clostridium

Positive

gram-_____ rods

Clostridium

• large (0.3-1.3×10um)

• Anaerobic

• Tends to decolorize

• Endospore-producing

C. spiroforme

All pathogenic species are straight rods except

C. perfringens

Most species ar motile by peritrichate flagella except

Clostridium

• oxidase-negative CN

• catalase-negative CN

• Fermentative

2-10% CO2

Prefers atmosphere containing

• amino acids

• Carbs

• Vitamins

• Blood/serum

Requires enriched media with

37C

Optimum growth of pathogenic clostridia is at

Endogenous

Many pathogenic clostridia are normal inhabitants of the intestinal tracts of animals and man

• Causes ______ infection

Exogenous

Other clostridia are commonly present in the soil

Causes ______ infections

• wound contamination

• Ingestion

• C. Tetanu

• C. botulinum

Neurotropic clostridia

• produces potent neurotoxins but are non-invasive

• Colonize the host to a very limited extent

• C. chauvoei

• C. septicum

• C. noyvi types A and B

• C. haemolyticum

• C. sordellii

• C. perfringens type A

Histotoxic clostridia

• prodces less potent toxins but invasive

• Includes gangrene-producing clostridia

• C. perfringens types A-E

• C. difficile

• C. colinum

• C. spiriforme

Enteropathogenic and enterotoxigenic clostridia

• enterotoxins are formed in the intestine then absorbed in the bloodstream then produces generalized toxemia

C. piliforme

Atypical clostridium

• causes Tyzzer’s disease in foals and laboratory animals

Tetanolysin

Clostridium tetani

• Which toxin causes hemolysin and enhanced tissue invasion?

Tetanospasmin

Clostridium tetani - which endotoxin is a:

• neurotoxin

• Plasmid-coded

• Responsible for signs of tetanus

Clostridium tetani

• endospores enter traumatic or surgical wounds via the neonatal umbilis or uterus following dystocia

Clostridium tetani

• Presence of facultative anaerobe and necrotic tissue

• creates anaerobic conditions

• Allows C.____ spores to germinate

Trismus

Tetanic Triad

• Lockjaw

Risus sardonicus

Tetanic Triad

• sardonic smile

Opisthotonos

Tetanic Triad

• backward arching of the trunk

4

Clostridium botulinum

• has ____ groups based on phenotype

Seven

Clostridium botulinum

• _____ types (A-G) based on antigenicity of toxin produced

C. botulinum type G

______ had been renamed C. argentinense

Eight

Clostridium botulinum

• produces ____ neurotoxins

Anaerobic situations

Clostridium botulinum

• germination of endospores, growth of vegetative cells and production of toxin occurs in _______

• foodborne

• Wound

• Infant

• Adult intestinal toxemia

• Iatrogenic

Five forms of botulism

Foodborne Botulism

• intoxication is usually caused by ingestion of performed toxin in food

• Toxin is absorbed in GI tract

• It is transported peripheral nerve cells via bloodstream

• It acts on neuromuscular junctions of cholinergic nerves and peripheral autonomic synapses

• Toxin binds to susceptible cells and it enters the cell through endocytosis

Soluble NSF attachment protein receptor

SNARE proteins

ACH

Release of ____ at myoneural junctions is prevented resulting to flaccid paralysis

Wound Botulism

• toxicoinfection

• Sporma are introduced through wounds where they germinate

• Toxin is formed at this localized site and spread throught the body

• Humans - common in drug addicts who use contaminated needles.

• Shaker foal’s syndrome

Infant Botulism

• intratestinal toxicoinfections

• Occurs when spores germinate in the intestines when the normal flora has not yet been fully established

• Human infants - floppy baby syndrome

• Do not feed babies under 12 months old honey

• Broiler chickens and thrkey - rare epidemics of type C

Adult intestinal toxemia

• rare

• Occurs when spores germinate in the intestines and produce toxins.

Iatrogenic botulism

• occurs when too much botulinum toxin is injected for cosmetic reason (wrinkles) or medical reason (migraine headaches)

Clostridium chauvei

• gas-gangrene clostridia

• Endogenous infections

• Often occur with blackleg in calves

• Endospores are ingested and normally pass harmlessly

• Occasionally, spores pass from the intestine via the lymphatics and bloodstream to the muscle masses (hindquarters) and sometimes cardiac muscles

• Trauma to the area causes tissue necroses

• Results to anaerobic conditions favoring germination of spores

• Followed by toxemia and bacteremia

Exogenous infections

Clostridium chauvei

• spores are introduced into wounds

• They will germinate in the anaerobic necrotic material

• Toxin is produced by vegetative cells

Toxin A (CCtA)

• responsible for much of the cytotoxicity and the hemolytic activity

• Pore-forming toxin

• Clostridium septicum

• Any spores present can germinate

• Replication of the bacterium leads to

• Toxin production

• Toxemia

• Rapid death

• Lesions at site of infection, edema at other body sites

C. noyvi type B

• common in soil and normal intestinal tract of herbivores

• Produced black disease (necrotic hepatitis) in dheep

• Animals usually found dead

C. noyvi type D

• found in rhminant digestive tract, liver and in the soil

• Cause if bacillary hemogloobinuria in cattle

C. noyvi type A

• associated with gas gangrene

Clostridium chauvoei

• cattle, sheep

• Blackleg (black quarter)

Clostridium septicum

• cattle sheep and pigs - malignant edema

• Sheep - braxy

• Chickens - necrotic dermatitis

c. noyvi type a

• sheep - big head of rams

• cattle and sheep - gas gangrene

C. noyvi type B

• sheep and cattle - black disease (necrotic hepatitis)

Clostridium sordellii

• cattle sheep horses - gas gangrene

Clostridium perfringens

• clostridial enterotoxemia

• Acute, high fatal introxicstions

• Affects sheep, lambs, calves, piglets and foals

A, B, C, D (and occasionally E)

Major exotoxins of C. perfringes types ____

C. perfringens exotoxin Type A

• occurs in intestinal tracts if humans and animals and soils

C. perfringens exotoxin Type B-E

• more adapted to survival in intestines

• In outbreaks, they survive long enough in soil to infect animals

C. perfringens exotoxin Type C

• does not appear to be a commensal in all instances or species

• True infections occur

c. perfringens toxin alpha

Lecithinase

• attacks cell membranes and results in necroses

C. perfringens toxin Beta

• pore-forming toxin

• lethal and necrotizing

• Binds to endothelial cells and caused necrosis

C. perfringens episilon toxin

• Pro-toxin (activated by proteolytic enzymes)

• Pore-forming

• Damages endothelium concentrates in brain and kidney

C. perfringens Iota toxin

• pro-toxin (activated by proteolyric enzymes)

• Dysregulates cytoskeleton leading to cell-death

CPE (C. perfringens enterotoxin)

• pore-forming toxin which also disrupts tight junctions

• Released during sporulation

• Responsible for food poisoning in people

NetB, NetF (necrotic enteritis toxins)

• pore-forming toxin

Clostridium spiriforme

• produces an exotoxin which is cytotoxic

• Similar in structure and function to iota toxin of C. perfringens type E

• Causes of spontaneous diarrhea in wenaling rabbits

• Frequently isolated from rabbits with mucoid enteritis

• Reported to cause naturally enterocolitis in foals and pigs

Clostridium difficile

• produces enterotoxin (A) and cytotoxin (B)

• Both belongs to group of large clostridial cytotoxins

• Cause of enterocolitis in humans, hamster, rabbit and guinea pig

• Occurs after antimicrobial therapy (clindamycin)

• Antimicrobial therapy alters the intestinal mocrobial population

• Natural diarrheal diseases have been described in foals and neonatal pigs

C. difficile vinary toxin

• 3rd toxin produced by some strains

• Role is unclear

Clostridium colinum

• quail disease

• Affected birds are inactive, sluggish, and anorexic

• May die within 1-2 days

Quail disease

• passes from the intestine via the portal curculation

• Lodges in the liver and causes liver necrosis

• Intestine becomes ulcerated

• Extensive spleen necrosis (in some birds)

• Excreted in feces of birds with chronic form of the condition

Clostridium piliforme

• obligate intracellular pathogen

• Filamentois

• Gram-variable

• Spore-forming

• Cannot be grown in conventional media, but can be cultured in fertile eggs or tissue culture

• Causes Tyzzer’s Disease

Tyzzer’s disease

• severe hepatic necrosis in foals and lab animals

• Can be demonstrated in hepatocytes using warthin-starry silver impregnation technique

• Transmission is spore jngestion (oral-fecal route)

Enterotoxemia

Specimens

• demonstration of toxin is required for diagnosis

• Contents of small intestine are collected from recently dead animal

• These should be submitted to laboratory ASAP as toxins are labile

Affected tissue and a piece of rib with bone marrow (14cm long)

For histotoxic clostridia

• specimen used

Cooked meat broth an thioglycollate medium

• liquid and semisolid media with low redox potential

Anaerobes

Modt pathogenic clostridia are strict ____ except C. perfringens which is aerotolerant

2-10% CO2

Atmosphere containing ____ enhances their growth

Clostridium tetani

• Necrotic tissue from wound ir wound exudate - heated to 80C for 20 mins

• Inoculated on blood agar plate and stiff agar

• Incubated at 37C for 3-4 days under atmosphere of H2 and CO2

C. botulinum

• food samples are macerated in NSS. The suspension is heat at 65,080C for 30 mins

• It will induce C. botulinum spores to germinate.

• Inoculate on blood agar plate with suspension and incubate at 35C for 5 days under atmosphere if H2 and CO2

C. perfringens and C. noyvi

• grow well on normal blood agar

• plates are incubated under strict anaerobic conditions with 10% CO2 at 37C for 2-4 days

Clostridium spiriforme

• non-hemolytic

• Convex

• Circular

• Shiny

• Whitish-grey colonies on blood afar under aerobic conditions at 47C

Clostridium difficile

• non-hemolytic

• Colonies are raised eith a rhizoid edge

• Special blood agar is required (contains yeast extract, haemin, bitamin K, cysteine, and antimicrobial agents.

Clostridium colinum

• fastidious and primary isolation is difficult

• Can be cultured using:

• Tryptose-phosphate-glucose broth with 8% sterile citrated horse plasm

• Thioglycollate broth with 3-10% horse serum

• 6-8 day fertile chicken eggs

• Polymyxin B can be added to isolation media to suppress contaminants.

Fluorescent Antibody

• widely employed technique for specific identification of histotoxic clostridial diseases

Clostridium botulinum

• straight rod

• Produces ival, subterminal soores

• Spores are very resistant - can be killed at 121C for 15 mins

Clostridium tetani

• drumstick form due to spherical spores being terminal and bulging the cell.

• have similar morphology w. c. Tetanomorphum

Clostridium spiriforme

• occurs as loosely-coiled, spirak gram-positive form

• Semi-circular form

• Spores are terminal

• Presence of semicircular gram-positive bacteria in feces or cecal contents is not sufficient for diagnosis

C. difficile

• gram-positive rod that forms ival, subterminal spores.

• Definitive diagnosis is made by enzyme immunoassays (ELISA) for detection of both toxins A and B in fecal specimens

C. colinum

• gram-positive rod that forms oval, subterminal spores

• Sporulation is frequent.

Clostridia with lecithinase activity

• produce an oplascent change around the colonies due to enzymatic action on the lecithin in the medium

Lipase-producing

Cause a pesrly layer or iridescent film

C. perfringens Litmus Milk Medium

• produced stormy-clot or stormy-fermentation

• Lactose is fermented producing acid which coagulates the casein

• Blue to pink

• Acid clot is broken up by gas formation

Vaccination

Best course of action for prevention of disease

tetracycline and metronidazole

C. perfeingens is resistant to ____ in DOGS

Bacitracin

C. perfringens is resistant to ____ in POULTRY

Ciprofloxacin

C. difficule is resitant to ___ in PIGS