anaerobes part i ut
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Anaerobes(Gram Neg)Anaerobes(Gram Neg)Victor S. Flauta, M.D.March 30, 2011
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Epidemiology
GNAR Mucosa of animals and humans Predominant species: oral cavity, GIT, vagina Infections acquired:
Endogenously (breached by trauma or disease)
Exception: clenched-fist wounds & bite wounds Iatrogenically (by surgery)
Aspiration pneumonia: mixed anaerobes
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Source: Uptodate
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GN Anaerobes
Organism ANA BAP BBE Comment
B. fragilis large Gray/black Regular GNR
F. nucleatum Bread crumb or opalescent
NG Indole +; pointed ends
B. wadsworthia Small, transluscent
Central black dot (H2S)
Cat +
Prev. intermedia Small Black on LKV Coccobacilli, IND +; lipase +; red fluorescence
Porphyromonas uniform NG Red fluorescence
Bacteroides ureolyticus
Flat transparent w/ pitting
NG Urea +, cat -
Veilonella Small, transparent
NG GNDC; red fluorescence
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Clues to probability of anaerobic
bacteria at infection site
Infection adjacent to surfaces that normally harbor anaerobes as normal flora
Abscess formation or tissue necrosis Putrid odor Gas formation Gram stain of exudate showing polymicrobial flora Organisms with morphologic features of anaerobes Classic features of histotoxic clostridial syndromes: tetanus,
botulism, C. perfringens food poisoning, gas gangrene, C. difficile-induced diarrhea or colitis, enteritis necroticans
Infections that, by prior experience, usually involve anaerobic bacteria
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Collection & Transport
Lower Resp Tract & endometrial samples are hard to obtain w/out contamination of resident flora
Swab: least desirable and should be discouraged small sample, prone to drying, intrinsically aerobic, can't
be quantitated
Transport immediately in proper container If transported in glass, it can stay at room temp
or refrigerated
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Isolation
Keep plates for 5 days to isolate Porphyromonas & Bilophila
Media: Brucella base is superior to CDC base & Schaedler base
for GNAR CDC is better for GPAC Fastidious anaerobe agar (Lab M): good for
Fusobacterium Ideal: 2 different basal media to maximize isolation
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Isolation
Media: LKV: for Bacteroides & Prevotella (rapid pigmentation) For Porphyromonas reduce LKV vancomycin
concentration (from 7.5 to 2 ug/ml) BBE: for Bacteroies, Bilophila, F. mortiferum/varium PEA: to prevent aerobic GNR & Clostridial swarming For Fusobacterium: neomycin-vancomycin agar metronidazole disk: R/O GPAR & facultative anaerobes
since they’ll show resistance
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Isolation
Must not expose to air during first 48 hrsTotal incubation period of at least 5 days
is recommended for primary plates.If shorter, may not detect Porphyromonas
& BilophilaBlood culture:
Controversial: only <5% of strict anaerobes Still used since some facultative anaerobes
grow faster in ANA blood culture media
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Identification
Vancomycin Resistant but Colistin & Kanamycin sensitive: Fusobacterium B. ureolyticus – NO3+ Bilophila – NO3+ Sutterella Leptotrichia (“brain surface” texture)
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Antibiotic B. fragilis B. frag. grp
Prevotella Pophyromonas F. nucleatum
F. mortiferum/ varium
Carbapenem S S S S S S
B lactamase inhibitor
S S S S S S
Penicillins R R R (50%) S S S
Cephalosporins(Cefoxitin;Ceftizoxime)
S (85%) S (70%)
S S S S
Chloramphenicol S S S S S S
Tetracycline(except tigecycline~ S to all)
R R R S S S
Lincosamides(Clindamycin)
S (80%) R (50%)
S S S S (80%)
Fluoroquinolones(Moxifloxacin)
S S (80%)
S S S S
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The susceptibility trends for the species of the Bacteroides fragilis group against various antibiotics from 1997 to 2004 were determined by using data for 5,225 isolates referred by 10 medical centers. The antibiotic test panel included ertapenem, imipenem, meropenem, ampicillin-sulbactam, piperacillin-tazobactam, cefoxitin, clindamycin, moxifloxacin, tigecycline, chloramphenicol, and metronidazole. From 1997 to 2004 there were decreases in the geometric mean (GM) MICs of imipenem, meropenem, piperacillin-tazobactam, and cefoxitin for many of the species within the group. B. distasonis showed the highest rates of resistance to most of the ß-lactams. B. fragilis, B. ovatus, and B. thetaiotaomicron showed significantly higher GM MICs and rates of resistance to clindamycin over time. The rate of resistance to moxifloxacin of B. vulgatus was very high (MIC range for the 8-year study period, 38% to 66%). B. fragilis, B. ovatus, and B. distasonis and other Bacteroides spp. exhibited significant increases in the rates of resistance to moxifloxacin over the 8 years. Resistance rates and GM MICs for tigecycline were low and stable during the 5-year period over which this agent was studied. All isolates were susceptible to chloramphenicol (MICs < 16 µg/ml). In 2002, one isolate resistant to metronidazole (MIC = 64 µg/ml) was noted. These data indicate changes in susceptibility over time; surprisingly, some antimicrobial agents are more active now than they were 5 years ago.
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New GNAR
Alistipes putredinis: appendicitis Alistipes finegoldii: appendicitis Bacteroides coprocola: human feces Bacteroides goldsteinii: infection (intestinal origin) Bacteroides nordii: infection (intestinal origin) Bacteroies salyersiae: infection (intestinal origin) Bacteroides plebeius: human feces Cetobacterium somerae: children’s feces Desulfovibrio piger: infection (intestinal origin) Dialister micraerophilus: human clinical samples Dialister propionicifaciens: human clinical smaples Faecalibacterium prausnitzii: feces Fusobacterium canifelinum: bite infections (dogs, cats) Fusobacterium equinum: oral cavity of horses
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New GNAR
Pophyromonas gulae: gingival sulcus of animals Porphyromonas somerae: human skin, soft tissue & bone
infections Porphyromonas uenonsis: non-oral human infections Prevotella baroniae: human oral cavity Prevotella marshii: human oral cavity Prevotella multiformis: human subgingival plaque Prevotella multisaccharivorax: human subgingival plaque Prevotella salivae: human oral cavity Prevotella shahii: human oral cavity Sneathia sanguinegens: human blood Tannerella forsythensis: human periodontal pockets
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B. fragilis
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B. fragilis
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Bacteroides fragilis
95% Confidence: Large colonies on ANA BAP Gray or black colonies on BBE GNR
Most common species in clinical specimens Nonmotile GNR with rounded ends Broth: pleomorphic with vacuoles CDC: nonhemolytic, gray with concentric whorls Significance of capsules are still unclear Key characteristics:
Growth enhanced by bile Resistant to KVC (kanamycin, vanc, colistin) & Penicillin Sensitive to Rifampin
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B. fragilis
Ability to tolerate oxygen : contain superoxide dismutase (protects against
the toxic effects of oxygen) the ability to survive exposure to oxygen
facilitates the survival and pathogenicity of the organism.
Bergan, T. Pathogenicity of anaerobic bacteria. Scand J Gastroenterol Suppl 1984; 91:1.
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B. fragilis
Normal constituent of colonic flora (abnormal in mouth, genital tract or URT)
Found as mixed infections in abscessProduce enterotoxin induces IL-8
inflammatory diarrheaAlso produces metalloproteases, LPS, capsular
polysaccharides periodontal disease and abscess formation
1st Rx: Metronidazole2nd Rx: Clindamycin
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B. thetaiotamicron
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B. thetaiotamicron
2nd most common species isolated in B. fragilis group
Infections: Peritonitis Intraabdominal abscess Hepatic abscess
Indole + (B. fragilis is indole neg)
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B. distasonis
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B. distasonis
B. distasonis showed the highest rates of resistance to most of the ß-lactams.
B. fragilis, B. ovatus, and B. distasonis and other Bacteroides spp. exhibited significant increases in the rates of resistance to moxifloxacin.
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B. vulgatus
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B. vulgatus
The rate of resistance to moxifloxacin of B. vulgatus was very high (MIC range for the 8-year study period, 38% to 66%).
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B. ureolyticus
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Bacteroides ureolyticus
Thin GNAR with rounded endsSmall colonies, flat transparent w/ pittingGreening of the agarMay resemble Bilophila phenotypically
(NO3+) but: Bilophila is strongly catalase + & resistant to bile Bacteroides ureolyticus is catalase neg & urea +
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B. ovatus
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Porphyromonas spp.
P. gingivalis Root canal infections, odontogenic sinusitis Produces phenylacetic acid Agglutinates sheep RBC Produces B-galactose-6-phosphate Produces N-acetyl-B-glucosaminidase
P. asaccharolytica Produces a-fucosidase Prevalent in urogenital or intestinal tract (important in
infections arising from these sources) P. endodontalis
Root canal infections, odontogenic sinusitis Not as either of the foregoing species
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Porphyromonas spp.
P. somerae: pleuropulmonary infections, skin & soft tissue infections, bacterial vaginosis
Pophyromonas gulae: gingival sulcus of animals
Porphyromonas somerae: human skin, soft tissue & bone infections
Porphyromonas uenonsis: non-oral human infections
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Bilophila wadsworthia
Reported by EJ Baron & colleagues in 1989 from appendicitis specimens and human feces
Present in small number in bowel flora3rd most common anaerobe recovered from
gangrenous or perforated appendixCommon constituent of the microbiota of intra-
abdominal infections Isolated from various clinical specimensEasily overlooked because of it’s fastidious
growth
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Bilophila spp.
GNR, non-spore-forming, nonmotile, pleomorphicGrows after 4 days on BBE as opaque black
coloniesGrows on Brucella agar in 4-7 days as translucent
gray coloniesCatalase +, asaccharolytic, urease +, H2S +, NO3
+, Negative: indole, esculin, oxidase, B lactamase
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Bilophila spp.
Major metabolic product: acetateLike B. fragilis, it grows in 20% bileUnlike B. fragilis, it does not ferment CHOUnlike Fusobacterium, it is strongly CAT +
& neg for butyric acidResistant to beta lactams
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Prevotella melaninogenica
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Prevotella melaninogenicaP. intermedia & others
Normal oral floraBrackish brown hematin pigmentAka P. melaninogenica group Infections:
Aspiration pneumonia Pulmonary abscess Pleural empyema Cerebral abscess
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Other Prevotellas
P. bivia, P. disiens, P. buccae, P. oralis, P. buccalisNormal flora of the urogenital tract & oropharynxAKA P. oralis group Infections:
Chronic otitis media & sinusitis Dental abscesses Ulcerating gingivostomatitis Female genital tract infections Cerebral abscesses
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Fusobacterium nucleatum
Slender, pointed ends
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F. nucleatum
Thin rod with tapering ends (“needle-shaped”) Capnocytophaga & Leptotrichia may also look like these
but both are indole neg.
Indole +Greening of agar when exposed to air (due to
production of H2O2)Has at least 3 different colony morphotypesPleuropulmonary infections
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F. necrophorum
Large, pleomorphic
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F. necrophorum
“Lipase positive fusobacterium”Bile sensitiveLong rod with round ends, pleomorphic
with bizarre formsIndole +Fluoresces chartreuseBeta hemolysis around gray-yellow
colonies
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F. necrophorum
Peritonsilar abscess: most common ANAPharyngotonsilitis in children or young
adults (as often as S. pyogenes) May be associated with infectious mono
Lemierre’s Disease Jugular vein septic thrombophlebitis Often complicated by sepsis & metastatic
abscess
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F. mortiferum
Bizarre, round bodies
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F. mortiferum
Indole negExtremely pleomorphicFilaments with swollen areas, round bodies &
irregular staining F. necrophorum may look similar but fewer round bodies
A bile resistant fusobacterium isolated from BBE is F. mortiferum or F. varium
ONPG + (F. varium is ONPG neg) Intraabdominal infections
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F. varium
Large, rounded ends
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F. varium
Intraabdominal infections
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Veilonella spp.
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Veilonella spp.
Small percentage isolated in human specimens
Rare infections: Meningitis, osteomyelitis, prosthetic joint
infections, pleuropulmonary infection, endocarditis, bacteremia
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Organism NO3 Catalase Glucose
Veilonella spp + V -
Acidaminococcusfermentans
- - -
Megasphaeraelsdenii
- - +
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