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 re-
activated, 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
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
SF Bacteria-Bacillus
Lab diagnosis :
Gram staining
Culture on Blood agar
Speciment :
Fluid, pus, blood, sputum
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
Clostridium tetani
C. botulinum
C.perfringens
C. tetani
C. difficile