CLOSTRIDIUM

Gram Positive, spore-Forming BacilliCLOSTRIDIA

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clostridium

habitat: Soil ,water , decaying animal , human and animal intestines. Type species: C. perfringens. (gas gangrene &food poisoning ) Other very important species are C. tetani (tetanus ) C. Botulinum (botulism) C. difficile.( pseudomembranous colitis)


GENERAL CHARACTERISTICS: 1. Anaerobic gram-positive bacilli 2. Spore formers 3. They decompose proteins 4. Exotoxin producers 5. Natural habitat; Soil or the intestinal tract of animals and man. 6. Some motile with peritrichous flagella (C.tetani ), others are non-motile. 7. Catalase negative. 8. Some species are capsulated (C.perfringens)
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Culture & identification

Grow anaerobically on blood agar or robertson’s cooked meat medium (liquid culture )or thioglycolate broth.

Clostridium perfringens

Spores are found in the soil ,and vegetative cell are normal flora of the colon and vagina. Two important diseases : Gas gangrene ( myonecrosis ) resulting from infection of dirty wound (war injuries). Food poisoning due to ingestion of food contaminated with enterotoxin producing strains.

Characteristics

Short fat bacilli , spores are not usually found as they are formed under nutritionally deficient conditions .more tolerant oxygen than other clostridia.


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Culture: Grows well on Blood Aagar anaerobically at 370C. Selectively isolated with gentamicin blood agar. Some strains are beta haemolytic and show double haemolysis on BA.

Micro & Macroscopic C. perfringens

NOTE: Large rectangular gram-positive bacilli
NOTE: Double zone of hemolysis



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Toxins

C.Perfringens produce a large number of toxins and enzymes that result in a spreading infection, many of these toxins have lethal ,necrotizing and hemolytic properties. Typing: Five types of C. perfringensA, B, C, D and E, Type A is the only human pathogen. The alpha toxin of C.Perfringens type A is lecithinase and its lethal action is proportionate to the rate at which it splits lecithin (an important constituent of the cell membranes ) to phosphorylcholine and diglyceride . -DNase -hyaluronidase -collagenase - some strain of C.Perfringens produce enterotoxin

* Identification: Based on the neutralisation of alpha toxin with specific antitoxin. This is known as Nagler reaction.

* Lactose egg yolk milk agar

Di glycride formation

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Pathogenicity: Causes:- Gas gangrene: wounds associated with overwhelming destruction of tissue.- Food poisoning: when ingested in large numbers, produce entero toxin resulting in diarrhoea.



Gas gangrene


Gas gangrene (myonecrosis): wound associated with traumatized tissue (especially muscle) may become infected with C.perfringens and other clostridia with sever ,life –threatening spreading infection. Activity of bacilli in injured tissue results in toxin and enzymes production ,allowing the bacteria multiply in the wound Characteristics signs and symptoms include: pain ,oedema and crepitation produced by gas in tissues.

* C. perfringens Pathogenesis food poisoning

Vegetative cell
Developing Spore
Toxin inclusion body
Cell wall lysis
Free Spore
Free toxin
Food
GI tract
Spore
Heat

Food Poisoning

Spore Contamination
Cooking
Germination
Slow cooling and/or storage at moderate temperature
Rapid proliferation
Ingestion
GI illness
Environment
Disease cycle

* Clostridium tetani

C. tetani

Habitat &transmission Present in the intestinal tract of animals ,and spores widespread in soil . Germenation of spores is promoted by poor blood supply and neccrotic tissue and debris in wound. Produce neurotoxin (tetanospasmin) by vegetative cells at the wound side, another toxin (tetanolysin) is haemolytic in nature.

Morphology: 1. Gram-positive anaerobic long thin bacilli 2. Motile with peritrichous flagella 3.Terminal rounded spores (Drumstick appearance). Media of Choice: Any medium with thioglycollate under anaerobic condition most especially Cooked Meat Media. Identification in vitro is by a toxin neutralization test on blood agar or in vivo by inoculation of culture filtrate into mice .
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Pathogenesis

The agent of tetanus (lockjaw) which is a typical toxin-mediated disease .the powerful,heat- labile neurotoxin is produced at the wound site and released during cell lysis .it carried by the peripheral nervous to the central nervous system where it blocks inhibitory mediators at spinal synapses . This cause muscle spasm and the characteristics signs of spasm of jaw muscles (lockjaw) , facial muscles and arching of the body .

Neonatal tetanus

Neonatal tetanus
This baby has neonatal tetanus. It is completely rigid. Tetanus kills most of the babies who get it. Infection usually happens when newly cut umbilical cord is exposed to dirt

This baby has tetanus. He cannot breast feed or open his mouth because the muscles in his face have become so tight

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Clostridium botulinum

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Clostridium botulinum

-Cause botulism -It found in the soil and in animals feces. -Types of Clostridium botulinum are distinguished by the antigenic type of toxin. -spores are highly resistant to heat ,spores stay at 100Cfor several hours ,resistant to acidic pH and high salt concentration.


Morphological Characteristics: -Spores are oval, and subterminal -Motile with peritrichous flagella
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* Toxicity: Botulinum toxin is heat resistant Botulinum toxin is the most powerful toxin known to man (1mg pure toxin has a 20 million lethal mice doses). Toxine Classification: 8 antigenic types has been identified (A-H) of toxin . Botox Only A, B, E, and F affects human. 1. Type A: Most common in western parts of US. More toxic than B. 2. Type B: Less toxic most frequent in occurrence. 3. Type E: Obtained from fish and fish products. 4. Type F: Similar to A and B. Has been isolated in Denmark.



Toxin composed of two chains(light and heavy )which linked by disulfide bond. Botulinum toxin is absorbed from the gut and bind to receptors of presynaptic membranes of motor neurons of the peripheral nervous system . Proteolysis -by the light chain of botulinum toxin-of the target SNARE protein in the neurons inhibits the release of acetylcholine at the synapse resulting in lack of muscle contraction and paralysis

Pathogenesis

The most common contaminated food are spiced ,smoked ,vacuum-packed or canned alkaline food that are eaten without cooking the spores germinate under anaerobic condition vegetative form grow and produce toxin. In infant botulism ,honey is the most cause of infection by ingests the spores and then germinate within intestinal tract and produce the toxin and the neurotoxin then gets absorbed into the blood stream and the toxin acts by blocking release of acetylcholine at synapses and neuromuscular junction .flaccid paralysis result


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C.difficile

C.difficile



-found in the faces of 3-6%adult and almost all healthy infants . -it is the agent of antibiotic-associated colitis which lead sometimes lethal pseudomembranous colitis. -it multiplies in the gut under the selective pressure of antibiotics. -the diarrhea may be watery or bloody and the patient frequently has associated abdominal cramps ,leukocytosis and fever.


-many antibiotics associated with this disease like: ampicillin,clindamycin. -then C.difficile produce two toxins : toxin A (enterotoxin)and also has some cytotoxic activity and toxin B is as cytotoxin -C.difficile responsible about 25% of cases of antibiotics associated diarrhea pseudomembranous colitis.