CLBACT - Anaerobes

Obligate aerobe

15%–21% O₂.

Microaerophile

5% O₂.

Facultative anaerobe

Multiplies well with or without O₂.

Aerotolerant anaerobe

Reduced concentrations of O₂.

Obligate anaerobe

Strict anaerobic environment (0% O₂).

Capnophile

5%–10% CO₂.

Superoxide anion

O₂ + e⁻ → O₂⁻ — toxic product of normal oxidation-reduction metabolism.

Hydrogen peroxide

O₂ + e⁻ + 2H⁺ → H₂O₂ — toxic product of normal oxidation-reduction metabolism.

Superoxide dismutase

4O₂⁻ + 4H⁺ → 2H₂O₂ + O₂. Present in aerobes and facultative anaerobes.

Catalase

2H₂O₂ → 2H₂O + O₂. Present in aerobes and facultative anaerobes.

Aerotolerant anaerobes

The protective enzymes are present in low concentrations; oxygen leads to cessation of growth.

Obligate anaerobes

Lack superoxide dismutase and/or catalase; oxygen leads to cell death. Require an environment with low redox potential.

Anaerobes as normal microbiota

Major portion of the normal microbiota of the skin, mucous membranes, GI and GU tract.

Endogenous infection

Spread from the organism's normal location in the body.

Exogenous infection

Trauma, surgical procedures, or consumption of contaminated food or water.

Common anaerobic infections

Oral infections, wound infections, abscesses, myonecrosis, gas gangrene, enteric diseases, joint infections, bloodstream infections.

Clinical significance

Consider the quality of the specimen and the presence and quantity of other organisms.

Gram-positive non-spore-forming anaerobic bacilli

Actinomyces, Mobiluncus, Lactobacillus, Propionibacterium; also Actinobaculum, Bifidobacterium, Eubacterium, Eggerthella, Collinsella, Paraeggerthella, Atopobium.

Gram-negative anaerobic bacilli

Bacteroides fragilis group, Fusobacterium, Porphyromonas, Prevotella.

Gram-positive anaerobic cocci

Peptococcus, Peptostreptococcus, Finegoldia, Staphylococcus saccharolyticus.

Gram-negative anaerobic cocci

Veillonella, Megasphaera, Anaeroglobus, Negativicoccus, Acidaminococcus.

Clostridium

• Obligate anaerobes, catalase-negative, gram-positive, spore-forming bacilli.
• MC exogenous anaerobic infections or intoxications.
• Toxins acquired via ingestion or open wounds contaminated with soil.
• Most species anaerobic; a few aerotolerant.

Clostridium virulence contributors

Collagenase, hyaluronidase, lecithinase, phospholipase.

Clostridium

Motile, swollen sporangia, non-encapsulated, single hemolytic reaction — EXCEPT C. perfringens. Carbohydrate fermenters — EXCEPT C. tetani.

Clostridium perfringens

• Gas gangrene bacillus.
• Reservoir: soil, sewage, dust, animal and human feces.
• Five types (A to E) based on quantities and types of exotoxins.
• MOT: puncture wound or ingestion.
• Normal flora of the GI tract and female genital tract.

C. perfringens diseases

Gas gangrene (myonecrosis), post-abortion sepsis, abdominal infections, enterocolitis.

α-Toxin

Phospholipase C, a lecithinase — necrotizing, hemolytic exotoxin of C. perfringens (gas gangrene).

Gas gangrene

• Myonecrosis by C. perfringens; wound contamination through trauma or surgery.
• Bullae, serous discharge, discoloration, tissue necrosis.
• CO₂ and H₂ accumulate in tissue — clinically detectable gas.

C. perfringens type A food poisoning

Mild; incubation 8–24 hours; enterotoxin ingestion in contaminated food; diarrhea and abdominal pain, self-limited. (The lecture lists β-toxin; type A food poisoning is caused by enterotoxin.)

Enteritis necroticans

• C. perfringens type C; clostridial necrotizing enteritis.
• Virulence factor: β-toxin.
• Incubation 5–6 hours.
• Bloody diarrhea, abdominal pain, bowel perforation.
• Fatality rate 15% to 25%.

C. perfringens bacteremia

Rapid hemolytic anemia may occur during severe sepsis due to hemolysis caused by the toxin.

C. perfringens microscopy

"Boxcar-shaped" bacilli; subterminal spores but difficult to induce.

C. perfringens culture

Dome-shaped (raised) colonies; double zone of β-hemolysis on SBA.

C. perfringens rapid tests

Lecithinase (+); motility (−). Ferments glucose, lactose, maltose, fructose.

Stormy fermentation

C. perfringens in litmus milk (+).

Direct Nagler test

• EYA or McClung agar with C. perfringens type A antitoxin.
• Lecithinase produces an opaque zone.
• (+) Inhibition of the lecithinase reaction by the antitoxin.

Reverse CAMP test (C. perfringens)

(+) Enhanced, "arrowhead-shaped" zone of hemolysis — α-toxin interacts with CAMP factor, producing synergistic hemolysis.

C. perfringens treatment

Clostridial myonecrosis: surgical debridement, radical amputation.

C. perfringens enteritis prevention

Adequate cooling and cold storage of cooked products.

Clostridium botulinum

• Reservoir: soil, marine and freshwater sediments; intestinal tract of fish, animals, birds, insects.
• Spores withstand 100 °C for several hours; heat resistance diminished by acid pH or high salt.
• Potential agent of bioterrorism (CDC).

Botulinum neurotoxin (BoNT)

• Binds the synapse of nerve fibers → acute flaccid paralysis and death.
• Seven antigenic varieties (A to G); types A, B, E and F cause human disease.
• Medical application: strabismus; beauty enhancer.

Forms of botulism

Foodborne; intestinal colonization by BoNT-producing clostridia; wound; iatrogenic; inhalational.

Botulism

• Incubation 12–36 hours (as early as 2 hours to 8 days).
• Cranial nerve palsy (double or blurred vision, impaired speech, difficulty swallowing).
• Bilateral descending muscle weakness.
• Sequelae: paralysis and respiratory failure leading to death.

Botulism key symptoms

Dry mouth → slurred speech → blurred vision → descending flaccid paralysis → respiratory failure → death.

Foodborne botulism

Ingestion of food with pre-formed toxin (vegetables, condiments, fish and meat products); absorbed through the small intestine into the circulation. MCC: toxin A.

Infant botulism

• Floppy baby syndrome (hypotonia).
• Ingestion of spores, germination, toxin production in vivo.
• Honey is a common vehicle.
• Lecture incubation: 18–36 hours (infant botulism incubates for days to weeks).

Wound botulism

Spores introduced into wounds; BoNT absorbed into the bloodstream. Incubation 4 to 14 days. Increase with injection drug use (black tar heroin).

C. botulinum microscopy

Oval/subterminal spores; "tennis rackets".

C. botulinum rapid tests

Lecithinase (−); lipase (+); motility (+). Ferments glucose.

Mouse bioassay

• Mouse neutralization test — recommended method for identification of BoNT; definitive ID of C. botulinum.
• Detects neurotoxin in serum or feces.
• Two samples injected into two mice: (−) mouse given the inactivated sample; (+) death.

C. botulinum treatment

Surgical debridement; botulinum antitoxin (BAT), serum-based — HBAT and BIG-IV; supportive care (mechanical ventilators).

C. botulinum prevention

Heat sterilization; acid preservation of food <pH 4.6, thorough cooking, refrigerated storage; no honey for infants; discard swollen cans; biosafety level 3 practices.

Clostridium tetani

Soil and environmental inhabitant; endospores in soil, dust and feces of farm animals. Spores enter the skin through puncture wounds and germinate. Causes tetanus.

Tetanospasmin (TeNT)

• Selectively cleaves synaptobrevin.
• Prevents release of inhibitory neurotransmitters (GABA, glycine). (The lecture prints "inhibitory and excitatory"; blocking acetylcholine is botulinum toxin.)
• Tension or cramping and twisting of skeletal muscles.

Tetanus incubation

7 days (range 3 to 21 days); length of incubation = distance from injury to CNS.

Tetanus symptoms

• Muscular rigidity (jaws, neck, lumbar region); difficulty swallowing; rigidity of abdomen, chest, back, limbs.
• Spastic paralysis.
• Lockjaw (trismus); distorted grin (risus sardonicus).
• Opisthotonus.

Trismus

Lockjaw — tetanus.

Risus sardonicus

Distorted grin — tetanus.

Opisthotonus

Spasm of muscles causing backward arching of head, neck and spine — tetanus.

Tetanus neonatorum

Use of contaminated instruments on newborns.

C. tetani microscopy

Round/terminal spores; "drumsticks".

C. tetani rapid tests

Lecithinase (−); lipase (−); motility (+) swarming. Ferments glucose.

Tetanus treatment and prevention

Antitoxin and vaccine. DTaP: diphtheria, tetanus, acellular pertussis trivalent vaccine. Adults: boosters every 10 years.

Clostridioides difficile

• Part of the GI biota in 5% of individuals.
• MCC of antibiotic-associated diarrhea.
• Most common HAI in North America — acquired in hospitals by patients receiving antibiotics; an "infection control dilemma".
• Increase in community-acquired cases.

C. difficile toxins

Enterotoxin (TcdA) and/or cytotoxin (TcdB) — work in concert to enter and damage the cytoskeleton of large-intestine cells, leading to apoptosis.

Pseudomembranous colitis

AKA C. difficile-associated disease (CDAD); bloody diarrhea with necrosis of colonic mucosa.

C. difficile microscopy

Subterminal oval spores.

C. difficile rapid tests

Lecithinase (−); lipase (−); motility (+); fluorescence (+). Ferments glucose and fructose.

CCFA

• Cycloserine-cefoxitin fructose agar — selective for C. difficile.
• Yellow colonies with a "ground-glass" appearance.
• Cycloserine and cefoxitin inhibit gram-negative coliforms.
• pH indicator: neutral red.

C. difficile on BAP

Barnyard or horse stable odor; fluoresces chartreuse (brilliant yellow-green).

C. difficile EIA

Enzyme immunoassay kits detecting toxin A, toxin B and glutamate dehydrogenase (GDH) in feces.

Cell culture cytotoxicity test

Gold standard for detection of C. difficile toxin type B; tissue culture; two to three days to a positive result.

Fecal microbiota transplant

C. difficile treatment — donor feces from a family member or fecal transplant bank, instilled through colonoscopy or endoscopy; cure rate 90%.

Actinomyces

• Endogenous microbiota of the oral cavity and female genital tract.
• Normal flora of animal and human mucous membranes.
• Etiologic agent of "lumpy jaw", a cervicofacial disease.

Actinomycosis

• MCC: Actinomyces israelii.
• Chronic, granulomatous infectious disease.
• Sinus tracts and fistulae that erupt to the surface and drain pus with sulfur granules.
• Sites: maxillary region (sinuses in neck and thorax); female genital tract (long-standing IUD).

Actinomyces microscopy

Filamentous gram-positive bacilli. (The lecture adds "some species are partially acid-fast such as Nocardia"; Actinomyces is not acid-fast.)

Sulfur granules

Dense clumps of bacteria in exudate — Actinomyces spp., Propionibacterium spp., Eubacterium nodatum.

Actinomyces colonies

Young: "spider-like" or "wooly" thin radiating filaments. Old: "molar tooth". May be red, pink, tan, yellow, white or grayish.

Lactobacillus

• Largest portion of the vaginal biota.
• Aerotolerant anaerobes.
• Maintains an acidic vaginal environment, protecting from urogenital infections.

Döderlein's bacillus

Lactobacillus acidophilus — most of the lactobacilli of the healthy vagina.

Lactobacillus infections

Endocarditis — most common; high mortality 23% to 27%. Also septicemia, intraabdominal abscesses, meningitis, oral infections, conjunctivitis.

Lactobacillus identification

Gram-positive pleomorphic rods; catalase (−); motility (−). SBA: small α-hemolytic colonies.

Tomato juice agar

Differential medium for Lactobacillus; pH 3 to 4.

Lactobacillus AST pattern

Resistant: vancomycin and cephalosporins.

Mobiluncus

• Associated with bacterial vaginosis — a clinical variant of the vaginosis associated with G. vaginalis.
• Curved, gram-variable bacilli.
• Catalase (−); motility (+); indole (−).

Gardnerella vaginalis

Gram-variable bacilli; bacterial vaginosis (BV).

Clue cells

Epithelial cells coated with bacteria, as seen in Pap smears — G. vaginalis.

Bacterial vaginosis

• Mobiluncus, Bacteroides, Prevotella, anaerobic GPC, G. vaginalis.
• Shift in the ecology of the vaginal microbiota.
• Gray-white, homogenous, malodorous vaginal discharge.
• Increased risk of HIV infection.

Cutibacterium acnes

• Former name: Propionibacterium acnes.
• Indigenous skin microbiota; rarely pathogenic.
• MC anaerobic contaminant of blood culture.
• Adhesion factors; forms biofilms.

C. acnes infections

Acne vulgaris, neurosurgical shunt infections, prosthetic joint infections, endophthalmitis after cataract surgery.

C. acnes identification

Pleomorphic rods with a diphtheroid appearance; catalase (+); indole (+).

Bacteroides fragilis group

• Most commonly isolated anaerobes from blood cultures.
• Major normal flora of the colon.
• Virulence factor: polysaccharide capsule.
• Enters sterile sites after surgery, trauma or disease.
• Significant cause of intra-abdominal abscess.

B. fragilis microscopy

Gram-negative bacilli with rounded ends; pleomorphic.

B. fragilis rapid tests

Catalase (+); motility (−); bile disk (+) 20% bile growth; lipase (−); lecithinase (−); gelatinase (−).

B. fragilis culture

BAP: non-hemolytic. BBE agar: gray colonies that blacken the agar.

B. fragilis AST

Susceptible: rifampicin. Resistant: penicillin, kanamycin, vancomycin, colistin.

Prevotella melaninogenica

• Pigmented saccharolytic gram-negative bacilli.
• Normal flora of oropharynx, nose, GI and urogenital tracts.
• Young colonies tan; brick-red fluorescence under UV light.

Porphyromonas

• Asaccharolytic and pigmented.
• Normal flora of oropharynx, nose, GI and urogenital tracts.
• Brick-red fluorescence under UV light.

Fusobacterium

Normal flora of the upper respiratory and GI tracts. F. nucleatum: pulmonary infections. F. necrophorum: lung and liver abscess.

Lemierre disease

• F. necrophorum; leukotoxin and plasminogen-binding toxin.
• Infectious thrombophlebitis in the internal jugular vein, commonly from head and neck infections.
• Complications: bacteremia and septic emboli.