Clostidium perfringens causes gas gangrene. Laboratory diagnosis can be made through gram staining of samples from infected tissues and wounds, which will show large numbers of gram positive bacilli. Cultures are also used to identify the bacteria, looking for characteristics such as B-hemolysis on blood agar. Proper treatment involves thorough debridement of damaged tissue from wounds, use of antibiotics that C. perfringens is susceptible to, administration of antitoxin for immunization, and hyperbaric oxygen therapy to reduce anaerobic conditions. Active immunization with toxiods has also been experimentally used.

1. CLOSTIDIUM PERFRINGENS
Laboratory diagnosis and
prophylaxis
Harshita thota
roll.no 108

2. Clostidium perfringens causes gas gangrene .
The diagnosis of gas gangrene must be made primarily on
clinical grounds as mere presence of clostridia in
wounds does not constitute gas gangrene.
Lab diagnosis can be done to Differentiate gas gangrene
from anaerobic streptococcal myositis , which may be
indistinguishable from it clinically in the early stages
In the later ,gram stained films show large numbers of
streptococci and pus cells but not bacilli, contrasting with
the scanty pus cells and diverse bacterial flora seen in
films from gas gangrene.

3. 1. SPECIMENS
1 Films from :
the muscles at the edge of the affected area.
the tissue in the necrotic area.
exudate in the deeper parts of the wound.
2 Exudates from:
the parts where the infection appears most active .
the depths of the wound .
They are collected with a capillary pipette or a swab.
3. Necrotic tissue and muscle fragments.

4. 2.MICROSCOPIC EXAMINATION
Gram staining is done to distinguish different species
of clostridia .
In the slide of :
Clostridium perfringens –
large numbers of regularly shaped, gram positive
bacilli without spores are seen.
Clostridium septicum-
‘citron bodies’ and boat- or leaf shaped pleomorphic
bacilli with irregular staining .

5.  Clostridium novyi –
Large bacilli with oval , sub terminal spores are seen.
 Clostridium tetani or C.tetanomorphum –
Slender bacilli with round, terminal spores are seen

7. 3 CULTURE
 Robertson’s Cooked meat broth-
Growth in cooked meat broth is sub cultured in blood
agar after 24-48 hrs
 Blood agar-
Aerobic and Anaerobic cultures are made on fresh
and heated blood agar , preferably on 5-6% agar to
prevent swarming .
Blood agar is incubated anaerobically for 48-72hrs.
most strains produce B- haemolysis on blood agar
and few are non haemolytic .
.

8.  A plate of serum and egg yolk agar , with
clostridium perfringens antitoxin spread on one half
, is used for Nagler reaction.

9.  Colonies on the half without antitoxin will be
surrounded by opacity while the colonies with
antitoxin will not show opacity due to neutralization
of alpha toxin.
 Alpha toxin ( lecithinase c ) splits lecithine into
phosphoryl choline and a diglycerate (lipid). The
lipid gets deposited around the colonies resulting in
opacity.
 Exceptions- cl.novyi ,some vibrios , some aerobic
spore bearers

10. Bacterial isolates can be identified by:
 Morphology
 Cultural characteristics
 Biochemical reactions
 Reverse CAMP test
 Toxigenicity of a strain can be done by animal
pathogenicity.

11. PROPHYLAXIS
1. Surgery :
All damaged tissue should be removed promptly and the
would should be irrigated with antiseptic solution to
remove blood clots , necrotic tissue and foreign
materials .
2.Antibiotics :
Gas gangrene organisms are susceptible to :
Metronidazole
Penicillin
Sulphonamide
Tetracyclin
Amoxycillin

12. 3. Antitoxin:
Passive immunization with anti gas gangrene serum
is used prophylactically in extensively soiled wound.
4. Hyperbaric oxygen :
it is introduced in the depth of wound to reduce
anaerobiosis .
5.Active immunization :
toxiods have been found .used expermentally
Not in practical use .