Anaerobic Bacteria

Category

Spore-forming:
rod, Gram (+)--- Clostridium
Nonspore-forming:
see next slides

Category
Sporerod, Gram (+)--- Clostridium
forming:
Nonspore-forming:
Rod, Gram (+)
Propionibacterium
Bifidobacterium

Lactobacillus
Eubacterium
Actinomyces

Rod, Gram (-) Bacteroides

Fusobacterium
Campylobacter
Cocci, Gram (+) Peptococcus
Cocci, Gram (-)

Peptostreptococcus
Veillonella

Clostridium Species

The clostridia are opportunistic

pathogens. Nonetheless, they are
responsible for some of the deadliest
diseases including gas gangrene,
tetanus and botulism. Less lifethreatening diseases include
pseudomembranous colitis (PC) and
food poisoning.
cause disease primarily through the
production of numerous exotoxins.
perfringens, tetani, botulinum, difficile

Clostridium Tetani

Pathogenesis of tetanus caused by C tetani

General introduction

C tetani is found worldwide.

Ubiquitous in soil, it is occasionally
found in intestinal flora of humans
and animals
C.tetani is the cause of tetanus,or
lockjaw. When spores are introduced
into wounds by contaminated soil or
foreign objects such as nails or glass
splinters

BIOCHEMICAL CHARACTERISTICS

Morphology: long and slender;

peritrichous flagella,no capsule,
terminal located round
spore(drum-stick apperance),
its diameter greater than
vegetative cell.
Culture:obligate anaerobic;
Gram(+); swarming occures on
blood agar, faint hemolysis.
Biochemical activities:does not
ferment any carbohydrate and
proteins.
Resistance: tolerate boiling for
60 min.alive several ten years in
soil.
Classification and Antigenic
Types: C tetani is the only species.
There are no serotypes

2-5 x 0.3-0.5um

Pathogenicity

No invasiveness; toxemia
(exogenous infection
produces two exotoxins:
tetanolysin, and
tetanospasmin(a kind of
neurotoxin, toxicity
strong)
The actions of
tetanospasmin are
complex and involve three
components of the
nervous system: central
motor control, autonomic
function, and the
neuromuscular junction.

retrograde transport
to (CNS)
delitescence a few
days to several
weeks
The two animal
species most
susceptible to this
toxemia are horses
and humans.

Clostridium tetani -Tetanospasmin

disseminates systemically
binds to ganglioside receptors
inhibitory neurones in CNS
glycine
neurotransmitter
stops nerve impulse to muscles
spastic paralysis
severe muscle contractions and
spasms
can be fatal

Tetanospasmin

Clinical Manifestations

The initial symptom is cramping and

twitching of muscles around a wound. The
patient usually has no fever but sweats
profusely and begins to experience pain,
especially in the area of the wound and
around the neck and jaw muscles (trismus).
Portions of the body may become extremely
rigid, and opisthotonos (a spasm
in which the head and heels are bent
backward and the body bowed forward) is
common.
Complications include fractures, bowel
impaction, intramuscular hematoma, muscle
ruptures, and pulmonary, renal, and cardiac
problems

Clinical Manifestations

DISEASE
CLINCAL MANIFESTATIONSA
Generalized Involvement of bulbar and paraspinal

Cephalic

Localized
Neonatal

muscles(trismus or lockjaw, risus sardonicus,

difficulty swallowing, irritability,
opisthotonos);involvement of autonomic
nervous system(sweating, hyper thermia,
cardiac arrhythmias, fluctuations in blood
pressure)
Primary infection in head,particularly
ear;isolated or combined involvement of cranial
nerves, particularly seventh cranial nerve; very
poor prognosis
Involvement of muscles in area of primary
injury; infection may precede generalized
disease; favorable prognosis
Generalized disease in neonates; infection
typically originates from umbilical
stump;very poor prognosis in infants whose
mothers are nonimmune

Epidemiology

1 million cases of tetanus occur annually in the

world,with a mortality rate ranging from20%
to 50%. But rare in most developed countries.
In some developing countries, tetanus is still
one of the ten leading causes of death, and
neonatal tetanus accounts for approximately
one-half of the cases worldwide.
In less developed countries, approximate
mortality rates remain 85% for neonatal
tetanus and 50% for nonneonatal tetanus.
In the United States, intravenous drug abusers
have become another population with an
increasing incidence of clinical tetanus
In untreated tetanus, the fatality rate is 90%
for the newborn and 40% for adults.

Immunity

Humoral immunity(antitoxin)
There is little, if any, inate immunity
and the disease does not produce
immunity in the patient.
Active immunity follows vaccination
with tetanus toxoid

Diagnosis

Diagnosis is primarily by the clinical

symptoms (above). The wound may not
be obvious.
C tetani can be recovered from the wound
in only about one-third of the cases.
It is important for the clinician to be
aware that toxigenic strains of C tetani
can grow actively in the wound of an
immunized person.
Numerous syndromes, including rabies
and meningitis, have symptoms similar to
those of tetanus and must be considered
in the differential diagnosis.

Vaccination

infant
DPT (diptheria, pertussis, tetanus)
tetanus toxoid
antigenic
no exotoxic activity

Control

The offending organism must be

removed by local debridemen

toxoid
TAT; Metronidazole (For more
serious wounds)
AIDS patients may not respond
to prophylactic injections of
tetanus toxoid

C. perfringens
soil, fecal contamination
gas gangrene
swelling of tissues
gas release
* fermentation products
wound contamination

Toxins
toxin Biological Feature Types of Toxins
A

lecithinase; increase the

vascular permeability;
hemolytic; produces
necrotizing activity

Necrotizing activity,
induces hypertension
by causing release of
catecholamines.

increase the
permeability of
gastrointestinal wall

Necrotizing activity;
increase the vascular
permeability

Toxins

Many of these toxins have lethal,

necrotizing, and hemolytic properties;
The alpha toxin produced by all types of C.
perfringens, is a lecithinase that lyses
erythrocytes, platelets, leukocytes, and
endothelial cells. And its lethal action is
proportionate to the rate at which it splits
lecithin to phosphorylcholine and
diglyceride.
The theta toxin has similar hemolytic and
necrotizing effects.
DNAase, hyaluronidase, a collagenase are
also produced

Enterotoxin

Many strains of type A produce

enterotoxin, which is a heat-labile protein
and destroyed immediately at 100 .
Trypsin treatment enhances the toxin
activity threefold.
The toxin is produced primarily by type A
strains but also by a few type C and D
strains.
It disrupts ion transport in the
ileum(primarily) and jejunum by inserting
into the cell membrane and altering
membrane permeability.
As superantigen.

Pathogenesis
Tissue degrading enzymes
lecithinase [ toxin]
proteolytic enzymes
saccharolytic enzymes
Destruction of blood vessels
Tissue necrosis
Anaerobic environment created
Organism spreads

Without treatment death

occurs within 2 days

effective antibiotic therapy

debridement
anti-toxin
amputation & death is rare

Gas gangrene

Gas gangrene is a life-threatening disease with

a poor prognosis and often fatal outcome.

Initial trauma to host tissue damages muscle and
impairs blood supply----lack of oxygenation
Initial symptoms : fever and pain in the infected
tissue.; more local tissue necrosis and systemic
toxemia. Infected muscle is discolored (purple
mottling) and edematous and produces a foulsmelling exudate; gas bubbles form from the
products of anaerobic fermentation.

Gas gangrene

As capillary permeability increases,

the accumulation of fluid increases,
and venous return eventually is
curtailed.
As more tissue becomes involved, the
clostridia multiply within the
increasing area of dead tissue,
releasing more toxins into the local
tissue and the systemic circulation.

Food poisoning

Enterotoxin producing strains.

These bacteria are found in

mammalian faeces and soil.
Small numbers of the bacteria may
also be found in foods and they may
propagate rapidly to dangerous
concentrations if the food is
improperly stored and handled.

Food poisoning

more than 108 vegetative cells are

ingested and sporulate in the gut, the
toxins can act rapidly in the body,
causing severe diarrhea in 6-18 hours,
dysentery, gangrene, muscle infections
The action of C. perfringens enterotoxin
involves marked hypersecretion in the
jejunum and ileum, with loss of fluids
and electrolytes in diarrhea.

Cellulitis, Fasciitis

Cellulitis, Fasciitis
Fasciitis : a rapidly progressive,
destructive process in which the organisms
spread through fascial plan es.
Fasciitis causes suppuration and the
formation of gas
Absense of muscle involvement

rapidity

Necrotizing Enteritis
Rare, acute necrotizing process in the
jejunum
Abdominal pain, bloody diarrhea, shock, and
peritonitis
Mortality: 50%
Beta-toxin-producing C. perfringens type C

Septicemia

Who is at risk?

Surgical patients; patient after

trauma with soil contamination.
People who ingest contaminated
meat products (without proper
refrigeration or reheating to
inactivate endotoxin)

Epidemiology

C. perfringens type A: the intestinal

tract of humans and animals, soil and
water contaminated with feces. forms
spores under adverse environmental
conditions and can survive for
prolonged periods.
Type B to E strains colonize the
intestinal tract of animals and
occasionally humans.

Epidemiology

Type A: gas gangrene, soft tissue

infections and food poisoning
Type C: enteritis; necroticans

Laboratory identification
lecithinase production

Double Hemolysis Circles

C. botulinum

Biological Features

Anaerobic
Gram-positive
rod-shaped
sporeformer
produces a protein neurotoxic.
soil, sediments of lakes, ponds,
decaying vegetation.
intestinal tracts of birds, mammals
and fish.

Division
---A, B, C1, D, E, F, and G.
---type A. 62%
---Not all produce toxin.
---C and D not
---G plasmid encoded.

Transmission
---spores heat resistant.
canning.
anaerobic environment
---Botulism
eating uncooked foods
spores
---GI, duodenum, blood stream,
neuromuscular synapses.

Virulence factors

---bacterial protease
---light chain,A,50 kDa;
heavy chain,100kDa.
---disulfide bond.
---A potent toxin

binds peripheral nerve receptors

acetylcholine neurotransmitter
inhibits nerve impulses
flaccid paralysis
death
Botulinum
respiratory
cardiac failure

toxin

Botulinum toxin

Bioterrorism
not an infection
resembles a chemical attack
10 ng can kill a normal adult

Epidemiology
---4: foodborne, infant, wound, undetermined.
---Certain foods; wound not.
---Foodborne botulism, consumption.
---Infant botulism, 1976, under 12m.
---ingestion, colonize and produce toxin in the
intestinal tract of infants.
honey.
---increased.
---internationally recognized.

Clinical syndromes
---18-36 hours:
---weakness, dizziness,dryness of the mouth.
---Nausea,vomiting.
---Neurologic features: blurred vision,
inability to swallow, difficulty in speech,
descending weakness of skeletal muscles,
respiratory paralysis.

Botulism( )

food poisoning
rare
fatal
germination of spore
inadequately sterilized canned food
home

not an infection

Infection with C. botulinum

Neonatal botulism
uncommon
the predominant form of
botulism
colonization occurs
no normal flora to compete
unlike adult

Wounds
extremely rare
an infection

Immunity
---specifically neutralized, antitoxin.
---toxoided, make good antigens.
---does not develop, amount toxic.
---Repeated occurrence.
---Once bound, unaffected by antitoxin.
---circulating toxin ,neutralized , injection
of antitoxin.
---treated immediately with antiserum.
---multivalent
toxoid,unjustified,infrequency.
experimental vaccine.

Diagnosis
---by clinical symptoms alone
---differentiation difficult.
--- most direct and effective: serum or
feces.
---most sensitive and widely used:
mouse neutralization test. 48h.
Culturing of specimens 5-7d.

Treatment

Individuals known to have ingested food

with botulism should be treated
immediately with antiserum.
antibiotic therapy (if infection)
Vaccination will not protect hosts from
botulism, however passive
immunisation with antibody is the
treatment of choice for cases of
botulism.

Prevention
---proper food handling and preparation.
--- spores survive boiling (100 degrees
at 1 atm) 1h.
---toxin heat-labile, boiling or intense
heating, inactivate the toxin.
---bulge, gas, spoiled.

C. difficile

After antibiotic use

Intestinal normal flora --greatly decreased
Colonization occurs
Enterotoxin secreted
Pseudomembanous colitis

Pseudomembranous Colitis

Pseudomembranous colitis (PC) results

predominantly as a consequence of the
elimination of normal intestinal flora
through antibiotic therapy.
Symptoms include abdominal pain with
a watery diarrhea and leukocytosis.
"Pseudomembranes" consisting of
fibrin, mucus and leukocytes can be
observed by colonoscopy.
Untreated pseudomembranous colitis
can be fatal in about 27-44%.

Therapy

Discontinuation of initial antibiotic

(e.g. ampicillin)
Specific antibiotic therapy (e.g.
vancomycin)

Obligate (strict) anaerobes

no oxidative phosphorylation
fermentation
killed by oxygen
lack certain enzymes
superoxide dismutase
* O2-+2H+ H2O2
catalase
* H2O2 H20 + O2
peroxidase
* H2O2 H20 /NAD to NADH

Strict anaerobe infectious

disease

Sites throughout body

Muscle, cutaneous/sub-cutaneous
necrosis
Abscesses

Bacterial Flora of the Body

Site

Total Bacteria
Ratio
(per/ml or gm) Anaerobes:Aerobes

Upper Airway
Nasal Washings
Saliva
Tooth Surface
Gingival Crevice

103-104
108-109
1010-1011
1011-1012

3-5:1
1:1
1:1
1000:1

Gastrointestinal Tract
Stomach
Small Bowel
Ileum
Colon

102-105
102-104
104-107
1011-1012

1:1
1:1
1:1
1000:1

Female Genital Tract

Endocervix
Vagina

108-109
108-109

3-5:1
3-5:1

Problems in identification of
anaerobic infections
air in sample (sampling, transportation)
no growth
identification takes several days or longer
limiting usefulness
often derived from normal flora
sample contamination can confuse

Virulence Factors
1.

Anti-phagocytic capsule
Also promote abscess formation

2.

Tissue destructive enzymes

B. fragilis produces variety of enzymes
(lipases, proteases, collagenases) that
destroy tissue Abscess Formation

3.

Beta-lactamase production
B. fragilis protect themselves and other
species in mixed infections

4.

Superoxide dismutase production

Protects bacteria from toxic O2 radicals as
they move out of usual niche

Characteristics of Anaerobic Infections

1.

Most pathogenic anaerobes are

usually commensals
Originate from our own flora

2.

Predisposing Conditions
Breeches in the mucocutaneous barrier
displace normal flora
Compromised vascular supply
Trauma with tissue destruction
Antecedent infection

Characteristics of Anaerobic
Infections
3. Complex Flora

Multiple species
Abdominal Infection Avg
of 5 species

3 anaerobic
2 aerobic

Less complex then nl flora

Fecal flora 400 different
species

Those predominant in stool

are not infecting species
Veillonella,
Bifidobacterium rarely
pathogenic

Species uniquely suited to

cause infection
predominate

4. Synergistic Mixture
of Aerobes &
Anaerobes

E. coli Consume O2
Allow growth
of
anaerobes

Anaerobes promote
growth of other
bacteria by being
antiphagocytic and
producing Blactamases

Clues to Anaerobic Infection

Infections in continuity to mucosal
surfaces
Infections with tissue necrosis and
abscess formation
Putrid odor
Gas in tissues
Polymicrobial flora
Failure to grow in the lab

1.

2.

3.
4.
5.
6.

BIOCHEMICAL KITS

e.g. API SYSTEM

GAS CHROMATOGRAPHY

volatile fermentation products

Bacteroides fragilis
Major disease causing strict anaerobic
after abdominal surgery
non-spore-former
Prominent capsule
anti-phagocytic
abscess formation
Endotoxin
low toxicity
structure different than other
lipolysaccharide

 Enterobacteriaceae (facultative anaerobes)

commonly cause disease
low numbers gut flora
Strict anaerobes
much less commonly cause disease
high numbers gut flora
.