does not hydrolyze ornithine or lysine.
aeromonas hydrophilia:
Aeromonas hydrophilia is an oxidase +, beta-hemolytic, Gram-negative bacilli found in fresh or stagnant water, and sometimes marine water. It is a facultative anaerobe, and it is linked to wound infections, particularly involving cuts of the feet or extremeties with exposure to contaminated water (fish tanks, the ocean, lakes, rivers, ponds, streams, etc...). They are Gram-negative rods that are straight with rounded ends, and are motile. In addition to causing a diarrheal illness, it can also cause wound infections, cellulitis, and necrotizing fasciitis and produces a cytotoxin enterotoxin known as aerolysin.
pseudomonas aeruginosa:
Burns: Burns are staged as 1st degree, 2nd degree, 3rd degree or 4th degree.
Decubitis: This is the official term for "pressure sore" or "bed sore" or "pressure ulcer". Ulcers are breaks in the skin with infection and pus and are also referred to as open skin or mucosal craters. Erythema nodosum is another term for a skin ulcer. Types of ulcers include the following:
- Serous (healing)
- Purulent (thick, green-yellow pus)
- Bloody
- Seropurulent (bloody and purulent)
- Serosanguinous (bloody, purulent but beginning to heal)
- Sulfur granules (seen in wounds of those with Actinomycosis)
Wound infections are most often caused by staphylococci, streptococci, or the Enterobacteriaceae, though a multitude of other organisms may cause infection, including things such as Eikenella (human bites), Bartonella (cat scratches), Pasteurella multocida (pet bites), Clostridium spp (tetanus, gas gangrene), and others...Wounds may be superficial, cutaneous, subcutaneous, deep, and may disseminate to deeper tissues, muscles, organs, bone, or bloodstream. Other organisms that can cause wound infections include Aeromonas, Plesiomonas, Mycobacterium spp, Serratia marcescens, Vibrio vulnificus, V. alginolyticus, Proteus, Pseudomonas, Streptomyces spp, Morganella morganii, S. lugdunensis, Acinetobacter spp, and others. Sometimes, mixed anaerobic infections occur.
There are many types of wounds that can become infected by bacteria:
There are many types of wounds that can become infected by bacteria:
- Abrasions/scratches
- Lacerations
- Cuts (Glass, Knives, Plastics, Other)
- Punctures (Stepping on a Nail, Staple Gun, Dart, Etc...)
- Blunt Force Trauma (Car Accidents, Falls, Etc...)
- Stabs
- Contusions (Closed Wounds)
- Bites (Human, Dog, Cat, Insects like Spiders, Snake, Bird, Other Animals)
- Tears (Skin Rips)
- Burns (1st, 2nd, 3rd, 4th degree)
- Rug Burns
- Avulsions
- Scrapes and scratches
- Penetrations (trauma)
- Gun Shot Wounds
- Splinters (wood, glass, plastic)
- Post-surgical and stitches
The skin is the first line of defense and protection. It is when this barrier is broken and bacteria is introduced that wound infections occur. The skin consists of the cutaneous layers (epidermis and dermis) and the underlying subcutaneous tissue (fats/lipids, blood supply, nerves, connective tissue). Once bacteria gets into the bloodstream, it can spread and seed to other areas, causing bacteremia, sepsis, soft tissue, muscle, bone and joint infections.
anaerobes bench (including facultative anaerobes and aerotolerant anaerobes):
Anaerobic bacteria are clinically significant because they are linked to serious, sometimes fatal, infections and they cause a wide variety of infections in humans, ranging from appendicitis, to cholecystitis (gallstones), otitis media (inner ear infection), oral and dental infections (Fusobacterium is a common cause, and Actinomycetes is a common cause), endometritis, endocarditis, brain and liver abscesses, myonecrosis (infection of the muscle), osteomyelitis (infection of the bone), peritonitis, empyema, salpingitis (infection of the ovaries), septic arthritis, sinusitis, post-surgical wound infections, especially following bowel surgery or trauma, perirectal abscess, tuboovarian abscess, bacteremia and sepsis. Basically anything internally, inside the body, may be infected by an anaerobe if the conditions are just right.
Anaerobic isolation may be easily missed if specimens are not properly collected and transported to the laboratory ASAP. Even a brief exposure to oxygen (10 minutes) is enough to kill and abolish some anaerobic organisms. Most anaerobic isolates require vitamin K and hemin for ideal growth. The clinical significance of this is that knowing which anaerobes have been isolated by the laboratory is critical in aiding physicians to diagnose diseases and treat them.
The BEST specimen for anaerobic isolation is an aspirate (pus, deep wounds, abscesses) obtained via needle and syringe, with the needle removed and the syringe capped. Tissue samples and biopsies or chunks of bone are also very good specimens for anaerobic culture. If a swab must be used to collect a specimen, ideally an anaerobe swab system should be used (the Port-A-Cult tube). Large volumes of purulent material and large chunks of tissue will maintain viability of anaerobes for many hours as long as extremes of heat or cold. If there is a delay, then specimens should be held at room temperature until processing unless it is >2 hours, at which point specimens should be refrigerated. Port-a-cults that have been exposed to oxygen will develop a purple color to them.
FACTORS LEADING TO ANAEROBIC INFECTIONS:
INDICATIONS OF POSSIBLE ANEROBIC INFECTION:
Anaerobic Media:
ANABAP (CDC Anaerobic Blood Agar): Recovery of all organisms
ANAPEA (CDC Anaerobic PEA Agar): Gram-Positive organisms
ANAKV (CDC Anaerobic KV Agar): Gram-Negative organisms
THIO Broth (Thioglycollate)
Aerobic/Oxygen Challenge: SBA or TSA, CHOC, MAC, CNA
Inoculate the media and place in an anaerobe pouch with a generator, tightly sealed and placed in a CO2 incubator. DO NOT open the anaerobe pouch until 48 hours. Cultures shouldn't be exposed to oxygen until after 48 hours incubation. Anaerobes are most sensitive to oxygen during their log phase.
THIO broths should be checked for cloudiness or turbidity, indicating growth. This should be Gram-stained and set up aerobically and anaerobically.
Anaerobic isolation may be easily missed if specimens are not properly collected and transported to the laboratory ASAP. Even a brief exposure to oxygen (10 minutes) is enough to kill and abolish some anaerobic organisms. Most anaerobic isolates require vitamin K and hemin for ideal growth. The clinical significance of this is that knowing which anaerobes have been isolated by the laboratory is critical in aiding physicians to diagnose diseases and treat them.
The BEST specimen for anaerobic isolation is an aspirate (pus, deep wounds, abscesses) obtained via needle and syringe, with the needle removed and the syringe capped. Tissue samples and biopsies or chunks of bone are also very good specimens for anaerobic culture. If a swab must be used to collect a specimen, ideally an anaerobe swab system should be used (the Port-A-Cult tube). Large volumes of purulent material and large chunks of tissue will maintain viability of anaerobes for many hours as long as extremes of heat or cold. If there is a delay, then specimens should be held at room temperature until processing unless it is >2 hours, at which point specimens should be refrigerated. Port-a-cults that have been exposed to oxygen will develop a purple color to them.
FACTORS LEADING TO ANAEROBIC INFECTIONS:
- Trauma to the mucous membranes, skin, vascular system
- Decreased oxygen to the tissues leading to necrosis
- Production of virulence factors, including toxins, enzymes that break down the tissue, capsule formation, inhibition of phagocytosis, evasion of the immune system, adherence factors to aid in attachment to mucosal surfaces
INDICATIONS OF POSSIBLE ANEROBIC INFECTION:
- Foul odor
- Presence of a large amount of gas
- Black color
- Brick-red fluorescence
- Sulfur granules present
- High white blood cell count or many seen
- Infection is close to a mucosal surface
Anaerobic Media:
ANABAP (CDC Anaerobic Blood Agar): Recovery of all organisms
ANAPEA (CDC Anaerobic PEA Agar): Gram-Positive organisms
ANAKV (CDC Anaerobic KV Agar): Gram-Negative organisms
THIO Broth (Thioglycollate)
Aerobic/Oxygen Challenge: SBA or TSA, CHOC, MAC, CNA
Inoculate the media and place in an anaerobe pouch with a generator, tightly sealed and placed in a CO2 incubator. DO NOT open the anaerobe pouch until 48 hours. Cultures shouldn't be exposed to oxygen until after 48 hours incubation. Anaerobes are most sensitive to oxygen during their log phase.
THIO broths should be checked for cloudiness or turbidity, indicating growth. This should be Gram-stained and set up aerobically and anaerobically.
ANAEROBIC BACTERIA TO WORK UP:
GRAM POSITIVE COCCI:
Linked to abscesses, meningitis, aspiration pneumonia, gingivitis, periodontal disease, bacteremia, sepsis, and are usually part of polymicrobial infections
-Clostridium spp (spore-forming)
-C. perfringens (gas gangrene or food-poisoning)
-Food Poisoning Type A (mild, self-limited, GI illness with diarrhea and cramping 8-12 hours after exposure; lasts 24 hours), enterotoxin production linked to sporulation
-Food Poisoning Type C (more serious but rarely seen), production of B-toxin; causes severe diarrhea, which may be bloody, and severe cramping 5-6 hours after exposure; may include vomiting; small intestine necrotic inflammation; may be fatal without treatment
-Enterotoxin production in contaminated food (meats, gravies common, improperly stored/jars)
-Myonecrosis or gas gangrene is due to contamination of a wound through trauma or surgery and the other types of Clostridium spp can cause this but this strain is the most common cause due to production of exotoxins (A-toxin)
-Bacteremia and sepsis (other strains can cause but this strain is the most common)
-C. botulinum (food poisoning)
-Results from ingestion of preformed botulinum toxin (type A, B or E), a very potent neurotoxin that causes paralysis and death
-C. tetani (tetanus)
-Results from production of neurotoxin tetanospasmin
-C. bifermentans (when present in blood cultures, this is a marker for serious underlying disease)
-C. sordelii
-C. septicum (when present in blood cultures, this is a marker for GI tract malignancy)
-C. difficile (pseudomembranous colitis)
-C. tertium (when present in blood cultures, this is a marker for serious underlying disease)
-C. ramosum
-Actinomyces spp (linked to actinomycosis)
-A. israelii
-A. naeslundii
-A. neuvenii
-A. odontolyticus
-Bifidobacterium spp (GI tract flora) (Linked to peritonitis)
-Eubacterium spp (GI tract flora) (Linked to peritonitis)
-Lactobacillus spp (urogenital tract flora) (Usually normal, but occasionally linked to UTI or endocarditis, intraabdominal abscess, meningitis, conjunctivitis, or oral infections)
-Propionibacterium spp (indigenous microflora of the skin; colonizer of sebaceous glands and hair follicles; frequiently isolated from blood cultures, but usually a contaminant from the patient's skin, which means the area was not properly disinfected prior to collection)
-Opportunistic infections: endocarditis, surgical wound infections, acne
-Arcanobacterium spp
-Opportunistic infections: pharyngitis, pharyngeal abscess, Vincent's angina, endocarditis
-Eggerthella spp
-Mobiluncus spp (linked to bacterial vaginosus)
GRAM NEGATIVE COCCI:
-Veillonella spp (normal flora of intestines and mouth)
-Associated with osteomyelitis and endocarditis or abscesses (polymicrobial)
GRAM NEGATIVE BACILLI/RODS:
-Bacteroides spp (Beta lactamase positive or variable, catalase +, esculin +, bile +, beta hemolysis) (GI tract flora) (Infections: linked to peritonitis, bacteremia, sepsis, soft tissue, abscess)
-B. fragilis
-B. urolyticus
-Fusobacterium spp (Beta lactamase negative), Fusiform GNR's
-F. nucleatum (linked to aspiration pneumonia, abscesses, bacteremia, sepsis)
-F. necrophorum
-Infections: periodontal disease, skin ulcers, Lemierre's disease; fluorishes with colon cancer cells; associated with ulcerative colitis; biofilm formation; lipopolysaccharide capsule can cause it to hide from the immune system and be difficult to treat; always treat as pathogenic
-Lemierre's disease is a complication of a bacterial sore throat leading to a peritonsillar abscess resulting in infectious thrombophlebitis, and can lead to bactermia, sepsis and septic embolism; ear, jaw, sinus, or salivary gland infections can also cause this
-Symptoms: sore throat, fever, malaise, body aches, lethargy, chills, swollen lymph nodes, nausea, vomiting, diarrhea, stomach pain, possible pulmonary involvement including cough, pleurisy, pleural effusion, shortness of breath, coughing up blood, and possible joint pain; may lead to septic shock, meningitis, pericarditis, and serious, even fatal, complications
-Porphyromonas spp (oral flora) (linked to aspiration pneumonia, abscesses, bacteremia, sepsis)
-Prevotella spp (Oral, vaginal and gut flora) (linked to aspiration pneumonia)
-P. intermedia
-P. loescheii
-P. melanogenica (On plate, grows as colonies that become pigmented brown with age)
-Other
-Infections caused by this species include polymicrobial, respiratory, pneumonia (aspiration), lung abscess, pulmonary empyema, chronic ear infection, chronic sinusitis, abscesses, burns, mouth infections, bites, UTI, paronychia, brain abscess, osteomyelitis, bacteremia, periodontal infections and abscesses and diseases, including gingivitis periodontitis, and also rheumatoid arthritis
-Bilophila wadsworthia
-Borrelia burgdorferi
-Capnocytophaga spp
-Campylobacter spp
-Mobiluncus spp
-Leptotrichia spp
GRAM POSITIVE COCCI:
Linked to abscesses, meningitis, aspiration pneumonia, gingivitis, periodontal disease, bacteremia, sepsis, and are usually part of polymicrobial infections
- Peptococcus niger (flora of mouth, GI tract, upper resp. tract)
- Mezlocillin is treatment for infections with this organism
- Finegoldia magna (flora of skin and mucous membranes) (Renamed from Peptostreptococcus magna) (Most pathogenic of the anaerobic cocci)
- Assoiated with biofilms on chronic ulcers, diabetic foot ulcers, and decubitis ulcers
- Virulent
- Tends to be resistant to Clindamycin
- Tends to be susceptible to Penicillins, Carbapenems, Metronidazole
- Named after American Microbiologist Sydney Finegold
- Magna is Latin for "large" (GPC's)
- On plate, grows as tiny pinpoint glistening clear colonies
- Schleiferella (Peptoniphilus) (flora of vagina and gut)
- Associated with diabetic skin and soft tissue infections, bone and joint infections, surgical site, blood infections and chorioamnionitis
- Tend to be involved in polymicrobial infections
- Peptostreptococcus spp (Some Renamed Peptoniphilus and Finegoldia magna, formerly P. magnus) (flora of lower reproductive tract of women and of the mouth, skin, lower GI, and urinary tract)
- Opportunistic infections in immunocompromised individuals or have traumatic injuries
- Can cause sepsis and abscesses of the brain, liver, breast or lung or necrotizing soft tissue infections
- Tend to be involved in polymicrobial infections
- Susceptible to beta lactams
- F. magna is associated with biofilm formation in ulcers such as diabetic ulcers and is named after American microbiologist Sydney Finegold
- Gemella spp (flora of mucus membranes)
- Associated with pulmonary infections of patients with CF, endocarditis and bacterial vaginosis
- Susceptible to penicillin-gentamicin therapy in combination
- Anaerobic (facultative) diplococci
- Negative oxidase and catalase reactions
- Streptococcus spp
- Staphylococcus spp
- Sarcina
- Parvimonas micra (associated with dental plaque and chronic periodonitis, intraabdominal abscess, sepsis, septic arthritis, osteomyelitis and discitis, especially following dental procedures)
-Clostridium spp (spore-forming)
-C. perfringens (gas gangrene or food-poisoning)
-Food Poisoning Type A (mild, self-limited, GI illness with diarrhea and cramping 8-12 hours after exposure; lasts 24 hours), enterotoxin production linked to sporulation
-Food Poisoning Type C (more serious but rarely seen), production of B-toxin; causes severe diarrhea, which may be bloody, and severe cramping 5-6 hours after exposure; may include vomiting; small intestine necrotic inflammation; may be fatal without treatment
-Enterotoxin production in contaminated food (meats, gravies common, improperly stored/jars)
-Myonecrosis or gas gangrene is due to contamination of a wound through trauma or surgery and the other types of Clostridium spp can cause this but this strain is the most common cause due to production of exotoxins (A-toxin)
-Bacteremia and sepsis (other strains can cause but this strain is the most common)
-C. botulinum (food poisoning)
-Results from ingestion of preformed botulinum toxin (type A, B or E), a very potent neurotoxin that causes paralysis and death
-C. tetani (tetanus)
-Results from production of neurotoxin tetanospasmin
-C. bifermentans (when present in blood cultures, this is a marker for serious underlying disease)
-C. sordelii
-C. septicum (when present in blood cultures, this is a marker for GI tract malignancy)
-C. difficile (pseudomembranous colitis)
-C. tertium (when present in blood cultures, this is a marker for serious underlying disease)
-C. ramosum
-Actinomyces spp (linked to actinomycosis)
-A. israelii
-A. naeslundii
-A. neuvenii
-A. odontolyticus
-Bifidobacterium spp (GI tract flora) (Linked to peritonitis)
-Eubacterium spp (GI tract flora) (Linked to peritonitis)
-Lactobacillus spp (urogenital tract flora) (Usually normal, but occasionally linked to UTI or endocarditis, intraabdominal abscess, meningitis, conjunctivitis, or oral infections)
-Propionibacterium spp (indigenous microflora of the skin; colonizer of sebaceous glands and hair follicles; frequiently isolated from blood cultures, but usually a contaminant from the patient's skin, which means the area was not properly disinfected prior to collection)
-Opportunistic infections: endocarditis, surgical wound infections, acne
-Arcanobacterium spp
-Opportunistic infections: pharyngitis, pharyngeal abscess, Vincent's angina, endocarditis
-Eggerthella spp
-Mobiluncus spp (linked to bacterial vaginosus)
GRAM NEGATIVE COCCI:
-Veillonella spp (normal flora of intestines and mouth)
-Associated with osteomyelitis and endocarditis or abscesses (polymicrobial)
GRAM NEGATIVE BACILLI/RODS:
-Bacteroides spp (Beta lactamase positive or variable, catalase +, esculin +, bile +, beta hemolysis) (GI tract flora) (Infections: linked to peritonitis, bacteremia, sepsis, soft tissue, abscess)
-B. fragilis
-B. urolyticus
-Fusobacterium spp (Beta lactamase negative), Fusiform GNR's
-F. nucleatum (linked to aspiration pneumonia, abscesses, bacteremia, sepsis)
-F. necrophorum
-Infections: periodontal disease, skin ulcers, Lemierre's disease; fluorishes with colon cancer cells; associated with ulcerative colitis; biofilm formation; lipopolysaccharide capsule can cause it to hide from the immune system and be difficult to treat; always treat as pathogenic
-Lemierre's disease is a complication of a bacterial sore throat leading to a peritonsillar abscess resulting in infectious thrombophlebitis, and can lead to bactermia, sepsis and septic embolism; ear, jaw, sinus, or salivary gland infections can also cause this
-Symptoms: sore throat, fever, malaise, body aches, lethargy, chills, swollen lymph nodes, nausea, vomiting, diarrhea, stomach pain, possible pulmonary involvement including cough, pleurisy, pleural effusion, shortness of breath, coughing up blood, and possible joint pain; may lead to septic shock, meningitis, pericarditis, and serious, even fatal, complications
-Porphyromonas spp (oral flora) (linked to aspiration pneumonia, abscesses, bacteremia, sepsis)
-Prevotella spp (Oral, vaginal and gut flora) (linked to aspiration pneumonia)
-P. intermedia
-P. loescheii
-P. melanogenica (On plate, grows as colonies that become pigmented brown with age)
-Other
-Infections caused by this species include polymicrobial, respiratory, pneumonia (aspiration), lung abscess, pulmonary empyema, chronic ear infection, chronic sinusitis, abscesses, burns, mouth infections, bites, UTI, paronychia, brain abscess, osteomyelitis, bacteremia, periodontal infections and abscesses and diseases, including gingivitis periodontitis, and also rheumatoid arthritis
-Bilophila wadsworthia
-Borrelia burgdorferi
-Capnocytophaga spp
-Campylobacter spp
-Mobiluncus spp
-Leptotrichia spp
bacterial growth and division of anaerobes, most sensitive at their exponential/log phase (b):
Bacteria divide and replicate by a process called binary fission, where they double at an exponential rate. In this growth cycle, bacteria divide by asexual reproduction into 2 new daughter cells, which then each divide, and so forth and so on. There are 4 phases to the bacterial growth cycle:
A) Lag Phase, B) Exponential/Log Phase, C) Stationary Phase, and D) Death Phase. This is how the bacteria increase their population. We observe this growth by the observation of colony forming units (CFU's) growing on the agar medium. 1 colony = 1,000 CFU/mL with the 0.001 loop, and 1 colony = 100 CFU/mL with the 0.01 larger loop. Log refers to the numbers of bacterial dividing and Time refers to the amount of time it takes for all 4 phases to be completed, and for the bacteria to use up all the nutrients and die. The reason we subculture bacteria and make purity plates is for this reason, to keep the bacteria alive, dividing, and keep them from using up all the nutrients on the agar and completely dying. Anaerobic bacteria are most sensitive to oxygen during the log phase/exponential phase, and this is why they are kept in anaerobic conditions, unopened until the first 48 hours have passed, and why they must be observed/worked up within 30 minutes then put back into anaerobic conditions as soon as possible, to keep them alive and dividing.
Bacterial Growth Phases:
A) Lag Phase - During this phase, bacteria are getting used to or adapting to their conditions (pH, moisture, temperature, agar nutrients, oxygen or CO2, etc...) and preparing for division by synthesizing RNA, enzymes, molecules, and proteins needed for division.
B) Exponential/Log Phase-During this phase, the logarithmic phase, bacterial cells divide exponentially and double. The log, or number of new bacteria dividing and appearing per unit of time, is directly proportional or equal to the amount of bacteria currently present in the population. With unlimited nutrients and growth, this doubling will continue at a constant rate. As seen in the diagrams below, plotting this phase is indicated by a straight, sloping line on the graphs (B). This is all based on the growth conditions available to the bacteria at the time. This cannot continue forever, as nutrients in the agar will be used up and depleted eventually by the bacterial population. Changes in moisture and temperature levels also affect bacterial growth. Eventually, toxins and wastes also accumulate within the medium from bacterial metabolism.
C) Stationary Phase - During this phase, a stagnancy occurs in which one or more of the growth factors is limited, an essential nutrient is depleted, and wastes begin to accumulate and inhibit growth. During this phase, the growth rate and the death rate are equal. This is represented by a horizontal line on the graphs below.
D) Death Phase - During this phase, the number of bacteria that are dying exceeds the amount that are dividing and the nutrients become depleted. Also referred to the decline phase, the bacteria die. It is during this phase that certain bacteria undergo changes in order to survive harsh conditions, such as the formation of or freeing of spores in some species, or formation of bacterial biofilms to try to keep some of the bacteria alive for survival.
A) Lag Phase, B) Exponential/Log Phase, C) Stationary Phase, and D) Death Phase. This is how the bacteria increase their population. We observe this growth by the observation of colony forming units (CFU's) growing on the agar medium. 1 colony = 1,000 CFU/mL with the 0.001 loop, and 1 colony = 100 CFU/mL with the 0.01 larger loop. Log refers to the numbers of bacterial dividing and Time refers to the amount of time it takes for all 4 phases to be completed, and for the bacteria to use up all the nutrients and die. The reason we subculture bacteria and make purity plates is for this reason, to keep the bacteria alive, dividing, and keep them from using up all the nutrients on the agar and completely dying. Anaerobic bacteria are most sensitive to oxygen during the log phase/exponential phase, and this is why they are kept in anaerobic conditions, unopened until the first 48 hours have passed, and why they must be observed/worked up within 30 minutes then put back into anaerobic conditions as soon as possible, to keep them alive and dividing.
Bacterial Growth Phases:
A) Lag Phase - During this phase, bacteria are getting used to or adapting to their conditions (pH, moisture, temperature, agar nutrients, oxygen or CO2, etc...) and preparing for division by synthesizing RNA, enzymes, molecules, and proteins needed for division.
B) Exponential/Log Phase-During this phase, the logarithmic phase, bacterial cells divide exponentially and double. The log, or number of new bacteria dividing and appearing per unit of time, is directly proportional or equal to the amount of bacteria currently present in the population. With unlimited nutrients and growth, this doubling will continue at a constant rate. As seen in the diagrams below, plotting this phase is indicated by a straight, sloping line on the graphs (B). This is all based on the growth conditions available to the bacteria at the time. This cannot continue forever, as nutrients in the agar will be used up and depleted eventually by the bacterial population. Changes in moisture and temperature levels also affect bacterial growth. Eventually, toxins and wastes also accumulate within the medium from bacterial metabolism.
C) Stationary Phase - During this phase, a stagnancy occurs in which one or more of the growth factors is limited, an essential nutrient is depleted, and wastes begin to accumulate and inhibit growth. During this phase, the growth rate and the death rate are equal. This is represented by a horizontal line on the graphs below.
D) Death Phase - During this phase, the number of bacteria that are dying exceeds the amount that are dividing and the nutrients become depleted. Also referred to the decline phase, the bacteria die. It is during this phase that certain bacteria undergo changes in order to survive harsh conditions, such as the formation of or freeing of spores in some species, or formation of bacterial biofilms to try to keep some of the bacteria alive for survival.
THIOGLYCOLLATE BROTH and MICROBIAL GROWTH VERSUS OXYGEN CONCENTRATION:
- AEROBIC
- Oxygen is only able to penetrate a short distance into the tube. Obligate aerobes grow ONLY at the surface in the oxic (pink) zone where oxygen is found. They require oxygen for metabolic activity and they grow well in ambient air incubators.
- Oxygen is only able to penetrate a short distance into the tube. Obligate aerobes grow ONLY at the surface in the oxic (pink) zone where oxygen is found. They require oxygen for metabolic activity and they grow well in ambient air incubators.
- ANAEROBIC
- Anaerobes are sensitive to oxygen, so the only grow AWAY from the surface. Since anaerobes are poisoned by oxygen, they grow the farthest away from it.
- FACULTATIVE AEROBE
- Facultative aerobes are able to grow in the presence OR in the absence of oxygen, so they grow throughout the tube, but the better growth occurs near the surface since the microbes are able to respire, so you'll have a few more organisms in the oxic zone.
- Facultative aerobes are able to grow in the presence OR in the absence of oxygen, so they grow throughout the tube, but the better growth occurs near the surface since the microbes are able to respire, so you'll have a few more organisms in the oxic zone.
- MICROAEROPHILES
- Microaerophiles grow away from the most oxic (pink) area of the tube. They need just a microscopic amount of oxygen, so you'll find the most growth just below the oxic zone at the interface of the pink (at the bottom) where it meets the anoxic zone . These organisms require the O2 level to be reduced to 5% or less.
- Microaerophiles grow away from the most oxic (pink) area of the tube. They need just a microscopic amount of oxygen, so you'll find the most growth just below the oxic zone at the interface of the pink (at the bottom) where it meets the anoxic zone . These organisms require the O2 level to be reduced to 5% or less.
- AEROTOLERANT ANAEROBE
- Aerotolerant anaerobes grow throughout the tube as well, but they only ferment so you will not observe better growth near the surface. These are able to survive some oxygen exposure, but need to be put in an anaerobic environment for metabolic processes. Most clinically significant anaerobes fall into this category.
- Aerotolerant anaerobes grow throughout the tube as well, but they only ferment so you will not observe better growth near the surface. These are able to survive some oxygen exposure, but need to be put in an anaerobic environment for metabolic processes. Most clinically significant anaerobes fall into this category.
CAPNOPHILIC microbes will grow best when the CO2 concentration is 5-10% in a CO2 incubator.
PEPTOSTREPTOCOCCUS SPP/Peptoniphilus:
Peptostreptococcus spp is an anaerobic small Gram-positive cocci found singly, in pairs, and in chains. It grows slowly. The most commonly isolated Peptostreptococcus spp is P. magnus, followed by P. anaerobius. This organism is associated with abscesses, particularly of the lung, brain, liver and breast. It is also associated with necrotizing soft tissue infections. It is commonly isolated in polymicrobial infections. They are the 2nd most commonly isolated anaerobes in clinical infections after Bacteroides spp. They are considered to be clinically significant and are similar to Veillonella, which are Gram-negative anaerobic cocci, and microaerophilic streptococci, which are aerotolerant. Other Peptostreptococcus spp isolated include the following: P. asaccharolyticus, P. anaerobius, P. prevotii, P. micros, P. hydrogenalis, P. saccharolyticus, P. intermedius, P. tetradius, P. vaginalis.
Trauma, immunodeficiency, chemotherapy, cancer patients, immunosuppression, poor circulation and decreased blood supply, surgery, and underlying medical conditions put patients at higher risk of infection from this organism.
This bacterium is part of the normal flora of the mouth, vagina, and GI tract.
Peptostreptococcus anaerobius is inhibited by sodium polyanethol sulfonate (SPS). After 24 hours on KV-laked ANA agar, this organism form small, peaked, round colonies.
Trauma, immunodeficiency, chemotherapy, cancer patients, immunosuppression, poor circulation and decreased blood supply, surgery, and underlying medical conditions put patients at higher risk of infection from this organism.
This bacterium is part of the normal flora of the mouth, vagina, and GI tract.
Peptostreptococcus anaerobius is inhibited by sodium polyanethol sulfonate (SPS). After 24 hours on KV-laked ANA agar, this organism form small, peaked, round colonies.
PEPTOCOCCUS:
Peptococcus spp are anaerobic Gram-positive cocci in pairs, chains, singles and tetrads.
P. niger produces colonies that are tiny, black, convex, shiny, smooth, circular, and light gray to olive-green upon exposure to air.
P. anaerobius often chains and colonies are medium, gray-white, opaque, with a sweet and fetid odor. These produce larger colonies than most anaerobes.
P. asaccharolyticus colonies are small, gray and translucent, and this species is indole +. This species has been renamed to Peptoniphilus asaccharolyticus. See image below.
P. magnus are large GPCs in pairs and clusters like staphylococci, and colonies are tiny, gray, translucent and nonhemolytic. This species has been renamed to Finegoldia magnus. See image below.
P. micros are large GPCs in packets and short chains, and colonies are tiny, white, opaque and nonhemolytic.
P. niger produces colonies that are tiny, black, convex, shiny, smooth, circular, and light gray to olive-green upon exposure to air.
P. anaerobius often chains and colonies are medium, gray-white, opaque, with a sweet and fetid odor. These produce larger colonies than most anaerobes.
P. asaccharolyticus colonies are small, gray and translucent, and this species is indole +. This species has been renamed to Peptoniphilus asaccharolyticus. See image below.
P. magnus are large GPCs in pairs and clusters like staphylococci, and colonies are tiny, gray, translucent and nonhemolytic. This species has been renamed to Finegoldia magnus. See image below.
P. micros are large GPCs in packets and short chains, and colonies are tiny, white, opaque and nonhemolytic.
FINEGOLDIA SPP
Finegoldia magna (formerly Peptostreptococcus magnus) is part of the normal flora of the genitourinary and gastrointestinal tracts. Sometimes it is also found on the skin or in the oral cavity, and may be a contaminant in laboratory cultures. It is the most frequently isolated anaerobic cocci, and is implicated in infections that are either monomicrobial or polymicrobial and typically involve the skin, bone, joints, prosthetics. Other infections it has been linked to include infective endocarditis, necrotizing pneumonia, meningitis, and mediastinitis. http://www.ncbi.nlm.nih.gov/pubmed/22571938
SCHLEIFERELLA SPP (PEPTONIPHILUS):
Schleiferella spp (Peptoniphilus) is part of the normal vaginal microbiota and gut microflora. It is Gram-positive, anaerobic cocci implicated in bloodstream infections, pneumonia, preterm delivery, soft-tissue infections, colon and bladder disease. Most infections are polymicrobial. Diabetic patients are particularly at risk for skin, bone, joint and soft-tissue infections with this organism. This organism has also been recovered from the amniotic fluid causing choramnionitis resulting in pre-term labor and neonatal sepsis. It is particularly linked to bloodstream infections. It is also commonly seen in polymicrobial pelvic or abdominal abscesses.
DNA testing has shown a 99.4% similarity between the strains P. asaccharolyticus and P. harei, which may make identification difficult. The MALDI-TOF does not currently have this organism in its database for ID. P. harei can only be differentiated from P. asaccharolyticus by its irregular colonies and cellular morphology.
Most species are susceptible to penicillin, metronidazole, and clindamycin.
http://onlinelibrary.wiley.com/doi/10.1111/1469-0691.12657/pdf; http://jcm.asm.org/content/49/3/1189.full
DNA testing has shown a 99.4% similarity between the strains P. asaccharolyticus and P. harei, which may make identification difficult. The MALDI-TOF does not currently have this organism in its database for ID. P. harei can only be differentiated from P. asaccharolyticus by its irregular colonies and cellular morphology.
Most species are susceptible to penicillin, metronidazole, and clindamycin.
http://onlinelibrary.wiley.com/doi/10.1111/1469-0691.12657/pdf; http://jcm.asm.org/content/49/3/1189.full
Porphyromonas spp:
Porphyromonas spp are Gram-negative coccobacili that produce small, translucent or opaque to dark brown-to-black colonies on agar that are more mucoid than those of Prevotella spp. Colonies fluoresce brick red under UV light and are indole +. This bacterium is part of the normal flora of the oral cavity, GI tract, colon, respiratory tract, and vaginal tract. It has been linked to bacterial vaginosis, periodontal disease, rheumatoid arthritis, and other polymicrobial infections.
PREVOTELLA SPP:
Prevotella spp are Gram-negative rods and coccobacilli in pairs or short chains. They are inhibited by the presence of bile, are resistant to vancomycin, and are catalase -, esculin -, indole -, nitrate -, and glucose/dextrose +, lactose +, sucrose +. They inhabit the oral cavity and the vaginal tract. They are often associated with polymicrobial anaerobic respiratory infections, lung abscesses, aspiration pneumonia, sinusitis, middle ear infections, and pulmonary empyema. They are also associated with oral abscesses, brain abscesses, Bartholin's cyst abscesses and infections, bites, paronychia, osteomyelitis, bacteremia, urinary tract infections, periodontal diseases, and periodontal disease.
P. melaninogenica colonies are dark-centered with gray-to-light brown edges, circular, entire, convex, smooth and shiny, and nonhemolytic. Colonies fluoresce brick red under UV light and produce a black pigment on Laked KV anaerobic agar after 72 hours. This bacterium is part of the normal upper respiratory flora. It is often found in polymicrobial infections.
P. disiens colonies are white, circular, entire, convex, transparent, opaque, smooth, shiny and nonhemolytic. They fluoresce brick red under UV light and produce a black pigment on LKV agar.
P. intermedia colonies are similar to that of the other Prevotella spp, small, translucent, opaque and are indole + and lipase +. This bacterium is linked to periodontal disease, periodontal abscesses, gingivities, and bacterial vaginosis.
GEMELLA:
Gemella spp are tiny, pinpoint, alpha-hemolytic, Gram-positive, facultative anaerobic cocci. They are catalase - and oxidase -. G. haemolysans ferments glucose in anaerobic conditions. This organism should be ruled out in cystic fibrosis patients experiencing exacerbations of their pulmonary condition. G. morbillorum and G. sanguinis are associated with acute invasive disease and with bacterial endocarditis. Gemella palaticanis is found in the mouths of dogs and may be associated with polymicrobial bite infections. This bacterium is found in the mucous membranes of humans in the GI tract and the oral cavity.
SARCINA:
Sarcina spp are Gram-positive cocci in a cuboidal arrangement, or a packet of 8 large cocci. This is part of the normal commensal skin flora and and part of the GI flora.
CLOSTRIDIUM SPECIES:
Clostridium spp bacteria are strict (obligate) anaerobes that are Gram-positive bacilli, catalase negative, motility +, spore-forming, capable of producing endospores. Many strains produce harmful toxins that damage tissues and other structures within the body.
CLOSTRIDIUM PERFRINGENS: GAS GANGRENE
Clostridium perfringens is associated with both gas gangrene and can also cause food poisoning. It is also associated with cholecysitis, bacteremia, sepsis and tissue necrosis. The production of gas gangrene is due to the bacteria's production of a-toxin, an enterotoxin. Food poisoning may be caused by ingestion of foods contaminated with this microbe, and incubation time is just 6-24 hours.
This organism is catalase -, spot indole +, lecithinase +, grows on egg-yolk agar, lipase - and is capable of producing endospores. The organism produces turbidity in THIO broth and produces double zones of beta-hemolysis on blood agar. The Gram-stain shows Gram-variable straight rods with blunt ends seen in singles or in pairs. Spores are not readily seen, however, they are large, central to subterminal, oval, and swell the cell. C. perfringens bacilli are thick and have a "boxcar" shape to them.
This organism is catalase -, spot indole +, lecithinase +, grows on egg-yolk agar, lipase - and is capable of producing endospores. The organism produces turbidity in THIO broth and produces double zones of beta-hemolysis on blood agar. The Gram-stain shows Gram-variable straight rods with blunt ends seen in singles or in pairs. Spores are not readily seen, however, they are large, central to subterminal, oval, and swell the cell. C. perfringens bacilli are thick and have a "boxcar" shape to them.
CLOSTRIDIUM SEPTICUM:
This organism can also cause gas gangrene, and it is associated with colorectal cancer and peritoneal abscesses. It causes muscle and tissue necrosis by the release of exotoxins, including alpha toxin, hemolytic toxin and lethal toxin. This Gram-positive, spore-forming (subterminal spores), Gram-positive rod has a drumstick appearance. It is motility +.
CLOSTRIDIUM TETANI: TETANUS/LOCK JAW
Clostridium tetani is the causative agent of tetanus. It is found in the soil and its spores are hardy and capable of surviving extreme conditions. This strict anaerobe and Gram-positive bacilli often produces terminal endospores, causing it to appear like "spoons", "drumsticks" or "tennis rackets" microscopically. This species produces a potent toxin known as tetanospasmin, which results in painful muscle spasms and even respiratory failure, and another potent toxin called tetanolysin. These toxins interrupt the central nervous system neuron signals and interfere with the release of neurotransmitters. Up to 40% of cases of tetanus result in mortality. There is a vaccination that protects against this infection and it is part of the DTP vaccination. It is motility +, may appear to stain Gram-negative because it is easily decolorized.
CLOSTRIDIUM TERTIUM:
Clostridium tertium is a rare anaerobe and spore-forming Gram-positive bacilli that appears "spoon-shaped" due to the terminal end spores it forms. It may appear Gram-negative on Gram stain because it is easily decolorized. It has been associated with cases of necrotizing fasciitis and gas gangrene, bacteremia, sepsis, meningitis, peritonitis, enterocolitis, septic arthritis, abscesses, and even pneumonia. It has also been linked to bone and soft tissue infections. Immunocompromised patients are at higher risk for infection with this organism. Because it produces no toxins, it directly damages tissues via colonization. Neutropenic patients, those who are on Beta-lactam antibiotics, and those with underlying illnesses or conditions are at greater risk of infection.
NOTE: Due to the nature of the spores, be aware that Gram stains may be tricky and Clostridium spp may appear as Gram-negative since the bacteria tend to not hold the Crystal violet stain very well and appear overdecolorized.
NOTE: Due to the nature of the spores, be aware that Gram stains may be tricky and Clostridium spp may appear as Gram-negative since the bacteria tend to not hold the Crystal violet stain very well and appear overdecolorized.
CLOSTRIDIUM BOTULINUM: BOTULISM
Clostridium botulinum is the causative agent of botulism, a potent and deadly form of food poisoning that is often fatal. In fact, the most potent toxin known to man is produced by this organism: botulinum, a neurotoxin and preformed toxin. A dose of just 1.3-2.1 ng/kg in humans is lethal and will paralyze the muscles of its host. The endospores produced by this organism are extremely hardy and survive extremes of temperature and conditions. It is also able to cause wound infections.
Foodborne botulism may be caused by the ingestion of improperly home-canned items or preserved foods, or by eating foods that were not properly processed or preserved. Symptoms begin within 18-36 hours after ingestion and include stomach ache, nausea, vomiting, blurred and double vision, drooping eyelids, difficulty speaking, swallowing, breathing, muscle weakness, constipation, and abnormal reflexes. Wound botulism results in the same symptoms and may also include paralysis and facial weakness. Infant botulism is caused by improperly processed infant formula or honey and produces symptoms such as a weak cry, constipation, muscle weakness, floppy movements, difficulty controlling head and neck movements, irritability, fussiness, fatigue, difficulty sucking or nursing, drooling, drooping eyelids, facial weakness, paralysis.
Foodborne botulism may be caused by the ingestion of improperly home-canned items or preserved foods, or by eating foods that were not properly processed or preserved. Symptoms begin within 18-36 hours after ingestion and include stomach ache, nausea, vomiting, blurred and double vision, drooping eyelids, difficulty speaking, swallowing, breathing, muscle weakness, constipation, and abnormal reflexes. Wound botulism results in the same symptoms and may also include paralysis and facial weakness. Infant botulism is caused by improperly processed infant formula or honey and produces symptoms such as a weak cry, constipation, muscle weakness, floppy movements, difficulty controlling head and neck movements, irritability, fussiness, fatigue, difficulty sucking or nursing, drooling, drooping eyelids, facial weakness, paralysis.
CLOSTRIDIUM DIFFICILE: PSEUDOMEMBRANOUS COLITIS
Clostridium difficile is a normal member of the gastrointestinal flora that is usually kept in check by other flora, however, when antibiotic therapy knocks out the other normal flora, this organism can overgrow in the GI tract. It produces 2 potent toxins, enterotoxin A and cytotoxin B. Cytotoxin B is associated with the production of pseudomembranous colitis.
CLOSTRIDIUM SORDELLII: POST-PARTUM INFECTIONS
Clostridium sordellii is a rare anaerobe that is motile and is associated with a variety of infections, including post-partum infections, muscle necrosis, endocarditis, peritonitis, septic arthritis, bacteremia, septicemia and pneumonia. It has also been associated with severe toxic shock syndrome. The mortality rate of some of these infections is somewhat high. Patients infected with this organism often have markedly high white blood cell counts, rapid heart rate, easy bleeding, and symptoms of shock.
ACTINOMYCES:
Actinomyces spp are branching, beaded, sometimes banded Gram-positive rods that are thin and filamentous. Colonies are small, smooth, flat, convex, gray-white and translucent. A. israelii and A. genecseriae produce white, opaque colonies with a "molar tooth" appearance. They are part of the normal microbiota of the oral, gastronintestinal and vaginal tract. A. odontolyticus turns red after a few days in ambient air and may be beta-hemolytic. Actinomyces spp are facultative anaerobes, other than A. meyeri, which is an obligate anaerobe. Actinomyces spp occasionally form endospores. They are normally soil bacteria, and some are part of the normal vaginal microbiota and oral microbiota. Therefore, the most common clinical infections associated with this bacterium are IUD infections and oral infections. In fact, it is the most common cause of infection in dental procedures and oral abscesses and is an opportunistic pathogen. A. naeslundii is an occupant of the oral cavity. Root and enamel cavities and periodontal disease are associated with this organism. It is also associated with bacterial vaginosis, along with other organisms. It is associated with biofilm formation, plaque, and may be found with tooth extractions. It is not normally pathogenic, but can invade the deeper tissues and cause opportunistic infections.
Actinomycosis is a more drastic infection caused by the bacterium, resulting in abscesses in the mouth, lungs or GI tract, or endocarditis. A. israelii is the most common genus of the species that causes these infections. Thoracic disease is also caused by this bacterium, resulting in a mass that extends to the chest wall due to aspiration of the bacterium. It is also associated with sinus tract infection with abdominal disease. It is also responsible for some cases of periodontal disease, oralcervicofacial disease, and lymphadenopathy. A. naesulundii, A. meyeri, A. israelii, A. viscosus, A. genecseriae, A. odontolyticus are associated with actinomycosis.
In thioglycollate broth, Actinomyces shows a zonal band of flocculance about 1 cm under the surface, with a zone of clearing above it. The species is catalase -.
Actinomycosis is a more drastic infection caused by the bacterium, resulting in abscesses in the mouth, lungs or GI tract, or endocarditis. A. israelii is the most common genus of the species that causes these infections. Thoracic disease is also caused by this bacterium, resulting in a mass that extends to the chest wall due to aspiration of the bacterium. It is also associated with sinus tract infection with abdominal disease. It is also responsible for some cases of periodontal disease, oralcervicofacial disease, and lymphadenopathy. A. naesulundii, A. meyeri, A. israelii, A. viscosus, A. genecseriae, A. odontolyticus are associated with actinomycosis.
In thioglycollate broth, Actinomyces shows a zonal band of flocculance about 1 cm under the surface, with a zone of clearing above it. The species is catalase -.
BIFIDOBACTERIUM:
Bifidobacterium spp are Gram-positive, nonmotile, anaerobic diptheroids or coccoid bacilli that are thin with pointed or rounded ends, highly irregular, some curved, some branching, with terminal ends like clubs or thick, bifurated (forked) ends ("dog bones"). Colonies are small, white, convex, shiny, with irregular edges. Growth is anaerobic and catalase -, nonmotile, and can ferment carbohydrates. They are part of the normal microbiota of the vaginal tract, oral cavity and gastrointestinal tract. They are one of the members of the colon microflora. They are normally beneficial probiotics. Occasionally, they are associated with bacterial vaginosis along with other organisms.
BILOPHILIA WADSWORTHIA:
Bilophila wadsworthia is an anaerobic, pale-staining, delicate Gram-negative rod. Colonies are small and translucent, resembling "fish eyes" and produce H2S, are bile esculin -, urease + and catalase +. This bacterium is part of the GI flora and carries out fermentation in the gut. It is occasionally found in patients who have appendicitis. It has been associated with intra-abdominal abscesses and periodontal abscesses.
EUBACTERIUM (Eggerthella):
Eubacterium (Eggerthella) spp are anaerobic Gram-positive rods or coccobacilli that are pleomorphic and often found in pairs or short chains. E. alactolyticum has a "seagull wing" shape similar to that of Campylobacter spp. E. nodatum is beading, branching and filamentous. E. lentum is small, straight rods with rounded ends. Colonies are small, gray, transparent-to-translucent, raised-to-convex. These bacterium have been associated with bacterial vaginosis. As part of the normal GI tract flora, they are the second most common gut flora after Bacteroides spp.
LACTOBACILLUS:
Lactobacillus spp are part of the normal commensal flora of the urinary, genital and digestive tracts. They are alpha-hemolytic, facultative anaerobic, lactic acid bacteria that are Gram-positive rods and may be found singularly or chains. They keep the homeostatic balance and pH balance of the urogenital tract and their presence helps to keep pathogenic bacteria at bay. They produce hydrogen peroxide, which is toxic to some pathogenic bacteria. Lactobacillus is naturally found in yogurt, cheese, buttermilk, kefir, sauerkraut, pickled foods, cider, alcoholic beverages such as beer and wine, fermented foods, and other products.
ARCANOBACTERIUM:
VEILLONELLA:
Veillonella spp are anaerobic, Gram-negative or Gram-variable cocci. They are part of the normal GI flora and oral flora and vaginal flora of humans. V. parvula is associated with endocarditis and rare osteomyelitis, as well as bacterial vaginosis. These bacterium are strong fermenters. Because of this, they are also associated with periodontal infections, dental cavities, and rarely, systemic infections.
BACTEROIDES:
Bacteroides spp are obligate anaerobes that will die after a short exposure to oxygen, therefore, anaerobes should not be open or exposed to oxygen for more than 30 minutes at a time. The Pigmented Bacteroides Group will fluoresce brick red with UV light and colonies will be brown or black. B. fragilis group is the most common of the Bacteroides spp recovered, and overall, Bacteroides spp are the most common anaerobes recovered clinically. There is no endotoxin and no lipid A in its cell membrane, but it does produce a capsule. It has low virulence but causes opportunistic infections due to trauma, surgery, bowel perforation, ischemia, and is associated in these cases with peritonitis and abscesses. This is also seen in obstetric and gynecoligic patients who have had a septic abortion, miscarriage, pelivc inflammatory disease, or an IUD infection. Bartholin's cyst abscesses and infections are also associated with this bacterium. Abscesses associated with this bacterium have to be surgically drained. It is often recovered on the KV-laked agar.
B. fragilis group are pale-staining Gram-negative rods that are pleomorphic with rounded ends, found in singles or pairs, and they sometimes resemble "safety pins" due to bipolar staining. Colonies are smooth, marbled, gray-white, circular, entire, convex, translucent-to-semiopaque, and nonhemolytic. This species is Beta-lactamase + and is bile esculin + at 48 hours. It is catalase + and indole -. It is not inhibited by colistin, kanamycin, or vancomycin.
B. melaninogenicus produce a black pigment when grown on blood agar. It is part of the normal flora of the mouth, vagina and GI tract. It is typically associated with necrotizing anaerobic pneumonia caused by aspiration (VAP, seizure, drunken state/vomit). It is also associated with periodontal disease.
B. distasonis are straight, round-ended GNRs found in singles and pairs. Colonies are gray-white, circular, convex, entire, smooth, translucent-to-opaque and nonhemolytic. They are bile esculin + at 48 hours.
B. ovatus are ovoid GNRs with rounded ends found in singles or pairs. Colonies are pale buff, circular, entire, convex, semi-opaque, often mucoid, and bile esculin + at 48 hours. In fact, 20% bile enhances the growth of this organism.
B. thetaiotaomicron are irregular-staining GNRs that are pleomorphic with rounded ends, found in singles and pairs. Colonies are circular, white, semi-opaque, shiny, punctiform and nonhemolytic. This species is indole +.
B. ureolyticus are pale, thin, delicate GNRs with rounded ends and are sometimes curved. Colonies are small, translucent, and produce greening of the agar upon exposure to oxygen/air. Colonies may "pit the agar" or may be smooth, convex or spreading. This species is urease +.
B. vulgatus are pleomorphic, GNRs with rounded ends seen in singles, pairs or chains and produce swellings or vacuoles in the cells. Colonies are gray, circular, convex, semi-opaque and nonhemolytic.
Bacteroides spp are linked to clinically significant infections and have been recovered in blood, bacteremia and sepsis, pleural fluid, peritoneal fluid, wounds, burns, brain abscesses and other abscesses, and are associated with colon inflammation and possible tumor formation.
B. fragilis group are pale-staining Gram-negative rods that are pleomorphic with rounded ends, found in singles or pairs, and they sometimes resemble "safety pins" due to bipolar staining. Colonies are smooth, marbled, gray-white, circular, entire, convex, translucent-to-semiopaque, and nonhemolytic. This species is Beta-lactamase + and is bile esculin + at 48 hours. It is catalase + and indole -. It is not inhibited by colistin, kanamycin, or vancomycin.
B. melaninogenicus produce a black pigment when grown on blood agar. It is part of the normal flora of the mouth, vagina and GI tract. It is typically associated with necrotizing anaerobic pneumonia caused by aspiration (VAP, seizure, drunken state/vomit). It is also associated with periodontal disease.
B. distasonis are straight, round-ended GNRs found in singles and pairs. Colonies are gray-white, circular, convex, entire, smooth, translucent-to-opaque and nonhemolytic. They are bile esculin + at 48 hours.
B. ovatus are ovoid GNRs with rounded ends found in singles or pairs. Colonies are pale buff, circular, entire, convex, semi-opaque, often mucoid, and bile esculin + at 48 hours. In fact, 20% bile enhances the growth of this organism.
B. thetaiotaomicron are irregular-staining GNRs that are pleomorphic with rounded ends, found in singles and pairs. Colonies are circular, white, semi-opaque, shiny, punctiform and nonhemolytic. This species is indole +.
B. ureolyticus are pale, thin, delicate GNRs with rounded ends and are sometimes curved. Colonies are small, translucent, and produce greening of the agar upon exposure to oxygen/air. Colonies may "pit the agar" or may be smooth, convex or spreading. This species is urease +.
B. vulgatus are pleomorphic, GNRs with rounded ends seen in singles, pairs or chains and produce swellings or vacuoles in the cells. Colonies are gray, circular, convex, semi-opaque and nonhemolytic.
Bacteroides spp are linked to clinically significant infections and have been recovered in blood, bacteremia and sepsis, pleural fluid, peritoneal fluid, wounds, burns, brain abscesses and other abscesses, and are associated with colon inflammation and possible tumor formation.
FUSOBACTERIUM SPP:
Fusobacterium spp are obligate anaerobes. They will die shortly after exposure to oxygen/air, and anaerobes should not be open/exposed to air for more than 30 minutes at a time. It is associated with periodontal disease or aspiration pneumonias, abdominal and pelvic abscesses, and middle ear infection (otitis media).
F. mortiferum are pale-staining, irregular, highly pleomorphic Gram-negative rods with swollen ends. They are large, filamentous, with bizarre, round bodies. Colonies are circular, entire, irregular, convex, umbonate, smooth and tranparent and are nonhemolytic. This spp is bile esculin + and indole variable.
F. necrophorum are pleomorphic Gram-negative rods with round-to-tapered ends and may be filamenouts or contain round bodies. Rods become more pleomorphic with age. Colonies are umbonate, ridged, tranparent-to-opaque, and fluoresce chartreuse under UV light. Some strains, as seen in the image above, are beta-hemolytic and other strains green the agar on exposure to air. This species is the causative agent of peritonsillar abscesses and Lemierre's Disease. It is lipase + and indole + and will die upon short exposure to oxygen/air.
F. nucleatum are long, slender Gram-negative rods with pointed ends, pale and spindle-shaped. There are 3 types of colonies:
1) White and bread-crumb-like, as seen in the image above on the right)
2) Speckled/"ground-glass"
3) Smooth, gray or gray-white
This species greens the agar upon exposure to air/oxygen and fluoresces chartreuse upon exposure to air/oxygen. It is usually nonhemolytic and produces a foul odor. It is indole +, lipase -, and inhibited by 20% bile.
F. varium is an uneven-staining, pleomorphic, Gram-negative rod found in singles or pairs. It is sometimes coccoid. Colonies are gray-white with a colorless edge and resemble a "fried egg". Colonies are nonhemolytic.
F. mortiferum are pale-staining, irregular, highly pleomorphic Gram-negative rods with swollen ends. They are large, filamentous, with bizarre, round bodies. Colonies are circular, entire, irregular, convex, umbonate, smooth and tranparent and are nonhemolytic. This spp is bile esculin + and indole variable.
F. necrophorum are pleomorphic Gram-negative rods with round-to-tapered ends and may be filamenouts or contain round bodies. Rods become more pleomorphic with age. Colonies are umbonate, ridged, tranparent-to-opaque, and fluoresce chartreuse under UV light. Some strains, as seen in the image above, are beta-hemolytic and other strains green the agar on exposure to air. This species is the causative agent of peritonsillar abscesses and Lemierre's Disease. It is lipase + and indole + and will die upon short exposure to oxygen/air.
F. nucleatum are long, slender Gram-negative rods with pointed ends, pale and spindle-shaped. There are 3 types of colonies:
1) White and bread-crumb-like, as seen in the image above on the right)
2) Speckled/"ground-glass"
3) Smooth, gray or gray-white
This species greens the agar upon exposure to air/oxygen and fluoresces chartreuse upon exposure to air/oxygen. It is usually nonhemolytic and produces a foul odor. It is indole +, lipase -, and inhibited by 20% bile.
F. varium is an uneven-staining, pleomorphic, Gram-negative rod found in singles or pairs. It is sometimes coccoid. Colonies are gray-white with a colorless edge and resemble a "fried egg". Colonies are nonhemolytic.
LEPTOTRICHIA SPP:
Leptotrichia spp are anaerobic, large, fusiform Gram-negative rods with one pointed end and one blunt end. Colonies are large and raspberry-like in appearance. This bacterium is part of the normal oral flora. Most of the time, if this organism causes infection, it is in neutropenic patients. L. buccalis is associated with post-dental work infections following root canals. It causes dental granulomas after a root canal filling.
ERYSIPELOTHRIX RHUSOPATHIAE:
These facultative anaerobe, non-sporulating Gram-positive bacilli are unique in that they produce H2S gas. They are catalase - and nonmotile. It is found in soil and water and it is a facultative anaerobe. It produces a disease called erysipeloid in humans and erysipelas in animals. Because it is a pathogen of pigs, rare disease in humans may occur in those who work closely with infected pigs, such as butchers or farmers and others. Erysipeloid is a mild cutaneous illness characterized by erythematous lesions on the back of the hands and fingers. Rarely, it may disseminate and cause sepsis and has also rarely been linked to endocarditis. Mild itching, pain, and fever that are self-limiting are typical. Other laboratory tests may reveal leukocytosis, and an increase in inflammatory markers such as C-reactive protein. It is resistant to Vancomycin and Penicillin is the treatment of choice for 7 days.
Mobiluncus spp:
Mobiluncus spp are Gram-negative or Gram-variable, anaerobic, small, thin, curved bacilli. Colonies are tiny and pinpoint at 48 hours, low, convex, and tranparent at 3-5 days. It is a causative agent of Bacterial Vaginosis (BV), along with Gardnerella vaginalis.
PROPIONIBACTERIUM SPP:
Propionibacterium spp are aerotolerant anaerobes, pleomorphic Gram-positive bacilli or diphtheroids shaped like clubs and are sometimes in palisade arrangement. Colonies are small, white-to-gray, larger and more yellow-tan with age, and some strains are beta-hemolytic (P. avidum). P. acnes is indole + and catalase +. All species are nonmotile and catalase +, except for P. proprionicum, which is catalase -.
Because this is part of the normal skin flora, it often causes contamination of blood cultures. It is also part of the normal GI flora. It is also linked to acne, eye infections, blepharitis, endolphthalmitis after eye surgery, folliculitis, corneal ulcers, endocarditis, septic arthritis, CSF shunt infections, suture infections, joint prostheses, and other opportunistic infections.
Because this is part of the normal skin flora, it often causes contamination of blood cultures. It is also part of the normal GI flora. It is also linked to acne, eye infections, blepharitis, endolphthalmitis after eye surgery, folliculitis, corneal ulcers, endocarditis, septic arthritis, CSF shunt infections, suture infections, joint prostheses, and other opportunistic infections.
ARCANOBACTERIUM HAEMOLYTICUM:
Arcanobacterium haemolyticum is a facultative anaerobe and a Gram-positive bacillus that is Beta-hemolytic, nonmotile, catalase -, and is part of the oropharyngeal flora, but is associated with polymicrobial infections and occasional pharyngitis, Vincent's angina, and peritonsillar abscesses.
BREVIBACTERIUM SPP:
Brevibacterium spp are Gram-positive bacilli in young cultures, but morph into cocci in older cultures. Colonies are nonhemolytic, but may green the agar after 48 hours of incubation on blood agar. They are nonmotile, tolerant of salt, aerobic, catalase + and urease -.
GARDNERELLA VAGINALIS:
Gardnerella vaginalis is a facultative anaerobe that stains Gram-variable as small, nonmotile, coccobacilli. It is linked to bacterial vaginosis.
Gardnerella vaginalis is a facultative anaerobe that stains Gram-variable as small, nonmotile, coccobacilli. It is linked to bacterial vaginosis.
Parvimonas micra:
This is an anaerobic, Gram-positive cocci, linked to aspiration pneumonia, septic arthritis, and other anaerobic infections.
sterile body fluids:
blood cultures bench
Microorganisms found in the bloodstream may either be transient or intermittent bacteremia, or more serious or continuous sepsis that can threaten a person's life and can affect every bodily organ. Bloodstream invasion of organisms, including bacteria, fungi, parasites, and others may have serious, immediate, even fatal consequences. Septic shock, multi-organ failure, disseminated intravascular coagulation, or death are possible. In fact, bacteremia, fungemia and sepsis may result in mortality rates ranging from 20-50% or higher. In addition to CSF infections, bloodstream infections rank as one of the most serious situations we see in the Microbiology laboratory and infectious disease, therefore, early detection, diagnosis and treatment is critical. Bacteria, fungi, viruses and parasites may all invade the blood. Positive blood cultures aid in the clinical diagnosis of many diseases.
The collection of blood using aseptic, sterile technique, the amount of blood collected, the timing of its arrival to the laboratory, the incubation temperature and time, the amount of blood cultures collected, the interpretation of the results by qualified laboratory professionals, the calling of those results, all contribute to diagnosis, treatment and patient care. Proper specimen collection and processing of blood collection begins with careful and cautious skin preparation prior to collecting the blood sample in order to reduce the risk of contaminants entering the blood culture bottles. This is accomplished by disinfecting the skin with 70% alcohol and applying an antiseptic to kill surface bacteria (application of iodine). If a patient has an existing IV line, the blood should be drawn below the IV line.
The amount of blood collected for bottles should be 10-20 mL per culture. For adults, 10 mL should be the minimum collected. For small children and infants, 1-5 mL are needed. In order to capture intermittent bacteremia, 2-3 blood culture sets should be collected at least 1 hour apart, preferably at different venipuncture sites prior to the patient beginning antibiotic therapy. The proper blood-to-broth ratio for blood cultures to reduce the antibacterial effect of serum in adults is 1:10. SPS is used as an anticoagulant for blood cultures because it inactivates neutrophils and components of serum complement.
The most common organism recovered from the bloodstream is Gram-positive cocci (CNS, Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium). Other commonly isolated organisms include Viridans streptococci, Gemella, Beta-hemolytic streptococci, Streptococcus pneumoniae, Escherichia spp, Klebsiella spp, Pseudomonas spp, Enterobacter spp, Proteus spp, Citrobacter spp, Bacteroides spp, Clostridium spp). Other recovered species include (Aeromonas, Plesiomonas, Campylobacter, Salmonella, Shigella, HACEK bacteria-Actinobacillus, Kingella, Cardiobacterium, Capnocytophaga), Listeria spp, Acinetobacter baumannii complex, Eikenella, Leptospira, Brucella, Haemophilus, Bartonella spp, Corynebacterium).
Septic shock, bacterial endocarditis, and endovascular infections are particularly dangerous and serious because in these cases, bacteria are continuously released into the bloodstream consistently at a pretty constant rate. Two types of bloodstream infections include intravascular (originate inside the cardiovascular system) and extravascular (originate in other areas outside the cardiovascular system and seed to other areas of the body).
1) Intravascular Infections
-Catheter-associated bacteremia
-Intravenous bacteremia
-S. epidermidis, CNS, S. aureus, Enterobacteriaceae, Pseudomonas, Corynebacterium, Candida, GNR are the most common causes
-Suppurative thrombophlebitis
-Infective endocarditis
-Viridans streptococci is the most common cause (ex: S. anginosus, S. mutans, S. sanguis, S. bovis, S. aureus, CNS, Enterobacteriaceae, Pseudomonas, Haemophilus, HACEK)
-Aneurysm
2) Extravascular Infections
-Bacteria enter the circulation through the lymphatic system
-Portals of entry:
-Genitourinary tract (25%)
-Respiratory tract (20%)
-Abscesses (10%)
-Surgical Wounds (5%)
-Biliary tract (5%)
-Misc. Sites (5%)
-Uncertain (25%)
-Most common causes:
-H. influenzae
-S. pneumoniae
-N. meningitidis
-Brucella spp
-Salmonella spp
-Listeria spp
-Enterobacteriaceae
-S. aureus
-N. gonorrhoeae
-Anaerobic cocci
-Bacteroides
-Clostridium
-Beta-hemolytic streptococci
-Pseudomonas
Fungemia is the presence of fungus in the blood. This is usually a serious condition that is typically seen in immunocompromised or immunosuppressed patients and people with serious illnesses. The most common fungus recovered is Candida albicans, followed by other Candida spp (tropicalis, glabrata, parapsilosis, lustinaniae, kruseii, guilliermondi, zeylanoides, pseudotropicalis, famata), Histoplasma capsulatum, Cryptococcus neoformans, Coccidioides immitis, Trichosporon cutaneum, Malessezia furfur, Rhodotorula spp, Nocardia asteroides, Fusarium spp, Cryptococcus albidus, Cryptococcus laurentii, Saccharomyces spp, Pichia burtonii, Blastomyces dermatiditis, Sporothrix schenckii, Blastoschizomyces capitatus, Trichphyton rubrum)
The collection of blood using aseptic, sterile technique, the amount of blood collected, the timing of its arrival to the laboratory, the incubation temperature and time, the amount of blood cultures collected, the interpretation of the results by qualified laboratory professionals, the calling of those results, all contribute to diagnosis, treatment and patient care. Proper specimen collection and processing of blood collection begins with careful and cautious skin preparation prior to collecting the blood sample in order to reduce the risk of contaminants entering the blood culture bottles. This is accomplished by disinfecting the skin with 70% alcohol and applying an antiseptic to kill surface bacteria (application of iodine). If a patient has an existing IV line, the blood should be drawn below the IV line.
The amount of blood collected for bottles should be 10-20 mL per culture. For adults, 10 mL should be the minimum collected. For small children and infants, 1-5 mL are needed. In order to capture intermittent bacteremia, 2-3 blood culture sets should be collected at least 1 hour apart, preferably at different venipuncture sites prior to the patient beginning antibiotic therapy. The proper blood-to-broth ratio for blood cultures to reduce the antibacterial effect of serum in adults is 1:10. SPS is used as an anticoagulant for blood cultures because it inactivates neutrophils and components of serum complement.
The most common organism recovered from the bloodstream is Gram-positive cocci (CNS, Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium). Other commonly isolated organisms include Viridans streptococci, Gemella, Beta-hemolytic streptococci, Streptococcus pneumoniae, Escherichia spp, Klebsiella spp, Pseudomonas spp, Enterobacter spp, Proteus spp, Citrobacter spp, Bacteroides spp, Clostridium spp). Other recovered species include (Aeromonas, Plesiomonas, Campylobacter, Salmonella, Shigella, HACEK bacteria-Actinobacillus, Kingella, Cardiobacterium, Capnocytophaga), Listeria spp, Acinetobacter baumannii complex, Eikenella, Leptospira, Brucella, Haemophilus, Bartonella spp, Corynebacterium).
Septic shock, bacterial endocarditis, and endovascular infections are particularly dangerous and serious because in these cases, bacteria are continuously released into the bloodstream consistently at a pretty constant rate. Two types of bloodstream infections include intravascular (originate inside the cardiovascular system) and extravascular (originate in other areas outside the cardiovascular system and seed to other areas of the body).
1) Intravascular Infections
-Catheter-associated bacteremia
-Intravenous bacteremia
-S. epidermidis, CNS, S. aureus, Enterobacteriaceae, Pseudomonas, Corynebacterium, Candida, GNR are the most common causes
-Suppurative thrombophlebitis
-Infective endocarditis
-Viridans streptococci is the most common cause (ex: S. anginosus, S. mutans, S. sanguis, S. bovis, S. aureus, CNS, Enterobacteriaceae, Pseudomonas, Haemophilus, HACEK)
-Aneurysm
2) Extravascular Infections
-Bacteria enter the circulation through the lymphatic system
-Portals of entry:
-Genitourinary tract (25%)
-Respiratory tract (20%)
-Abscesses (10%)
-Surgical Wounds (5%)
-Biliary tract (5%)
-Misc. Sites (5%)
-Uncertain (25%)
-Most common causes:
-H. influenzae
-S. pneumoniae
-N. meningitidis
-Brucella spp
-Salmonella spp
-Listeria spp
-Enterobacteriaceae
-S. aureus
-N. gonorrhoeae
-Anaerobic cocci
-Bacteroides
-Clostridium
-Beta-hemolytic streptococci
-Pseudomonas
Fungemia is the presence of fungus in the blood. This is usually a serious condition that is typically seen in immunocompromised or immunosuppressed patients and people with serious illnesses. The most common fungus recovered is Candida albicans, followed by other Candida spp (tropicalis, glabrata, parapsilosis, lustinaniae, kruseii, guilliermondi, zeylanoides, pseudotropicalis, famata), Histoplasma capsulatum, Cryptococcus neoformans, Coccidioides immitis, Trichosporon cutaneum, Malessezia furfur, Rhodotorula spp, Nocardia asteroides, Fusarium spp, Cryptococcus albidus, Cryptococcus laurentii, Saccharomyces spp, Pichia burtonii, Blastomyces dermatiditis, Sporothrix schenckii, Blastoschizomyces capitatus, Trichphyton rubrum)
Listeria monocytogenes
Listeria monocytogenes is a catalase +, non-branching, non-spore-forming Gram-positive rod found singly or in short-chains. Rods may be short or appear as coccobacilli. It is also sometimes seen in Coryneform, as palisades (side-by-side like a "picket fence") or "V" forms. It may colonize the human gastrointestinal tract in some, but it causes infection via the fecal-oral route by ingestion of contaminated foods, such as meat and dairy products, or fruit such as cantaloupe. If a mother is colonized, they may pass it to their neonate. It may get into the bloodstream and cause sepsis or meningitis. It is a potent pathogen. One of the virulence factors that makes this pathogen so potent is listeriolysin, which is a cytotoxic and hemolytic toxin that enables it to survive within phagocytic cells. Infections typically occur in neonates or in patients that are immunosuppressed. In utero, an infection may occur that is referred to as granulomatosis infantiseptica, and it causes stillbirth.
Listeria monocytogenes can be presumptively identified by observation of motility by direct wet mount, or by observation of umbrella motility in motility or SIM agars after overnight incubation. It exhibits a characteristic tumbling motility if incubated in nutrient broth at 25 degrees Celsius for 1-2 hours. It ferments glucose and is bile-esculin positive. If a small, Gram-positive, catalase + rod exhibiting a small zone of beta-hemolysis on blood agar is isolated from the blood or CSF, listeriosis should be ruled out. It is also CAMP + and lipophilic test negative. It is hippurate +. It also ferments glucose/dextrose.
FLOW CHART FOR LISTERIA:
Blood Plate: Beta-hemolytic
Perform Gram Stain: Gram-positive, straight, non-spore-forming, short rods in singles, or side-by-side
Perform Slide Motility Test at 25 degrees Celsius AND at 37 degrees Celsius:
-Get out TSB broth x2 and inoculate heavily with organism from your plate with a sterile swab
-Loosen the caps and incubate one broth at 25 degrees Celsius and the other at 37 degrees Celsius for 1-2 hours
-Label 2 slides and place of drop of each tube on each slide with a coverslip
-Observe for "tumbling" motility (a tumbling leaf-like motion) with the condenser down and the light on low under the microscope on 10x, 20x and 40x objectives
-Listeria monocytogenes is motile at 25 degrees Celsius, but not at 37 degrees Celsius
Perform a Tube Motility Test at 25 degrees Celsius AND at 37 degrees Celsius:
-Liquify the tube media and cool back down to room temperature to re-gel
-Stab down the center of the media with a sterile inoculating loop and your organism to about 3/4 down to the bottom of the tube
-Incubate overnight for 18-24 hours; Incubate one tube at 25 degrees Celsius and the other at 37 degrees Celsius with the caps loose
-Observe for characteristic "umbrella" motility
-Listeria monocytogenes is motile at 25 degrees Celsius but not at 37 degrees Celsius
Perform a Bile Esculin Test:
-Bring the tube agar to room temperature
-Lightly stab just under the media with your organism, and streak across the top with a sterile loop
-Incubate at 37 degrees Celsius with the cap lightly loosened for 1 hour to 24 hours until the agar turns black
-Listeria monocytogenes hydrolyzes bile in the presence of esculin and will blacken the agar. This is a positive test.
Perform a CAMP test:
-Streak a straight line down the middle of a blood plate with the QC Staphylococcus aureus organism
-Perpendicular (at a right angle to) the Staph streak, but not touching it, streak across with your organism
-Incubate for 18-24 hours and observe for a positive CAMP test, with the formation of "arrow" patterns facing the Staph streak
Perform a Glucose Test:
-Liquify the glucose/dextrose slant
-Bring back to room temperature to re-gel
-Stab straight down the middle about 1/4 of the way down the tube beneath the agar with your organism
-Loosen the cap and incubate overnight for 18-24 hours
-Observe for yellowing of the agar due to fermentation of the sugar and production of lactic acid
Perform a Hippurate Test:
-Inoculate the tube with hippurate powder and sterile water as directed for 2 hours
-Add drops of ninhydrin reagent as directed and reincubate for another 30 minutes
-Observe for purple color, indicating a positive Hippurate Test
Treatment usually consists of ampicillin or penicillin with or without an aminoglycoside.
Listeria monocytogenes can be presumptively identified by observation of motility by direct wet mount, or by observation of umbrella motility in motility or SIM agars after overnight incubation. It exhibits a characteristic tumbling motility if incubated in nutrient broth at 25 degrees Celsius for 1-2 hours. It ferments glucose and is bile-esculin positive. If a small, Gram-positive, catalase + rod exhibiting a small zone of beta-hemolysis on blood agar is isolated from the blood or CSF, listeriosis should be ruled out. It is also CAMP + and lipophilic test negative. It is hippurate +. It also ferments glucose/dextrose.
FLOW CHART FOR LISTERIA:
Blood Plate: Beta-hemolytic
Perform Gram Stain: Gram-positive, straight, non-spore-forming, short rods in singles, or side-by-side
Perform Slide Motility Test at 25 degrees Celsius AND at 37 degrees Celsius:
-Get out TSB broth x2 and inoculate heavily with organism from your plate with a sterile swab
-Loosen the caps and incubate one broth at 25 degrees Celsius and the other at 37 degrees Celsius for 1-2 hours
-Label 2 slides and place of drop of each tube on each slide with a coverslip
-Observe for "tumbling" motility (a tumbling leaf-like motion) with the condenser down and the light on low under the microscope on 10x, 20x and 40x objectives
-Listeria monocytogenes is motile at 25 degrees Celsius, but not at 37 degrees Celsius
Perform a Tube Motility Test at 25 degrees Celsius AND at 37 degrees Celsius:
-Liquify the tube media and cool back down to room temperature to re-gel
-Stab down the center of the media with a sterile inoculating loop and your organism to about 3/4 down to the bottom of the tube
-Incubate overnight for 18-24 hours; Incubate one tube at 25 degrees Celsius and the other at 37 degrees Celsius with the caps loose
-Observe for characteristic "umbrella" motility
-Listeria monocytogenes is motile at 25 degrees Celsius but not at 37 degrees Celsius
Perform a Bile Esculin Test:
-Bring the tube agar to room temperature
-Lightly stab just under the media with your organism, and streak across the top with a sterile loop
-Incubate at 37 degrees Celsius with the cap lightly loosened for 1 hour to 24 hours until the agar turns black
-Listeria monocytogenes hydrolyzes bile in the presence of esculin and will blacken the agar. This is a positive test.
Perform a CAMP test:
-Streak a straight line down the middle of a blood plate with the QC Staphylococcus aureus organism
-Perpendicular (at a right angle to) the Staph streak, but not touching it, streak across with your organism
-Incubate for 18-24 hours and observe for a positive CAMP test, with the formation of "arrow" patterns facing the Staph streak
Perform a Glucose Test:
-Liquify the glucose/dextrose slant
-Bring back to room temperature to re-gel
-Stab straight down the middle about 1/4 of the way down the tube beneath the agar with your organism
-Loosen the cap and incubate overnight for 18-24 hours
-Observe for yellowing of the agar due to fermentation of the sugar and production of lactic acid
Perform a Hippurate Test:
-Inoculate the tube with hippurate powder and sterile water as directed for 2 hours
-Add drops of ninhydrin reagent as directed and reincubate for another 30 minutes
-Observe for purple color, indicating a positive Hippurate Test
Treatment usually consists of ampicillin or penicillin with or without an aminoglycoside.
Corynebacterium spp
Corynebacterium spp are catalase +, non-branching, non-spore-forming, Coryneform, Gram-positive rods/bacilli. They are often seen in a variety of forms, including resembling "drumsticks", looking like "Chinese letters", "V" or "L" forms, or palisades (side-by-side like a "picket fence"). They are also referred to as "diphtheroids". Corynebacterium spp are common inhabitants of the normal human flora, but some of them may be pathogenic or opportunists associated with disease in humans.
Corynebacterium diphtheriae-Inhabitant of the human nasopharynx in a carrier state; NOT considered part of the the normal flora; Very rare cause of infection by person-to-person direct contact or exposure to respiratory droplets with infected cutaneous lesions or via contamintaed objects/fomites; Causative agent of diphtheria.
Corynebacterium jeikeium-Found in the skin flora of patients that are hospitalized (axillary, inguinal, rectal regions); Usually causes infection via contaminated IV catheters and is associated with septicemia, wound infections, and endocarditis in patients who are immunocompromised.
Corynebacterium urealyticum-Normal skin flora that may gain accidental entry to a normally sterile site; This type is UREASE +; Associated with UTI or wound infections in immunocompromised patients
Corynebacterium ulcerans-Normal flora of humans, but infections may occur due to close animal contact during the summer and have been associated with a diphtheria-like sore throat/pharyngitis
Corynebacterium haemolyticum-a Corynebacterium that produces small raised colonies surrounded by medium zones of hemolysis
Corynebacterium striatum-Normal skin flora, but may cause infection via accidental entry to a nromally sterile site and is associated with bacteremia, lung abscesses and pneumonia in immunocompromised patients. The images show colonies that are white, small, raised, shiny, round and moist.
Corynebacterium pseudotuberculosis-May cause infection with close animal contact, though, rare, since this is associated with infections in animals (sheep, horses, goats); Associated with granulomatous lymphadenitis
Corynebacterium pseudodiphtheriticum-May cause infection by the patient's own normal flora entering a normally sterile site; Inhabitant of the normal pharyngeal and skin flora; Associated with endocarditis, pneumonia, lung abscesses
Corynebacterium minutissimum-Normal skin flora; may cause infection by the patient's own normal flora entering a normally sterile site; Associated with pruritic skin infections (erythrasma), septicemia, or abscesses
Corynebacterium aquaticum-Found in fresh water; Associated with rare bacteremia
Corynebacterium xerosis-Part of the normal flora of the human conjunctiva, nasopharynx, and skin that may gain inadvertant access to a normally sterile site; Associated with rare septicemia or endocarditis
Corynebacterium amycolatum-Normal flora of the conjunctiva, nasopharynx and skin that may gain inadvertant access to a normally sterile site; Associated with endocarditis, septicemia, or pneumonia in patients who are immunocompromised
Corynebacterium auris-Normal human flora that is rarely implicated in human infections; Associated with otitis media
Corynebacterium pseudodiphtheriticum-May cause infection by the patient's own normal flora entering a normally sterile site; Inhabitant of the normal pharyngeal and skin flora; Associated with endocarditis, pneumonia, lung abscesses
Corynebacterium minutissimum-Normal skin flora; may cause infection by the patient's own normal flora entering a normally sterile site; Associated with pruritic skin infections (erythrasma), septicemia, or abscesses
Corynebacterium aquaticum-Found in fresh water; Associated with rare bacteremia
Corynebacterium xerosis-Part of the normal flora of the human conjunctiva, nasopharynx, and skin that may gain inadvertant access to a normally sterile site; Associated with rare septicemia or endocarditis
Corynebacterium amycolatum-Normal flora of the conjunctiva, nasopharynx and skin that may gain inadvertant access to a normally sterile site; Associated with endocarditis, septicemia, or pneumonia in patients who are immunocompromised
Corynebacterium auris-Normal human flora that is rarely implicated in human infections; Associated with otitis media
Gram stains of Coryneform bacteria:
Coryneform bacteria are Gram-positive, short, or slightly curved rods/bacilli. They may have swollen, rounded ends like drumsticks or chicken legs, and some have somewhat rudimentary branching or spiral forms. Rods may be arranged singly in pairs, in palisades form (side-by-side like a picket fence), parallel forms, or in "V", "L" or Chinese Letters shapes/foms. Examples are shown in the Gram stains below:
Most Corynebacterium spp grow on blood and chocolate agars and some are lipophilic and will produce a + lipophilic test. Colonies appear as small to medium, gray to white, nonhemolytic, and often dry on blood agar. They are nonmotile. C. ulcerans and urealyticum are urease +.
sterile body fluids bench
4 Tubes of cerebrospinal fluid (CSF) collected for laboratory analysis. In this image, all 4 tubes are clear. Tube 2 goes to the Microbiology laboratory and MUST BE CYTOCENTRIFUGED and centrifuged prior to smear and Gram stain. Always check the Hematology and Chemistry results for WBC and diff. results and for protein and glucose results. In bacterial meningitis, glucose is decreased in about 50% of patients who have this condition, whereas patients with viral meningitis tend to have normal glucose values. Patients with bacterial meningitis also tend to have increased polymorphonuclear neutrophils (PMNs) in their CSF, whereas those with viral meningitis tend to have more monocytes and lymphocytes, many of which may be "reactive". Protein levels are usually high in patients with bacterial meningitis and it is normal to high in patients with viral meningitis.
CSF fluid is normally a sterile body fluid, uncontaminated with organisms. Most organisms do not cross the blood-brain barrier, however, if they do, it can result in a traumatic infection called meningitis and or meningoencephalitis. Patients with purulent bacterial meningitis will exhibit acute inflammatory exudate, or cloudy CSF, with a large amount of polymorphonuclear neutrophils (PMNs) on the Gram stain. Additionally, glucose levels will be decreased and protein levels will be increased.
Aseptic meningitis, on the otherhand, is characterized by an increase of lympocytes and monocytes and is usually associated with viral infections or syphilis or spirochete infections, such as leptospirosis. The primary agents causing viral meningitis include the enteroviruses (Coxsackieviruses A and B and echoviruses), mumps virus, herpes simplex virus (HSV), and arboviruses (togavirus, bunyavirus, equine encephalitis, S. Louis encephalitis, West Nile virus, etc..). Measles, cytomegalovirus, Epstein-barr virus, Hepatitis, Varicella-zoster virus (VZV), rabies virus, myxoviruses and paramyxoviruses, BK virus, JC virus are less common causes of meningoencephalitis.
Parasites can also cause meningoencephalitis or brain abscess. These include Naegleria fowleri, Acanthamoeba spp, Entamoeba histolytica, Strongyloides stercoralis, Taenia solium, toxoplasmosis, or others.
Aseptic meningitis, on the otherhand, is characterized by an increase of lympocytes and monocytes and is usually associated with viral infections or syphilis or spirochete infections, such as leptospirosis. The primary agents causing viral meningitis include the enteroviruses (Coxsackieviruses A and B and echoviruses), mumps virus, herpes simplex virus (HSV), and arboviruses (togavirus, bunyavirus, equine encephalitis, S. Louis encephalitis, West Nile virus, etc..). Measles, cytomegalovirus, Epstein-barr virus, Hepatitis, Varicella-zoster virus (VZV), rabies virus, myxoviruses and paramyxoviruses, BK virus, JC virus are less common causes of meningoencephalitis.
Parasites can also cause meningoencephalitis or brain abscess. These include Naegleria fowleri, Acanthamoeba spp, Entamoeba histolytica, Strongyloides stercoralis, Taenia solium, toxoplasmosis, or others.
N. meningitidis has been associated with outbreaks of epidemic meningitis amongst adolescents and young adults in overcrowded conditions and cramped quarters, such as hospitals, military quarters, and college dormitories.
H. influenzae type b in kids has been linked with an increased incidence of meningitis. The Hib vaccination has decreased these incidences and provides a degree of protection by age 5. Its presence in the CSF can be rapidly detected by bacterial antigen testing (example: DIRECTIGEN latex agglutination test).
CSF should be hand-delivered STAT to the laboratory, and NEVER REFRIGERATED because various agents such as S. pneumoniae may not be detectable after an hour unless a special antigen detection method is used.
Streptococcus epidermidis is the main bacterial cause of acute infective endocarditis, particularly of heart valves and prosthetics. Enterococci spp are also a cause of infective endocarditis, and a penicillin derivative and aminoglycoside may result in synergistic action to treat acute endocarditis due to infection with this organism. Enterococcal isolated from patients with endocarditis should be tested for routine susceptibility and screened for high-level aminoglycoside resistance.
respiratory bench
lower respiratory tract infections
RESPIRATORY CULTURE: SPUTUM AND TRANSTRACHEAL:
This is the portion of the respiratory tract below the larynx:
-Gram stain predominant organism
-ID and perform susceptibility tests on predominant organisms
-If normal flora, ID as light, moderate or heavy normal oropharyngeal flora
BAL or MINI-BAL:
- Expectorated sputum (early morning specimens are best, after gargling with just water and before brushing the teeth or using mouthwash or eating; take 3-5 deep breaths first; collect in sterile container)
- Aerosol-induced sputum
- Tracheal aspirate (suction)
- Naso-tracheal aspirate
- Pleural fluid
- Transtracheal aspirate
- Transthoracic lung aspirates
- Tissue biopsies
This is the portion of the respiratory tract below the larynx:
- Trachea
- Bronchi
- Bronchioles
- Alveoli
-Gram stain predominant organism
-ID and perform susceptibility tests on predominant organisms
-If normal flora, ID as light, moderate or heavy normal oropharyngeal flora
BAL or MINI-BAL:
- Catheter telescoping down endotracheal tube in place of bronchoscopy collects aspirate
- Performed when ventilator-associated pneumonia (VAP) is suspected, which is the leading cause of death from nosocomial infections in patients on mechanical ventilation, which usually occurs approximately within 48 hours post-intubation
- Looking for pneumonia-causing pathogens, fungi (yeasts, molds), mycobacteria, Legionella, viruses
- These specimens are never rejected because it is an invasive procedure
- 1 colony = 100 cfu/mL with the 0.01 (10 uL) inoculating loop
- Preliminary no growth: PNAG
- Cultures with colony growth: Quantitate each pathogen found
- Mixed or normal flora: mixed or normal respiratory flora with total colony count
- Yeast: perform urease test, germ tube and cornmeal agar
- GPR: R/O Nocardia, B. cereus, B. anthracis, Rhodococcus equi, Coryneforms if predominant or rapidly urease + or patient is intubated
Definite Respiratory Tract Pathogens:
-Corynebacterium diphtheriae -Mycobacterium tuberculosis (AFB) -Mycoplasma pneumoniae -Chlamydia trachomatis -Chlamydia pneumoniae -Bordetella pertussis -Legionella spp -Pneumocystis carinii -Nocardia spp -Histoplasma capsulatum (Mycology) -Coccidioides immitis (Mycology) -Cryptococcus neoformans (Mycology) -Blastomyces dermatitidis (Mycology) -Viruses (RSV, Influenza, Adenoviruses, Noroviruses, Rhinoviruses, Enteroviruses, HSV, Parainfluenzae virus) Rare Respiratory Tract Viruses: -Francisella tularensis -Bacillus anthracis -Yersinia pestis -Burkholderia pseudomallei and B. cepacia -Coxiella burnetti -Chlamydia psittaci -Brucella -Salmonella -Pasteurella multocida -Klebsiella rhinoscleromatis -VZV -Parasites -Acinetobacter -Stenotrophomonas (inherent multi-drug resistance) -HACEK bacteria: (fastidious GNRs) HACEK is an abbreviation of the initials of the genera of this group of bacteria: Haemophilus, Aggregatibacter (previously Actinobacillus), Cardiobacterium, Eikenella corrodens, Elizabethkingia, Kingella. Colonizers, Opportunists, and Possible Pathogens: -Acinetobacter spp -Viridans streptococci -Beta-hemolytic streptococci -Streptococci pneumoniae -Staphylococcus aureus -Nesseria meningitidis -Mycoplasma spp -Haemophilus influenzae -H. influenzae aegyptius -Haemophilus parainfluenzae -Haemophilus aphrophilus -Moraxella catarrhalis -Candida albicans -HSV -Enterobacteriaceae (GNRs) -Mycobacterium spp -Pseudomonas spp (mucoid and non-mucoid morphotypes) -Burkholderia cepacia -Filamentous fungi -Klebsiella pnemoniae -Klebsiella ozaenae -Eikenella corrodens -Bacteroides -Peptostreptococcus -Actinomyces spp -Capnocytophaga -Actinobacillus spp -Haemophilus aphrophilus -Entamocha gingivalis -Trichomonas tenax -Staphylococcus -Pseudomonas aeruginosa -Pasteurella multocida -Enterococcus spp -Achromobacter spp |
Haemophilus influenzae
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The X and V factor test may be performed by inoculating the H. QUAD plate. Use a different sterile loop to inoculate each quadrant with specimen. H. influenzae grows only in quadrants 3-4, engraved on the tope of each quadrant. If there is growth in other quadrants, it is not H. influenzae. Perform the Vitek NH card to determine which type of Haemophilus spp you have isolated.
Haemophilus influenzae is a small Gram-negative short rod or coccobacillus that is resistant to Bacitracin disk on the Chocolate agar (colonies will show resistance around the disk) and requires X and V factors to grow. Colonies will satellite around a streak of S. aureus on blood agar. H. influenzae is usually B-lactamase + (resistant to penicillin and ampicillin), oxidase +, and catalase +. On Gram-stain, this microbe is Gram-negative and coccobacilli in shape (egg-shaped to small rods). H. influenzae has a pungent odor. H. parainfluenzae can sometimes cause similar infections as H. influenzae, and it is ornithine decarboxylase +.
Infections:
H. influenzae type b (Hib) produces the following infections:
1) Bacteremia
2) Pneumonia
3) Epiglottitis
4) Acute bacterial meningitis
5) Cellulitis
6) Osteomyelitis
7) Infectious arthritis
8) Otitis media (ear infection) in children
9) Eye infections in children (conjunctivitis)
10) Sinusitis in children
Colony Morphology:
-Small, convex, transluscent-to-tan-to-grayish glistening colonies
Infections:
H. influenzae type b (Hib) produces the following infections:
1) Bacteremia
2) Pneumonia
3) Epiglottitis
4) Acute bacterial meningitis
5) Cellulitis
6) Osteomyelitis
7) Infectious arthritis
8) Otitis media (ear infection) in children
9) Eye infections in children (conjunctivitis)
10) Sinusitis in children
Colony Morphology:
-Small, convex, transluscent-to-tan-to-grayish glistening colonies
Haemophilus aphrophilus
This organism forms transparent, pinpoint, nonhemolytic colonies on blood agar and it is associated with bacterial endocarditis.
Streptococcus pneumoniae
Streptococcus pneumoniae is a Gram-positive diplococcus (lancet-shaped with pointed ends), also found in chains, that may cause bacterial pneumonia. In a CF patient, it may occur along with or secondary to P. aeruginosa, S. aureus, and/or B. cepacia. It is susceptible to the optochin disk (Taxo P) on Neomycin blood agar and is alpha-hemolytic. It is also pneumoslide + and bile solubility +. There are both mucoid and non-mucoid strains/morphotypes. The direct quellung reaction using specific antiserum is useful in testing CSF suspected of having this organism.
Klebsiella pneumoniae-gnr, enterobacteriaceae (rare is normal oropharyngeal flora)
Klebsiella pneumoniae is a Gram-negative bacilli that appears as a lactose-fermenter on MacConkey agar. It is not motile, and it is oxidase and indole negative, and it is very mucoid and slimy. Patients who are experiencing bacterial pneumoniae due to this organism tend to produce sputum that is thick, bloody, mucoid, and looks like currant jelly. Fatality rates may be as high as 50%, even with antimicrobial treatment, and this organism tends to infect individuals who are immunocompromised. Klebsiella oxytoca, on the other hand, is indole positive, except for a few rare strains (1%). On blood agar, colonies appear as gray, mucoid, and stringy. On MacConkey agar, colonies are mucoid, and dark pink. K. pneumoniae is ONPG +, indole -, glucose +, oxidase -, citrate +, and VP +. It gives a TSI reaction of acid slant/acid butt (yellow/yellow), no H2S, + production of gas (bubbles/cracks)
pseudomonas aeruginosa-non-mucoid and mucoid strains
Pseudomonas aeruginosa may exist in a variety of morphotypes, even in a single specimen, and may have a variety of susceptibility patterns as well. Mucoid and non-mucoid morphotypes exist. The mucoid strain is more commonly isolated in Cystic Fibrosis (CF) patients, and are slow-growing strains that may take >48 hours to mature and up to 7 days to react to biochemicals. It is strongly oxidase +, indole -, catalase +, motility +, nitrate +, and a straight, GNR. Many strains produce a green or blue-green, shiny, diffusible pigment known as pyocyanin. On blood agar, it forms colonies with feathered edges and a grape-like odor. It is a water organism and has been associated with contamianted hot tubs, whirlpools, contact lens solutions, and hospital environments (sinks, showers, respiratory equipment), medical equipment. It may colonize the respiratory tract or GI tract. It has been associated with respiratory infections, UTIs, burn and wound infections. Cystic Fibrosis patients are particularly at risk for infection. Some species grow at 42 degrees Celsius and produce gelatinase. It is usually susceptible to gentamicin.
moraxella catarrhalis
Moraxella catarrhalis:
Oxidase +, MCAT (butyrate) disk +, B-lactamase +, Catalase +, DNAse +, nonmotile, fastidious, Aerobic, GNCB, doesn't ferment any sugars in the sugars test
Virulence Factor: Trimeric Autotransporter Adhesins for sticking to mucosal surfaces. Colonies are pink-brown, rough, friable and opaque on CHOC agar and like "hockey-pucks", able to be moved/slid across the surface of the agar.
Infections:
1) Middle ear (otitis media) in children
2) Eye
3) Central Nervous System (bacteremia, meningitis and septicemia on rare occasions)
4) Joints (septic arthritis)
5) Respiratory, particularly the upper respiratory tract:
-Chronic bronchitis
-Acute and maxillary sinusitis
-Acute laryngitis
-Bronchopneumonia in the elderly, smokers, immunocompromised, those with COPD; occurs usually by pulmonary aspiration of the microbe
6) Urethritis (rarely)
7) Endocarditis (rare)
Colonizations:
-Common by age 2 and high in number; Lower in number in adults
-Emerging infectious pathogen, especially in patients with recurring and chronic respiratory conditions
-Pharyngeal colonizations predispose an individual in increased susceptibility of pulmonary parenchyma to infection
Colony Morphology:
-Colonies are able to be moved across the agar with a wooden stick and slide like a hockey puck
Antimicrobial Resistance:
-Commonly resistant to penicillin, ampicillin, amoxicillin, trimethoprim, trimethoprim-sulfaoxazole, clindamycin, tetracycline
Other species of Moraxella, such as M. osloensis, may be mistaken for Neisseria spp, because they Gram-stain as small, Gram-negative diplococci versus coccobacilli.
Oxidase +, MCAT (butyrate) disk +, B-lactamase +, Catalase +, DNAse +, nonmotile, fastidious, Aerobic, GNCB, doesn't ferment any sugars in the sugars test
Virulence Factor: Trimeric Autotransporter Adhesins for sticking to mucosal surfaces. Colonies are pink-brown, rough, friable and opaque on CHOC agar and like "hockey-pucks", able to be moved/slid across the surface of the agar.
Infections:
1) Middle ear (otitis media) in children
2) Eye
3) Central Nervous System (bacteremia, meningitis and septicemia on rare occasions)
4) Joints (septic arthritis)
5) Respiratory, particularly the upper respiratory tract:
-Chronic bronchitis
-Acute and maxillary sinusitis
-Acute laryngitis
-Bronchopneumonia in the elderly, smokers, immunocompromised, those with COPD; occurs usually by pulmonary aspiration of the microbe
6) Urethritis (rarely)
7) Endocarditis (rare)
Colonizations:
-Common by age 2 and high in number; Lower in number in adults
-Emerging infectious pathogen, especially in patients with recurring and chronic respiratory conditions
-Pharyngeal colonizations predispose an individual in increased susceptibility of pulmonary parenchyma to infection
Colony Morphology:
-Colonies are able to be moved across the agar with a wooden stick and slide like a hockey puck
Antimicrobial Resistance:
-Commonly resistant to penicillin, ampicillin, amoxicillin, trimethoprim, trimethoprim-sulfaoxazole, clindamycin, tetracycline
Other species of Moraxella, such as M. osloensis, may be mistaken for Neisseria spp, because they Gram-stain as small, Gram-negative diplococci versus coccobacilli.
burkholderia cepacia
Burkholderia cepacia is significant if isolated in Cystic Fibrosis (CF) patients. It grows on BCSA agar as colonies that are grayish-white with a purple tint to them and fermentation will cause a color change in the agar to pink. This is a screening tool only. B. cepacia is motility +, oxidase + (14% are oxi -), LDC + in 80% of strains (20% are LDC-), and is an environmental bacterium that is never part of the normal oropharyngeal flora, other than that it may colonize the respiratory tract of CF patients. If isolated from respiratory cultures, it may mean that medical and/or respiratory equipment is heavily contaminated with the organism. Medical devices may become colonized and contaminated with the organism, which thrives in moist environments and clinical settings. It is inherently multi-drug-resistant. On MacConkey agar, it is a non-lactose-fermenter. On blood agar, it appears a smooth and mucoid to wrinkled and dry, resembling P. stutzeri. Some morphotypes are bright yellow in color.
stenotrophomonas maltophilia
Stenotrophomonas (Xanthomonas) maltophilia is an inherent multi-drug-resistant organism. It is an oxi -, catalase +, motility +, pigmented, short Gram-negative rod. It is found in nosocomial infections and has been associated with IV and urinary catheter infections, respiratory infections, bacteremia, wound, pneumonia, and other miscellaneous infections. It may colonize the skin and/or respiratory tract, particularly of CF patients and is the 3rd most common GNR isolated in clinical specimens such as this one. Debilitated patients are most at risk for infection with this organism, particularly those in the ICU or who are on broad spectrum antibiotic therapy or otherwise immunocompromised. It thrives in moist hospital environments and appears as large, smooth, mucoid, glistening colonies with uneven edges on blood agar, and may produce a lavender-green to light purple pigment with a green discoloration on the agar. One unique identifier of this organism is that it smells like ammonia. The best treatment for those infected with this organism is SXT.
sphingomonas paucimobilis
Sphingomonas paucimobilis is a soil organism that doesn't usually cause infection in humans unless they are immunocompromised in some way. It is an inherent multi-drug-resistant organism (MDRO) that is oxidase +, motility +, glucose +, sucrose +, and bile esculin +. It is a straight GNR. On blood agar, it produces a yellow-to-orange pigment. If it grows on MacConkey agar (most fail to grow), it will appear as a NLF. This is an environmental bacterium that contaminates water and is occasionally associated with hospital water systems. It is never a part of the normal flora. Susceptible patients may become infected with the microbe upon exposure to contaminated medical devices or solutions. Additionally, the organism has been associated with catheter-related bacteremia, wound infections and UTIs. It is susceptible to SXT, chloramphenicol, ciprofloxacin and aminoglycosides.
elizabethkingia meningoseptica:
Elizabethkingia meningoseptica is a Gram-negative bacilli that is found in the environment. It is found in soil, wet soil, water, salt water, and is typically not a part of the normal human flora. It may be a laboratory contaminant, or it may be associated with cases of meningitis and sepsis in pediatric patients, particularly outbreaks in neonatal NICUs and amongst premature newborns. Some strains are linked to invasive meningitis, whereas others rarely cause endocarditis, pneumonia, bacteremia, soft tissue infections, sepsis and meningitis in immunocompromised adults with underlying medical conditions. More often than not, however, it is linked to contaminated lipid and water solutions, tap water, and venous catheter tubing. It grows well on blood and chocolate agars, but does not grow at all on CNA and may not grow on MacConkey agar. It is an obligate aerobe, non-fastidious, nonmotile, catalase +, oxidase +, indole +, urease -, mannitol +, PYR +. Colonies are either nonpigmented or a slight yellow color, shiny, and umbonate. Chryseobacterium spp, similar to this organism, produces an orange -to-yellow pigment. Vitek 2 sometimes mistakes this organism (E. meningoseptica) with Sphingobacterium spp.
Chryseobacterium spp:
Sphingobacterum spp:
acinetobacter spp:
Acinetobacter Spp:
Acinetobacter species are Gram-negative coccobacilli, nonmotile, oxidase -, strictly aerobic, nonfermenters. Some strains are beta-lactamase producers. Most of these are opportunistic pathogens, especially in immunocompromised patients, and are associated with nosocomial infections. This is an environmental microbe and has been isolated from water and soil. They are motile via twitching motility or swarming motility. This bacterium is hardy and somewhat mucoid due to the production of exopolysaccharide. This enables the bacteria to form a slimy biofilm and causes it to be instrinsicly resistant to many antimicrobials. In fact, this group of microbes is equiped with a couple of highly effective efflux pumps that enable it to "vomit out" many of the antimicrobials used to treat it with. Because of the biofilm production and presence of capsules, this organism can survive on surfaces such as countertops and plastics and catheters for long periods of time.
-Acinetobacter baumanii complex-glucose +, nonhemolytic; Can cause life-threatening infections; Tends to be multi-drug resistant; Associated with nosocomial infections, especially in the ICU; Can survive on the skin or dry surfaces for weeks at a time; Can form biofilms; Plasmid-mediated gene transfer and development of multi-drug resistance; Virulence factors (toxins) cause it to produce necrosis; Associated with VAP (ventilator-associated pneumonia)
-A. radioresistens-Radiation-resistant bacteria; GNCB; nonmotile; non-spore-forming; aerobic; nonfermenter; pleomorphic
-A. lwoffii-glucose -, nonhemolytic; linked to cases of meningitis; catheter-associated infections
-A. haemolyticus-hemolytic and potentially pathogenic
-A. calcoaceticus-causative agent of nosocomial pneumonia; Small, Gram-negative coccobacilli; mucoid, hemolytic colonies on blood agar; colorless on MacConkey agar; oxidase -, catalase +, arginine -, ornithine -, nitrate -, lysine -, ONPG -; opportunistic pathogen and part of the ABC (Acinetobacter baumannii complex)
Acinetobacter species are Gram-negative coccobacilli, nonmotile, oxidase -, strictly aerobic, nonfermenters. Some strains are beta-lactamase producers. Most of these are opportunistic pathogens, especially in immunocompromised patients, and are associated with nosocomial infections. This is an environmental microbe and has been isolated from water and soil. They are motile via twitching motility or swarming motility. This bacterium is hardy and somewhat mucoid due to the production of exopolysaccharide. This enables the bacteria to form a slimy biofilm and causes it to be instrinsicly resistant to many antimicrobials. In fact, this group of microbes is equiped with a couple of highly effective efflux pumps that enable it to "vomit out" many of the antimicrobials used to treat it with. Because of the biofilm production and presence of capsules, this organism can survive on surfaces such as countertops and plastics and catheters for long periods of time.
-Acinetobacter baumanii complex-glucose +, nonhemolytic; Can cause life-threatening infections; Tends to be multi-drug resistant; Associated with nosocomial infections, especially in the ICU; Can survive on the skin or dry surfaces for weeks at a time; Can form biofilms; Plasmid-mediated gene transfer and development of multi-drug resistance; Virulence factors (toxins) cause it to produce necrosis; Associated with VAP (ventilator-associated pneumonia)
-A. radioresistens-Radiation-resistant bacteria; GNCB; nonmotile; non-spore-forming; aerobic; nonfermenter; pleomorphic
-A. lwoffii-glucose -, nonhemolytic; linked to cases of meningitis; catheter-associated infections
-A. haemolyticus-hemolytic and potentially pathogenic
-A. calcoaceticus-causative agent of nosocomial pneumonia; Small, Gram-negative coccobacilli; mucoid, hemolytic colonies on blood agar; colorless on MacConkey agar; oxidase -, catalase +, arginine -, ornithine -, nitrate -, lysine -, ONPG -; opportunistic pathogen and part of the ABC (Acinetobacter baumannii complex)
Other non-lactose-fermenting, gram-negative rods/bacilli that may rarely cause respiratory and/or other types of infections:
Ralstonia spp:
Ralstonia spp are oxidase +, motility +, urease +, NLF GNRs that are inherently multi-drug resistant. This organism is associated with heavily contaminated medical equipment and solutions, and CF patients are especially at risk for infection. This slow-grower may take up to 72 hours to produce visible colonies on agar.
Acinetobacter spp:
Acinetobacter spp are plump Gram-negative coccobacilli that resist alcohol decolorization during the Gram staining process. They are oxidase -, smooth, opaque, raised, creamy, tiny colonies that may or may not be beta-hemolytic on blood agar and that are NLF on MacConkey agar. Colonies produce a rotton odor like a gym locker. This species may colonize the skin and respiratory tract, medical instruments, IV or UT catheters, and may result in nosocomial infection during warm seasons. Also an MDRO, infections with this organism are treated best with SXT. CF patients are at higher risk for infection.
Chryseomonas spp and Flavimonas spp:
Chryseomonas and Flavimonas spp are uncommon GNRs that are both oxidase -, NLF, short-to-medium, thick, often paired Gram-negative rods that may contaminate moist hospital environments, such as water and respiratory equipment, catheters, and may result in respiratory and other infections (sepsis, catheter, wound, peritonitis). CF patients are at higher risk of infection. Colonies appear wrinkled and stick to the surface of the agar with resistance to move. Colonies are rare, smooth and often yellow in color. Esculin hydrolysis may be used to differentiate these two organisms. Chryseomonas spp is bile esculin +, whereas Flavimonas spp is bile esculin -.
Achromobacter spp:
Achromobacter spp grows well on BA, CHOC, MAC, BHI broth, THIO and in blood culture bottles. It has been found on contaminated medical devices and is oxidase +, a NLF on MAC agar, glucose +, and is a medium, straight Gram-negative rod. It has been linked to respiratory infections and rare septicemia, especially in CF patients. Colonies appear as smooth and glistening.
Agrobacterium ochrobacter:
Agrobacterium spp is a short, NLF GNR that produces small colonies. It is most commonly encountered in the clinical setting as a nosocomial infection due to contaminated medical devices and is associated with water and a moist environment. It may colonize the GI tract, IV catheters, foreign bodies, peritoneal dialysis equipment, or respiratory tract. CF patients are at higher risk of infection. Animals are host reservoirs of this organism as well, and infection has occurred through dog and cat bites and scratches. It produces a capsule and is able to form biofilms.
Ralstonia spp are oxidase +, motility +, urease +, NLF GNRs that are inherently multi-drug resistant. This organism is associated with heavily contaminated medical equipment and solutions, and CF patients are especially at risk for infection. This slow-grower may take up to 72 hours to produce visible colonies on agar.
Acinetobacter spp:
Acinetobacter spp are plump Gram-negative coccobacilli that resist alcohol decolorization during the Gram staining process. They are oxidase -, smooth, opaque, raised, creamy, tiny colonies that may or may not be beta-hemolytic on blood agar and that are NLF on MacConkey agar. Colonies produce a rotton odor like a gym locker. This species may colonize the skin and respiratory tract, medical instruments, IV or UT catheters, and may result in nosocomial infection during warm seasons. Also an MDRO, infections with this organism are treated best with SXT. CF patients are at higher risk for infection.
Chryseomonas spp and Flavimonas spp:
Chryseomonas and Flavimonas spp are uncommon GNRs that are both oxidase -, NLF, short-to-medium, thick, often paired Gram-negative rods that may contaminate moist hospital environments, such as water and respiratory equipment, catheters, and may result in respiratory and other infections (sepsis, catheter, wound, peritonitis). CF patients are at higher risk of infection. Colonies appear wrinkled and stick to the surface of the agar with resistance to move. Colonies are rare, smooth and often yellow in color. Esculin hydrolysis may be used to differentiate these two organisms. Chryseomonas spp is bile esculin +, whereas Flavimonas spp is bile esculin -.
Achromobacter spp:
Achromobacter spp grows well on BA, CHOC, MAC, BHI broth, THIO and in blood culture bottles. It has been found on contaminated medical devices and is oxidase +, a NLF on MAC agar, glucose +, and is a medium, straight Gram-negative rod. It has been linked to respiratory infections and rare septicemia, especially in CF patients. Colonies appear as smooth and glistening.
Agrobacterium ochrobacter:
Agrobacterium spp is a short, NLF GNR that produces small colonies. It is most commonly encountered in the clinical setting as a nosocomial infection due to contaminated medical devices and is associated with water and a moist environment. It may colonize the GI tract, IV catheters, foreign bodies, peritoneal dialysis equipment, or respiratory tract. CF patients are at higher risk of infection. Animals are host reservoirs of this organism as well, and infection has occurred through dog and cat bites and scratches. It produces a capsule and is able to form biofilms.
staphylococcus aureus and mrsa:
S. aureus may cause respiratory and other infections, as may MRSA. On blood agar, S. aureus produces tan-golden colonies with large zones of Beta-hemolysis. It is catalase + and coagulase + and beta-lactamase +. PBP2 + organisms are considered presumptive MRSA. If the Vitek results show Oxacillin resistance, this is confirmation of MRSA. If there is also Vancomycin resistance, this is a VRSA. Infection control should be notified of positive MRSAs and they should be frozen and kept for future studies and research.
S. aureus is a catalase +, coagulase +, beta-lactamase (Cefinase) +, Gram-positive cocci in clusters. It grows on Mannitol Salt Agar (MSA). MRSA is PBP2 positive and resistant to Oxacillin. This is one of the top causes of chronic respiratory infections, particularly in CF patients.
bordetella pertussis and parapertussis (whooping cough):
Bordetella pertussis is the causative agent of "whooping cough". With the Tdap vaccination series and booster shots in the USA, cases have dropped significantly, however, in recent years, there have been sporadic outbreaks because shots that were thought to last a long time, if not a lifetime, have been found not to provide definitive lifelong immunity in some persons as once thought. Please refer to the CDC website for more information on endemic outbreaks in the USA. <http://www.cdc.gov/pertussis/outbreaks/about.html>. It has been recommended that cephalexin be used instead of penicillin in Bordet-Gengou charcoal agar because penicillin may actually inhibit some strains of B. pertussis.
Bordetella parapertussis is an aerobic, Gram-negative coccobacillus that grows on blood agar and causes pertussis in infants, similar to B. pertussis, and is urease negative at 4 hours and positive at 18 hours, oxidase negative, catalase +, citrate +, is nonmotile, forms small zones of beta-hemolysis on blood agar with a slight brownish coloration of the medium.
Bordetella bronchiseptica is also a rare cause of respiratory infections in immunocompromised patients who have had close contact with a pet, such as a dog, because it causes infectious bronchitis and acute tracheobronchitis in dogs (Kennel Cough). Cats and rabbits may also carry it. It stains as small, Gram-negative bacilli that look like "safety pins". It is highly motile due to peritrichous flagella, rapid urease +, oxidase +, catalase +.
Bordetella parapertussis is an aerobic, Gram-negative coccobacillus that grows on blood agar and causes pertussis in infants, similar to B. pertussis, and is urease negative at 4 hours and positive at 18 hours, oxidase negative, catalase +, citrate +, is nonmotile, forms small zones of beta-hemolysis on blood agar with a slight brownish coloration of the medium.
Bordetella bronchiseptica is also a rare cause of respiratory infections in immunocompromised patients who have had close contact with a pet, such as a dog, because it causes infectious bronchitis and acute tracheobronchitis in dogs (Kennel Cough). Cats and rabbits may also carry it. It stains as small, Gram-negative bacilli that look like "safety pins". It is highly motile due to peritrichous flagella, rapid urease +, oxidase +, catalase +.
Legionella: causative agent of legionella, legionairre's disease and pontiac fever
Corynebacterium diphtheriae: causative agent of diphtheria
Corynebacterium diphtheriae is the causative agent of diphtheria, a pseudomembrane formation on the back of the throat and tonsils, which can cause severe pharyngitis, difficulty breathing, forward breathing, closing up/blocking of the throat, and death. Today, it is part of the DTaP vaccination, so cases are extremely rare. Corynebacterium spp produces small, matte, white colonies on blood agar. It is a Gram-positive, catalase +, non-spore-forming, non-motile, lipophilic test +, aerobic bacilli/rod. It is often in pallisades formation (side-by-side), or looks like Chinese letters or drumsticks.
respiratory fungal infections: yeasts and moulds
Candida spp produce convex, white, globular colonies on blood agar, with or without "feet". Tests that should be performed incude the following: Germ tube test (if +, the yeast can be identified as C. albians. If germ tube -, a Vitek yeast card or Vitek 20 C API should be performed to identify the yeast). To perform the Germ Tube test, take a frozen germ tube vial (small, green top) out of the freezer and bring to room temperature. Make a heavy inoculum with a sterile (packaged) swab and touch at least 5 well-isolated colonies. Incubate in ambient air at 37 degrees Celsius for 2 -2.5 hours. Make a wet mount on a clean slide with 1 drop of inoculum and a coverslip. A positive Germ-tube test will look like the following image:
Gram stain of yeast will appear like this:
If you have a yeast, you also need to subculture it to a Cornmeal agar. Perform a regular 4-quadrant streak, and stab the agar 4x in the first quadrant where the 2nd quadrant overlays the first, placing a coverslip on top of this area to cover your stab marks. This encourages the growth of pseudohyphae and chlamydospores, which aids in the identification of the yeast, and which is used to correlate with the Vitek or API identification results.
1) Make sure you bring the Cornmeal agar to room temperature prior to inoculation
2) Remove the moisture from the lid of the agar with a piece of sterile gauze
3) Perform inoculation steps as written in the paragraph above, with stabs and coverslip
4) Incubate in the fungal incubator at 30 degrees Celsius for at least 24 hours, cover-side down to prevent condensation from dripping down onto the agar
5) Either leave in incubator or take out and place at room temperature
6) Pseudohyphae and chlamydospores take 3-5 days on average to appear. Remove the stage clip from the microscope, turn the light and the condenser down, and place the open plate on the flat stage. Read under 20x and 40x objective lenses and view around the area and edges of the coverslip and stab marks in the agar, where pseudohyphae production is enhanced.
7) Confirm results with reference books, API and/or Vitek results. See also the Mycology section on this website for other yeasts and their appearances. If unable to identify the organism in Bacteriology section, please refer to Mycology section for further aids on identification.
1) Make sure you bring the Cornmeal agar to room temperature prior to inoculation
2) Remove the moisture from the lid of the agar with a piece of sterile gauze
3) Perform inoculation steps as written in the paragraph above, with stabs and coverslip
4) Incubate in the fungal incubator at 30 degrees Celsius for at least 24 hours, cover-side down to prevent condensation from dripping down onto the agar
5) Either leave in incubator or take out and place at room temperature
6) Pseudohyphae and chlamydospores take 3-5 days on average to appear. Remove the stage clip from the microscope, turn the light and the condenser down, and place the open plate on the flat stage. Read under 20x and 40x objective lenses and view around the area and edges of the coverslip and stab marks in the agar, where pseudohyphae production is enhanced.
7) Confirm results with reference books, API and/or Vitek results. See also the Mycology section on this website for other yeasts and their appearances. If unable to identify the organism in Bacteriology section, please refer to Mycology section for further aids on identification.
Cystic fibrosis patients
Cystic fibrosis is a hereditary disorder that affects neonates, kids and adolescents. Throughout the body, disruption and dysfunction of the exocrine glands (secretory glands) occurs. Because excess, thick mucus is produced in the respiratory tract, it results in chronic pulmonary disease with chronic bacterial infections seen. The most common pathogens associated with infections in CF patients include the following:
Respiratory specimens collected for testing include the following:
- Pseudomonas aeruginosa-mucoid or nonmucoid
- Staphylococcus aureus
- Haemophilus influenzae
- Burkholderia cepacia complex
- Stenotrophomonas (xanthomonas) maltophilia
- Streptococcus pneumoniae
Respiratory specimens collected for testing include the following:
- Sputum
- Gagged sputum
- Induced sputum
- Pleural fluid
- Transtracheal aspirate
- Transthoracic lung aspirate
- Tissue biopsy
upper respiratory tract infections, other than strep throat (Nasopharyngeal culture)
This type of culture is utilized to isolate pneumococci, meningococci, haemophilus, and moraxella spp, and to detect the presence of S. pyogenes, S. aureus, C. diphtheriae, B. pertussis, B-hemolytic streptococci, S. pneumoniae, H. influenzae and parainfluenzae, N. meningitidis.
- Laryngitis (viruses)
- Laryngotracheobronchitis (M. pneumoniae, Rhinoviruses, Enteroviruses)
- Epiglottitis (H. influenzae type b)
- Abscesses (Fusobacterium, Bacteroides, Anaerobic cocci, S. pyogenes, Viridans streptococci)
- Diptheria (C. diptheriae)
- Whooping Cough (Bordetella pertussis)
- Rhinoscleroma (K. rhinoscleromatis, K. ozaenae)
- Thrush (Candida albicans)
- Periodontal (Eikenella corrodens, S. milleri, B. fragilis, Peptostreptococcus, Veillonella, Prevotella, Porphyromonas, Fusobacterium, Actinomyces israelii)
- Salivary Gland and Jaw (Staphylococcus aureus)
- Neck (Peptostreptococccus, Bacteroides, Prevotella, Porphyromonas, Fusobacterium, Actinomyces, Streptococci)
- Sinuses and Nasopharyngeal (Staphylococcus aureus, Molds, Viruses)
miscellaneous bench:
eye cultures:
Eye infections may be caused by bacteria, fungi, viruses or parasites.
Bacterial Conjunctivitis:
-Accounts for 20% of cases of conjunctivitis are caused by adenoviruses in kids and 14% in adults and are contagious;
-Inflammation of the conjunctiva
Blepharitis:
Staphylococcus epidermidis and S. aureus are the main causes, but Moraxella lacunata is also a cause, Viruses, Lice
-Symptoms include burning, itching, feeling that something is in the eye, crusting of the eyelids, inflammation of the margins/edges of the eyelids, irritation, redness
-Viral causes may result in a blisterlike eruption on the eyelids
-Chronic inflammation of the eyelid where the eyelashes grow
-The sebaceous glands near the eyelid margin are not functioning properly
-Caused by excess growth of bacteria that normally live on the skin that block the sebaceous (oil) glands or by allergies, viruses, or parasites
-Staphylococci
-Rosacea
-Allergic, Contact or Seborrheic Dermatitis
-Demodex spp eyelash mite (parasite) infestation/overgrowth/allergy
-Lice
-HSV
-Varicella-Zoster
-Molluscum contagiosum
-Infection typically lasts 2-4 weeks or it may be chronic
-Triggered by a horomone imbalance
Keratitis:
S. epidermidis, S. aureus, S. pneumoniae, P. aeruginosa
-Inflammation of the cornea with moderate-to-intense pain, decrease in vision, photophobia (light sensitivity), "gritty" sensation in the eye, and occasional discharge from the eye
-Acute or Chronic
-VIral (HSV, Shingles)
-Bacterial, following injury or contact lens use (contaminated lens cleaner)
-Fungal (Fusarium, Aspergillus spp, Alternaria spp, Candida spp)
-Amoebic (Acanthamoeba) from tap water or contaminated contact lens solution
-Parasitic (Onchocerca volvularis-following a bite by a blackfly; River Blindness)
-Superficial or ulcerative
Keratoconjunctivitis:
Same agents cause this that cause keratitis, and O. neonatorum, N. gonorrhoeae, and C. trachomatis can also cause
-Infection involoving the cornea and conjunctiva
-Dry eyes
Endophthalmitis:
S. aureus, S. epidermidis, S. pneumoniae, Other streptococci, N. meningitidis, P. aeruginosa, Other GNRs, Fungi, Virus: HSV, Parasites: Toxoplasma gondii, Toxocara
-Infection of the aqueous or vitreous humor of the eye that develops quickly and progresses quickly, and may lead to blindness; pain while moving the eye is common, as well as decreased vision
-May be a complication of all types of intraocular surgeries (example: cataract surgery)
-May be a result of a penetration trauma to the eye (example: sawdust, sauder, metal, etc...)
Lacrimal Infections:
Actinomyces, Propionibacterium propionicum
-Canaliculitis: inflammation of the canaliculus
-Rare, chronic inflammation of the lacrimal canals in which the eyelid swells up; thick, mucopurulent discharge
Dacryocystitis:
S. pneumoniae, S. aureus, S. pyogenes, H. influenzae
-Inflammation and infection of the lacrimal sac with pain, swelling, tenderness
-Secondary to blockage of nasolacrimal duct where it meets the lacrimal sac
-Complications may include corneal ulceration, orbital cellulitis, optic neuritis, proptosis, blindness
-May recur about 60% of the time
Dacryoadenitis:
S. pneumoniae, S. aureus, S. pyogenes, N. gonorroheae, Viruses (Epstein-Barr Virus, Mumps)
-Acute infection of the lacrimal gland (tear-producing gland) with pain, redness, swelling of the upper eyelid, conjunctival discharge
-Chronic inflammation due to other disorders in the body, such as autoimmune disorders like sarcoidosis, thyroid problems, orbital tumors
Folliculitis (Eyelid/Eyelashes):
S. epidermidis, S. saprophyticus, S. aureus, P. aeruginosa
-Infection of the hair follicles on the edges of the eyelid
Stye:
S. epidermidis, S. aureus, S. saprophyticus (rarely)
-Infection of the hair follicles on the edges of the eyelid
-An external stye (or sty) is also called a hordeolum
-At the base of the eyelid lie sebaceous (oil-producing) glands called the sebaceous glands of Zeis. These may become infected with bacteria, and this is the most common cause of a stye (blocked oil gland)
-At the base of the eyelid lie apocrine (sweat-producing) glands called the sweat glands of Moll, which may also become infected with bacteria.
-Styes are small red bumps that form on the outside of the eyelid.
-Internal styes form on the inside of the eyelid and are due to an infection of the apocrine sweat glands lining the inside of the eyelid (Meibomian sweat glands).
-Styes are small swellings and they are painful red bumps with tenderness and crusting or burning of the eye and they are a mixture of water, pus and bacteria, which can spread if the stye ruptures.
-Styes usually resolve on their own in 7-10 days (self-limiting)
-Chalzia is a type of chronic stye that does not resolve without antibiotics.
-Styes are usually caused by S. epidermidis or S. aureus bacteria
-Recommended treatment is warm compresses and cleansing the eye with mild soap, such as baby shampoo. Erythromycin ointment, bacitracin ointment or moxifloxacin eye drops are typically good antibiotic choices to resolve this infection.
Bacterial Conjunctivitis:
- Many bacteria can cause conjunctivitis, mainly Staphylococcus aureus, Staphylococcus epidermidis
- Others include: H. influenzae (kids), S. pneumoniae (kids), H. influenzae aegyptius (kids), Moraxella lacunata, Y. enterocolitica, F. tularensis, T. pallidum (syphilis), C. trachomatis, N. gonorrhoeae, S. pyogenes, Corynebacterium macginleyi
- Rarely: Propionibacterium, CNS
- Bacterial conjunctivitis is referred to as "pink eye"
- Infections may be secondary to viruses or other spread of other bacterial infections
- Infections may be secondary to traumatic wounds, scratches, abrasions, chemical accidents, and other conditions
- Yeasts, molds and mycobacterium may also cause non-bacterial conjunctivitis and most of the other conditions below
- In neonates, nessierial and chlamydial infections may occur during passage through an infected vaginal canal (trachoma is one of the leading causes of blindness in the world)
-Accounts for 20% of cases of conjunctivitis are caused by adenoviruses in kids and 14% in adults and are contagious;
-Inflammation of the conjunctiva
Blepharitis:
Staphylococcus epidermidis and S. aureus are the main causes, but Moraxella lacunata is also a cause, Viruses, Lice
-Symptoms include burning, itching, feeling that something is in the eye, crusting of the eyelids, inflammation of the margins/edges of the eyelids, irritation, redness
-Viral causes may result in a blisterlike eruption on the eyelids
-Chronic inflammation of the eyelid where the eyelashes grow
-The sebaceous glands near the eyelid margin are not functioning properly
-Caused by excess growth of bacteria that normally live on the skin that block the sebaceous (oil) glands or by allergies, viruses, or parasites
-Staphylococci
-Rosacea
-Allergic, Contact or Seborrheic Dermatitis
-Demodex spp eyelash mite (parasite) infestation/overgrowth/allergy
-Lice
-HSV
-Varicella-Zoster
-Molluscum contagiosum
-Infection typically lasts 2-4 weeks or it may be chronic
-Triggered by a horomone imbalance
Keratitis:
S. epidermidis, S. aureus, S. pneumoniae, P. aeruginosa
-Inflammation of the cornea with moderate-to-intense pain, decrease in vision, photophobia (light sensitivity), "gritty" sensation in the eye, and occasional discharge from the eye
-Acute or Chronic
-VIral (HSV, Shingles)
-Bacterial, following injury or contact lens use (contaminated lens cleaner)
-Fungal (Fusarium, Aspergillus spp, Alternaria spp, Candida spp)
-Amoebic (Acanthamoeba) from tap water or contaminated contact lens solution
-Parasitic (Onchocerca volvularis-following a bite by a blackfly; River Blindness)
-Superficial or ulcerative
Keratoconjunctivitis:
Same agents cause this that cause keratitis, and O. neonatorum, N. gonorrhoeae, and C. trachomatis can also cause
-Infection involoving the cornea and conjunctiva
-Dry eyes
Endophthalmitis:
S. aureus, S. epidermidis, S. pneumoniae, Other streptococci, N. meningitidis, P. aeruginosa, Other GNRs, Fungi, Virus: HSV, Parasites: Toxoplasma gondii, Toxocara
-Infection of the aqueous or vitreous humor of the eye that develops quickly and progresses quickly, and may lead to blindness; pain while moving the eye is common, as well as decreased vision
-May be a complication of all types of intraocular surgeries (example: cataract surgery)
-May be a result of a penetration trauma to the eye (example: sawdust, sauder, metal, etc...)
Lacrimal Infections:
Actinomyces, Propionibacterium propionicum
-Canaliculitis: inflammation of the canaliculus
-Rare, chronic inflammation of the lacrimal canals in which the eyelid swells up; thick, mucopurulent discharge
Dacryocystitis:
S. pneumoniae, S. aureus, S. pyogenes, H. influenzae
-Inflammation and infection of the lacrimal sac with pain, swelling, tenderness
-Secondary to blockage of nasolacrimal duct where it meets the lacrimal sac
-Complications may include corneal ulceration, orbital cellulitis, optic neuritis, proptosis, blindness
-May recur about 60% of the time
Dacryoadenitis:
S. pneumoniae, S. aureus, S. pyogenes, N. gonorroheae, Viruses (Epstein-Barr Virus, Mumps)
-Acute infection of the lacrimal gland (tear-producing gland) with pain, redness, swelling of the upper eyelid, conjunctival discharge
-Chronic inflammation due to other disorders in the body, such as autoimmune disorders like sarcoidosis, thyroid problems, orbital tumors
Folliculitis (Eyelid/Eyelashes):
S. epidermidis, S. saprophyticus, S. aureus, P. aeruginosa
-Infection of the hair follicles on the edges of the eyelid
Stye:
S. epidermidis, S. aureus, S. saprophyticus (rarely)
-Infection of the hair follicles on the edges of the eyelid
-An external stye (or sty) is also called a hordeolum
-At the base of the eyelid lie sebaceous (oil-producing) glands called the sebaceous glands of Zeis. These may become infected with bacteria, and this is the most common cause of a stye (blocked oil gland)
-At the base of the eyelid lie apocrine (sweat-producing) glands called the sweat glands of Moll, which may also become infected with bacteria.
-Styes are small red bumps that form on the outside of the eyelid.
-Internal styes form on the inside of the eyelid and are due to an infection of the apocrine sweat glands lining the inside of the eyelid (Meibomian sweat glands).
-Styes are small swellings and they are painful red bumps with tenderness and crusting or burning of the eye and they are a mixture of water, pus and bacteria, which can spread if the stye ruptures.
-Styes usually resolve on their own in 7-10 days (self-limiting)
-Chalzia is a type of chronic stye that does not resolve without antibiotics.
-Styes are usually caused by S. epidermidis or S. aureus bacteria
-Recommended treatment is warm compresses and cleansing the eye with mild soap, such as baby shampoo. Erythromycin ointment, bacitracin ointment or moxifloxacin eye drops are typically good antibiotic choices to resolve this infection.
Streptococcus pneumoniae (courtesty ASM, http://www.asmmicrobelibrary.org"); Alpha-hemolytic colonies with depressed centers like "donuts", entire, circular, often "flat", mucoid or non-mucoid, susceptible to optochin disk (14 mm or more), Gram-positive cocci in lancet-shaped pairs and chains, facultative anaerobe, bile esculin test +
ear cultures:
Otitis Externa:
Otitis Media:
- Outer ear infection, involving the auditory canal and outer skin
- May be acute or chronic, localized or diffuse
- May consist of a pustule or furuncle, erysipelas, or diffuse spreading (swimmer's ear)
- Chronic infections are due to irritation of drainage from the middle ear due to a burst eardrum with a suppurative discharge
- Malignant otitis externa is a necrotizing infection that spreads to other areas of soft tissue, cartilage and bone and can spread to the CNS, particularly in diabetic patients who are immunocompromised
- Pseudomonas aeruginosa, Proteus main causes and linked to chronic external ear infections
- Other causes: S. aureus, S. pyogenes, Other GNRs, V. alginolyticus, V. haemolyticus, Corynebacterium spp
- Chronic ear infections may be linked to the mold Aspergillus niger
- Candida albicans can also cause outer ear infection
- Also known as "Swimmer's Ear"
Otitis Media:
- Inner ear infection, involving the eustachian tube and the eardrum
- Streptococcus pneumoniae and S. pyogenes are a main cause and can travel from the throat (pharyngitis) up through the eustachian tube, causing inner ear infection
- Other causes include: H. influenzae, Nocardia, anaerobes, M. catarrhalis, Alloiococcus otitis, Turicella otitidis, Corynebacterium auris, GNRs, S. aureus, P. aeruginosa, Proteus
- Anaerobes linked to chronic infections: Fusobacterium spp, Bacteroides spp, Spirochetes, Peptostreptococcus, Prevotella spp
- Chronic ear infections may be linked to the mold Aspergillus niger
- Viruses such as RSV and Influenza may also cause
Culture swabs for outer ear infections and culture tympanocentesis fluid by needle aspiration for inner ear infections.
otitis externa (outer ear infections):
otitis media (inner/middle ear infection:
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Corynebacterium auris-Rarely linked to otitis media (coryneform, GPR)
Turicella otiditis-Rarely linked to otitis media (coryneform, irregular GPRs)
Turicella otiditis-Rarely linked to otitis media (coryneform, irregular GPRs)
acute and chronic sinusitis:
Adults and Adolescents:
S. pneumoniae, H. influenzae, M. catarrhalis, Rhinovirus (15%), Influenza (5%), Parainfluenza (3%), Adenovirus (<1%)
- H. influenzae (50%), S. pneumoniae (19%), S. pyogenes, M. catarrhalis, P. acnes, Anaerobes (2%), Enterobacteriaceae, Molds such as Aspergillus spp, etc...
S. pneumoniae, H. influenzae, M. catarrhalis, Rhinovirus (15%), Influenza (5%), Parainfluenza (3%), Adenovirus (<1%)
Facts:
-The sinuses include: sphenoidal, maxillary, paranasal, frontal and blockage of just one may cause infection in one sinus, pair of sinuses, or multiple sinuses.
-Also called "sinus infection" or "rhinosinusitis"
-Acute: Lasts <4 weeks
-Chronic: Lasts >12 weeks
-Bacterial, viral or fungal infection may cause
-May be caused by allergies
-May be caused by air pollution
-Patients who are immunocompromised, suffer from asthma, or who have Cystic Fibrosis are at risk for recurrent episodes
-The sinuses include: sphenoidal, maxillary, paranasal, frontal and blockage of just one may cause infection in one sinus, pair of sinuses, or multiple sinuses.
-Also called "sinus infection" or "rhinosinusitis"
-Acute: Lasts <4 weeks
-Chronic: Lasts >12 weeks
-Bacterial, viral or fungal infection may cause
-May be caused by allergies
-May be caused by air pollution
-Patients who are immunocompromised, suffer from asthma, or who have Cystic Fibrosis are at risk for recurrent episodes
iuds:
Infections due to biofilms formed on intrauterine devices are not uncommon, and typically occur during the insertion process, where anaerobes and microbiota may be pushed deep into the uterine canal, and are associated with pelvic inflammatory disease. Infections are typically polymicrobial, and involve anaerobes such as Gardnerella vaginalis and/or Mobiluncus spp. Infections may also occur along with STD infections such as Neisseria gonorrhoeae and/or Chlamydia trachomatis. Infection is referred to as bacterial vaginosis and may include organisms such as Prevotella spp, Streptococcus spp, Bacteroides spp, Peptostreptococcus spp, Enterobacteriaceae, Atopobium vaginae, Gemella spp, Fusobacterium spp, Leptotrichia spp, Mogibacterium spp, Porphyromonas spp, Proprionibacterium spp, Sphingomonas spp, Veillonella spp, Mycoplasma spp and Ureaplasma spp.
iv/cannulas: (See biofilm tab for more information)
There are a variety of catheters and cannulas. On blood agar, the catheter tip should be either rolled across the plate 4 times or placed on the plate with the lid replaced and shaken well across the plate.
Catheter infections and indwelling device infections may be caused by biofilm-producing organisms such as S. epidermidis, other types of staphylococci and CNS, and yeast such as C. albicans. IV and urinary catheters are 2 of the most common causes of nosocomial (hospital-acquired) bloodstream infections in the USA and cases are on the rise. In a recent research study performed on ICUs in the USA, research revealed that approximately 87% of primary catheter-associated bloodstream infections are in caused by central lines, and that of all urinary tract infections combined, about 95% are due to urinary catheters. About 3-8% of inserted central venous catheters result in bloodstream infection in the USA each year. Of these infections, the mortality rate from bloodstream infection ranges from 12-25%, which is high. In contrast, catheter-assocated infections due to urinary catheters is <5%. Many of the organisms that cause catheter-related UTI are highly resistant to antimicrobial drugs, and this scenario is occurring more and more and incidences are on the rise.
Organisms that commonly cause IV-catheter-associated infections include S. aureus, Stenotrophomonas maltophilia, CNS, S. epidermidis, Pseudomonas aeruginosa, C. albicans. Organisms that commonly cause UTI-catheter-associated infections include many of the Enterobacteriaceae, such as E. coli, Enterococcus, Pseudomonas, Klebsiella, Enterobacter, Citrobacter, Candida albicans.
References:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2963580/
See the tab on Bacterial biofilms for further information
Catheter infections and indwelling device infections may be caused by biofilm-producing organisms such as S. epidermidis, other types of staphylococci and CNS, and yeast such as C. albicans. IV and urinary catheters are 2 of the most common causes of nosocomial (hospital-acquired) bloodstream infections in the USA and cases are on the rise. In a recent research study performed on ICUs in the USA, research revealed that approximately 87% of primary catheter-associated bloodstream infections are in caused by central lines, and that of all urinary tract infections combined, about 95% are due to urinary catheters. About 3-8% of inserted central venous catheters result in bloodstream infection in the USA each year. Of these infections, the mortality rate from bloodstream infection ranges from 12-25%, which is high. In contrast, catheter-assocated infections due to urinary catheters is <5%. Many of the organisms that cause catheter-related UTI are highly resistant to antimicrobial drugs, and this scenario is occurring more and more and incidences are on the rise.
Organisms that commonly cause IV-catheter-associated infections include S. aureus, Stenotrophomonas maltophilia, CNS, S. epidermidis, Pseudomonas aeruginosa, C. albicans. Organisms that commonly cause UTI-catheter-associated infections include many of the Enterobacteriaceae, such as E. coli, Enterococcus, Pseudomonas, Klebsiella, Enterobacter, Citrobacter, Candida albicans.
References:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2963580/
See the tab on Bacterial biofilms for further information
Staphylococcus epidermidis, a common skin flora inhabitant, may cause an opportunistic infection, such as a urinary catheter or IV catheter infection, if introduced into a sterile area inadvertently. Organisms such as this have the ability to produce a sticky, adherent biofilm, making it difficult to treat and eradicate.
mrsa/esbl/mdro organisms:
MRSA stands for "Methicillin-Resistant Staphylococcus aureus. This bacterium is resistant to oxacillin. MRSA is resistant because it has developed resistance genes able to be passed on to daughter cells via resistance mechanisms such as bacterial conjugation, plasmids, transposons, binary fission (division). These genes render the baterium resistant to the beta-lactam antibiotics. These include the penicillins and the cephalosporins. This makes infections more difficult to treat and can be dangerous. MRSA mostly occurs as nosocomial infections, however, community-acquired infections are also on the rise. Individuals may also be colonized with MRSA without experiencing any symptoms of illness, therefore, many hospitals are now performing surveiilance cultures to see if an individual may be carrying MRSA in the nares (nostrils), axillary areas (armpits) or groin. It is estimated that at least 30% of healthcare workers do carry MRSA in their nares due to exposure to it. It is important to know if one is a carrier of MRSA, as it can be spread to immunocompromised individuals, who may become ill, and it can cause opportunistic infections if the conditions are just right, such as illness, hospitalization, surgery, or a break in the skin or mucosa due to injury or surgery.
Elderly patients, hospitalized patients, those in nursing homes and care centers, diabetics, those in prisons, the military, in tight quarters, and those who are otherwise immunocompromised are at higher risk for infections such as respiratory, urinary tract, wound, IV catheter or cannula, or post-surgical. After initial infection with MRSA, symptoms may rapidly progress in just 24-48 hours, and may invade the tissues and become resistant to treatment in just 72 hours. At this point, toxic shock syndrome, low blood pressure, shock, and even necrotic pneumonia may occur. A fever, followed by a rash that resembles pimples, spider bites or boils are typical initial symptoms. The bumps can spread, grow larger and more painful, and develop from smaller boils (furuncles) into larger, deeper, more painful ones (carbuncles). A serious complication of MRSA is sepsis (widespread infection) and/or endocarditis.
Community-acquired MRSA infections commonly manifest as smaller boils, impetigo, folliculitis, abscesses, ulcers, and cellulitis. More virulent strains are able to penetrate and invade the deeper tissues, and some lead to necrotizing fasciitis. Some even cause bone and joint infections. Surgical incision and drainage may be required to drain deep abscesses that have formed due to a virulent strain of MRSA.
Diagnosis is based on signs and symptoms, bacterial culture, susceptibility patterns, PCR (polymerase chain reaction), biochemical testing, and a rapid latex agglutination test called the PBP2 test that detects the PBP2 protein (a penicillin-binding protein) that causes S. aureus to be resistant to oxacillin. Another gene, the mecA gene, causes antibiotic resistance. This gene blocks B-lactam antibiotics and methicillin from penetrating the cell wall. See the tab on enzymes for more information about this mechanism. This group of antibiotics normally inactivates the enzymes needed for cell wall synthesis. Some strains are also resistant to Vancomycin (VRSA), but this is very rare. Cases are on the rise, however.
Elderly patients, hospitalized patients, those in nursing homes and care centers, diabetics, those in prisons, the military, in tight quarters, and those who are otherwise immunocompromised are at higher risk for infections such as respiratory, urinary tract, wound, IV catheter or cannula, or post-surgical. After initial infection with MRSA, symptoms may rapidly progress in just 24-48 hours, and may invade the tissues and become resistant to treatment in just 72 hours. At this point, toxic shock syndrome, low blood pressure, shock, and even necrotic pneumonia may occur. A fever, followed by a rash that resembles pimples, spider bites or boils are typical initial symptoms. The bumps can spread, grow larger and more painful, and develop from smaller boils (furuncles) into larger, deeper, more painful ones (carbuncles). A serious complication of MRSA is sepsis (widespread infection) and/or endocarditis.
Community-acquired MRSA infections commonly manifest as smaller boils, impetigo, folliculitis, abscesses, ulcers, and cellulitis. More virulent strains are able to penetrate and invade the deeper tissues, and some lead to necrotizing fasciitis. Some even cause bone and joint infections. Surgical incision and drainage may be required to drain deep abscesses that have formed due to a virulent strain of MRSA.
Diagnosis is based on signs and symptoms, bacterial culture, susceptibility patterns, PCR (polymerase chain reaction), biochemical testing, and a rapid latex agglutination test called the PBP2 test that detects the PBP2 protein (a penicillin-binding protein) that causes S. aureus to be resistant to oxacillin. Another gene, the mecA gene, causes antibiotic resistance. This gene blocks B-lactam antibiotics and methicillin from penetrating the cell wall. See the tab on enzymes for more information about this mechanism. This group of antibiotics normally inactivates the enzymes needed for cell wall synthesis. Some strains are also resistant to Vancomycin (VRSA), but this is very rare. Cases are on the rise, however.
dialysis water (dialysate):
Dialysate fluid should be sterile. This test is performed on the dialysate fluid to make sure that it isn't contaminated with unwanted microbes that could potential make a patient sick with infection or even be dangerous or deadly. Many laboratories test their deionized water weekly or monthly, and many laboratories test dialysate fluid for sterility for Nephrology departments in hospitals.
anaerobes:
Obligate anaerobes are strict anaerobes that will die if they are exposed to oxygen.
Facultative anaerobes are those that will tolerate oxygen if need be and can grow anaerobically and/or aerobically.
Microaerophilic organisms are those that need a minute amount of oxygen, increased amount of CO2, and nitrogen.
Anaerobic organisms are plated on anaerobic medium, such as CDC ANA blood agar, CDC ANA PEA agar, and KV-laked agar, and placed in a bag or box with an anaerobic generator (it creates an anaerobic environment and an indicator will change color if it is exposed to oxygen). QC is usually performed each week to make sure that an anaerobic environment is truly being achieved. The catalyst pouches should only be used once becuase they become inactivated after repeated use. Body fluids for anaerobic culture are also put into a broth medium, such as thioglycollate broth. Thioglycollate broth is stored at room temperature and in the dark so that there is a decreased absorption of oxygen by the medium.
Anaerobic infections are unique from aerobic infections in that they are usually polymicrobial. Some infections are synergistic, such as Vincent's angina.
Optimal specimens for culture of anaerobic organisms should be normally sterile body fluids, aspirates, or a syringe filled with pus, obtained prior to the administration of antibiotics. Swabs are the least desirable specimen for anaerobic culture. Tissue samples are also good choices for anaerobic culture. Body fluids include things such as CSF, pleural fluid, peritoneal fluid, pericardial fluid, amniotic fluid, bile, abscess pus/material, synovial fluid. If the initial Gram-stain shows Gram-positive, spore-forming rods, or the specimen looks green, yellow, or filled with pus, and was only ordered aerobically, add anaerobic media to the culture and order an anaerobic culture as well.
Normally, anerobic plates are only first opened after 48 hours of incubation, however, for example, if you notice that an aspirate of a deep wound is plated on blood agar plates aerobically and anaerobically, but at 24 hours there is growth on the anaerobic plate only, it is OK to go ahead and begin organism identification. The characteristic most commonly associated with the presence of strict, obligate anaerobes is the presence of a foul, putrid odor from cultures and tissue specimens.
Anaerobic susceptibility tests are helpful in the management of patients who have synovial infections.
Facultative anaerobes are those that will tolerate oxygen if need be and can grow anaerobically and/or aerobically.
Microaerophilic organisms are those that need a minute amount of oxygen, increased amount of CO2, and nitrogen.
Anaerobic organisms are plated on anaerobic medium, such as CDC ANA blood agar, CDC ANA PEA agar, and KV-laked agar, and placed in a bag or box with an anaerobic generator (it creates an anaerobic environment and an indicator will change color if it is exposed to oxygen). QC is usually performed each week to make sure that an anaerobic environment is truly being achieved. The catalyst pouches should only be used once becuase they become inactivated after repeated use. Body fluids for anaerobic culture are also put into a broth medium, such as thioglycollate broth. Thioglycollate broth is stored at room temperature and in the dark so that there is a decreased absorption of oxygen by the medium.
Anaerobic infections are unique from aerobic infections in that they are usually polymicrobial. Some infections are synergistic, such as Vincent's angina.
Optimal specimens for culture of anaerobic organisms should be normally sterile body fluids, aspirates, or a syringe filled with pus, obtained prior to the administration of antibiotics. Swabs are the least desirable specimen for anaerobic culture. Tissue samples are also good choices for anaerobic culture. Body fluids include things such as CSF, pleural fluid, peritoneal fluid, pericardial fluid, amniotic fluid, bile, abscess pus/material, synovial fluid. If the initial Gram-stain shows Gram-positive, spore-forming rods, or the specimen looks green, yellow, or filled with pus, and was only ordered aerobically, add anaerobic media to the culture and order an anaerobic culture as well.
Normally, anerobic plates are only first opened after 48 hours of incubation, however, for example, if you notice that an aspirate of a deep wound is plated on blood agar plates aerobically and anaerobically, but at 24 hours there is growth on the anaerobic plate only, it is OK to go ahead and begin organism identification. The characteristic most commonly associated with the presence of strict, obligate anaerobes is the presence of a foul, putrid odor from cultures and tissue specimens.
Anaerobic susceptibility tests are helpful in the management of patients who have synovial infections.
1: OBLIGATE AEROBE: Obligate aerobes have to have oxygen in order to grow because they cannot perform anaerobic respiration or fermentation. Because oxygen concentration is highest at the top of the THIO tube, that is where they will gather.
2: OBLIGATE ANAEROBE: Oxygen concentration is the lowest at the bottom of the THIO tube, therefore, this is where anaerobic bacteria will gather, thriving by anaerobic respiration and fermentation.
3: FACULTATIVE ANAEROBE: Many organisms are facultative anaerobes. This means that they will grow in the presence of or absence of oxygen. Most of these organisms will settle at the top, because more ATP energy is able to be generated there due to the presence of oxygen. But they will often form a veil-like turbidity throughout the THIO medium.
4: MICROAEROPHILIC ORGANISMS: These organisms need just a tiny bit of oxygen. Because of this, they will not gather at the very top, rather, just in the top half of the tube but not at the surface. They are poisoned by too much oxygen, and they cannot perform fermentation or anaerobic respiration.
5: AEROTOLERANT ORGANISMS: These organisms perform anaerobic respiration, but are not poisoned by oxygen either, so they are able to "tolerate" it and it does not have a negative effect on them. The organisms will be spread evenly throughout the THIO tube.
2: OBLIGATE ANAEROBE: Oxygen concentration is the lowest at the bottom of the THIO tube, therefore, this is where anaerobic bacteria will gather, thriving by anaerobic respiration and fermentation.
3: FACULTATIVE ANAEROBE: Many organisms are facultative anaerobes. This means that they will grow in the presence of or absence of oxygen. Most of these organisms will settle at the top, because more ATP energy is able to be generated there due to the presence of oxygen. But they will often form a veil-like turbidity throughout the THIO medium.
4: MICROAEROPHILIC ORGANISMS: These organisms need just a tiny bit of oxygen. Because of this, they will not gather at the very top, rather, just in the top half of the tube but not at the surface. They are poisoned by too much oxygen, and they cannot perform fermentation or anaerobic respiration.
5: AEROTOLERANT ORGANISMS: These organisms perform anaerobic respiration, but are not poisoned by oxygen either, so they are able to "tolerate" it and it does not have a negative effect on them. The organisms will be spread evenly throughout the THIO tube.
Bacteroides fragilis is an obligate anaerobe. It is a Gram-negative bacilli that resides in the colon as a part of the commensal flora, but can become an opportunistic pathogen if displaced into other areas of the body due to cancer or trauma or abscess. Surgery, underlying diseases, and peritonitis may introduce this bacterium into the bloodstream and cause infection and sepsis. In anaerobic infections, this is the most common anaerobe that is isolated. B. fragilis is part of the B. fragilis group, and accounts for the majority of Bacteroides spp within this group. It is beta-lactamase + and therefore resistant to penicillin.
This bacterium is associated with bloodstream infections, abscesses, wounds, peritoneal fluid and pleural fluid. Some strains produce colon inflammation and are linked to cancer.
This bacterium is associated with bloodstream infections, abscesses, wounds, peritoneal fluid and pleural fluid. Some strains produce colon inflammation and are linked to cancer.
Peptostreptococcus spp are anaerobic Gram-positive cocci. They may appear elongated if recovered from broth, such as blood or thioglycollate. They are part of the normal commensal flora of the human mouth, GI tract, and urogenital tract. They can be opportunistic pathogens if the conditions are right and are associated with sepsis. They are associated with the formation of abscesses in multiple areas of the body, such as the lung, liver, brain, breast, intraabdominal area, and others. They are typically recovered from polymicrobial infections, and may also cause necrotizing fasciitis. They are the second most commonly isolated anaerobes after Bacteroides fragilis. Peptostreptococcus magnus is the most common of this species to be isolated. P. tetradius is clinically significant, as are P. asaccharolyticus, P. anaerobius, P. prevotii, and P. micros.
Infections with this organism may occur in/on any body site and they are very difficult to isolate. They occur frequently in chronic infections which may include the following types of infections:
Fusobacterium spp are anaerobic, Gram-negative, fusiform bacilli. They tend to be longer and have pointed or rounded ends. They are associated with periodontal disease, Skin ulcers, and Lemierre's Disease. Some strains may be associated with colon cancer and ulcerative colitis. It produces a capsule, which makes it capable of producing bacterial biofilm.
F. necrophorum
F. nucleatum
Infections with this organism may occur in/on any body site and they are very difficult to isolate. They occur frequently in chronic infections which may include the following types of infections:
- Sinus, Tooth, Mastoiditis, Periodontitis
- Polymicrobial skin and soft tissue (gangrene, necrotizing fasciitis, ulcers, burns, bites, breast, rectal, anal infections)
- Abscesses
- Bacteremia, Endocarditis, Pericarditis
- Throat infections
- Vaginal infections
- Lower respiratory tract infections
- Post-surgical
- Post-dental
- Aspiration pneumonia
- Empyema
- Peritonitis
- Pelvic
- Bone and joint
- Trauma
- Immunocompromised
- Immunodeficient
- Immunosuppressed
- Those with cancer or malignancies
- Those with vascular disease
- Sickle cell anemics
- Diabetics
- Those with prosthetics or indwelling catheters or artificial heart valves
- Post-surgical patients
Fusobacterium spp are anaerobic, Gram-negative, fusiform bacilli. They tend to be longer and have pointed or rounded ends. They are associated with periodontal disease, Skin ulcers, and Lemierre's Disease. Some strains may be associated with colon cancer and ulcerative colitis. It produces a capsule, which makes it capable of producing bacterial biofilm.
F. necrophorum
F. nucleatum
Prevotella spp are Gram-negative anaerobes (bacilli) that are part of the commensal flora of the mouth, upper respiratory tract and urovaginal tract. They are sometimes isolated in mixed anaerobic infections of the respiratory tract, including:
P. intermedia is linked to periodontitis, necrotizing ulcerative gingivitis, and dentoalveolar abscesses. It is also associated with bacterial vaginosis.
P. bivia is associated with pelvic inflammatory disease.
P. brevis is linked to dental and oral infections.
P. bryantii is linked to gingivitis and periodontal disease.
P. nigrescens is linked to gingivitis and periodontal disease.
P. albensis is linked to gingivitis and periodontal disease.
- Aspiration pneumonia
- Empyema
- Lung abscess
- Sinusitis
- Chronic otitis media
- Burns
- Bites
- Nail infections (paronychia, infection of the folds of the nail)
- Brain abscesses
- Bone infections
- Bacteremia
- Periodontal disease
- Periodontal abscesses
- Rheumatoid arthritis
P. intermedia is linked to periodontitis, necrotizing ulcerative gingivitis, and dentoalveolar abscesses. It is also associated with bacterial vaginosis.
P. bivia is associated with pelvic inflammatory disease.
P. brevis is linked to dental and oral infections.
P. bryantii is linked to gingivitis and periodontal disease.
P. nigrescens is linked to gingivitis and periodontal disease.
P. albensis is linked to gingivitis and periodontal disease.
quality assurance/quality control (QA/QC) and antimicrobial testing by kirby-bauer disk diffusion and by etest, and antibiotic classes:
Quality Control and Assurance is performed daily, weekly and monthly to make sure that all reagents and disks are up-to-date and properly working, that kits are ready for use, and that paperwork is in compliance with regulations. It makes sure that the antibiotic-infused disks are working and that bugs are not becoming antibiotic-resistant and if they are, we can keep track of it. The goal of all of this is excellent patient care that is not compromised in any way. All of the methods and reagents we use to help us identify the organisms and their susceptibility for patient diagnosis and treatment need to work, so we make sure that things are working properly, are stored at the proper temperature and moisture and pH levels, and change out stains and reagents that may be getting ready to expire.
One advantage of the antimicrobial dilution tests is that they can detect varying degrees of organism sensitivity and resistance. The disk diffusion test is very specific and must be performed exactly as described. The media must be poured to a specific width. A McFarland standard of 0.5-0.63 turbidity must be made with the organism and saline prior to inoculation to the Mueller-Hinton agar plates in a specific manner to create a bacterial lawn of growth. Disks must be added to the agar within 15 minutes of plating the organism. And the plates must be incubated ASAP. And when testing Pseudomonas aeruginosa by KB testing, the cation content of the medium is critical when testing for antibiotic susceptibility. All of these components are critical or the test will not come out correctly and will yield inaccurate susceptibility results.
In a disk diffusion (Kirby-Bauer) susceptibility test, if the disks are placed on the inoculated medium and left at room temperature for >15 minutes before incubation, zones of larger diameter may result, because the antimicrobial-permeated disks begin to diffuse into the medium immediately once placed onto the agar. Additionally, if the poured medium in the KB plate is too THIN, this will also yield zone sizes that are too large for all antibiotics tested. On the other hand, if the bacterial suspension is not diluted to the proper concentration (>0.63 McFarland turbidity standard), disk zone sizes will appear too small. Too many organisms in the inoculum, or a >0.63 McFarland turbidity standard, is a factor that will make an organism appear to be more resistant on a disk diffusion susceptibility test than it really is. Media poured too THICK will also yield zone sizes that are too small for all antibiotics tested. The pH in the medium is also important. For example, if the pH is too low, then aminoglycoside zones will be too small and penicillin zones will be too large. It is also important to check and make sure that your disks are not expired. For example, if you use an outdated disk that has broken down, then the zone sizes will be too small.
In the agar disk diffusion susceptibility test (KB), as an antimicrobic agent diffuses away from the disk, the concentration gradient is decreased. In using this method of determining antimicrobial susceptibility, the size of the inhibition zone used to indicate susceptibility by correlating the zone size with minimum inhibitory concentrations. Zone diameters are measured in millimeters (mm) and compared to the CLSI antimicrobial standards.
The stock cultures needed for quality control (QC) testing of DNAse production are: Serratia marcescens/E. coli
The stock cultures needed for quality control (QC) testing of deamination activity are: Proteus mirabilis/E. coli
The stock cultures needed for quality control (QC) testing of oxidase production are: Pseudomonas aeruginosa/E. coli
The stock cultures needed for quality control (QC) testing of indole production are: E. coli/Klebsiella pneumoniae or Proteus mirabilis
The stock cultures needed for quality control (QC) testing of motility are: E. coli/K. pneumoniae
The stock cultures needed for quality control (QC) testing of catalase production are: Staphylococcus aureus/Streptococcus agalactiae
The stock cultures needed for quality control (QC) testing of coagulase production are: Staphylococcus aureus/Staphylococcus epidermidis
The stock cultures needed for quality control (QC) testing of novobiocin disk are: Staphylococcus epidermidis/Staphylococcus saprophyticus
The stock cultures needed for quality control (QC) testing of beta-lactamase production are: Staphylococcus aureus/Streptococcus agalactiae
The stock cultures needed for quality control (QC) testing of PYR are: Enterococcus faecalis/Streptococcus agalactiae
ANTIMICROBIALS:
-Penicillins (Inhibit cell wall synthesis)
The penicillins are bacteriacidal (they kill bacteria). They inhibit the transpeptidase enzyme/penicillin binding protein thereby blocking cell wall synthesis.
-Penicillins:
1) Penicilllin G, the original penicillin, was discovered by Alexander Fleming, and it was first used in 1941. It is in oral, intramuscular injection, or IV forms. It comes in
an aqueous (crystalline) form, as Procaine penicillin G or Benzathine penicillin G.
2) Penicillin V, an oral form, is stable in the stomach and treats strep throat/pharyngitis caused by Group A beta-hemolytic streptococci. Group B beta-hemolytic
streptococci is also universally susceptible to this penicillin.
-Aminopenicillins
The aminopenicillins provide better coverage of Gram-negative bacteria and are considered "broad-spectrum". They offer better penetration through the outer membranes of the cell plus enhanced binding to the transpeptidase enzyme, however, they are inhibited by bacteria that produce the enzyme penicillinase.
1) Amoxicillin (oral form) is better absorbed than ampicillin and is used to treat bronchitis, otitis media, sinusitis, and infections caused by Listeria spp.
2) Ampicillin, in oral or IV form, is often combined with other drugs such as one of the aminoglycosides (ex: gentamicin) for even broader-spectrum Gram-negative
coverage. Serious urinary tract infectons caused by the Enterobacteriaceae or those caused by Enterococcus spp are often successfully treated with this antibiotic
provided they are not resistant to it. It is also a good choice for treatment of infections caused by Haemophilus spp.
-Penicillinase-Resistant Antimicrobials
These antimicrobials are useful against beta-lactamase-producers. Beta-lactamase is an enzyme produced by some bacteria that destroys the beta-lactam rings of penicillin. Examples of microbes that may produce beta-lactamase include S. aureus, Neisseria spp, Haemophilus spp, Moraxella spp, and others. These provide good Gram-positive coverage (not good for Gram-negative infections), and they are good for treatment of streptococcal infections.
-Azlocillin
-Cloxacillin (Oral form)
-Dicloxacillin (Oral form)
-Flucloxacillin
-Methicillin (IV form) is highly effective against staphylcococcal infections. It is administered with caution, however, because it is linked to cases of interstitial nephritis.
-Nafcillin (IV form)
-Oxacillin (IV form)
-Temocillin
-Anti-Pseudomonal, Including Carboxypenicillins, Ureidopenicillins, Monobactams
These antimicrobials provide a wider coverage against Gram-negative infections, including those caused by Pseudomonas aeruginosa, as well as many infections caused by Gram-positive organisms.
A. Carboxypenicillins
1) Ticarcillin
2) Carbenicillin- This medication is administered in higher doses to be effective, because it offers lower activity. It must be administered with caution, because it produces
a high sodium load, cases of platelet dysfunction, and hypokalemia.
B. Ureidopenicillins
1) Piperacillin
2) Mezlocillin
-Aminoglycosides (Cover Gram-negative, aerobic bacterial infections; Given by IV, given IM or topically; Bind to the 30S ribosomal subunit or 50S ribosomal subunit, which is detrimental to protein synthesis of bacterial proteins needed for growth and survival)
This group of drugs diffuses across the cell wall and is often used in combination with penicillin to facilitate that diffusion. Aerobic Gram-negative Enterobacteriaceae infections are treated with these drugs (ex: E. coli, P. aeruginosa) and side effects may include renal toxicity or neuromuscular blockade with high doses).
-Amikacin (This drug provides the broadest spectrum of its class and is good for nosocomial infections that have developed resistance to other drugs).
-Gentamicin (This drug is the most commonly used and it is given in combination with the penicillins to treat nosocomial infections).
-Kanamycin
-Neomycin (This drug provides broad coverage, but is very toxic. It is used topically for skin infections).
-Netilmicin (This drug is used for preoperative coverage prior to GI surgery and is good because it is not absorbed by the GI tract. It is an oral antibiotic).
-Tobramycin (Given to treat infections with P. aeruginosa).
-Paromomycin
-Spectinomycin (This drug is given by intramuscular injection and treats gonorrhea. It is used with Doxycycline to treat Chlamydia trachomatis infections. There is no serious toxicity reported with this drug).
-Streptomycin (This is the oldest drug of this class and many organisms are now resistant to it).
-Anisamycin
-Rifaximin
-Penicillin Combinations (Synergism) (Help prevent resistance to the antibiotics); Beta-Lactamase Inhibitors
These antimicrobials provide broad-spectrum coverage against beta-lactamase producing Gram-positive organisms, Gram-negative organisms, and anaerobes such as B. fragilis.
-Amoxicillin/Clavulanate (Augmentin)
-Ampicillin/Sulbactam (Unasyn)
-Piperacillin/Tazobactam (Zosyn)
-Ticarcillin/Clavulanate (Timentin)
-The Cephalosporins (Disrupt cell wall synthesis)
This is a widely used group of antibiotics that have a beta-lactam ring, are resistant to beta-lactamase, and cover a broad-spectrum of Gram-negative and Gram-positive microbes (>20).
-First Generation (Streptococci and Staphylococci are more susceptible to this antimicrobial and it is an effective alternative to penicillin. One of these medications if often given before surgery to prevent infection from commensal skin flora.
-Cefazolin
-Cefalothin
-Cefalexin
-Cephalothin
-Cephaprin
-Cephradine
-Cephalexin
-Cefadroxil
-Second Generation (Streptococci and Staphylococci are more susceptible to these antimicrobials. They also cover more Gram-negative bacilli, as well as anaerobes such as B. fragilis).
-Cefoxitin (anaerobes)
-Cefuroxime (S. pneumoniae, H. influenzae, M. catarrhalis, community-acquired pneumonia, sinusitis, otitis media)
-Cefaclor
-Cefamandole
-Cefprozil
-Cefmetazole (anaerobes)
-Cefotetan (anaerobes)
-Cefonicid
-Cefdinir
-Loracarbef
-Third Generation (These provide both good Gram-negative and Gram-positive coverage, particularly for inpatient treatment of pneumonia and for community acquired pneumonia, meningitis, and pyelonephritis).
-Cefixime
-Cefdinir
-Cefoperazone
-Cefotaxime
-Cefpodoxime
-Ceftazadime
-Ceftibuten
-Ceftizoxime
-Ceftriaxone (This one has the best CSF penetration and covers the bacteria that cause meningitis. It is the first-line drug of choice for meningitis in neonates, children and adults. It is administered via an intramuscular injection to treat gonorrheal infections).
-Fourth Generation (This drug provides great Gram-negative coverage and good Gram-positive coverage. It is an extended spectrum 3rd generation cephalosporin).
-Cefepime (Good for treating infections caused by P. aeruginosa).
-Fifth Generation
-Ceftaroline fosamil
-Ceftobiprole
-Polypeptides (Block synthesis of isoprenyl pyrophosphate, a building block of the peptidoglycan cell wall)
-Bacitracin
-Colistin
-Polymyxin B
-Quinolones/Fluoroquinolones (Block the function of the DNA gyrase)
This class of drugs is a large and important group of antibiotics because they tend to be a safer group of drugs, they achieve high blood levels with oral absorption, and they penetrate extremely well into the tissues. They act by inhibiting the DNA gyrase, resulting in breakage of the DNA structure itself. They contain both a fluorine group and a nalidixic acid. Resistance has emerged due to point mutations in DNA gyrase subunits. There are very few side effects, but they may include GI irritation, cartilage damage, tendonitis, rupture involving the Achille's tendon, CNS side effects, overgrowth of C. difficile with severe diarrhea, hyperglycemia or hypoglycemia, or seizures in those with renal insufficiency. A high drug level in target tissue promotes bactericidal effects. One downside to this group of drugs is that most do not cover anaerobic infections. The class does, however, provide good coverage for Gram-negative infections such as those caused by P. aeruginosa, the Enterobacteriaceae, Salmonella, Shigella, Campylobacter, Aeromonas, Plesiomonas, Legionella, Brucella, Mycobacteria, and complicated UTIs, prostatitis, or epididymitis.
A) FIRST GENERATION
1) Nalidixic Acid
B) SECOND GENERATION
1) Norfloxacin
2) Ciprofloxacin (This provides poor Gram-positive coverage, except for S. aureus associated with osteomyelitis and B. anthracis).
3) Enoxacin
4) Ofloxacin
5) Levofloxacin (This drug provides expanded Gram-positive coverage, including S. pneumoniae, Enterococcis spp, Streptococcus, and Gram-negative coverage, including P. aeruginosa. This drug is used to treat skin infections and community-acquired pneumonia).
C) THIRD GENERATION
1) Gatifloxacin (This drug is used to treat respiratory infections, pneumonia, and anaerobic infections).
D) FOURTH GENERATION
1) Trovafloxacin (This drug provides good broad-spectrum activity, but there is a risk of liver toxicity, so it is rarely used).
2) Gemifloxacin
3) Moxifloxacin (This drug is used to treat respiratory infections, pneumonia, anaerobic infections, and intra-abdominal enteric infections such as peritonitis).
-Tetracyclines (Bind the 30S ribosomal subunits)
-Tetracycline (These oral antibiotics pass through the intestine without being absorbed. They provide broad-spectrum coverage).
-Tigecycline (Derived from Tetracycline, this drug is a glycycline, has similar broad-spectrum coverage, and is offered via IV. It is used for complicated skin and soft-tissue infections and provides activity against MRSA and VRE. A side effect includes GI upset).
-Minocycline
-Doxycycline (This drug is better absorbed with food. It is used to treat STDs caused by C. trachomatis, walking pneumonia caused by M. pneumoniae, an is an alternative to erythromycin. Animal and tick-borne diseases caused by Brucella and the Rickettsiae are treated with this drug. Sometimes this drug is used to treat bad cases of acne as well. Side effects may include GI irritation with nausea, vomiting, diarrhea, phototoxic dermatitis, renal and hepatic toxicity, discolored teeth, or depressed bone growth).
-Oxytetracycline
-Demeclocycline
-Macrolides (Bind to the 50S ribosomal subunit)
These are amongst the safest antibiotics and are typically well-tolerated by patients. They provide broad-spectrum coverage against Gram-positives, some Gram-negatives and atypical pathogens such as Legionella, Chlamydia pneumoniae, Mycoplasma spp. They have excellent absorption and fewer side effects than other drugs. Should side effects occur, they often involve mild abdominal pain due to intestinal peristalsis, rare cholestatic hepatitis, or prolonged QT syndrome. They are commonly given as second-line agents for skin and soft-tissue infections caused by organisms other than MRSA. They are also a good choice for urinary tract infections, sinusitis, otitis media, bronchitis, S. pneumoniae infections becoming resistant to other drugs.
-Azithromycin (Zithromax) (This drug is also called the "Z-Pack". It is an alternative treatment for syphilis but resistance has been emerging. Atypical mycobacterial infections may be treated with this drug, as well as Legionnaires disease caused by Legionella spp.)
-Clarithromycin (Biaxin) (This drug is used to treat a variety infections, including atypical mycobacterial infections).
-Dirithromycin
-Erythromycin (This was previously the drug of choice for community-acquired pneumonia. It may cause GI irritation, may cause prolonged QT syndrome, and therefore is not recommended for those with heart arrhythmias).
-Roxithromycin
-Telithromycin (This drug is effective against S. pneumoniae).
-Spiramycin
-Clindamycin (Oral or Cream; This blocks bacterial toxin production, but is only good for Gram-positive coverage, not Gram-negative coverage. It also covers anaerobes, such as B. fragilis, and is often given in combination with an aminoglycoside to cover aerobic Gram-negatives for penetrating wound infections of the abdomen. It is a good choice in combination with another drug to treat peritonitis. It is also used to treat Bacterial Vaginosis, acne vulgaris, rosacea, toxic shock syndrome, anaerobic aspiration pneumonia, lung abscesses. Use of this drug may result in overgrowth of C. difficile and/or pseudomembranous colitis with severe diarrhea because it also destroys the normal flora of the GI tract.
-Carbapenems (Inhibit cell wall synthesis)
These antimicrobials provide one of the broadest coverage for Gram-positive, Gram-negative and anaerobic microbes. They remain stable to beta-lactamases, and they are very small molecules that can easily pass through porin channels, reducing the wear-and-tear on the kidneys and are a safer choice of therapy. Given with Cilastin, they inhibit a natural enzyme found in the kidneys (dihydropeptidase) that would normally otherwise break down Imipenem.
-Imipenem (Given with caution, as it may cause seizures in some patients)
-Ertapenem (Requires only 1 daily dose of IV administration and drug of choice for empiric coverage of severe diabetic food infections that tend to be polymicrobial)
-Meropenem (This drug is as powerful as Imipenem and is stable against dihydropeptidase on its own, so Cilastin is not needed; It has a reduced potential for causing seizures)
-Doripenem (Newer)
-Lipopeptides (Disrupt cell membrane potential/charge)
-Daptomycin (This drug is a cyclic lipopeptide that kills Gram-positive bacteria by altering the microbe cell-membrane electrical potential charge and disrupting transport. It only provides Gram-positive coverage, NO Gram-negative coverage. It is active against MRSA and VRE. Its main side effect is myopathy with CPK elevation, so its use is monitored closely).
-Lincosamides (Bind to the 50S ribosomal subunit)
-Clindamycin
-Lincomycin
-Glycopeptides (Inhibit peptidoglycan synthesis)
-Teicoplanin
-Vancomycin (This drug is given by IV only and covers ALL Gram-positive microbes, including MRSA and VRE. It is given to treat S. epidermidis catheter infections, staphylococci and streptococci-associated endocarditis, and inhibits peptidoglycan synthesis earlier than penicillin. It also acts synergistically with the aminoglycosides).
-Telavancin
-Dalbavancin
-Oritavancin
-Oxazolidinones (Inhibit cell wall synthesis)
-Linezolid (Covers Vancomycin-Resistant Enterococcus; It is given in IV or Oral form, and is efective against resistant Gram-positive bugs. It is often given in combination with a broad-spectrum beta-lactam drug for empiric coverage of hospital-acquired pneumonia and for infections caused by S. aureus or MRSA. It is expensive and it may cause bone marrow suppression, thrombocytopenia, anemia or neutropenia. GI irritation, or headaches as side effects.
-Monobactams (Inhibit cell wall synthesis)
These antimicrobials provide great Gram-negative coverage.
-Aztreonam (Used to treat nosocomial infections, multi-drug-resistant organisms, and Gram-negative bacteria, including P. aeruginosa; Given in combination with other drugs, such as Vancomycin or Clindamycin).
-Nitrofurans (Treat UTIs/kidney infections)
-Nitrofurantoin (Macrobid)
-Furazolidone
-Other, Including AntiRibosomal Drugs:
The antiribosomal drugs block ribosomal action, inhibiting cellular growth and survival. Some bind the 50S subunit (large) of the ribosome, whereas others bind the 30S subunit (small) of the ribosome.
-Chloramphenicol (This is a very broad-spectrum drug that is inexpensive and kills most clinically significant bacteria, including Gram-positives, Gram-negatives and anaerobes such as B. fragilis. It may have rare but severe side effects, such as bone marrow depression, aplastic anemia, toxicity to the liver and kidneys, abdominal distention, or cyanosis. This is typically given as a last alternative, when there is no alternate antibiotic. It is given to cover bacterial meningitis when the organism is not known because of its excellent penetration into the CSF. It is also a good alternative for patients who are allergic to penicillin or for those who cannot take tetracycline).
-Fosfomycin
-Metronidazole (Flagyl)
-Mupirocin (Bactroban)
-Quinupristin/Dalfopristin (Synercid) (This is an older antibiotic effective in treating some drug-resistant organisms. It is used only when absolutely necessary and is active against Gram-positive microbes. It is used to treat life-threatening infections with VRE or complicated skin infections caused by MRSA or S. pyogenes. Side effects may include hyperbilirubinemia, pain at the infusion site, arthralgia or myalgias).
-Thiamphenicol
-Tinidazole
-Trimethoprim and Trimethoprim/Sulfamethoxazole (This drug is an antimetabolite that blocks nucleotide and DNA formation by blocking Th4, which bacteria require, make themselves, and use in PABA. Bacteria end up utilizing the sulfa drug, thinking it is PABA, which competitively inhibits production of TH4, disrupting the bacteria. It acts synergistically to inhibit many Gram-positives and Gram-negatives. It is an oral antibiotic, and it is excreted in the urine. Side effects may include nausea, vomiting, diarrhea, and skin rashes and itching. This drug is commonly used to treat respiratory infections, ear infections, sinusitis, bronchitis, pneumonia, GI tract infections, urinary tract infections, prostatitis, urethritis, and pneumonia).
-Sulfanomides (Inhibit folate synthesis)
-Mafenide
-Sulfacetamide
-Sulfadiazine
-Silver sulfadiazine
-Sulfadimethoxine
-Sulfamethizole
-Sulfamethoxazole
-Sulfanilimide
-Sulfasalazine
-Sulfisoxazole
-Trimethoprim-Sulfamethoxazole (Bactrim/Septra)
-Sulfonamidochrysoidine (Prontosil)
Terms:
A. Bactericidal-Kills bacteria
B. Bacteristatic-Inhibits bacterial growth/Reduces bacteria
C. Synergism-This term describes when combined antimicrobial drugs are clearly more effective than the most active drug alone; The effect of combined antimicrobials is greater than the sum of the effects observed with the two drugs independently. Synergism can be determined by the following in vitro methods: a) kill-curve method, b) checkerboard method using an MIC endpoint, c) checkerboard method using an MBC endpoint. Synergism should be assessed when combining antibiotic therapy.
D. Antagonism-This term describes when combined antimicrobial drugs are clearly less effective than the most active drug alone
E. Minimum Bactericidal Concentration (MIC)-This is the smallest concentration of antimicrobial agent that prevents growth in subculture or results in a 99.9% decrease of the initial inoculum
F. Minimal Inhibitory Concentration (MIC)-This is the smallest amount of test antimicrobial that will inhibit visible growth of a microbe. It is also the end point of the tube dilution test for antibiotic susceptibility.
G. Prophylaxis-This is preventative medicine, and includes antimicrobials (usually broad-spectrum) given as a measure to prevent disease, as opposed to specific disease treatment. These are antimicrobials given before surgery or dental procedures, particularly in susceptible patients, as a means to prevent opportunistic pathogens from causing a post-procedure infection. This term also describes antimicrobials that are given to treat a wide variety of microbes (Gram-positive, Gram-negative and anaerobes) until a definitive diagnosis can be made while laboratory cultures and susceptibility tests are being performed, until the drug can be changed to a more specific drug to target a specific microbe.
H. Empiric-Empiric therapy is the term for when antibiotics or antimicrobials are administered to known microbes. For example, once laboratory cultures and susceptibility testing provide definitive identification and susceptibility patterns, specific drugs may be administered to treat/target specific microbes.
I. ETEST-This is the Epsilometer Test. It is an in vitro test used by the microbiology laboratory to test susceptibilities by determining the MIC (minimal inhibitory concentration) to see if a microbe is sensitive to, or susceptible to, a specific antimicrobial. This aids physicians in finding out what are the best drugs to treat an individual's particular infection. In this test, a ready-to-use strip impregnated with a specific predefined gradient of antibiotic is applied to the surface of an agar plate inoculated with a dilution of microbe and saline (the test strain). From the second the strip touches the agar, there is an instantaneous diffusion of the drug to the agar. The agar plate must be incubated for 18-24 hours (sometimes for up to 48 hours for slower-growing strains) and the plate observed the following day for a symmetrical inhibition ellipse. The MIC is read/interpreted where the edge of the ellipse intersects the ETEST strip.
J. MDRO-Multi-drug resistant organism; This is a microbe that is resistant to many of the commonly used antimicrobials normally used to treat it; Typically an organism that is resistant to more than 3 classes of antimicrobials, or susceptible to <2 classes of drugs
K. ESBL-Extended spectrum beta-lactamase; Beta-lactamase is an enzyme produced by some microbes that render them resistant to the penicillins via the enzyme's ability to break the beta-lactam ring present in the penicillins and the cephalosporins
L. Kirby-Bauer Disk Diffusion-In this test, a bacterial lawn is swabbed onto the surface of a Mueller-Hinton agar or a Sheep's blood-based Mueller-Hinton agar with a dilution of test organism. Antibiotic-impregnated filter paper disks are added to the surface of the agar and it is incubated for 18-24 hours. Zones of clearing are measured in millimeters the following day, and compared with CSM or CLSI standards and interpreted as Susceptible, Intermediate or Resistant.
A test that can be used to monitor bactericidal activity during antimicrobial therapy in cases of endocarditis is the Schilchter test.
GC and Group B Streptococcus Screens:
The B-hemolytic streptococci found in Lancefield Group B streptococci, Streptococcus agalactiae, are part of the normal vaginal flora and mucosa in some females, and have been shown to be associated with peurperal fever (childbirth fever) and sometimes fatal neonatal meningitis (neonatal purulent meningitis) and endocarditis. In fact, this organism is the most common organism associated with neonatal purulent meningitis. S. agalactiae (beta-hemolytic, Group B streptococci) are facultative anaerobes, catalase -, hippurate hydrolysis +, bile esculin hydrolysis -, bacitracin susceptibility disk resistant, and optochin susceptibility disk resistant. They are also CAMP-positive. On Gram-stain, they form Gram-positive cocci in chains and pairs. They will agglutinate in the PathoDX streptococci latex agglutination test with the Group B latex test reagent. This organism is universally susceptible to penicillin. Susceptibility tests are typically not performed unless requested by the doctor, or if the patient is allergic to penicillin.
S. agalactiae is found in the urogenital tract of about 30% of women. Because this organism is the most common organism associated with neonatal purulent meningitis, endocervical swabs are used to swab the cervix of a pregnant female at 35-37 weeks gestation and sent to microbiology for culture to rule out the presence of the organism. Swabs are placed into LIM broth and incubated in CO2 for the first 24 hours. After 24 hours, the broth is inoculated onto Neomycin blood agar and incubated in CO2 for another 24 hours. Plates are then observed at 24 and 48 hours after incubation for beta-hemolysis and tested with a streptococcal latex agglutination test for the presence of S. agalactiae. S. agalactiae presents as small, beta-hemolytic colonies with narrow zones of beta hemolysis.
S. agalactiae is found in the urogenital tract of about 30% of women. Because this organism is the most common organism associated with neonatal purulent meningitis, endocervical swabs are used to swab the cervix of a pregnant female at 35-37 weeks gestation and sent to microbiology for culture to rule out the presence of the organism. Swabs are placed into LIM broth and incubated in CO2 for the first 24 hours. After 24 hours, the broth is inoculated onto Neomycin blood agar and incubated in CO2 for another 24 hours. Plates are then observed at 24 and 48 hours after incubation for beta-hemolysis and tested with a streptococcal latex agglutination test for the presence of S. agalactiae. S. agalactiae presents as small, beta-hemolytic colonies with narrow zones of beta hemolysis.
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Streptococcus agalactiae is positive with the PathoDX latex reagent for Group B. Place 2 drops of reagent 1 and 2 drops of reagent 2 in a glass tube. With a wooden stick applicator, touch 5 colonies and mix into the tube. Add 4 drops of reagent 3 and mix again. Place 1-2 drops in 1 circle on the card. Place 1 drop of Group B latex reagent (pink cap) in the same circle. Mix and rock the card for 15-60 seconds. If agglutination (clumping) appears, the test is positive for Group B strep.
It is important to identify Group B streptococci, or S. agalactiae, or N. gonorrhoeae, especially that a pregnant woman or woman of child-bearing age may be carrying because if passed to an infant during the pregnancy or birthing process, the infant could become very ill with meningitis due to the Group B streptococcus, or develop conjunctivits or other illness due to N. gonorrhoeae.
THE HACEK BACTERIA:
The HACEK bacteria are all fastidious, slow-growing bacteria that require CO2 for growth, do not grow on MAC agar, and all ferment glucose/dextrose. Some strains require 2-7 days of growth for colonies to appear on the agar. They grow well on CHOC agar.
These bacterium are all part of the normal oropharyngeal flora, however, they are all associated with acute infective endocarditis, bacteremia, abscesses, peritonitis, middle ear infections, conjunctivitis, pneumonia, arthritis, osteomyelitis, and periodontal infections.
Haemophilus spp: (Facultative Anaerobes)
These are Gram-negative coccobacilli that are catalase +, oxidase +, and many produce beta-lactamase.
1) H. influenzae-
H. influenzae does not grow well, if at all, on the blood or MAC agars, but grows well on CHOC agar. In fact, the only time it grows on BAP is as a satellite phenomenon around other bacterium such as S. aureus, where it grows in the hemolytic zone around that bacterum. This is due to the fact that S. aureus releases factor V, needed for the growth of H. influenzae. Colonies are convex, pale, smooth, transparent and gray-to-tan/orange with a flowery scent to them. It needs both X and V factors to grow, and will grow only in quadrants III and IV of the H. Quad plate, without producing hemolysis. It is catalase +, oxidase + and B-lactamase + or variable.
2) H. parainfluenzae-
H. parainfluenzae is similar to H. influenzae but it does not need factor X to grow, just factor V, and does not produce hemolysis. Therefore, it grows in quadrants II, III and IV without hemolysis.
3) H. haemolyticus-
This one needs both factors X and V to grow, and produces hemolysis, therefore, on the H. Quad plate, it grows in quadrants III and IV but also produces hemolysis, unlike H. influenzae.
4) H. parahaemolyticus-
This one only requires factor V to grow, and also produces hemolysis, therefore, it grows in quadrants II, III and IV and produces hemolysis.
Aggregatibacter (Formerly Actinobacillus spp:
Aggregatibacter spp are bipolar-staining Gram-negative rods in singles, pairs and chains. They grow better with moisture (sterile gauze, moistened and added).
1) A. actinomycetemcomitans-
This species produces pinpoint colonies that are rough, sticky, adhering to the agar with a slight greenish tinge, and star-like in appearance. It is catalase +, lactose -, indole - and urease -.
2) A. ureae-
This species resembles the Pasteurellae and is urease +.
3) A. suis-
This species resembles A. lignieresii and A. equuli, but the colonies are beta-hemolytic and urease +, bile esculin +, and lactose +.
4) A. ligniersii-
This species resembles A. equuli and is urease +.
5) A. hominis-
6) A. equuli-
This species produces small colonies at 24 hours that are sticky and adherent to the agar, smooth and rough, nonhemolytic, and urease +.
7) A. aphrophilus-
8) A. paraphrophilus-
Cardiobacterium hominis:
This species grows better with a piece of moistened sterile gauze added to the bag. It produces pleomorphic Gram-negative bacilli with one rounded end and one tapered end making them look like "teardrops" that may form clusters or rosettes. Colonies are small, slightly alpha-hemolytic, smooth, round, glistening and opaque and may pit the agar. It is indole +, catalase -, mannitol +, sorbitol +, and oxidase +. It may also produce beta-lactamase. It is part of the normal oropharyngeal and upper respiratory flora, but it may cause endocarditis or infection of the heart valves.
C. valvarum-
Eikenella corrodens-
This organism is a fastidious, pleomorphic Gram-negative bacillus/coccobacillus. It is a facultative anaerobe. About half of the strains "pit the agar", whereas the other half do not. Colonies are small, gray, and produce a slight greening of the agar underneath them. This bacteria smells faintly of bleach. It is oxidase +, indole -, catalase -, urease -. It is linked to periodontal disease and, rarely, acute bacterial endocarditis and infection of the heart valves. It is part of the commensal mouth and upper respiratory tract flora, and therefore is also linked to human bite infections. Diabetic and immunocompromised patients are at higher risk for infection with this microbe. Infections may produce a very foul odor, similar to that of anaerobic infections. It is typically susceptible to penicillin, cephalosporins, the fluoroquinolones and tetracycline, however, it is intrinsicly resistant to the macrolides, clindamycin and metronidazole.
Kingella spp:
1) K. denitrificans-This species produces short, plump Gram-negative coccobacilli that are catalase -. Colonies are small, nonhemolytic, and pit the agar. Colonies will also grow on Thayer-Martin or Martin-Lewis agar. It is nitrate reduction +.
2) K. kingae-This species produces short, plump Gram-negative coccobacilli that are catalase -. Colonies are small with a narrow zone of beta-hemolysis and may pit the agar. This organism is associated with acute bacterial endocarditis, meningitis, bacteremia, septic arthritis, osteomyelitis, spinal infections and lower respiratory tract infections. It is part of the normal throat flora, but is an opportunistic pathogen.
These bacterium are all part of the normal oropharyngeal flora, however, they are all associated with acute infective endocarditis, bacteremia, abscesses, peritonitis, middle ear infections, conjunctivitis, pneumonia, arthritis, osteomyelitis, and periodontal infections.
Haemophilus spp: (Facultative Anaerobes)
These are Gram-negative coccobacilli that are catalase +, oxidase +, and many produce beta-lactamase.
1) H. influenzae-
H. influenzae does not grow well, if at all, on the blood or MAC agars, but grows well on CHOC agar. In fact, the only time it grows on BAP is as a satellite phenomenon around other bacterium such as S. aureus, where it grows in the hemolytic zone around that bacterum. This is due to the fact that S. aureus releases factor V, needed for the growth of H. influenzae. Colonies are convex, pale, smooth, transparent and gray-to-tan/orange with a flowery scent to them. It needs both X and V factors to grow, and will grow only in quadrants III and IV of the H. Quad plate, without producing hemolysis. It is catalase +, oxidase + and B-lactamase + or variable.
2) H. parainfluenzae-
H. parainfluenzae is similar to H. influenzae but it does not need factor X to grow, just factor V, and does not produce hemolysis. Therefore, it grows in quadrants II, III and IV without hemolysis.
3) H. haemolyticus-
This one needs both factors X and V to grow, and produces hemolysis, therefore, on the H. Quad plate, it grows in quadrants III and IV but also produces hemolysis, unlike H. influenzae.
4) H. parahaemolyticus-
This one only requires factor V to grow, and also produces hemolysis, therefore, it grows in quadrants II, III and IV and produces hemolysis.
Aggregatibacter (Formerly Actinobacillus spp:
Aggregatibacter spp are bipolar-staining Gram-negative rods in singles, pairs and chains. They grow better with moisture (sterile gauze, moistened and added).
1) A. actinomycetemcomitans-
This species produces pinpoint colonies that are rough, sticky, adhering to the agar with a slight greenish tinge, and star-like in appearance. It is catalase +, lactose -, indole - and urease -.
2) A. ureae-
This species resembles the Pasteurellae and is urease +.
3) A. suis-
This species resembles A. lignieresii and A. equuli, but the colonies are beta-hemolytic and urease +, bile esculin +, and lactose +.
4) A. ligniersii-
This species resembles A. equuli and is urease +.
5) A. hominis-
6) A. equuli-
This species produces small colonies at 24 hours that are sticky and adherent to the agar, smooth and rough, nonhemolytic, and urease +.
7) A. aphrophilus-
8) A. paraphrophilus-
Cardiobacterium hominis:
This species grows better with a piece of moistened sterile gauze added to the bag. It produces pleomorphic Gram-negative bacilli with one rounded end and one tapered end making them look like "teardrops" that may form clusters or rosettes. Colonies are small, slightly alpha-hemolytic, smooth, round, glistening and opaque and may pit the agar. It is indole +, catalase -, mannitol +, sorbitol +, and oxidase +. It may also produce beta-lactamase. It is part of the normal oropharyngeal and upper respiratory flora, but it may cause endocarditis or infection of the heart valves.
C. valvarum-
Eikenella corrodens-
This organism is a fastidious, pleomorphic Gram-negative bacillus/coccobacillus. It is a facultative anaerobe. About half of the strains "pit the agar", whereas the other half do not. Colonies are small, gray, and produce a slight greening of the agar underneath them. This bacteria smells faintly of bleach. It is oxidase +, indole -, catalase -, urease -. It is linked to periodontal disease and, rarely, acute bacterial endocarditis and infection of the heart valves. It is part of the commensal mouth and upper respiratory tract flora, and therefore is also linked to human bite infections. Diabetic and immunocompromised patients are at higher risk for infection with this microbe. Infections may produce a very foul odor, similar to that of anaerobic infections. It is typically susceptible to penicillin, cephalosporins, the fluoroquinolones and tetracycline, however, it is intrinsicly resistant to the macrolides, clindamycin and metronidazole.
Kingella spp:
1) K. denitrificans-This species produces short, plump Gram-negative coccobacilli that are catalase -. Colonies are small, nonhemolytic, and pit the agar. Colonies will also grow on Thayer-Martin or Martin-Lewis agar. It is nitrate reduction +.
2) K. kingae-This species produces short, plump Gram-negative coccobacilli that are catalase -. Colonies are small with a narrow zone of beta-hemolysis and may pit the agar. This organism is associated with acute bacterial endocarditis, meningitis, bacteremia, septic arthritis, osteomyelitis, spinal infections and lower respiratory tract infections. It is part of the normal throat flora, but is an opportunistic pathogen.
The "satellite test" is used as a tool to help in the identification of H. influenzae. Streaks of S. aureus are made on sheep's blood agar. The hemolysis of the RBCs in the agar releases factor V, or NAD, needed for the growth of H. influenzae. Therefore, if you streak across the streak of S. aureus with your unknown organism suspected of being a possible Haemophilus spp, if it is H. influenzae, colonies will "satellite" around the S. aureus by growing near the zone of clearing or hemolysis where factor V or NAD has been released. It will not grow outside the hemolytic zone.
nasopharyngeal colonization and opportunistic pathogens
neisseria spp:
Neisseria spp-
Pathogenic:
-Neisseria meningitidis: oxidase +, beta-lactamase +, catalase +, Gram-negative cocci in pairs (diplococci) resembling kidney beans or coffee beans paired and facing each other, intracellular, replicate inside neutrophils, avoid phagocytosis, avoid complement, evade the immune system, pili, twitching motility, express porins, ferments glucose/dextrose and maltose, possesses a polysaccharide capsule, sequesters iron, requires CO2 for growth, associated with meningitis and Waterhouse-Friderichsen syndrome; WBC count in bacterial meningitis is increased, glucose is decreased, and protein is increased
-N. gonorrhoeae: all of the above and ferments only glucose/dextrose, no capsule, resistant to colistin, associated with gonorrhea; coagglutination is assocciated with this organism due to proteinemia; beta-lactamase production renders this organism resistant to penicillin; diagnosis in females is best made from a cervical culture
Non-Pathogenic/Rarely Pathogenic/Commensal:
-N. cinerea-oxidase +, catalase +, GNDC, similar to N. gonorrhoeae and M. catarrhalis biochemically, associated with similar infections, more pigmented and golden-brown on agar than N. gonorrhoeae, ferments glucose/dextrose, susceptible to colistin, can grow on sheep blood/chocolate agar/mueller hinton agar; has also rarely caused meningitis
-N. sicca-rarely causes sepsis; glucose +, maltose +, sucrose +
-N. lactamica-beta-lactamase +, oxidase +, catalase +, ferments lactose, colonization with this organism may increase susceptibility for colonization with N. meningitidis, predisposing an individual to a higher risk of infection and meningitis
-N. dentrificans-found in the mouths of mammals, such as cats and dogs; rarely associated with dog/cat bite infections
-N. elongata-rod-shaped, oxidase +, catalase -
-N. flavescens-golden yellow colonies; rarely associated with rheumatic heart disease and infection of heart valves
-N. subflava-rarely associated with development of meningitis following neurological surgery
-N. weaveri-nonmotile, GNR associated with dog bite infections
-N. mucosa-sucrose-fermenter and associated with endocarditis
-N. pharyngis-nonpathogen; normal flora
-N. polysaccharea-nonpathogen; normal flora
-N. macaecae-nonpathogen; normal flora
-N. bacilliformis-rarely associated with endocarditis in immunocompromised patients, Gram-negative rod, nonmotile, aerobic, can grow on blood agar/chocolate agar/martin lewis agar, nonfermenter, oxidase +, not all strains are catalase +
Neisseria spp all have the enzyme to oxidize tetramethyl-phenylenediamine. Special media called Thayer-Martin, Modified Thayer Martin, or Martin-Lewis chocolate-based agars are media selective for isolation of Neisseria from cervical exudates. Proper media for culture of a urethral discharge from a man includes chocolate agar and modified Thayer-Martin agars, and the specimen is obtained via a special urethral swab. The primary isolation of Neisseria spp, especially gonorrhoeae, requires CO2 and will die after 30 minutes if not in a system containing CO2 in 35-37 degrees Celcius. Beta-lactamase-producers are penicillin-resistant.
Media Used to Recover Neisseria spp:
CHOCOLATE AGAR: This agar is a non-selective, enriched medium containing horse's blood. Red blood cells within this medium have been slowly lysed by slowing heating to 80 degrees Celsius. It is used to grow fastidious bacteria requiring special nutrients for growth. Since blood cells are lysed, this medium contains both X (Hemin) and V (NAD) factors. (Use this as a primary isolation plate for both male and female exudates)
THAYER-MARTIN AGAR: This agar is a mueller-hinton based agar with 5% chocolate sheep's blood and the antibiotics vancomycin, colistin, nystatin and trimethoprim (Use this to plate an endocervical exudate from a female)
MODIFIED THAYER-MARTIN AGAR: Chocolate agar base containing vancomycin, colistin, nystatin, and trimethoprim (Use this to plate urethral discharge from a man)
MARTIN-LEWIS AGAR: This is a chocolate blood-based agar containing the antibiotic anisomycin
JEMBEC SYSTEM: Sodium bicarbonate and sodium citrate, CO2 tablets to create a CO2 atmosphere, and resealable bag or film
Haemophilus spp:
Haemophilus spp:
Haeomphilus spp are Gram-negative coccobacilli and small rods that are facultative anaerobes and require 5-10% CO2 incubation at 35-37 degrees Celsius. Most will not grow on sheep blood agar, but will grow well on chocolate agar because they are fastidious (they require special nutrients in order to survive/thrive/grow). They are oxidase +, catalase +, beta-lactamase +, and produce a flowery or musty/mousy odor on CHOC agar. This species grows well when performing the satellite test or Staph-Streak test. All but H. aphrophilus require hemin (X factor) and nicotine adenine dinucleotide (NAD or V factor) for in vitro growth. Most are normal inhabitants of the upper respiratory tract of humans. This bacterium family is very susceptible to drying and to temperature extremes, so specimens have to be plated immediately, particularly if they have not been placed in a suitable transport medium. This species will grow in THIO broth or in BHI broth and in blood culture bottles, however, the solutions may only produce weak turbidity. Rabbit or horse blood agars are used to detect hemolysis by hemolysin-producing strains that will not grow on sheep's blood. For example, the H QUAD plate blood segment consists of rabbit blood to detect hemolysis. Organisms typically grow within 24 hours but should be held 72 hours before being discarded as negative.
-H. influenzae-requires factors X and V and is nonhemolytic; causative agent of sepsis and bacterial meningitis in children and a wide range of other infections, particularly the encapsulated strains, which can be life-threatening and may include meningitis, epiglottitis, cellulitis, bacteremia, septic arthritis, pneumonia; Nonencapsulated strains may cause otitis media, sinusitis, conjunctivitis, exacerbations of chronic bronchitis, pneumonia, bacteremia in patients with underlying medical conditions but are typically normal flora of the human upper respiratory tract; Spread is person-to-person by inhalation of contaminated respiratory droplets or by a person's own endogenous strains; Colonies are small, smooth, and translucent at 24 hours (nonencapsulated) or larger, more mucoid colonies with a mousy odor at 24 hours (encapsulated) and are nonhemolytic on rabbit blood agar
*Encapsulated Strains: Type b is most common; Capsule is antiphagocytic and highly associated with virulence; Cell envelope factors facilitate attachment to host cells; Capsule is a sugar-alcohol phosphate; There are six groups: a, b, c, d, e, f but type b is the most commonly encountered serious clinical infection in humans; The Hib vaccination has reduced cases and infections with this type and provides immunity
*Nonencapsulated Strains: Pili and other cell surface factors are associated with attachment to host cells; Normal inhabitants of the respiratory tract
*To increase sensitivity of the direct Gram stain performed on CSF, specimens should be centrifuged at 2,000 rpm for 10 minutes and the smear should be prepared from the resulting pellet to concentrate the specimen and to increase the sensitivity of the direct microscopic exam by 5-10x or 100x (cytocentrifuge). This bacterium tends to stain a very pale pink and may be difficult to differentiate from the pink background of proteinaceous material.
*Treat with ceftriaxone or cefotaxime for life-threatening infections
*Treat with cephalosporins (several) for localized infections, beta-lactam inhibitor combinations, macrolides, trimethoprim/sulfa, or quinolones
-H. parainfluenzae-requires factor V and is nonhemolytic; These Gram-stain as small pleomorphic rods or long filamentous forms; Colonies are medium-to-large, smooth, translucent, and nonhemolytic on rabbit blood agar; Ferments mannitol
-H. haemolyticus-requires factors X and V and is hemolytic; nonpathogenic; Colonies resemble those of H. influenzae, except that they are beta-hemolytic on rabbit blood agar
-H. parahaemolyticus-requires factor V and is hemolytic; These Gram-stain as short to medium bacilli; Colonies resemble H. parainfluenzae but are beta-hemolytic on rabbit blood agar
-H. aphrophilus and H. paraphprophilus-causative agent of infective endocarditis (part of the HACEK bacteria); These Gram-stain as very short bacilli or occasional filamentous forms; These do not require X or V factors for growth and will grow on sheep's blood agar; Catalase - and ferments sucrose or lactose; Colonies are round, convex, and have an opaque zone near the middle; H. paraphrophilus ferments lactose and mannitol
-H. aegyptius-associated with purulent conjunctivitis (pink eye), rarely causes sepsis, biochemically similar to H. influenzae; requires factors X and V and is nonhemolytic; These Gram-stain as long, slender, Gram-negative bacilli; Colonies are tiny at 48 hours
-H. ducreyi-requires factor X and is nonhemolytic; causative agent of chancroid with regional lymphadenitis (STD); Gram stain as slender or coccobacillary rods appearing as "schools of fish"; May require 7 days to grow at 33-35 degrees Celsius in high humidity (place a sterile gauze pad moistened with sterile water inside the CO2 pouch); Colonies are flat, small, smooth, translucent-to-opaque at 48-72 hours, and can be pushed across the agar intact; Treat with erythromycin, ceftriaxone or ciprofloxacin
-H. felis-found in the mouths of cats and rarely associated with cat bite infections
-H. pittmaniae-may be associated with respiratory infections in patients with underlying lung diseases or conditions
-H. segnis-Gram-stain as pleomorphic rods; Colonies are convex, gray-white, smooth, or granular at 48 hours
Haeomphilus spp are Gram-negative coccobacilli and small rods that are facultative anaerobes and require 5-10% CO2 incubation at 35-37 degrees Celsius. Most will not grow on sheep blood agar, but will grow well on chocolate agar because they are fastidious (they require special nutrients in order to survive/thrive/grow). They are oxidase +, catalase +, beta-lactamase +, and produce a flowery or musty/mousy odor on CHOC agar. This species grows well when performing the satellite test or Staph-Streak test. All but H. aphrophilus require hemin (X factor) and nicotine adenine dinucleotide (NAD or V factor) for in vitro growth. Most are normal inhabitants of the upper respiratory tract of humans. This bacterium family is very susceptible to drying and to temperature extremes, so specimens have to be plated immediately, particularly if they have not been placed in a suitable transport medium. This species will grow in THIO broth or in BHI broth and in blood culture bottles, however, the solutions may only produce weak turbidity. Rabbit or horse blood agars are used to detect hemolysis by hemolysin-producing strains that will not grow on sheep's blood. For example, the H QUAD plate blood segment consists of rabbit blood to detect hemolysis. Organisms typically grow within 24 hours but should be held 72 hours before being discarded as negative.
-H. influenzae-requires factors X and V and is nonhemolytic; causative agent of sepsis and bacterial meningitis in children and a wide range of other infections, particularly the encapsulated strains, which can be life-threatening and may include meningitis, epiglottitis, cellulitis, bacteremia, septic arthritis, pneumonia; Nonencapsulated strains may cause otitis media, sinusitis, conjunctivitis, exacerbations of chronic bronchitis, pneumonia, bacteremia in patients with underlying medical conditions but are typically normal flora of the human upper respiratory tract; Spread is person-to-person by inhalation of contaminated respiratory droplets or by a person's own endogenous strains; Colonies are small, smooth, and translucent at 24 hours (nonencapsulated) or larger, more mucoid colonies with a mousy odor at 24 hours (encapsulated) and are nonhemolytic on rabbit blood agar
*Encapsulated Strains: Type b is most common; Capsule is antiphagocytic and highly associated with virulence; Cell envelope factors facilitate attachment to host cells; Capsule is a sugar-alcohol phosphate; There are six groups: a, b, c, d, e, f but type b is the most commonly encountered serious clinical infection in humans; The Hib vaccination has reduced cases and infections with this type and provides immunity
*Nonencapsulated Strains: Pili and other cell surface factors are associated with attachment to host cells; Normal inhabitants of the respiratory tract
*To increase sensitivity of the direct Gram stain performed on CSF, specimens should be centrifuged at 2,000 rpm for 10 minutes and the smear should be prepared from the resulting pellet to concentrate the specimen and to increase the sensitivity of the direct microscopic exam by 5-10x or 100x (cytocentrifuge). This bacterium tends to stain a very pale pink and may be difficult to differentiate from the pink background of proteinaceous material.
*Treat with ceftriaxone or cefotaxime for life-threatening infections
*Treat with cephalosporins (several) for localized infections, beta-lactam inhibitor combinations, macrolides, trimethoprim/sulfa, or quinolones
-H. parainfluenzae-requires factor V and is nonhemolytic; These Gram-stain as small pleomorphic rods or long filamentous forms; Colonies are medium-to-large, smooth, translucent, and nonhemolytic on rabbit blood agar; Ferments mannitol
-H. haemolyticus-requires factors X and V and is hemolytic; nonpathogenic; Colonies resemble those of H. influenzae, except that they are beta-hemolytic on rabbit blood agar
-H. parahaemolyticus-requires factor V and is hemolytic; These Gram-stain as short to medium bacilli; Colonies resemble H. parainfluenzae but are beta-hemolytic on rabbit blood agar
-H. aphrophilus and H. paraphprophilus-causative agent of infective endocarditis (part of the HACEK bacteria); These Gram-stain as very short bacilli or occasional filamentous forms; These do not require X or V factors for growth and will grow on sheep's blood agar; Catalase - and ferments sucrose or lactose; Colonies are round, convex, and have an opaque zone near the middle; H. paraphrophilus ferments lactose and mannitol
-H. aegyptius-associated with purulent conjunctivitis (pink eye), rarely causes sepsis, biochemically similar to H. influenzae; requires factors X and V and is nonhemolytic; These Gram-stain as long, slender, Gram-negative bacilli; Colonies are tiny at 48 hours
-H. ducreyi-requires factor X and is nonhemolytic; causative agent of chancroid with regional lymphadenitis (STD); Gram stain as slender or coccobacillary rods appearing as "schools of fish"; May require 7 days to grow at 33-35 degrees Celsius in high humidity (place a sterile gauze pad moistened with sterile water inside the CO2 pouch); Colonies are flat, small, smooth, translucent-to-opaque at 48-72 hours, and can be pushed across the agar intact; Treat with erythromycin, ceftriaxone or ciprofloxacin
-H. felis-found in the mouths of cats and rarely associated with cat bite infections
-H. pittmaniae-may be associated with respiratory infections in patients with underlying lung diseases or conditions
-H. segnis-Gram-stain as pleomorphic rods; Colonies are convex, gray-white, smooth, or granular at 48 hours
There is a special case in which H. influenzae will grow on 5% sheep blood agar, and that is during the satellite test. Tiny colonies of H. influenzae are observed growing very close to the colonies of S. aureus that have lysed the red blood cells in the agar, releasing V factor. This phenomenon is known as satelliting.
Moraxella spp:
Moraxella spp:
-M. catarrhalis-commensal, but can cause pneumonia, tracheobronchitis, ear infection, acute and chronic sinusitis, and, rarely, meningitis
-M. lacunata-associated with blepharoconjunctivitis
-M. atlantae-rarely causes bacteremia/sepsis in immunocompromised patients (cancer), and is an oxidase -, catalase +, GNR
-M. canis-rarely associated with dog bite wounds and ulcerated lymph nodes
-M. lincolnii-nonpathogenic; found in respiratory tract
-M. nonliqufaciens-nonpathogenic; found in respiratory tract
-M. saccharolytica-associated with rare cases of meningitis in children
-M. osloensis-similar to N. gonorrhoeae in Gram-stained smears of uterine cervix exudates
Moraxella spp are associated with nosocomial transmission and all species are carbohydrates negative. Gram-stains performed on sputum often reveal large, Gram-negative diplococci within polymorphonuclear neutrophils.
Moraxella catarrhalis-
-M. catarrhalis-commensal, but can cause pneumonia, tracheobronchitis, ear infection, acute and chronic sinusitis, and, rarely, meningitis
-M. lacunata-associated with blepharoconjunctivitis
-M. atlantae-rarely causes bacteremia/sepsis in immunocompromised patients (cancer), and is an oxidase -, catalase +, GNR
-M. canis-rarely associated with dog bite wounds and ulcerated lymph nodes
-M. lincolnii-nonpathogenic; found in respiratory tract
-M. nonliqufaciens-nonpathogenic; found in respiratory tract
-M. saccharolytica-associated with rare cases of meningitis in children
-M. osloensis-similar to N. gonorrhoeae in Gram-stained smears of uterine cervix exudates
Moraxella spp are associated with nosocomial transmission and all species are carbohydrates negative. Gram-stains performed on sputum often reveal large, Gram-negative diplococci within polymorphonuclear neutrophils.
Moraxella catarrhalis-
- Large kidney-shaped diplococci (sometimes mixed with some coccobacilli)
- Nonmotile
- Catalase +
- Oxidase +
- Aerobic
- MCAT disk + (blue within 2 minutes) (butyrate esterase test)
- "Hockey puck" colonies that can be moved across the surface of the agar
- Putrid/strong odor
- Gray-white or tan colonies, circular, glistening, nonhemolytic, waxy, fragile/friable, easily break
- Fastidious
- Grows well on chocolate agar and on blood agar, but does not grow on MacConkey agar
- Do not ferment any of the sugars in the CTA sugars test (lactose, glucose/dextrose, maltose, sucrose)
- DNAase +
- Nitrate reduction +
- Beta-lactamase +
- Part of the normal oropharyngeal flora, however, can cause infections if conditions are right (opportunistic pathogen)
- Infections it can cause: respiratory tract, ear, eye, acute and chronic sinusitis, maxillary sinusitis, bronchitis, laryngitis, bronchopneumonia, bacterial pneumonia, bacteremia, meningitis, sepsis, urethritis, septic arthritis, endocarditis (rare)