MMSC438 EXAM 3 ANAEROBIC BACTERIOLOGY

Obligate anaerobe

Organisms killed by oxygen due to lacking enzymes for toxic oxygen products.

Aerotolerant anaerobe

Survives short oxygen exposure, but requires an anaerobic environment to perform metabolism.

Facultative anaerobe

Preferentially uses oxygen when available, but can grow without it at a slower rate.

Clostridium

obligate anaerobic spore-forming GPR

Bacteroides

obligate anaerobic GNR

Porphyromonas

Clinically significant anaerobic GNR

Prevotella

anaerobic GNR involved in oral and pelvic infections.

Propionibacterium (Cutibacterium acnes)

Clinically significant anaerobic Gram-positive rod associated with skin conditions like acne.

Anaerobic cocci (Peptostreptococcus)

group of anaerobic GPC

Fusobacterium

Clinically significant anaerobic Gram-negative rod with tapered ends.

Campylobacter ureolyticus group

Gram-negative microaerophilic or anaerobic organism group.

Bilophila wadsworthia

Clinically significant anaerobic GNR associated with gangrenous appendicitis.

Sutterella wadsworthensis

anaerobic microaerophilic GNR found in the human GI tract.

Exogenous anaerobic infection (definition)

Infection caused by anaerobic organisms that exist outside the body.

Exogenous anaerobic infection (route of infection)

Occurs via ingestion or trauma.

Exogenous anaerobic pathogen (example)

Clostridium (GPR, spore-forming).

botulism, tetanus, food poisoning

Endogenous anaerobic infection (definition)

Infection caused by organisms inside the body that become infectious when displaced.

Endogenous anaerobic pathogens (examples)

Bacteroides, Porphyromonas, Prevotella, Clostridium, Propionibacterium, and anaerobic cocci.

Key difference: Exogenous vs. Endogenous anaerobic infection

Exogenous originates outside the body; endogenous originates from displaced internal normal flora.

Clostridium (common infections)

Bloodstream infections and gas gangrene.

Bacteroides (common infections)

Bloodstream, abscesses, and female GU infections.

Prevotella (common infections)

Oral infections and female GU infections.

Porphyromonas (common infections)

Oral infections.

Fusobacterium (common infections)

Oral infections and bloodstream infections.

Cutibacterium acnes (common infections)

Bloodstream and prosthetic device infections.

Anaerobic culture: Swab specimens

Poor quality/unacceptable unless placed in proper transport media.

Collection: Large tissue

Sterile container.

Collection: Small tissue

Anaerobic tubes with agar.

Collection: Blood for anaerobic culture

Anaerobic bottle.

Preferred specimen type for anaerobic culture

Aspirates.

Anaerobic transport gas mixture

H₂ (5-10%), CO₂ (5-10%), and N₂ (80-90%).

Role of palladium pellets in anaerobic transport

Acts as a catalyst to remove residual oxygen.

Processing steps for anaerobic specimen

Direct Gram stain and Gram stain of ground up tissue.

Nutritional requirements for most anaerobic bacteria

Horse or sheep RBCs, Vitamin K, Hemin, and yeast extract.

Best media for recovery of anaerobic bacteria

CDC blood agar or Brucella blood agar.

PEA (Phenylethyl alcohol agar) for anaerobes

Selective primary plating media for gram-positive anaerobes.

KVLB (Kanamycin-Vancomycin Laked Blood agar)

Selective primary media for Bacteroides, Prevotella, and Porphyromonas.

KVLB agar (fluorescence test)

Prevotella and Porphyromonas emit brick-red fluorescence on KVLB plates.

BBE (Bacteroides Bile Esculin agar)

Selective for Bacteroides fragilis group; growth of black colonies indicates bile tolerance.

CCFA (Cycloserine-Cefoxitin Fructose Agar)

Specialized, non-routine medium used for anaerobic culture.

Clostridium species (Gram stain morphology)

Large gram-positive rods with a "boxcar" appearance.

Cutibacterium acnes (Gram stain and biochemical key features)

GPR; indole/catalase positive.

Fusobacterium nucleatum (Gram stain morphology)

Thin, pointed gram-negative rods.

Fusobacterium mortiferum (Gram stain morphology)

Thicker gram-negative rods.

Anaerobic jar/chamber gas mixture

5-10% H₂, 5-10% CO₂, and 80-90% N₂.

Palladium catalyst function in anaerobic incubation

Removes residual oxygen in anaerobic jars or chambers.

C. perfringens (colony presentation on anaerobic BAP)

Double zone hemolysis.

C. perfringens (reaction on EYA)

Lecithinase positive on egg-yolk agar (EYA).

Bacteroides fragilis group (colony presentation on BBE)

Growth of black colonies (bile esculin positive).

Prevotella, Porphyromonas, and Fusobacterium (KVLB fluorescence)

Brick-red fluorescence on KVLB.

Fusobacterium (fluorescence feature)

Chartreuse fluorescence.

Why full identification of anaerobic bacteria is not always cost-effective

Full ID requires expensive methods, many are reported presumptively, and full ID is often clinically unnecessary.

Aerotolerance test (procedure)

Inoculate CHOC in CO₂ and BAP anaerobically, then compare growth.

Aerotolerance test (interpretation)

Obligate anaerobes grow only on the anaerobic BAP, not CHOC in CO₂.

Disk and chemical tests for anaerobic GPC and GNR

Uses antimicrobial disks and spot chemical tests like indole for identification.

Egg-yolk agar (EYA) role in anaerobic ID

Detects lecithinase activity, producing an opaque zone around colonies.

Methods for definitive identification of anaerobes

Enzyme systems, 16S rRNA sequencing, and MALDI-TOF MS.

Definitive anaerobic ID methods (turnaround time)

Take longer than presumptive methods, but provide definitive identification.

Antimicrobial susceptibility testing guidelines for anaerobes

Governed by CLSI M11 guidelines.

Indications for anaerobic susceptibility testing

Serious infections including brain abscess, endocarditis, bacteremia, osteomyelitis, and prosthetic/joint infections.

Easiest susceptibility test method for anaerobes

Etest using Mueller-Hinton agar supplemented with 5% sheep RBCs.

Five approaches to treating anaerobic infections

Surgical therapy (debridement), hyperbaric oxygen, antimicrobial therapy, antitoxins, and fecal microbiota transplant (FMT).

Metronidazole (anaerobic coverage)

Excellent for Bacteroides fragilis with good tissue penetration, including the CNS.

Clindamycin (anaerobic coverage)

Active against many anaerobes with good tissue penetration (except CNS); resistance is increasing.

Penicillin + β-lactamase inhibitor (anaerobic coverage)

Covers β-lactamase producing anaerobes (e.g., amoxicillin/clavulanate, ampicillin/sulbactam, piperacillin/tazobactam).

Second-generation cephalosporins for anaerobes

Cefoxitin, cefotetan, cefmetazole; active against B. fragilis (resistance increasing) and used for surgical prophylaxis.

Carbapenems (anaerobic coverage)

Excellent broad anaerobic coverage; meropenem is slightly better for GNRs.

Quinolones for anaerobes (levofloxacin, moxifloxacin)

Good tissue penetration; reserved for β-lactam allergy; resistance is increasing.