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Bacilli anthracis

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12/19/2025 1 Spore-Forming Gram-Positive Bacilli: Bacillus and Clostridium Species Presenter : Dr. Evelyn Godfrey MMED 2 M& I University of Dodoma
12/19/2025 2 Table of contents: 1. Introduction 2. Bacillus anthracis 3. Bacillus cereus 4. Clostridium botulinum 5. Clostridium tetani 6. Clostridia That Produce Invasive Infections 7. Clostridium difficile And Diarrheal Disease
12/19/2025 3 Introduction • The Gram-positive spore-forming bacilli are the Bacillus and Clostridium species. • These bacilli are ubiquitous and because they form spores, they can survive in the environment for many years. • Bacillus species are aerobes and the Clostridium species are anaerobes . • Species of Bacillus and related genera most do not cause disease and are not well characterized in medical microbiology.
12/19/2025 4 Cont.. • Anthrax, a classical disease in the history of microbiology is caused by Bacillus anthracis. • Anthrax remains an important disease of animals and occasionally of humans B. anthracis is a major potential agent of bioterrorism and biologic warfare. • B. cereus causes food poisoning and occasionally eye or other localized infections.
12/19/2025 5 Cont.. • The genus Clostridium is extremely heterogeneous and more than 200 species have been described. • Clostridia cause several important toxin-mediated diseases, including tetanus (Clostridium tetani), botulism (C. botulinum), gas gangrene (C. perfringens), antibiotic-associated diarrhea and pseudomembranous colitis (C. difficile).
12/19/2025 6 Bacillus spp • The genus Bacillus includes large aerobic, Gram-positive rods occurring in chains. • The members of the genus Bacillus are closely related but differ both phenotypically and in terms of pathogenesis. Pathogenic species possess virulence plasmids. • Most members of the genus Bacillus are saprophytic organisms prevalent in soil, water, and air, and on vegetation (e.g. B. subtilis).
12/19/2025 7 Cont.. • B. cereus can grow in foods and cause food poisoning by producing either an enterotoxin (diarrhea) or an emetic toxin (vomiting). • B. cereus may occasionally produce disease in immunocompromised humans (eg, meningitis, endocarditis, endophthalmitis, conjunctivitis, or acute gastroenteritis). • B. anthracis, which causes anthrax, is the principal pathogen of the genus.
12/19/2025 8 Bacillus anthracis • B. anthracis, the causative agent of anthrax, has a worldwide distribution. • Anthrax is caused by inhalation, skin exposure or by gastrointestinal (GI) absorption. • B. anthracis was the first pathogenic bacterium to be seen under microscope. • B. anthracis was the first bacterium shown to be the cause of a disease.
12/19/2025 9 Cont.. • It was from studies on anthrax that Koch established his famous postulates in 1876. In 1877, Robert Koch grew the organism in pure culture, demonstrated its ability to form endospores and produced experimental anthrax by injecting it into animals. • It was the first bacterium to be isolated in pure culture and shown to possess spores. • For the first time, spores were discovered from B. anthracis and Bacillus subtilis by Koch and Cohn hence they established the germ theory of disease. • B. antracis was the first bacterium used for the preparation of attenuated vaccine by Louis Pasteur.
12/19/2025 10 Morphology • B. anthracis is a Gram-positive spore-forming bacillus. • It is a very large bacillus measuring 1–1.2 m in width and 3–5 m in length. • In smears from infected tissues, the bacteria are found as single, in pairs, and in short chains, the entire chain being surrounded by a capsule. • In culture, B. anthracis grows as long chains and may appear similar to streptobacilli.
12/19/2025 11 Cont.. • The bacilli are arranged end-to-end and the ends of the bacilli are truncated, not rounded or often concave and somewhat swollen. • This gives the chain of bacilli a “bamboo-stick” appearance . • The bacterium forms the capsule only when grown on nutrient agar containing 0.7% sodium bicarbonate in the presence of 5–20% carbon dioxide. • The capsule is polypeptide (polymer of D-glutamic acid) in nature. • B. anthracis is nonmotile and nonacid fast.
12/19/2025 12 chain of bacilli a “bamboo-stick” appearance of B. anthracis
12/19/2025 13 Culture • B. anthracis is an aerobe and facultative anaerobe. The bacteria grow at a temperature range of 12–45°C, optimum temperature being 37°C. • B. anthracis grow on a wide range of media including ordinary nutrient media and several selective media. • Nutrient agar: On nutrient agar after 24 hours of incubation, B. anthracis produces grayish and granular colonies measuring 2–3 mm in diameter. Under low-power microscopy, the edges of the colony appear as long, interlacing chains of bacilli resembling locks of matted hair, which gives them a “medusa head” appearance with an uneven surface and wavy margin.
12/19/2025 14 A “medusa head” appearance with an uneven surface and wavy margin on nutrient agar.
12/19/2025 15 Cont.. • Blood agar: On horse or sheep blood agar, B. anthracis colonies are gray or white, typically nonhemolytic, with a dry, ground-glass appearance. The colonies are at least 3 mm in diameter and sometimes have tails. • Solid medium containing penicillin: B. anthracis on a solid medium containing 0.05–0.5 U of penicillin/mL produces large, spherical colonies within 3–6 hours and occurs in chains on the surface of the agar, resembling a string of pearls. This property is known as string of pearls reaction and is useful in differentiation of B. anthracis from B. cereus and other aerobic spore bearers.
12/19/2025 16 Cont.. • Gelatin medium: In a gelatin stab, there is growth down the stab line with lateral spikes, longer near the surface, giving an ‘inverted fir tree’ appearance. The process of liquefaction is slow and late, which occurs after 7 days at 20°C and starts at the surface.
12/19/2025 17 Cont.. • Selective medium: Knisely’s Polymyxin B-Lysozyme-EDTA Thallous acetate (PLET) agar medium is a selective medium used for isolation of B. anthracis from mixtures containing other spore-bearing bacilli. The medium is composed of heart infusion agar, polymyxin, lysozyme, ethylene diamine tetra acetic acid (EDTA) and thallous acetate.
12/19/2025 18 Cont.. Production of capsular material is associated with the formation of a characteristic mucoid or “smooth” (S) colony type. Capsulated bacteria on serum or bicarbonate medium produce smooth or mucoid colonies. Smooth variants that form capsule are virulent strains of B. anthracis. Rough (R) variants that lack capsule are relatively avirulent.
12/19/2025 19 Biochemical reactions • B. anthracis produces acid from glucose, maltose, sucrose, trehalose and dextrin but not from lactose, arabinose, D-xylose, or D-mannitol. • They reduce nitrate to nitrite. • They demonstrate a weak lecithinase reaction on egg-yolk agar, which gives a narrow zone of opalescence around the colonies. • They are also catalase positive.
12/19/2025 20 Pathogenesis • Anthrax is primarily a disease of herbivores goats, sheep, cattle and horses, Humans become infected incidentally by contact with infected animals or their products. • Spores from contaminated soil find easy access when ingested with spiny or irritating vegetation. • In humans, the infection is usually acquired by the entry of spores through injured skin (cutaneous anthrax) or rarely the mucous membranes (gastrointestinal anthrax) or by inhalation of spores into the lung (inhalation anthrax)
12/19/2025 21 Cont.. • The spores germinate in the tissue at the site of entry and growth of the vegetative organisms results in formation of a gelatinous edema and congestion. • Bacilli spread via lymphatics to the bloodstream, and they multiply freely in the blood and tissues shortly before and after the animal’s death. • B. anthracis isolates that do not produce a capsule are not virulent and do not induce anthrax in test animals. • The poly-γ-d-glutamic acid capsule is ant phagocytic. The capsule gene is present on a plasmid.
12/19/2025 22 Cont.. • Anthrax toxins are made up of three proteins: a) Protective antigen (PA) b) Edema factor (EF) c) Lethal factor (LF). • PA is a protein that binds to specific cell receptors and after proteolytic activation, it forms a membrane channel that mediates entry of EF and LF into the cell. • Edema factor is an adenylate cyclase with protective factor it forms a toxin known as edema toxin. Edema toxin is responsible for cell and tissue edema.
12/19/2025 23 Cont.. • Lethal factor plus protective factor form lethal toxin, which is a major virulence factor and cause of death in infected animals and humans. • The lethal toxin can quickly kill the animals by impairing both innate and adaptive immunity allowing organism proliferation and cell death. • The anthrax toxin genes are encoded on another plasmid, pXO1.
12/19/2025 24 B. anthracis pathogenesis
12/19/2025 25 Cont.. • In inhalation anthrax (wool sorters’ disease), the spores from the dust of wool, hair or hides are inhaled, phagocytosed in the lungs and transported by the lymphatic drainage to the mediastinal lymph nodes, where germination occurs. • This is followed by toxin production and the development of hemorrhagic mediastinitis and sepsis which are usually rapidly fatal. • In anthrax sepsis, the number of organisms in the blood exceeds 107 /mL just before death.
12/19/2025 26 Pathology • In susceptible animals and humans, the organisms proliferate at the site of entry. • The capsules remain intact and the organisms are surrounded by a large amount of proteinaceous fluid containing few leukocytes from which they rapidly disseminate and reach the bloodstream. • In resistant animals, the organisms proliferate for a few hours, by which time there is massive accumulation of leukocytes. • The capsules gradually disintegrate and disappear. The organisms remain localized.
12/19/2025 27 Clinical Findings • In humans, approximately 95% of cases are cutaneous anthrax, and 5% are inhalation. Gastrointestinal anthrax is very rare. • Cutaneous anthrax generally occurs on exposed surfaces of the arms or hands followed in frequency by the face and neck. • A pruritic papule develops 1–7 days after entry of the organisms or spores through a scratch.
12/19/2025 28 A pruritic papule of B. anthracis
12/19/2025 29 Cont.. • Initially, it resembles an insect bite. The papule rapidly changes into a vesicle or small ring of vesicles that coalesce, and a necrotic ulcer develops. • The lesions typically are 1–3 cm in diameter and have a characteristic central black eschar. • Lymphangitis, lymphadenopathy and systemic signs and symptoms of fever, malaise and headache.
12/19/2025 30 Cont.. • After 7–10 days, the eschar is fully developed. Eventually it dries, loosens and separates; healing is by granulation and leaves a scar. • It may take many weeks for the lesion to heal and the edema to subside. • Antibiotic therapy does not appear to change the natural progression of the disease but prevents dissemination.
12/19/2025 31 Cont.. • Cutaneous anthrax can lead to sepsis, the consequences of systemic infection, including meningitis and death. • The incubation period in inhalation anthrax may be as long as 6 weeks. • The early clinical manifestations are associated with marked hemorrhagic necrosis and edema of the mediastinum.
12/19/2025 32 Cont.. • Substernal pain may be prominent and there is pronounced mediastinal widening visible on chest radiographs. • Hemorrhagic pleural effusions follow involvement of the pleura; cough is secondary to the effects on the trachea. • Sepsis occurs and there may be hematogenous spread to the gastrointestinal tract, causing bowel ulceration, or to the meninges, causing hemorrhagic meningitis. • The fatality rate in inhalation anthrax is high in the setting of known exposure, it is higher when the diagnosis is not initially suspected.
12/19/2025 33 Diagnostic Laboratory Tests • Specimens to be examined are fluid or pus from a local lesion, blood, pleural fluid and cerebrospinal fluid in inhalational anthrax associated with sepsis and stool or other intestinal contents in the case of gastrointestinal anthrax. • Stained smears from the local lesion or of blood from dead animals often show chains of large Gram-positive rods. • Anthrax can be identified in dried smears by immunofluorescence staining techniques.
12/19/2025 34 Cont.. • B. anthracis grown on blood agar plates, the organisms produce nonhemolytic gray to white, tenacious colonies with a rough texture and a ground-glass appearance. Commas shaped outgrowths (Medusa head, “curled hair”) may project from the colony. • Demonstration of capsule requires growth on bicarbonate-containing medium in 5–7% carbon dioxide. Gram-stain shows large Gram- positive rods.
12/19/2025 35 Cont.. • In semisolid medium, anthrax bacilli are always nonmotile, but related organisms (eg, B. cereus) exhibit motility by “swarming.” • Clinical laboratories that recover large Gram-positive rods from blood, cerebrospinal fluid, or suspicious skin lesions, which phenotypically match the description of B. anthracis as mentioned, should immediately contact their public health laboratory and send the organism for confirmation. • Definitive identification requires lysis by a specific anthrax γ- bacteriophage, detection of the capsule by fluorescent antibody or identification of toxin genes by polymerase chain reaction (PCR).
12/19/2025 36 Resistance and Immunity • Immunization to prevent anthrax is based on the classic experiments of Louis Pasteur. • In 1881 he proved that cultures grown in broth at 42–52°C for several months lost much of their virulence and could be injected live into sheep and cattle without causing disease subsequently such animals proved to be immune. • Active immunity to anthrax can be induced in susceptible animals by vaccination with live attenuated bacilli, with spore suspensions or with protective antigen from culture filtrates.
12/19/2025 37 Treatment • Ciprofloxacin is recommended for treatment, other agents with activity include penicillin G, doxycycline, erythromycin and vancomycin. • Setting of potential exposure to B. anthracis as an agent of biologic warfare, prophylaxis with ciprofloxacin or doxycycline should be given for 60 days and three doses of vaccine (AVA Bio Thrax) should be administered. • AIGIV is a human polyclonal antiserum that also inhibits binding of PA to its receptors. Like Raxibacumab it is used as an adjunct to antimicrobial agents for the treatment of severe forms of anthrax.
12/19/2025 38 Epidemiology, Prevention and Control • Soil is contaminated with anthrax spores from the carcasses of dead animals. These spores remain viable for decades. Perhaps, spores can germinate in soil at a pH of 6.5 at proper temperature. • Grazing animals infected through injured mucous membranes serve to perpetuate the chain of infection. • Contact with infected animals or with their hides, hair and bristles is the source of infection in humans.
12/19/2025 39 Cont.. Control measures include: 1) Disposal of animal carcasses by burning or by deep burial in lime pits. 2) Decontamination (usually by autoclaving) of animal products. 3) Protective clothing and gloves for handling potentially infected materials. 4) Active immunization of domestic animals with live attenuated vaccines. 5) Persons with high occupational risk should be immunized.
12/19/2025 40 Bacillus cereus • Food poisoning caused by B. cereus has two distinct forms. The emetic type which is associated with fried rice, milk and pasta and the diarrheal type which is associated with meat dishes and sauces. • The emetic form is manifested by nausea, vomiting, abdominal cramps and occasionally diarrhea and is self-limiting with recovery occurring within 24 hours. It begins 1–5 hours after ingestion of a plasmid-encoded preformed cyclic peptide (emetic toxin) in the contaminated food products.
12/19/2025 41 Cont.. • B. cereus is a soil organism that commonly contaminates rice. When large amounts of rice are cooked and allowed to cool slowly, the B. cereus spores germinate and the vegetative cells produce the toxin during log-phase growth or during sporulation. • B. cereus produces toxins that cause disease that is more of intoxication than a food-borne infection.
12/19/2025 42 Cont.. • The diarrheal form has an incubation period of 1–24 hours and is manifested by profuse diarrhea with abdominal pain and cramps fever and vomiting are uncommon. • In this syndrome, ingested spores that develop into vegetative cells of B. cereus secrete one of three possible enterotoxins which induce fluid accumulation and other physiological responses in the small intestine.
12/19/2025 43 Cont.. • The presence of B. cereus in a patient’s stool is not sufficient to make a diagnosis of B. cereus disease because the bacteria may be present in normal stool specimens; a concentration of 105 bacteria or more per gram of food is considered diagnostic. • B. cereus is an important cause of eye infections such as severe keratitis and endophthalmitis. • Typically the organisms are introduced into the eye by foreign bodies associated with trauma but infections can also occur after surgery.
12/19/2025 44 Cont.. • B. cereus has also been associated with localized infections such as wound infections and with systemic infection including endocarditis, catheter-associated bacteremia, central nervous system infections, osteomyelitis and pneumonia; the presence of a medical device or intravenous drug use predisposes to these infections. • Outbreaks of bacteremia in neonatal intensive care units and other hospital units during construction in health care facilities have been reported.
12/19/2025 45 Cont.. • B. cereus is resistant to a variety of antimicrobial agents, including penicillins and cephalosporins. • Serious nonfoodborne infections should be treated with vancomycin or clindamycin with or without an aminoglycoside. • Ciprofloxacin has been useful for the treatment of wound infections.
12/19/2025 46 Clostridium spp • The clostridia are large anaerobic, Gram-positive, motile rods. Many decompose proteins or form toxins, and some do both. • Their natural habitat is the soil, marine sediments, sewage or the intestinal tract of animals and humans where they live as saprophytes. • The clostridia continue to increase in number as new species are discovered and several species have been sequenced. • There are 19 clusters based on 16SrRNA gene sequence analysis. Most clinically related species are in RNA Cluster I. Among the pathogens in this cluster are the organisms causing botulism, tetanus, gas gangrene and pseudomembranous colitis.
12/19/2025 47 Clostridium spp
12/19/2025 48 Morphology • Most species are anaerobic but few species such as Clostridium tertium, Clostridium histolyticum, etc. are aero tolerant and hence can grow on agar even in presence of air. • Clostridia are rod-shaped bacilli measuring 3–8 0.4–1.2 m in size. • The bacilli are highly pleomorphic and show long filaments and involution forms.
12/19/2025 49 Cont.. • They are Gram-positive but old cultures may appear Gram variable and even Gram-negative. • Most Clostridium species with few exceptions (e.g., Clostridium perfringens, Clostridium tetani type VI) are motile due to the presence of peritrichous flagella. • Most clostridia except C. perfringens and Clostridium butyricum are non capsulated • Clostridia has ability to produce endospores.
12/19/2025 50 Culture • Most species are anaerobic but few species, such as C. tertium, C. histolyticum, are aero tolerant and hence can grow on agar even in presence of air. • Presence of adequate quantity of low redox potential (Eh) substances in the media is important for the growth of these anaerobic bacteria. • Use of reducing substances, such as ascorbic acid, glutathione, unsaturated fatty acids, cysteine, thioglycolic acid, alkaline glucose or metallic iron maintain low redox potentials in the medium.
12/19/2025 51 Cont.. • Most clostridia grow at an optimum temperature of 37°C and pH of 7– 7.4. Growth on solid media is variable. Some clostridial species produce hemolysis on blood agar. • Robertson’s cooked meat (RCM) broth is a useful medium for the growth of clostridia. • The medium contains unsaturated fatty acids, digested meat, and sulfhydryl compounds. Clostridia grow well in the medium, rendering the broth turbid. • Saccharolytic clostridia turn the meat pink while proteolytic species turn the meat black and produce foul smell. Most clostridia produce gas.
12/19/2025 52 Clostridium botulinum • C. botulinum which causes the disease botulism, is worldwide in distribution and it is found in soil and occasionally in animal feces. • Types of C. botulinum are distinguished by the antigenic type of toxin they produce. • Spores of the organism are highly resistant to heat, withstanding 100°C for several hours. • Heat resistance is diminished at acid pH or high salt concentration.
12/19/2025 53 Toxins • During the growth of C. botulinum and during autolysis of the bacteria, toxin is liberated into the environment. • Seven antigenic varieties of toxin (serotypes A–G) are known. • Types A, B, E, and F are the principal causes of human illness. Types A and B have been associated with a variety of foods and type E predominantly with fish products. • Botulinum toxins have three domains. Two of the domains facilitate binding to and entry of toxin into the nerve cell.
12/19/2025 54 Cont.. • The third domain is the toxin which is a 150 kDa protein that is cleaved into a heavy chain (H, 100 kDa) and a light chain (L, 50 kDa) that are linked by a disulfide bond. • Botulinum toxin is absorbed from the gut, enters the blood circulation and binds to receptors of presynaptic membranes of motor neurons of the peripheral nervous system and cranial nerves. • The toxin does not cross the blood brain barrier or affect the central nervous system.
12/19/2025 55 Cont.. • Proteolysis by the L chain of botulinum toxin: of the target SNARE proteins (soluble-N-ethyl maleimide-sensitive factor attachment protein) in the neurons inhibits the release of acetylcholine at the synapse, resulting in lack of muscle contraction and paralysis.
12/19/2025 56 Cont.. • The SNARE proteins are synaptobrevin (also known as vesicle- associated membrane protein or VAMP) SNAP 25 and syntaxin. • The toxins of C. botulinum types A, C, and E cleave the 25,000 kDa SNAP 25. • Type C also cleaves syntaxin. Types B, D, F, and G toxins cleave only synaptobrevin.
12/19/2025 57 Cont.. • C. botulinum toxins are among the most toxic substances known: The lethal dose for a human is probably about 1–2 µg/kg. • The toxins are destroyed by heating for 20 minutes at 100°C. • Strains that produce toxins A, B, or F are associated with infant botulism.
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12/19/2025 59 Pathogenesis • The most common offenders are spiced, smoked, vacuum packed or canned alkaline foods that are eaten without cooking. In such foods, spores of C. botulinum germinate under anaerobic conditions, vegetative forms grow and produce toxin. • The pathogenesis differs from the way that adults acquire infection. • The infant ingests the spores of C. botulinum and the spores germinate within the intestinal tract. The vegetative cells produce toxin as they multiply; the neurotoxin then gets absorbed into the bloodstream.
12/19/2025 60 Cont.. • In rare instances adults with gastrointestinal anatomical abnormalities or functional disorders may develop “infant botulism.” • Wound botulism is the result of tissue contamination with spores and is seen primarily in injection drug users. • Very rarely, inhalational botulism occurs when toxin enters the respiratory tract. The toxin acts by blocking release of acetylcholine at synapses and neuromuscular junctions . • The result is flaccid paralysis. The electromyogram and edrophonium strength test results are typical.
12/19/2025 61
12/19/2025 62 Infant botulism
12/19/2025 63 Clinical Findings. • Symptoms begin 18–24 hours after ingestion of the toxic food, with visual disturbances (incoordination of eye muscles, double vision), inability to swallow and speech difficulty, signs of bulbar paralysis are progressive and death occurs from respiratory paralysis or cardiac arrest. • Gastrointestinal symptoms are not prominent. There is no fever. The patient remains fully conscious until shortly before death. The mortality rate is high. Patients who recover do not develop antitoxin in the blood.
12/19/2025 64 Cont.. • The infants in the first months of life develop poor feeding, weakness, and signs of paralysis (floppy baby). • Infant botulism may be one of the causes of sudden infant death syndrome. • C. botulinum and botulinum toxin are found in feces but not in serum.
12/19/2025 65 Diagnostic Laboratory Tests • Clinicians who suspect a case of botulism should contact the appropriate public health authorities before submitting specimens to the laboratory. • Detection of toxin and not the organism is required for definitive diagnosis. Toxin can often be demonstrated in serum, gastric secretions or stool from the patient and toxin may be found in leftover food.
12/19/2025 66 Cont.. • Clinical swabs or other specimens obtained from patients should be transported using anaerobe containers. • Suspect foods should be left in their original containers. This mouse bioassay is the test of choice for the confirmation of botulism. • C. botulinum may be grown from food remains and tested for toxin production, but this is rarely done and is of questionable significance. • In infant botulism, C. botulinum and toxin can be demonstrated in bowel contents but not in serum. Other methods used to detect toxin include ELISAs and PCR, but the latter may detect organisms that carry the gene but do not express toxin.
12/19/2025 67 Treatment • Supportive care, especially intensive care, is key in the management of patients with botulism. • Adequate respiration must be maintained by mechanical ventilation if necessary and in severe cases may need to be maintained for up to 8 week. • Antitoxin does not reverse the paralysis but if administered early it can prevent its advancement. Although most infants with botulism recover with supportive care alone treatment with human derived botulinum immune globulin (BIG) is recommended
12/19/2025 68 Epidemiology and Prevention • Spores of C. botulinum are widely distributed in soil, they often contaminate vegetables, fruits and other materials. • A chief risk factor for botulism lies in home-canned foods, particularly string beans, corn, peppers, olives, peas and smoked fish or vacuum- packed fresh fish in plastic bags. • The risk from home-canned foods can be reduced if the food is boiled for more than 20 minutes before consumption. • Botulinum toxin is considered to be a major potential agent for bioterrorism and biologic warfare.
12/19/2025 69 Clostridium tetani • C. tetani, which causes tetanus, is worldwide in distribution in the soil and in the feces of horses and other animals. • Several types of C. tetani can be distinguished by specific flagellar antigens. All share a common O (somatic) antigen, which may be masked and all produce the same antigenic type of neurotoxin, tetanospasmin.
12/19/2025 70 Toxin • The vegetative cells of C. tetani produce the plasmid-encoded toxin tetanospasmin (150 kDa) that is cleaved by a bacterial protease into two peptides (50 and 100 kDa) linked by a disulfide bond. • The larger peptide initially binds to receptors on the presynaptic membranes of motor neurons. It then migrates by the retrograde axonal transport system to the cell bodies of these neurons to the spinal cord and brainstem. • The toxin diffuses to terminals of inhibitory cells, including both glycinergic interneurons and γ-aminobutyric acid (GABA)– secreting neurons from the brainstem.
12/19/2025 71 Cont.. • The smaller peptide degrades synaptobrevin a protein required for docking of neurotransmitter vesicles on the presynaptic membrane. Release of the inhibitory glycine and GABA is blocked, and the motor neurons are not inhibited. • Hyperreflexia, muscle spasms, and spastic paralysis result. • Extremely small amounts of toxin can be lethal for humans.
12/19/2025 72 C. tetani toxin
12/19/2025 73 Pathogenesis • C. tetani is not an invasive organism. The infection remains strictly localized in the area of devitalized tissue (wound, burn, injury, umbilical stump, surgical suture) into which the spores have been introduced. • The volume of infected tissue is small, and the disease is almost entirely a toxemia.
12/19/2025 74 Cont.. • Germination of the spore and development of vegetative organisms that produce toxin are aided by necrotic tissue, calcium salts and associated pyogenic infections all of which aid establishment of low oxidation reduction potential. • The toxin released from vegetative cells reaches the central nervous system and rapidly becomes fixed to receptors in the spinal cord and brainstem and exerts the actions.
12/19/2025 75 Clinical Findings • The incubation period may range from 4 to 5 days up to 3 weeks. • The disease is characterized by tonic contraction of voluntary muscles. Muscular spasms often involve first the area of injury and infection and then the muscles of the jaw (trismus, lockjaw), which contract so that the mouth cannot be opened. • Gradually, other voluntary muscles become involved, resulting in tonic spasms.
12/19/2025 76 Cont.. • Any external stimulus may precipitate a tetanic generalized muscle spasm. • The patient is fully conscious and pain may be intense. • Death usually results from interference with the mechanics of respiration. • The mortality rate in generalized tetanus is very high.
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12/19/2025 78 Diagnosis • The diagnosis rests on the clinical picture and a history of injury although only 50% of patients with tetanus have an injury for which they seek medical attention. • The primary differential diagnosis of tetanus is strychnine poisoning. • Anaerobic culture of tissues from contaminated wounds may yield C. tetani, but neither preventive nor therapeutic use of antitoxin should ever be withheld pending such demonstration. • Proof of isolation of C. tetani must rest on production of toxin and its neutralization by specific antitoxin.
12/19/2025 79 Prevention Treatment • Prevention of tetanus depends on active immunization with toxoids, aggressive wound care, prophylactic use of antitoxin and administration of penicillin. • The intramuscular administration of 250–500 units of human antitoxin (tetanus immune globulin) gives adequate systemic protection (0.01 unit or more per milliliter of serum) for 2–4 weeks , It neutralizes the toxin that has not been fixed to nervous tissue. • Active immunization with tetanus toxoid should accompany antitoxin prophylaxis.
12/19/2025 80 Cont.. • Patients who develop symptoms of tetanus should receive muscle relaxants, sedation, and assisted ventilation. • Surgical debridement is vitally important because it removes the necrotic tissue that is essential for proliferation of the organisms. • Penicillin strongly inhibits the growth of C. tetani and stops further toxin production.
12/19/2025 81 Cont.. • When a previously immunized individual sustains a potentially dangerous wound, an additional dose of toxoid should be injected to restimulate antitoxin production. • This “recall” injection of toxoid may be accompanied by a dose of antitoxin if the patient has not had current immunization or boosters or if the history of immunization is unknown. • Initial immunization should be carried out in all children during the first year of life.
12/19/2025 82 Clostridia That Produce Invasive Infections. • Many different toxin-producing clostridia (C. perfringens and related clostridia) can produce invasive infection (including myonecrosis and gas gangrene) if introduced into damaged tissue. • An enterotoxin of C. perfringens is a common cause of food poisoning.
12/19/2025 83 Toxins • The invasive clostridia produce a large variety of toxins and enzymes that result in a spreading infection. • Many of these toxins have lethal, necrotizing and hemolytic properties. In some cases, these are different properties of a single substance in other instances, they are attributable to different chemical entities. • The alpha toxin of C. perfringens type A is a lecithinase, and its lethal action is proportionate to the rate at which it splits lecithin (an important constituent of cell membranes) to phosphorylcholine and diglyceride.
12/19/2025 84 Cont.. • Alpha toxin also aggregates platelets, thereby leading to formation of thrombi in small blood vessels and adding to poor tissue profusion and extending the consequences of anaerobiosis, namely, destruction of viable tissue (gas gangrene). • The theta toxin has similar hemolytic and necrotizing effects but is not a lecithinase. It is a member of the cholesterol-dependent cytolysins that act by forming pores in cell membranes.
12/19/2025 85 Cont.. • Epsilon toxin is a protein that causes edema and hemorrhage is very potent. • DNase and hyaluronidase, a collagenase that digests collagen of subcutaneous tissue and muscle, are also produced. Some strains of C. perfringens produce a powerful enterotoxin (C. perfringens enterotoxin, CPE), especially when grown in meat dishes. When more than 108 vegetative cells are ingested and sporulate in the gut, CPE is formed. • CPE is a protein (35 kDa) that may be a nonessential component of the spore coat; it is distinct from other clostridial toxins. It induces intense diarrhea in 7–30 hours.
12/19/2025 86 Cont.. • The action of C. perfringens enterotoxin involves marked hypersecretion in the jejunum and ileum, with loss of fluids and electrolytes in diarrhea eg nausea, vomiting, and fever. • This illness is similar to that produced by B. cereus and tends to be self-limited. • Enterotoxin-producing strains of C. perfringens may also play a role in antibiotic associated diarrhea and necrotizing enterocolitis in infants.
12/19/2025 87
12/19/2025 88 Pathogenesis • In invasive clostridial infections, spores reach tissue either by contamination of traumatized areas (soil, feces) or from the intestinal tract. • The spores germinate at low oxidation - reduction potential; vegetative cells multiply, ferment carbohydrates present in tissue, and produce gas. • The distention of tissue and interference with blood supply, together with the secretion of necrotizing toxins and hyaluronidase, favor the spread of infection.
12/19/2025 89 Cont.. • Tissue necrosis extends providing an opportunity for increased bacterial growth, hemolytic anemia and ultimately, severe toxemia and death. • In gas gangrene (clostridial myonecrosis) a mixed infection is the rule. In addition to the toxigenic clostridia, proteolytic clostridia and various cocci and Gram-negative organisms are also usually present. • C. perfringens occurs in the genital tracts of 5% of women. Before legalization of abortion in the United States, clostridial uterine infections followed instrumented abortions.
12/19/2025 90 Cont.. • C. sordellii has many of the properties of C. perfringens. • C. sordellii has been reported to cause a toxic shock syndrome after medical abortion with mifepristone and intravaginal misoprostol. • Endometrial infection with C. sordellii is implicated. Clostridial bacteremia especially that caused by C. septicum, is a frequent occurrence in patients with neoplasms. • In New Guinea, C. perfringens type C produces necrotizing enteritis (pigbel) that can be highly fatal in children. • Immunization with type C toxoid appears to have preventive value.
12/19/2025 91 Clinical Findings and Treatment • From a contaminated wound (e.g., a compound fracture, postpartum uterus) the infection spreads in 1–3 days to produce crepitation in the subcutaneous tissue and muscle, foul-smelling discharge, rapidly progressing necrosis, fever, hemolysis, toxemia, shock and death. • Treatment is with early surgery (amputation) and antibiotic administration. Advent of specific therapy, early amputation was the only treatment
12/19/2025 92 Cont.. • At times the infection results only in anaerobic fasciitis or cellulitis. • C. perfringens food poisoning usually follows the ingestion of large numbers of clostridia that have grown in warmed meat dishes. • The toxin forms when the organisms sporulate in the gut, with the onset of diarrhea; usually without vomiting or fever—in 7–30 hours. The illness lasts only 1–2 days.
12/19/2025 93 Diagnostic Laboratory Tests • Specimens consist of material from wounds, pus, and tissue. • The presence of large Gram-positive rods in Gram-stained smears suggests gas gangrene clostridia; spores are not regularly present. • Material is inoculated into chopped meat–glucose medium and thioglycolate medium and onto blood agar plates incubated anaerobically.
12/19/2025 94 Cont.. • After pure cultures have been obtained by selecting colonies from anaerobically incubated blood plates, they are identified by biochemical reactions (various sugars in thioglycolate, action on milk), hemolysis and colony morphology.
12/19/2025 95 Cont.. • Lecithinase activity is evaluated by the precipitate formed around colonies on egg yolk media. • Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a rapid and sensitive method for identification of invasive Clostridium species recovered in culture. • C. perfringens rarely produces spores when cultured on agar in the laboratory.
12/19/2025 96 Treatment • Extensive surgical debridement of the involved area and excision of all devitalized tissue, in which the organisms are prone to grow. • Administration of antimicrobial drugs, particularly penicillin, is begun at the same time. • Hyperbaric oxygen may be of help in the medical management of clostridial tissue infections. It is said to “detoxify” patients rapidly.
12/19/2025 97 Cont.. • Antitoxins are available against the toxins of C. perfringens, C. novyi, C. histolyticum, and C. septicum, usually in the form of concentrated immune globulins. • Polyvalent antitoxin (containing antibodies to several toxins) has been used.
12/19/2025 98 Prevention and Control • Early and adequate cleansing of contaminated wounds and surgical debridement, together with the administration of antimicrobial drugs directed against clostridia (eg, penicillin), are the best available preventive measures.
12/19/2025 99 Clostridium difficile And Diarrheal Disease • Pseudomembranous Colitis is diagnosed by detection of one or both C. difficile toxins in stool and by endoscopic observation of pseudo membranes or micro abscesses in patients who have diarrhea and have been given antibiotics. • Plaques and micro abscesses may be localized to one area of the bowel. • The diarrhea may be watery or bloody, and the patient frequently has associated abdominal cramps, leukocytosis, and fever.
12/19/2025 100 Cont.. • Although many antibiotics have been associated with pseudomembranous colitis the most common are ampicillin and clindamycin and more recently the fluoroquinolones. • The disease is treated by discontinuing administration of the offending antibiotic and orally giving metronidazole, vancomycin, or fidaxomicin. • Fecal transplantation has become a successful and routine method for recurrent and refractory disease. • This usually involves administration of the feces of a healthy related donor by way of colonoscopy or less commonly via a nasogastric tube into the gastrointestinal tract of the patient.
12/19/2025 101 Cont.. • Administration of antibiotics results in proliferation of drug-resistant C. difficile that produces two toxins. • Toxin A, a potent enterotoxin that also has some cytotoxic activity, binds to the brush border membranes of the gut at receptor sites. • Toxin B is a potent cytotoxin C. difficile toxins have glycosyltransferase activity and act by modifying signaling molecules that control various cellular functions.
12/19/2025 102 Cont.. • This results in apoptosis, capillary leakage, cytokine stimulation, and other consequences that lead to colitis. • Both toxins are usually found in the stools of patients with pseudomembranous colitis. However toxin A- negative, toxin B- positive infections have been described. • Not all strains of C. difficile produce the toxins, and the toxin genes are found on a large chromosomal pathogenicity island along with three other genes that regulate toxin expression
12/19/2025 103 Cont.. • Diagnosis is made clinically and supported by demonstration of toxin in the stool by a variety of methods that includes anaerobic toxigenic culture, enzyme immunoassay, and molecular tests that detect the genes that encode toxins A or B. • The responsible host factors include the aging population, the increase in survival of immunocompromised susceptible individuals, and the increase in administration of antibiotics and gastric acid suppressant agents. • Organism factors relate primarily to emergence of certain strain types that are more virulent due to mutations in the pathogenicity locus.
12/19/2025 104 Cont.. • Antibiotic-Associated Diarrhea: The administration of antibiotics frequently leads to a mild to moderate form of diarrhea, termed antibiotic-associated diarrhea. • This disease is generally less severe than the classic form of pseudomembranous colitis. • As many as 25% of cases of antibiotic-associated diarrhea are caused by C. difficile infection.
12/19/2025 105 References: • Sharma, Anil K., Girish Kumar Gupta, and Mukesh Yadav. 2022. Medical Microbiology. • Pitt, Sarah J. 2017. Clinical Microbiology for Diagnostic Laboratory Scientists. • Geo F.B, Janet S.B., and Stephen A.M. (2010). Jawetz, Melnick and Adelberg’s Review of Medical Microbiology. Publ. Lange Medical Publications, New York.
12/19/2025 106 • The End!

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Bacilli anthracis and Clostridia spp.pptx

  • 1.
    12/19/2025 1 Spore-Forming Gram-Positive Bacilli:Bacillus and Clostridium Species Presenter : Dr. Evelyn Godfrey MMED 2 M& I University of Dodoma
  • 2.
    12/19/2025 2 Table ofcontents: 1. Introduction 2. Bacillus anthracis 3. Bacillus cereus 4. Clostridium botulinum 5. Clostridium tetani 6. Clostridia That Produce Invasive Infections 7. Clostridium difficile And Diarrheal Disease
  • 3.
    12/19/2025 3 Introduction • TheGram-positive spore-forming bacilli are the Bacillus and Clostridium species. • These bacilli are ubiquitous and because they form spores, they can survive in the environment for many years. • Bacillus species are aerobes and the Clostridium species are anaerobes . • Species of Bacillus and related genera most do not cause disease and are not well characterized in medical microbiology.
  • 4.
    12/19/2025 4 Cont.. • Anthrax,a classical disease in the history of microbiology is caused by Bacillus anthracis. • Anthrax remains an important disease of animals and occasionally of humans B. anthracis is a major potential agent of bioterrorism and biologic warfare. • B. cereus causes food poisoning and occasionally eye or other localized infections.
  • 5.
    12/19/2025 5 Cont.. • Thegenus Clostridium is extremely heterogeneous and more than 200 species have been described. • Clostridia cause several important toxin-mediated diseases, including tetanus (Clostridium tetani), botulism (C. botulinum), gas gangrene (C. perfringens), antibiotic-associated diarrhea and pseudomembranous colitis (C. difficile).
  • 6.
    12/19/2025 6 Bacillus spp •The genus Bacillus includes large aerobic, Gram-positive rods occurring in chains. • The members of the genus Bacillus are closely related but differ both phenotypically and in terms of pathogenesis. Pathogenic species possess virulence plasmids. • Most members of the genus Bacillus are saprophytic organisms prevalent in soil, water, and air, and on vegetation (e.g. B. subtilis).
  • 7.
    12/19/2025 7 Cont.. • B.cereus can grow in foods and cause food poisoning by producing either an enterotoxin (diarrhea) or an emetic toxin (vomiting). • B. cereus may occasionally produce disease in immunocompromised humans (eg, meningitis, endocarditis, endophthalmitis, conjunctivitis, or acute gastroenteritis). • B. anthracis, which causes anthrax, is the principal pathogen of the genus.
  • 8.
    12/19/2025 8 Bacillus anthracis •B. anthracis, the causative agent of anthrax, has a worldwide distribution. • Anthrax is caused by inhalation, skin exposure or by gastrointestinal (GI) absorption. • B. anthracis was the first pathogenic bacterium to be seen under microscope. • B. anthracis was the first bacterium shown to be the cause of a disease.
  • 9.
    12/19/2025 9 Cont.. • Itwas from studies on anthrax that Koch established his famous postulates in 1876. In 1877, Robert Koch grew the organism in pure culture, demonstrated its ability to form endospores and produced experimental anthrax by injecting it into animals. • It was the first bacterium to be isolated in pure culture and shown to possess spores. • For the first time, spores were discovered from B. anthracis and Bacillus subtilis by Koch and Cohn hence they established the germ theory of disease. • B. antracis was the first bacterium used for the preparation of attenuated vaccine by Louis Pasteur.
  • 10.
    12/19/2025 10 Morphology • B.anthracis is a Gram-positive spore-forming bacillus. • It is a very large bacillus measuring 1–1.2 m in width and 3–5 m in length. • In smears from infected tissues, the bacteria are found as single, in pairs, and in short chains, the entire chain being surrounded by a capsule. • In culture, B. anthracis grows as long chains and may appear similar to streptobacilli.
  • 11.
    12/19/2025 11 Cont.. • Thebacilli are arranged end-to-end and the ends of the bacilli are truncated, not rounded or often concave and somewhat swollen. • This gives the chain of bacilli a “bamboo-stick” appearance . • The bacterium forms the capsule only when grown on nutrient agar containing 0.7% sodium bicarbonate in the presence of 5–20% carbon dioxide. • The capsule is polypeptide (polymer of D-glutamic acid) in nature. • B. anthracis is nonmotile and nonacid fast.
  • 12.
    12/19/2025 12 chain ofbacilli a “bamboo-stick” appearance of B. anthracis
  • 13.
    12/19/2025 13 Culture • B.anthracis is an aerobe and facultative anaerobe. The bacteria grow at a temperature range of 12–45°C, optimum temperature being 37°C. • B. anthracis grow on a wide range of media including ordinary nutrient media and several selective media. • Nutrient agar: On nutrient agar after 24 hours of incubation, B. anthracis produces grayish and granular colonies measuring 2–3 mm in diameter. Under low-power microscopy, the edges of the colony appear as long, interlacing chains of bacilli resembling locks of matted hair, which gives them a “medusa head” appearance with an uneven surface and wavy margin.
  • 14.
    12/19/2025 14 A “medusahead” appearance with an uneven surface and wavy margin on nutrient agar.
  • 15.
    12/19/2025 15 Cont.. • Bloodagar: On horse or sheep blood agar, B. anthracis colonies are gray or white, typically nonhemolytic, with a dry, ground-glass appearance. The colonies are at least 3 mm in diameter and sometimes have tails. • Solid medium containing penicillin: B. anthracis on a solid medium containing 0.05–0.5 U of penicillin/mL produces large, spherical colonies within 3–6 hours and occurs in chains on the surface of the agar, resembling a string of pearls. This property is known as string of pearls reaction and is useful in differentiation of B. anthracis from B. cereus and other aerobic spore bearers.
  • 16.
    12/19/2025 16 Cont.. • Gelatinmedium: In a gelatin stab, there is growth down the stab line with lateral spikes, longer near the surface, giving an ‘inverted fir tree’ appearance. The process of liquefaction is slow and late, which occurs after 7 days at 20°C and starts at the surface.
  • 17.
    12/19/2025 17 Cont.. • Selectivemedium: Knisely’s Polymyxin B-Lysozyme-EDTA Thallous acetate (PLET) agar medium is a selective medium used for isolation of B. anthracis from mixtures containing other spore-bearing bacilli. The medium is composed of heart infusion agar, polymyxin, lysozyme, ethylene diamine tetra acetic acid (EDTA) and thallous acetate.
  • 18.
    12/19/2025 18 Cont.. Production ofcapsular material is associated with the formation of a characteristic mucoid or “smooth” (S) colony type. Capsulated bacteria on serum or bicarbonate medium produce smooth or mucoid colonies. Smooth variants that form capsule are virulent strains of B. anthracis. Rough (R) variants that lack capsule are relatively avirulent.
  • 19.
    12/19/2025 19 Biochemical reactions •B. anthracis produces acid from glucose, maltose, sucrose, trehalose and dextrin but not from lactose, arabinose, D-xylose, or D-mannitol. • They reduce nitrate to nitrite. • They demonstrate a weak lecithinase reaction on egg-yolk agar, which gives a narrow zone of opalescence around the colonies. • They are also catalase positive.
  • 20.
    12/19/2025 20 Pathogenesis • Anthraxis primarily a disease of herbivores goats, sheep, cattle and horses, Humans become infected incidentally by contact with infected animals or their products. • Spores from contaminated soil find easy access when ingested with spiny or irritating vegetation. • In humans, the infection is usually acquired by the entry of spores through injured skin (cutaneous anthrax) or rarely the mucous membranes (gastrointestinal anthrax) or by inhalation of spores into the lung (inhalation anthrax)
  • 21.
    12/19/2025 21 Cont.. • Thespores germinate in the tissue at the site of entry and growth of the vegetative organisms results in formation of a gelatinous edema and congestion. • Bacilli spread via lymphatics to the bloodstream, and they multiply freely in the blood and tissues shortly before and after the animal’s death. • B. anthracis isolates that do not produce a capsule are not virulent and do not induce anthrax in test animals. • The poly-γ-d-glutamic acid capsule is ant phagocytic. The capsule gene is present on a plasmid.
  • 22.
    12/19/2025 22 Cont.. • Anthraxtoxins are made up of three proteins: a) Protective antigen (PA) b) Edema factor (EF) c) Lethal factor (LF). • PA is a protein that binds to specific cell receptors and after proteolytic activation, it forms a membrane channel that mediates entry of EF and LF into the cell. • Edema factor is an adenylate cyclase with protective factor it forms a toxin known as edema toxin. Edema toxin is responsible for cell and tissue edema.
  • 23.
    12/19/2025 23 Cont.. • Lethalfactor plus protective factor form lethal toxin, which is a major virulence factor and cause of death in infected animals and humans. • The lethal toxin can quickly kill the animals by impairing both innate and adaptive immunity allowing organism proliferation and cell death. • The anthrax toxin genes are encoded on another plasmid, pXO1.
  • 24.
  • 25.
    12/19/2025 25 Cont.. • Ininhalation anthrax (wool sorters’ disease), the spores from the dust of wool, hair or hides are inhaled, phagocytosed in the lungs and transported by the lymphatic drainage to the mediastinal lymph nodes, where germination occurs. • This is followed by toxin production and the development of hemorrhagic mediastinitis and sepsis which are usually rapidly fatal. • In anthrax sepsis, the number of organisms in the blood exceeds 107 /mL just before death.
  • 26.
    12/19/2025 26 Pathology • Insusceptible animals and humans, the organisms proliferate at the site of entry. • The capsules remain intact and the organisms are surrounded by a large amount of proteinaceous fluid containing few leukocytes from which they rapidly disseminate and reach the bloodstream. • In resistant animals, the organisms proliferate for a few hours, by which time there is massive accumulation of leukocytes. • The capsules gradually disintegrate and disappear. The organisms remain localized.
  • 27.
    12/19/2025 27 Clinical Findings •In humans, approximately 95% of cases are cutaneous anthrax, and 5% are inhalation. Gastrointestinal anthrax is very rare. • Cutaneous anthrax generally occurs on exposed surfaces of the arms or hands followed in frequency by the face and neck. • A pruritic papule develops 1–7 days after entry of the organisms or spores through a scratch.
  • 28.
    12/19/2025 28 A pruriticpapule of B. anthracis
  • 29.
    12/19/2025 29 Cont.. • Initially,it resembles an insect bite. The papule rapidly changes into a vesicle or small ring of vesicles that coalesce, and a necrotic ulcer develops. • The lesions typically are 1–3 cm in diameter and have a characteristic central black eschar. • Lymphangitis, lymphadenopathy and systemic signs and symptoms of fever, malaise and headache.
  • 30.
    12/19/2025 30 Cont.. • After7–10 days, the eschar is fully developed. Eventually it dries, loosens and separates; healing is by granulation and leaves a scar. • It may take many weeks for the lesion to heal and the edema to subside. • Antibiotic therapy does not appear to change the natural progression of the disease but prevents dissemination.
  • 31.
    12/19/2025 31 Cont.. • Cutaneousanthrax can lead to sepsis, the consequences of systemic infection, including meningitis and death. • The incubation period in inhalation anthrax may be as long as 6 weeks. • The early clinical manifestations are associated with marked hemorrhagic necrosis and edema of the mediastinum.
  • 32.
    12/19/2025 32 Cont.. • Substernalpain may be prominent and there is pronounced mediastinal widening visible on chest radiographs. • Hemorrhagic pleural effusions follow involvement of the pleura; cough is secondary to the effects on the trachea. • Sepsis occurs and there may be hematogenous spread to the gastrointestinal tract, causing bowel ulceration, or to the meninges, causing hemorrhagic meningitis. • The fatality rate in inhalation anthrax is high in the setting of known exposure, it is higher when the diagnosis is not initially suspected.
  • 33.
    12/19/2025 33 Diagnostic LaboratoryTests • Specimens to be examined are fluid or pus from a local lesion, blood, pleural fluid and cerebrospinal fluid in inhalational anthrax associated with sepsis and stool or other intestinal contents in the case of gastrointestinal anthrax. • Stained smears from the local lesion or of blood from dead animals often show chains of large Gram-positive rods. • Anthrax can be identified in dried smears by immunofluorescence staining techniques.
  • 34.
    12/19/2025 34 Cont.. • B.anthracis grown on blood agar plates, the organisms produce nonhemolytic gray to white, tenacious colonies with a rough texture and a ground-glass appearance. Commas shaped outgrowths (Medusa head, “curled hair”) may project from the colony. • Demonstration of capsule requires growth on bicarbonate-containing medium in 5–7% carbon dioxide. Gram-stain shows large Gram- positive rods.
  • 35.
    12/19/2025 35 Cont.. • Insemisolid medium, anthrax bacilli are always nonmotile, but related organisms (eg, B. cereus) exhibit motility by “swarming.” • Clinical laboratories that recover large Gram-positive rods from blood, cerebrospinal fluid, or suspicious skin lesions, which phenotypically match the description of B. anthracis as mentioned, should immediately contact their public health laboratory and send the organism for confirmation. • Definitive identification requires lysis by a specific anthrax γ- bacteriophage, detection of the capsule by fluorescent antibody or identification of toxin genes by polymerase chain reaction (PCR).
  • 36.
    12/19/2025 36 Resistance andImmunity • Immunization to prevent anthrax is based on the classic experiments of Louis Pasteur. • In 1881 he proved that cultures grown in broth at 42–52°C for several months lost much of their virulence and could be injected live into sheep and cattle without causing disease subsequently such animals proved to be immune. • Active immunity to anthrax can be induced in susceptible animals by vaccination with live attenuated bacilli, with spore suspensions or with protective antigen from culture filtrates.
  • 37.
    12/19/2025 37 Treatment • Ciprofloxacinis recommended for treatment, other agents with activity include penicillin G, doxycycline, erythromycin and vancomycin. • Setting of potential exposure to B. anthracis as an agent of biologic warfare, prophylaxis with ciprofloxacin or doxycycline should be given for 60 days and three doses of vaccine (AVA Bio Thrax) should be administered. • AIGIV is a human polyclonal antiserum that also inhibits binding of PA to its receptors. Like Raxibacumab it is used as an adjunct to antimicrobial agents for the treatment of severe forms of anthrax.
  • 38.
    12/19/2025 38 Epidemiology, Preventionand Control • Soil is contaminated with anthrax spores from the carcasses of dead animals. These spores remain viable for decades. Perhaps, spores can germinate in soil at a pH of 6.5 at proper temperature. • Grazing animals infected through injured mucous membranes serve to perpetuate the chain of infection. • Contact with infected animals or with their hides, hair and bristles is the source of infection in humans.
  • 39.
    12/19/2025 39 Cont.. Control measuresinclude: 1) Disposal of animal carcasses by burning or by deep burial in lime pits. 2) Decontamination (usually by autoclaving) of animal products. 3) Protective clothing and gloves for handling potentially infected materials. 4) Active immunization of domestic animals with live attenuated vaccines. 5) Persons with high occupational risk should be immunized.
  • 40.
    12/19/2025 40 Bacillus cereus •Food poisoning caused by B. cereus has two distinct forms. The emetic type which is associated with fried rice, milk and pasta and the diarrheal type which is associated with meat dishes and sauces. • The emetic form is manifested by nausea, vomiting, abdominal cramps and occasionally diarrhea and is self-limiting with recovery occurring within 24 hours. It begins 1–5 hours after ingestion of a plasmid-encoded preformed cyclic peptide (emetic toxin) in the contaminated food products.
  • 41.
    12/19/2025 41 Cont.. • B.cereus is a soil organism that commonly contaminates rice. When large amounts of rice are cooked and allowed to cool slowly, the B. cereus spores germinate and the vegetative cells produce the toxin during log-phase growth or during sporulation. • B. cereus produces toxins that cause disease that is more of intoxication than a food-borne infection.
  • 42.
    12/19/2025 42 Cont.. • Thediarrheal form has an incubation period of 1–24 hours and is manifested by profuse diarrhea with abdominal pain and cramps fever and vomiting are uncommon. • In this syndrome, ingested spores that develop into vegetative cells of B. cereus secrete one of three possible enterotoxins which induce fluid accumulation and other physiological responses in the small intestine.
  • 43.
    12/19/2025 43 Cont.. • Thepresence of B. cereus in a patient’s stool is not sufficient to make a diagnosis of B. cereus disease because the bacteria may be present in normal stool specimens; a concentration of 105 bacteria or more per gram of food is considered diagnostic. • B. cereus is an important cause of eye infections such as severe keratitis and endophthalmitis. • Typically the organisms are introduced into the eye by foreign bodies associated with trauma but infections can also occur after surgery.
  • 44.
    12/19/2025 44 Cont.. • B.cereus has also been associated with localized infections such as wound infections and with systemic infection including endocarditis, catheter-associated bacteremia, central nervous system infections, osteomyelitis and pneumonia; the presence of a medical device or intravenous drug use predisposes to these infections. • Outbreaks of bacteremia in neonatal intensive care units and other hospital units during construction in health care facilities have been reported.
  • 45.
    12/19/2025 45 Cont.. • B.cereus is resistant to a variety of antimicrobial agents, including penicillins and cephalosporins. • Serious nonfoodborne infections should be treated with vancomycin or clindamycin with or without an aminoglycoside. • Ciprofloxacin has been useful for the treatment of wound infections.
  • 46.
    12/19/2025 46 Clostridium spp •The clostridia are large anaerobic, Gram-positive, motile rods. Many decompose proteins or form toxins, and some do both. • Their natural habitat is the soil, marine sediments, sewage or the intestinal tract of animals and humans where they live as saprophytes. • The clostridia continue to increase in number as new species are discovered and several species have been sequenced. • There are 19 clusters based on 16SrRNA gene sequence analysis. Most clinically related species are in RNA Cluster I. Among the pathogens in this cluster are the organisms causing botulism, tetanus, gas gangrene and pseudomembranous colitis.
  • 47.
  • 48.
    12/19/2025 48 Morphology • Mostspecies are anaerobic but few species such as Clostridium tertium, Clostridium histolyticum, etc. are aero tolerant and hence can grow on agar even in presence of air. • Clostridia are rod-shaped bacilli measuring 3–8 0.4–1.2 m in size. • The bacilli are highly pleomorphic and show long filaments and involution forms.
  • 49.
    12/19/2025 49 Cont.. • Theyare Gram-positive but old cultures may appear Gram variable and even Gram-negative. • Most Clostridium species with few exceptions (e.g., Clostridium perfringens, Clostridium tetani type VI) are motile due to the presence of peritrichous flagella. • Most clostridia except C. perfringens and Clostridium butyricum are non capsulated • Clostridia has ability to produce endospores.
  • 50.
    12/19/2025 50 Culture • Mostspecies are anaerobic but few species, such as C. tertium, C. histolyticum, are aero tolerant and hence can grow on agar even in presence of air. • Presence of adequate quantity of low redox potential (Eh) substances in the media is important for the growth of these anaerobic bacteria. • Use of reducing substances, such as ascorbic acid, glutathione, unsaturated fatty acids, cysteine, thioglycolic acid, alkaline glucose or metallic iron maintain low redox potentials in the medium.
  • 51.
    12/19/2025 51 Cont.. • Mostclostridia grow at an optimum temperature of 37°C and pH of 7– 7.4. Growth on solid media is variable. Some clostridial species produce hemolysis on blood agar. • Robertson’s cooked meat (RCM) broth is a useful medium for the growth of clostridia. • The medium contains unsaturated fatty acids, digested meat, and sulfhydryl compounds. Clostridia grow well in the medium, rendering the broth turbid. • Saccharolytic clostridia turn the meat pink while proteolytic species turn the meat black and produce foul smell. Most clostridia produce gas.
  • 52.
    12/19/2025 52 Clostridium botulinum •C. botulinum which causes the disease botulism, is worldwide in distribution and it is found in soil and occasionally in animal feces. • Types of C. botulinum are distinguished by the antigenic type of toxin they produce. • Spores of the organism are highly resistant to heat, withstanding 100°C for several hours. • Heat resistance is diminished at acid pH or high salt concentration.
  • 53.
    12/19/2025 53 Toxins • Duringthe growth of C. botulinum and during autolysis of the bacteria, toxin is liberated into the environment. • Seven antigenic varieties of toxin (serotypes A–G) are known. • Types A, B, E, and F are the principal causes of human illness. Types A and B have been associated with a variety of foods and type E predominantly with fish products. • Botulinum toxins have three domains. Two of the domains facilitate binding to and entry of toxin into the nerve cell.
  • 54.
    12/19/2025 54 Cont.. • Thethird domain is the toxin which is a 150 kDa protein that is cleaved into a heavy chain (H, 100 kDa) and a light chain (L, 50 kDa) that are linked by a disulfide bond. • Botulinum toxin is absorbed from the gut, enters the blood circulation and binds to receptors of presynaptic membranes of motor neurons of the peripheral nervous system and cranial nerves. • The toxin does not cross the blood brain barrier or affect the central nervous system.
  • 55.
    12/19/2025 55 Cont.. • Proteolysisby the L chain of botulinum toxin: of the target SNARE proteins (soluble-N-ethyl maleimide-sensitive factor attachment protein) in the neurons inhibits the release of acetylcholine at the synapse, resulting in lack of muscle contraction and paralysis.
  • 56.
    12/19/2025 56 Cont.. • TheSNARE proteins are synaptobrevin (also known as vesicle- associated membrane protein or VAMP) SNAP 25 and syntaxin. • The toxins of C. botulinum types A, C, and E cleave the 25,000 kDa SNAP 25. • Type C also cleaves syntaxin. Types B, D, F, and G toxins cleave only synaptobrevin.
  • 57.
    12/19/2025 57 Cont.. • C.botulinum toxins are among the most toxic substances known: The lethal dose for a human is probably about 1–2 µg/kg. • The toxins are destroyed by heating for 20 minutes at 100°C. • Strains that produce toxins A, B, or F are associated with infant botulism.
  • 58.
  • 59.
    12/19/2025 59 Pathogenesis • Themost common offenders are spiced, smoked, vacuum packed or canned alkaline foods that are eaten without cooking. In such foods, spores of C. botulinum germinate under anaerobic conditions, vegetative forms grow and produce toxin. • The pathogenesis differs from the way that adults acquire infection. • The infant ingests the spores of C. botulinum and the spores germinate within the intestinal tract. The vegetative cells produce toxin as they multiply; the neurotoxin then gets absorbed into the bloodstream.
  • 60.
    12/19/2025 60 Cont.. • Inrare instances adults with gastrointestinal anatomical abnormalities or functional disorders may develop “infant botulism.” • Wound botulism is the result of tissue contamination with spores and is seen primarily in injection drug users. • Very rarely, inhalational botulism occurs when toxin enters the respiratory tract. The toxin acts by blocking release of acetylcholine at synapses and neuromuscular junctions . • The result is flaccid paralysis. The electromyogram and edrophonium strength test results are typical.
  • 61.
  • 62.
  • 63.
    12/19/2025 63 Clinical Findings. •Symptoms begin 18–24 hours after ingestion of the toxic food, with visual disturbances (incoordination of eye muscles, double vision), inability to swallow and speech difficulty, signs of bulbar paralysis are progressive and death occurs from respiratory paralysis or cardiac arrest. • Gastrointestinal symptoms are not prominent. There is no fever. The patient remains fully conscious until shortly before death. The mortality rate is high. Patients who recover do not develop antitoxin in the blood.
  • 64.
    12/19/2025 64 Cont.. • Theinfants in the first months of life develop poor feeding, weakness, and signs of paralysis (floppy baby). • Infant botulism may be one of the causes of sudden infant death syndrome. • C. botulinum and botulinum toxin are found in feces but not in serum.
  • 65.
    12/19/2025 65 Diagnostic LaboratoryTests • Clinicians who suspect a case of botulism should contact the appropriate public health authorities before submitting specimens to the laboratory. • Detection of toxin and not the organism is required for definitive diagnosis. Toxin can often be demonstrated in serum, gastric secretions or stool from the patient and toxin may be found in leftover food.
  • 66.
    12/19/2025 66 Cont.. • Clinicalswabs or other specimens obtained from patients should be transported using anaerobe containers. • Suspect foods should be left in their original containers. This mouse bioassay is the test of choice for the confirmation of botulism. • C. botulinum may be grown from food remains and tested for toxin production, but this is rarely done and is of questionable significance. • In infant botulism, C. botulinum and toxin can be demonstrated in bowel contents but not in serum. Other methods used to detect toxin include ELISAs and PCR, but the latter may detect organisms that carry the gene but do not express toxin.
  • 67.
    12/19/2025 67 Treatment • Supportivecare, especially intensive care, is key in the management of patients with botulism. • Adequate respiration must be maintained by mechanical ventilation if necessary and in severe cases may need to be maintained for up to 8 week. • Antitoxin does not reverse the paralysis but if administered early it can prevent its advancement. Although most infants with botulism recover with supportive care alone treatment with human derived botulinum immune globulin (BIG) is recommended
  • 68.
    12/19/2025 68 Epidemiology andPrevention • Spores of C. botulinum are widely distributed in soil, they often contaminate vegetables, fruits and other materials. • A chief risk factor for botulism lies in home-canned foods, particularly string beans, corn, peppers, olives, peas and smoked fish or vacuum- packed fresh fish in plastic bags. • The risk from home-canned foods can be reduced if the food is boiled for more than 20 minutes before consumption. • Botulinum toxin is considered to be a major potential agent for bioterrorism and biologic warfare.
  • 69.
    12/19/2025 69 Clostridium tetani •C. tetani, which causes tetanus, is worldwide in distribution in the soil and in the feces of horses and other animals. • Several types of C. tetani can be distinguished by specific flagellar antigens. All share a common O (somatic) antigen, which may be masked and all produce the same antigenic type of neurotoxin, tetanospasmin.
  • 70.
    12/19/2025 70 Toxin • Thevegetative cells of C. tetani produce the plasmid-encoded toxin tetanospasmin (150 kDa) that is cleaved by a bacterial protease into two peptides (50 and 100 kDa) linked by a disulfide bond. • The larger peptide initially binds to receptors on the presynaptic membranes of motor neurons. It then migrates by the retrograde axonal transport system to the cell bodies of these neurons to the spinal cord and brainstem. • The toxin diffuses to terminals of inhibitory cells, including both glycinergic interneurons and γ-aminobutyric acid (GABA)– secreting neurons from the brainstem.
  • 71.
    12/19/2025 71 Cont.. • Thesmaller peptide degrades synaptobrevin a protein required for docking of neurotransmitter vesicles on the presynaptic membrane. Release of the inhibitory glycine and GABA is blocked, and the motor neurons are not inhibited. • Hyperreflexia, muscle spasms, and spastic paralysis result. • Extremely small amounts of toxin can be lethal for humans.
  • 72.
  • 73.
    12/19/2025 73 Pathogenesis • C.tetani is not an invasive organism. The infection remains strictly localized in the area of devitalized tissue (wound, burn, injury, umbilical stump, surgical suture) into which the spores have been introduced. • The volume of infected tissue is small, and the disease is almost entirely a toxemia.
  • 74.
    12/19/2025 74 Cont.. • Germinationof the spore and development of vegetative organisms that produce toxin are aided by necrotic tissue, calcium salts and associated pyogenic infections all of which aid establishment of low oxidation reduction potential. • The toxin released from vegetative cells reaches the central nervous system and rapidly becomes fixed to receptors in the spinal cord and brainstem and exerts the actions.
  • 75.
    12/19/2025 75 Clinical Findings •The incubation period may range from 4 to 5 days up to 3 weeks. • The disease is characterized by tonic contraction of voluntary muscles. Muscular spasms often involve first the area of injury and infection and then the muscles of the jaw (trismus, lockjaw), which contract so that the mouth cannot be opened. • Gradually, other voluntary muscles become involved, resulting in tonic spasms.
  • 76.
    12/19/2025 76 Cont.. • Anyexternal stimulus may precipitate a tetanic generalized muscle spasm. • The patient is fully conscious and pain may be intense. • Death usually results from interference with the mechanics of respiration. • The mortality rate in generalized tetanus is very high.
  • 77.
  • 78.
    12/19/2025 78 Diagnosis • Thediagnosis rests on the clinical picture and a history of injury although only 50% of patients with tetanus have an injury for which they seek medical attention. • The primary differential diagnosis of tetanus is strychnine poisoning. • Anaerobic culture of tissues from contaminated wounds may yield C. tetani, but neither preventive nor therapeutic use of antitoxin should ever be withheld pending such demonstration. • Proof of isolation of C. tetani must rest on production of toxin and its neutralization by specific antitoxin.
  • 79.
    12/19/2025 79 Prevention Treatment •Prevention of tetanus depends on active immunization with toxoids, aggressive wound care, prophylactic use of antitoxin and administration of penicillin. • The intramuscular administration of 250–500 units of human antitoxin (tetanus immune globulin) gives adequate systemic protection (0.01 unit or more per milliliter of serum) for 2–4 weeks , It neutralizes the toxin that has not been fixed to nervous tissue. • Active immunization with tetanus toxoid should accompany antitoxin prophylaxis.
  • 80.
    12/19/2025 80 Cont.. • Patientswho develop symptoms of tetanus should receive muscle relaxants, sedation, and assisted ventilation. • Surgical debridement is vitally important because it removes the necrotic tissue that is essential for proliferation of the organisms. • Penicillin strongly inhibits the growth of C. tetani and stops further toxin production.
  • 81.
    12/19/2025 81 Cont.. • Whena previously immunized individual sustains a potentially dangerous wound, an additional dose of toxoid should be injected to restimulate antitoxin production. • This “recall” injection of toxoid may be accompanied by a dose of antitoxin if the patient has not had current immunization or boosters or if the history of immunization is unknown. • Initial immunization should be carried out in all children during the first year of life.
  • 82.
    12/19/2025 82 Clostridia ThatProduce Invasive Infections. • Many different toxin-producing clostridia (C. perfringens and related clostridia) can produce invasive infection (including myonecrosis and gas gangrene) if introduced into damaged tissue. • An enterotoxin of C. perfringens is a common cause of food poisoning.
  • 83.
    12/19/2025 83 Toxins • Theinvasive clostridia produce a large variety of toxins and enzymes that result in a spreading infection. • Many of these toxins have lethal, necrotizing and hemolytic properties. In some cases, these are different properties of a single substance in other instances, they are attributable to different chemical entities. • The alpha toxin of C. perfringens type A is a lecithinase, and its lethal action is proportionate to the rate at which it splits lecithin (an important constituent of cell membranes) to phosphorylcholine and diglyceride.
  • 84.
    12/19/2025 84 Cont.. • Alphatoxin also aggregates platelets, thereby leading to formation of thrombi in small blood vessels and adding to poor tissue profusion and extending the consequences of anaerobiosis, namely, destruction of viable tissue (gas gangrene). • The theta toxin has similar hemolytic and necrotizing effects but is not a lecithinase. It is a member of the cholesterol-dependent cytolysins that act by forming pores in cell membranes.
  • 85.
    12/19/2025 85 Cont.. • Epsilontoxin is a protein that causes edema and hemorrhage is very potent. • DNase and hyaluronidase, a collagenase that digests collagen of subcutaneous tissue and muscle, are also produced. Some strains of C. perfringens produce a powerful enterotoxin (C. perfringens enterotoxin, CPE), especially when grown in meat dishes. When more than 108 vegetative cells are ingested and sporulate in the gut, CPE is formed. • CPE is a protein (35 kDa) that may be a nonessential component of the spore coat; it is distinct from other clostridial toxins. It induces intense diarrhea in 7–30 hours.
  • 86.
    12/19/2025 86 Cont.. • Theaction of C. perfringens enterotoxin involves marked hypersecretion in the jejunum and ileum, with loss of fluids and electrolytes in diarrhea eg nausea, vomiting, and fever. • This illness is similar to that produced by B. cereus and tends to be self-limited. • Enterotoxin-producing strains of C. perfringens may also play a role in antibiotic associated diarrhea and necrotizing enterocolitis in infants.
  • 87.
  • 88.
    12/19/2025 88 Pathogenesis • Ininvasive clostridial infections, spores reach tissue either by contamination of traumatized areas (soil, feces) or from the intestinal tract. • The spores germinate at low oxidation - reduction potential; vegetative cells multiply, ferment carbohydrates present in tissue, and produce gas. • The distention of tissue and interference with blood supply, together with the secretion of necrotizing toxins and hyaluronidase, favor the spread of infection.
  • 89.
    12/19/2025 89 Cont.. • Tissuenecrosis extends providing an opportunity for increased bacterial growth, hemolytic anemia and ultimately, severe toxemia and death. • In gas gangrene (clostridial myonecrosis) a mixed infection is the rule. In addition to the toxigenic clostridia, proteolytic clostridia and various cocci and Gram-negative organisms are also usually present. • C. perfringens occurs in the genital tracts of 5% of women. Before legalization of abortion in the United States, clostridial uterine infections followed instrumented abortions.
  • 90.
    12/19/2025 90 Cont.. • C.sordellii has many of the properties of C. perfringens. • C. sordellii has been reported to cause a toxic shock syndrome after medical abortion with mifepristone and intravaginal misoprostol. • Endometrial infection with C. sordellii is implicated. Clostridial bacteremia especially that caused by C. septicum, is a frequent occurrence in patients with neoplasms. • In New Guinea, C. perfringens type C produces necrotizing enteritis (pigbel) that can be highly fatal in children. • Immunization with type C toxoid appears to have preventive value.
  • 91.
    12/19/2025 91 Clinical Findingsand Treatment • From a contaminated wound (e.g., a compound fracture, postpartum uterus) the infection spreads in 1–3 days to produce crepitation in the subcutaneous tissue and muscle, foul-smelling discharge, rapidly progressing necrosis, fever, hemolysis, toxemia, shock and death. • Treatment is with early surgery (amputation) and antibiotic administration. Advent of specific therapy, early amputation was the only treatment
  • 92.
    12/19/2025 92 Cont.. • Attimes the infection results only in anaerobic fasciitis or cellulitis. • C. perfringens food poisoning usually follows the ingestion of large numbers of clostridia that have grown in warmed meat dishes. • The toxin forms when the organisms sporulate in the gut, with the onset of diarrhea; usually without vomiting or fever—in 7–30 hours. The illness lasts only 1–2 days.
  • 93.
    12/19/2025 93 Diagnostic LaboratoryTests • Specimens consist of material from wounds, pus, and tissue. • The presence of large Gram-positive rods in Gram-stained smears suggests gas gangrene clostridia; spores are not regularly present. • Material is inoculated into chopped meat–glucose medium and thioglycolate medium and onto blood agar plates incubated anaerobically.
  • 94.
    12/19/2025 94 Cont.. • Afterpure cultures have been obtained by selecting colonies from anaerobically incubated blood plates, they are identified by biochemical reactions (various sugars in thioglycolate, action on milk), hemolysis and colony morphology.
  • 95.
    12/19/2025 95 Cont.. • Lecithinaseactivity is evaluated by the precipitate formed around colonies on egg yolk media. • Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a rapid and sensitive method for identification of invasive Clostridium species recovered in culture. • C. perfringens rarely produces spores when cultured on agar in the laboratory.
  • 96.
    12/19/2025 96 Treatment • Extensivesurgical debridement of the involved area and excision of all devitalized tissue, in which the organisms are prone to grow. • Administration of antimicrobial drugs, particularly penicillin, is begun at the same time. • Hyperbaric oxygen may be of help in the medical management of clostridial tissue infections. It is said to “detoxify” patients rapidly.
  • 97.
    12/19/2025 97 Cont.. • Antitoxinsare available against the toxins of C. perfringens, C. novyi, C. histolyticum, and C. septicum, usually in the form of concentrated immune globulins. • Polyvalent antitoxin (containing antibodies to several toxins) has been used.
  • 98.
    12/19/2025 98 Prevention andControl • Early and adequate cleansing of contaminated wounds and surgical debridement, together with the administration of antimicrobial drugs directed against clostridia (eg, penicillin), are the best available preventive measures.
  • 99.
    12/19/2025 99 Clostridium difficileAnd Diarrheal Disease • Pseudomembranous Colitis is diagnosed by detection of one or both C. difficile toxins in stool and by endoscopic observation of pseudo membranes or micro abscesses in patients who have diarrhea and have been given antibiotics. • Plaques and micro abscesses may be localized to one area of the bowel. • The diarrhea may be watery or bloody, and the patient frequently has associated abdominal cramps, leukocytosis, and fever.
  • 100.
    12/19/2025 100 Cont.. • Althoughmany antibiotics have been associated with pseudomembranous colitis the most common are ampicillin and clindamycin and more recently the fluoroquinolones. • The disease is treated by discontinuing administration of the offending antibiotic and orally giving metronidazole, vancomycin, or fidaxomicin. • Fecal transplantation has become a successful and routine method for recurrent and refractory disease. • This usually involves administration of the feces of a healthy related donor by way of colonoscopy or less commonly via a nasogastric tube into the gastrointestinal tract of the patient.
  • 101.
    12/19/2025 101 Cont.. • Administrationof antibiotics results in proliferation of drug-resistant C. difficile that produces two toxins. • Toxin A, a potent enterotoxin that also has some cytotoxic activity, binds to the brush border membranes of the gut at receptor sites. • Toxin B is a potent cytotoxin C. difficile toxins have glycosyltransferase activity and act by modifying signaling molecules that control various cellular functions.
  • 102.
    12/19/2025 102 Cont.. • Thisresults in apoptosis, capillary leakage, cytokine stimulation, and other consequences that lead to colitis. • Both toxins are usually found in the stools of patients with pseudomembranous colitis. However toxin A- negative, toxin B- positive infections have been described. • Not all strains of C. difficile produce the toxins, and the toxin genes are found on a large chromosomal pathogenicity island along with three other genes that regulate toxin expression
  • 103.
    12/19/2025 103 Cont.. • Diagnosisis made clinically and supported by demonstration of toxin in the stool by a variety of methods that includes anaerobic toxigenic culture, enzyme immunoassay, and molecular tests that detect the genes that encode toxins A or B. • The responsible host factors include the aging population, the increase in survival of immunocompromised susceptible individuals, and the increase in administration of antibiotics and gastric acid suppressant agents. • Organism factors relate primarily to emergence of certain strain types that are more virulent due to mutations in the pathogenicity locus.
  • 104.
    12/19/2025 104 Cont.. • Antibiotic-AssociatedDiarrhea: The administration of antibiotics frequently leads to a mild to moderate form of diarrhea, termed antibiotic-associated diarrhea. • This disease is generally less severe than the classic form of pseudomembranous colitis. • As many as 25% of cases of antibiotic-associated diarrhea are caused by C. difficile infection.
  • 105.
    12/19/2025 105 References: • Sharma,Anil K., Girish Kumar Gupta, and Mukesh Yadav. 2022. Medical Microbiology. • Pitt, Sarah J. 2017. Clinical Microbiology for Diagnostic Laboratory Scientists. • Geo F.B, Janet S.B., and Stephen A.M. (2010). Jawetz, Melnick and Adelberg’s Review of Medical Microbiology. Publ. Lange Medical Publications, New York.
  • 106.