Clostridium perfringens The agent of - Gangrene - Food poisoning Bacteria features: GPB, Capsulate, non-motile. Spores: bulging Rapidly growing (doubling time ~ 8min) specially on soil.

Colonies: Colonies: On blood agar: β-haemolytic, flat, spreading, rough, translucent colonies with irregular margins.blood agarβ-haemolytic

Determinants of pathogenicity 12 exotoxins and an enterotoxin The major exotoxins: alpha toxin, beta toxin, epsilon-toxin and Theta-toxin. Types of the organism (A through E) can be differentiated according to the toxin repertoire. Alpha toxin (Lecithinase) is the primary toxin and common to all types of C. perfringens but mostly by type A.

Determinants of pathogenicity PhosphorylcholineDiglyceride Lecithinase (or phospholipase) degrades Lecithin in cell membrane into Phosphorylcholine and Diglyceride. Hyaluronidase produced by C. perfringens breaks down the intraceullar cytoskeleton. Collagenase and other proteinases break down muscles and liquiefy. Dnase Enterotoxin

Nagler reaction It is done in: A Nagler agar plate, containing 5-10% egg yolk. It is used to identify strains produceing α- toxin (phospholipase). C. perfringens phospholipase causes turbidity (characteristic precipitate) around the colonies on egg-yolk medium Inhibited by specific antiserum.

One-half of the plate is inoculated with antitoxin to act as a control in the identification.antitoxin

Heat resistancy Type A strains (causing gas gangrene): produce heat inactivated spores quickly. Food poisoning strains: produce heat resistant spores.

Transmission A saprophyte, occuring in soil, H 2 O, decomposing plant, human and animal feces. Vegetative cells are members of normal flora in colon and 5% of female genital systems (vagina).

Gangrene Gangrene is necrosis and putrefaction of tissues. Gas production forms bubbles of gas in muscle (crepitus) and smell in decomposing tissue. Gangrene is mostly a mix infection mainly by C. perfringens but also C. septicum, D. sordelli & other proteolitic clusteridums, cocci, gram negative bacilli…. may be found.

Clinical findings Glycogen metabolism: Myonecrosis Bad smelling exuda, inflamation, very progressive necrosis, fever, hemolysis, Crepitation, hemolytic anemia, severe toxemia, shock.

Crepitation

war and truma wounds. Gangrene is associated with war and truma wounds. Also after: Also after: Illegal abortion Gangrene Toxic shock syndrome) - Illegal abortion (causing Gangrene or Toxic shock syndrome) infant delivery days after infant delivery (Endometric infection).

Anaerobic cellulitis This is infection of the dermis and subcutaneous tissue.

Treatment - Antibiotic trapy: Penicillin - Surgery (Ampotasion): Wounds should be debrided -H2O2 -Hyperbaric oxygen therapy (HBOT) -Antitoxin (polyvalent against different sub-species.)

Hyperbaric oxygen therapy (HBOT) Hyperbaric oxygen therapy (HBOT) is a medical treatment in which the patient breathes 100% oxygen while inside a pressurized chamber.

Hyperbaric oxygen

Food poisoning

Soil and food. Survives cooking and grows to large numbers in reheated food, especially meat. Enterotoxin is produced when more than 10 8 vegetative cells is eaten. The cells convert into spores in GI. Enterotoxin is a part of spore coat. Clinical findings: Clinical findings: Incubation: 8-16 hours Symptoms: watery diarrhea, cramps, little vomiting. Resolves in one or two days.

Treatment and prevention of food poisoning Treatment: Symptomatic – No antimicrobial drugs Prevention Cooking well

Lab diagnosis Smear of necrotic tissue and exudate samples: large positive G+ rods. Cultured anaerobically I. Blood agar II.Thyoglicolate (identified with fermentation reactions) Food poisoning: isolation of similar strains in large numbers from food sources and faeces.

Clostridium difficile GPB Motile Long, irregularly (often "drumstick" or "spindle") shaped cells Spores: Terminal, bulging Ubiquitous in nature and especially prevalent in soil

Transmission Ubiquitous in nature and are especially prevalent in soil. Commensal bacteriumCommensal bacterium of the human intestine in 2-5% of the population. intestine

Disease Disease Clostridium difficile infection (CDI) Antibiotic-associated diarrhea (25% of antibiotic- related diarrhea) Antibiotic-associated pseudomembranous colitis Nosocomial infection

Determinants of pathogenicity Enterotoxin (toxin A) and Cytotoxin (toxin B) (heat and acid-labile proteins) EnterotoxinCytotoxin Both found in patients’ feces with pseudomembranous colitis. Attach to the intestinal epithelial cells at the microvilli level and are responsible for the diarrhea and inflammation. diarrheainflammation

Pathogenesis of C. difficile: Pseudomembranous colitis Pseudomembranous colitis A severe infection of the colon, often resulting from eradication of the normal gut flora by antibiotics.colongut flora antibiotics Antibiotic (Clindamycin and Ampicillin) supress drug-sensitive normal flora, allowing C. difficile.

Pseudomembranous colitis (PMC) Acute colitis characterized by the formation of an adherent inflammatory membrane overlying sites of mucosal injury. Exudates composed of fibrin, mucin, bacteria and polymorphonuclear leukocytes attach to the mucosal surface.

Clinical findings Diarrhea Pseudomembranes (yellow-white plaques) on the colonic mucosa. Visualised by colonoscopy.

Lab diagnosis Culture and isolation of C. difficile on Blood agar Identification of toxins by specific antibody using Enzyme immunoassays (EIAs) for the detection of toxins A and B in fecal material. Identification of exotoxins by cell cultures: Cytopatic effects (CPEs) PCR

Treatment Withdrew the antibiotics Clindamycin and Ampicillin. Oral metronidazole or vancomycin instead along with fluids.