Spore-Forming Gram-positive Bacilli

Flashcards on Spore-Forming Gram-positive Bacilli: Bacillus & Clostridium species

Bacillus species

Large gram-positive rods; ubiquitous bacilli; aerobes; catalase (+); occurring in chains.

Clostridium species

Large gram-positive rods; obligate anaerobes; catalase (-).

Catalase

Enzyme that catalyzes the decomposition of hydrogen peroxide to water and oxygen.

Spore-forming bacilli

Bacilli that can survive in the environment for many years; have square ends and are arranged in long chains; spores are located in the center of the bacilli.

Clostridium species motility

Large anaerobic, motile rods with peritrichous flagella (except C. perfringens, which lacks flagella).

Bacillus species motility

Large aerobic motile rods with peritrichous flagella (except B. anthracis, which is nonmotile).

Bacillus species general characteristics

Gram-positive rods occurring in chains; saprophytic organisms prevalent in soil, water, air, and vegetation; produce endospores; principal pathogen of the genus.

Anthrax transmission

Anthrax is usually transmitted to humans from animals; acquired through injured skin (cutaneous anthrax), inhalation of spores (inhalation anthrax), ingestion (gastrointestinal anthrax), or injection of contaminated heroin (injection anthrax).

Epidemiology of Anthrax

Herbivores ingest contaminated vegetation or contact with contaminated animal products (hides, wool, hair); humans are accidental hosts through ingestion, inhalation, or contact.

Cutaneous anthrax

95% of human cases; occurs on exposed surfaces of the arms or hands; contact with infected animals or their products.

Clinical Presentation of Cutaneous Anthrax

Small, non-painful pruritic papule at inoculation site that develops into a hemorrhagic vesicle & ruptures, slow-healing painless ulcer with black eschar in the center – malignant pustule - is formed surrounded by edema

Inhalation Anthrax

Spores from the dust of wool, hair, or hides are inhaled; early clinical manifestation includes breathing difficulties and edema of the mediastinum; later, hemorrhagic mediastinitis and sepsis, which are usually rapidly fatal.

Gastrointestinal Anthrax

Very rare in humans; abdominal pain, severe vomiting and diarrhea; hemorrhagic ascites.

Pathogenesis of Anthrax

B. anthracis produces antiphagocytic capsule (pXO2 plasmid encoded); bacilli spread via lymphatics to the bloodstream and multiply freely; toxin is made of protective antigen, edema factor, and lethal factor (pXO1 plasmid encoded).

Multicomponent anthrax toxin mechanism

Protective antigen (PA) binds to cell receptor and forms a channel; Edema Factor (EF, adenylate cyclase) and Lethal Factor (LF, protease) are transferred into the cell.

Capsule of Bacillus anthracis

Composed of poly-D-glutamic acid, serves as one of the principal virulence factors; inhibits phagocytosis of the vegetative cells by macrophages.

Diagnostic laboratory tests for Anthrax

Gram stain, culture, immunofluorescence staining, ELISA; definitive diagnosis includes lysis by specific bacteriophage, detection of the capsule by fluorescent antibody, or identification of toxin genes by PCR.

Culture of Bacillus anthracis

Colonies of B anthracis are round and have a “cut glass” appearance in transmitted light; comma-shaped outgrowths (Medusa head, “curled hair”, “comet’s tail”) may project from the colony.

Lysis of Bacillus anthracis by the lytic phage gamma

Clear area in the region of confluent growth where the γ-phage was applied; results from the phage's ability to lyse the bacterial cells; specific for B. anthracis.

Resistance and immunity to Anthrax

Natural exposure to cutaneous anthrax gives long-lasting T cell immunity; vaccination includes live attenuated bacilli or supernatant of cell-free culture filtrate containing PA; persons at risk are immunized.

Treatment for Anthrax

Ciprofloxacin, penicillin G, doxycycline, erythromycin, and vancomycin are effective; Ciprofloxacin or Doxycycline - for prophylaxis

Prevention and Control of Anthrax

Proper disposal of animal carcasses, decontamination of animal products, protective clothing and gloves, and active immunization of domestic animals.

B. anthracis as bioweapon

Letters containing anthrax spores were mailed to several news media offices and to Senators offices in the US beginning on September 18, 2001; resulted in 22 cases of anthrax—11 inhalation and 11 cutaneous.

Bacillus cereus

Soil organism; can cause food poisoning by producing either an emetic toxin (vomiting) or an enterotoxin (profuse diarrhea with cramps); produces beta-lactamases, and so is resistant to beta-lactam antibiotics

Gastroenteritis Bacillus cereus clinical presentation

Severe vomiting (emetic form) associated with fried rice, milk, pasta or diarrhea (diarrheal form) associated with meat dishes and sauces

Clinically related Clostridium species

C. perfringens (gas gangrene; food poisoning), C. tetani (tetanus), C. botulinum (botulism), C. difficile (pseudomembranous colitis).

Clostridium Species

Anaerobic; large gram-positive rods; spores are usually wider than the rods and located terminally or subterminally; most are motile by peritrichous flagella; most are saprophytes in soil or intestinal tract.

Culture of Clostridium

Grow under anaerobic conditions; grow well on blood-enriched media and media used to culture anaerobes; some produce large raised colonies, others smaller colonies; many produce hemolysis on blood agar.

Growth Characteristics of Clostridium

Ferment variety of sugars; many digest proteins; milk is turned into acid, digested or undergoes “stormy fermentation” – clot torn by gas (C. perfringens).

Clostridium botulinum

Causative agent of botulism; found in soil and occasionally in animal feces; spores are highly heat resistant.

Botulinum Toxin

Produce seven (A-G) antigenically distinct toxins (botulinum toxins; Botox); liberated during the growth and during autolysis of the bacteria; A, B, E and F are the principal causes of human illness; destroyed by heating at 100oC for 20 mins

C. botulinum Pathogenicity

An intoxication resulting from the ingestion of food in which C. botulinum has produced toxin; prevents the release of the neurotransmitter acetylcholine from axon endings at the neuromuscular junction → causes flaccid paralysis.

Foodborne botulism

Double vision, inability to swallow, speech difficulty, bulbar paralysis, constipation, and abdominal pain; no fever; bilateral descending weakness of peripheral muscle; death occurs from respiratory paralysis or cardiac arrest.

C.botulinum – clinical findings

Infant botulism- weakness, signs of paralysis, C. botulinum and its toxin are found in feces; wound botulism – symptoms similar to those of foodborne botulism with longer incubation time, less GI symptoms.

Food-borne botulism

Toxin produced-->Toxin ingested; infant botulism - C. botulinum spores ingested-->C. botulinum grows in gastrointestinal tract-->Toxin produced; Wound botulism - C. botulinum spores in wound-->C. botulinum grows in wound-->Toxin produced-->Toxin in bloodstream

Botulism

Food-borne intoxication (most common) spiced, smoked, vacuum packed, or canned alkaline foods that are eaten without cooking; Infant botulism (common in the US) honey is the most frequent vehicle of infection; Wound botulism tissue contamination with spores associated with skin-popping using contaminated “black tar” heroin; Inhalation botulism (very rarely) when pure form of the toxin enters the respiratory tract.

C. botulinum Laboratory Diagnosis

Detection of toxin (not the organism) in feces or serum from the patient and in leftover food; typing of toxin is done by neutralization with specific antitoxin Note: Rare strains of Clostridium butyricum and Clostridium baratii produce botulinum neurotoxin and cause botulism in humans .

C.botulinum ▪ Treatment

Stomach lavage and high enemas; Trivalent (A, B, E) antitoxin or heptavalent antitoxin administered intravenously promptly; Adequate ventilation by mechanical respirator.Main danger lies in home-canned foods; The risk from home-canned food can be reduced by boiling the food for 20 min.

Clostridium tetani Physiology and Structure

Small, motile; spore-forming, “tennis racket” appearance; extremely sensitive to oxygen toxicity; all produce the same antigenic type of neurotoxin – tetanospasmin – plasmid-encoded.

Toxin

Inhibits release of an inhibitory mediator (e.g., gamma-aminobutyric acid or glycine) which acts on postsynaptic spinal neurons; motor neurons are not inhibited – spastic paralysis results.

Botulism and tetanus are induced by neurotoxin - producing bacteria

Botulinum toxin prevents nerve transmission (flaccid paralysis); Tetanus toxin prevents inhibitory nerve transmission (spastic paralysis)

Pathogenesis of C. tetani

Disease is almost entirely toxemia; germination of the spore and development of vegetative organisms that produce toxin - aided by establishment of low oxidation-reduction potential by (1) necrotic tissue (2) calcium salts (3) associated pyogenic infections; The toxin released from vegetative cells reaches the CNS and rapidly becomes fixed to receptors in the spinal cord and brainstem.

C.tetani Clinical Diseases

Convulsive tonic contraction of voluntary muscles; first the area of injury then the muscles of the jaw (trismus, lockjaw) mouth cannot be opened gradually, other voluntary muscles become involved

C. tetani

Trismus (lockjaw), Sardonic Smile (risus sardonicus), Opisthotonos (backward arching)

C.tetani Laboratory Diagnosis

Diagnosis depends on the clinical picture and a history of injury

Clostridium tetani Control

Universal active immunization with tetanus toxoid should be mandatory; tetanus toxoid produced by detoxifying the toxin with formalin; three injections comprise the initial course of immunization.

CLOSTRIDIA THAT PRODUCE INVASIVE INFECTIONS

C perfringens - the most common (90%)

Clostridia that produce invasive infections

Large gram-positive bacilli; spores are rarely observed; non-motile; capsulated; hemolytic; metabolically active; subdivided into 5 types: A, B, C, D, E based on the four major lethal toxins they produce. Type A causes most of the human infections.

C.perfringens Pathogenicity and Immunity

Spectrum of diseases primarily by producing toxins and enzymes: a-toxin: lecithinase (phospholipase C) lyses a variety of cells endotheliel cells, leukocytes, platelets, erythrocytes, Other types produce theta toxin – perfringolysin O & epsilon toxin – causes edema and hemorrhage

C.perfringens

Strains of C. perfringens are widely distributed in nature and inhabit the intestine of humans and animals, the genital tracts of 5% of women. Portal of entry - Trauma Due to toxins and enzymes tissue is necrotized Fermentation of tissue carbohydrates, lipids, and amino acids yields gas, and an accumulation of gas bubbles in the subcutaneous spaces. Hemolytic anemia, Toxemia, Death

Laboratory Diagnosis for C. perfringens

PUS, necrotic tissue, Smears: large gram-positive rods with or without spores Culture: anaerobic culture on blood plate Identification: • “Storming fermentation”• Lecithinase test• Hemolysis on blood agar “double zone”• Biochemical tests • MALDI-TOF MS

Clostridium perfringens food poisoning

C perfringens is one of the most common causes of food poisoning in the United States Onset: 7-30 hours after ingestion• Typical symptoms: abdominal pain and diarrhea Prevention: reheated to at least 74 oC before serving

CLOSTRIDIA THAT PRODUCE INVASIVE INFECTIONS

C. perfringens (endometrial infections ); C. sordellii (cause a toxic shock syndrome after medical abortion); Clostridial bacteremia (especially by C. septicum - a frequent occurrence in patients with neoplasms; In New Guinea, C. perfringens type C produces necrotizing enteritis food poisoning

C.perfringens Laboratory Diagnosis

Storming fermentation, double zone of hemolysis Lecithinase test opaque halo with egg yolk

C.perfringens Treatment

prompt and extensive surgical debridement, Antibiotics (penicillin) administration, Hyperbaric oxygen may detoxify” patients rapidly; Prevention and Control: Preventive measures: surgical debridement and prophylactic antibiotics.

Clostridium difficile and diarrheal diseases

Clostridium difficile is a bacterium that can be found in the colon 25% of all cases of Antibiotic - associated diarrhea Complications: pseudomembranous colitis, toxic megacolon, sepsis C. difficile produces two toxins Frequently implicated antibiotics metronidazole, vancomycin, or fidaxomicin Fecal transplantation for recurrent and refractory disease