Spore Forming Gram-positive Bacilli

Titik Nuryastuti Microbiology Department, Fac. of Medicine, UGM

Spores
Why do bacteria produce spores? Survival Classification Definition = a resting cell, highly resistant to dessication, heat, and chemical agents; when returned to favourable conditions bacteria reactivated, the spores germinate to produce single vegetative cells.

SF Bacteria- Bacillus

Aerobic, G+ rods in chains, spores are located in center of the non-motile bacilli Found in soil, water, air and vegetation Spores are viable for decades. B. cereus produce enterotoxin and cause food poisoning. B. anthracis infection in human through injured skin (cutaneous anthrax), mucous membranes (GI anthrax), or inhalation of spores into lung.

Bacillus anthracis

SF Bacteria- Bacillus

Spores germinate in the tissue of entry, and growth of vegetative organisms result in formation of a gelatinous oedema and congestion. Spread via lymphatics to bloodstream and multiply freely in blood and tissues. Capsulated, poly-D-glutamic acid capsule is antiphagocytic

SF Bacteria- Bacillus

Anthrax toxin is made up of three proteins: Protective antigen (PA), edema factor (EF) and lethal factor (LF). Clinical finding : Cutaneous Anthrax(malignant pustule): Generally occurs on exposed surfaces of the arms, face and neck through wound contamination by the spores of the organism. About 95% of the cases with amortality rate 20% . Inhalation Anthrax(wool sorter disease): About 5% of the cases with 85-90% mortality. Treatment: ciprofloxacin, penicillin G along with gentamicin and streptomycin.

SF Bacteria-Bacillus

Lab diagnosis : Gram staining Culture on Blood agar Speciment :

SF Bacteria - Clostridium

Anaerobic, G+, motile rods Their natural habitat is the soil or the intestinal tract of human and animals, where they live as saprophytes Found in soil, animal faeces. Spores is placed centrally, subterminally or terminally; most species are motile with flagella.

SF Bacteria - Clostridium

Many decompose proteins of form toxins, some do both Among the pathogens are the organisms causing botulism, tetanus, gas gangrene, and pseudomembranous colitis.

C. botulism, C. tetani, C. perfringens, C. difficile

SF Bacteria - Clostridium

Many form colonies with a zone of haemolysis on blood agar. C perfringens typically produce multiple zones of haemolysis around colonies.

Identification

In most species, the spores are located centrally, subterminally or terminally. Most species of Clostridia are motile with peritrichous flagella

Clostridium

Epidemiology Ubiquitous Present in soil, water, sewage Normal flora in GI tracts of animals and humans Pathogenesis Spore formation resistant to heat, dessication, and disinfectants can survive for years in adverse environments Rapid growth in oxygen deprived, nutritionally enriched environment Toxin elaboration (histolytic toxins, enterotoxins, neurotoxins)

Clostridium botulinum

Epidemiology Commonly isolated in soil and water Human disease associated with botulinum toxin A, B, E, F Pathogenesis Blocks neurotransmission at peripheral cholinergic synapses Prevents release of acetylcholine, resulting in muscle relaxation Recovery depends upon regeneration of nerve endings

SF Bacteria C.botulinum

C botulinum causes botulism -Distinguished by antigenic type of toxin Spores are resistant to 100C for many hours, diminished at acid pH or high salt. Toxin - 7 antigenic varieties (A G). A, B, E (F) mainly harmful to human. Botulinum toxin is absorbed from gut and binds to receptors of presynaptic nervous system and cranial nerves. Lethal dose to human 1-2 g.

SF Bacteria - Clostridium
Pathogenesis Most cases, through ingestion of uncooked food. Toxin acts by blocking release of acetylcholine at synapses and neuromuscular junctions flacid paralysis. Symptoms such as visual disturbances, inability to swallow, speech problem; seldom with no apparent GI symptoms; no fever.

Botulism

Clinical Syndromes Foodborne botulism Associated with consumption of preformed toxin Home-canned foods (toxin A, B) Preserved fish (toxin E) Onset of symptoms 1-2 days Blurred vision vision, dilated pupils, dry mouth, constipation Bilateral descending weakness of peripheral muscles; death related to respiratory failure Infant botulism Consumption of foods contaminated with botulinum spores 6-10% of syrups or honeys Disease associated with neurotoxin produced in vivo Onset of symptoms in 3-10 days Wound botulism (skin popping) Asymptomatic adult carriage

Botulism: Treatment

Treatment Supportive care Elimination of organism from GI tract Gastric lavage Metronidazole or penicillin Botulinum Immunoglobulin (BIG): pooled plasma from adults immunized with pentavalent (ABCDE) botulinum toxoid Trivalent equine Immunoglobulin (ABE) Prevention Prevention of spore germination (Storage <4C, high sugar content, acid PH) Destruction of preformed toxin (20 min at 80C)

SF Bacteria - Clostridium

floppy baby = infant botulism. C botulinum spores in babies food. Treatment antitoxins raised in horses. Trivalent (A, B, E) antitoxin must be promptly administered intravenously with precautions; plus adequate ventilations.

Clostridium tetani

Epidemiology Spores found in most soils, GI tracts of animals Disease in un-vaccinated or inadequately immunized Disease does not induce immunity Pathogenesis Spore inoculated into wound Tetanospasmin Heat-labile neurotoxin Retrograde axonal transport to CNS Blocks release of inhibitory neurotransmitters (eg. GABA) into synapses, allowing excitatory synapses to be unregulated. This results in muscle spasms Binding is irreversible Tetanolysin Oxygen labile hemolysin, unclear clinical significance

SF Bacteria C.tetani

Clostridium tetani cause tetanus. Distinguishable by specific flagellar antigens. Pathogenesis: Wound contamination, not an invasive organism. The toxins released from vegetative cells reaches the CNS and rapidly becomes fixed to receptors in the spinal cord and brain stem and exerts their action.

C. tetani

Toxins: Tetanospasmin binds to receptors on the presynaptic membranes of motor neurons. Clinical Findings: Incubation period: 4-5 days to many weeks. The disease is chacterized by tonic contraction of voluntary muscles.

Tetanus

Treatment Debridement of wound Metronidazole Tetanus immunoglobulin Prevention Vaccination with a series of 3 tetanus toxoid Booster dose every 10 years

Clostridium perfringens

Epidemiology GI tract of humans and animals Type A responsible for most human infections, is widely distributed in soil and water contaminated with feces Type B-E do not survive in soil but colonize the intestinal tracts of animals and occasionally humans Pathogenesis -toxin: lecithinase (phospholipase C) that lyses erythrocytes, platelets and endothelial cells resulting in increased vascular permeability and hemolysis -toxin: necrotizing activity Enterotoxin: binds to brush borders and disrupts small intestinal transport resulting in increased membrane permeability Clinical manifestations Self-limited gastroenteritis Soft tissue infections: cellulitis, fascitis or myonecrosis (gas gangrene)

C. perfringens

Many different- toxin producing clostridia can produce invasive infections(including myonecrosis and gas gangrene) if introduced into damaged tissue. About 30 species of clostridia may produce such an infection, but the most common in invasive disease is C. perfringens(90%). An enterotoxin of C. perfringens is a common cause of food poisoning. Toxins: produce different types of toxins and enzymes that result in spreading infection. They have lethal, necrotizing, and hemolytic properties. Pathogenesis: Wound contamination. Clinical Findings: Infection spreads in 1-3 days. Crepitation in subcutaneous tissue and muscle, fever, tissue necrosis, hemolytic anemia, severe toxemia and death.

Clostridial soft tissue infections

Crepitant cellulitis Fascitis Myonecrosis

Clostridial myonecrosis

Clinical course Symptoms begin 1-4 days after inoculation and progresses rapidly to extensive muscle necrosis and shock Local area with marked pain, swelling, serosanguinous discharge, bullae, slight crepitance May be associated with increased CPK Treatment Surgical debridement Antibiotics Hyperbaric oxygen

C. difficile

Pseudomembranous colitis (Antibiotic Associated Diarrhea)

Clostridium difficile

Epidemiology Endogenous infection Colonizes GI tract in 5% healthy individuals Antibiotic exposure associated with overgrowth of C. difficile Cephalosporins, clindamycin, ampicllin/amoxicillin Other contributing factors: agents altering GI motility, surgery, age, underlying illness Exogenous infection Spores detected in hospital rooms of infected patients Pathogenesis Enterotoxin (toxin A) produces chemotaxis, induces cytokine production and hypersecretion of fluid, development of hemorrhagic necrosis Cytotoxin (toxin B) Induces polymerization of actin with loss of cellular cytoskeleton

C. difficile colitis
Clinical syndromes Asymptomatic colonization Antibiotic-associated diarrhea Pseudomembranous colitis Diagnosis Isolation of toxin Culture Treatment Discontinue antibiotics Metronidazole or oral vancomycin Pooled human IVIG for severe disease Probiotics (saccharomyces boulardii) New drugs (nitazoxanide, tolevamer) Relapse in 20-30% (spores are resistant)

The Genus

Clostridium
Left. Stained pus from a mixed anaerobic infection. At least three different clostridia are apparent. Right. Electron micrograph of C.perfringens

Clostridium tetani

C. difficile

C. botulinum

C. tetani