I. TYPING METHODS FOR CLOSTRIDIUM
3
biotic sensitivity testing, and defined volumes of the different bacteriocins
are dropped on to sectors of the seeded plate. The pattern of inhibition
which develops after subsequent growth of the lawn defines the bacteriocin
type of the organism.
There are factors in favour of each method and further detail may be
found in the literature (Shannon, 1957; MacPherson and Gillies, 1969).
111. GROWTH AND IDENTIFICATION O F
CLOSTRIDIA
Many excellent references may be found on the growth and identification
of the genus Clostridium (Dowell and Hawkins, 1974; Holdeman and
Moore, 1972; Smith, 1975; Smith and Holdeman, 1968; Willis, 1969).
In most laboratories, cooked meat or thioglycollate-containing broths and
the Gaspak anaerobic jar (Bioquest, B.B.L.) are used in cultivation. I t is
beyond the scope of this Chapter to discuss the many variations in growth
techniques and identification procedures.
The clostridia are usually Gram-positive, spore-forming, catalasenegative anaerobic bacteria. Some species are much more aerotolerant than
others, but anaerobic techniques must be applied to assure the isolation of
any member of the genus. C. perfringens is often reluctant to produce
spores and is also non-motile. I n addition, a double zone of haemolysis
surrounds the smooth, entire-edged colonies of this organism on human
or sheep blood agar plates; the inner zone of beta haemolysis is produced
by the theta toxin while the outer weaker zone is caused by the alpha toxin.
Some isolates, however, may lack the theta toxin or produce rough, irregular
colonies which will confuse identification. The organism can be specifically
identified by its saccharolytic activity on various sugars and neutralisation
of its toxins with specific antisera. Some clostridia may only be specifically
identified by toxin neutralisation tests, e.g. C. botulism and C. tetani.
Newer analytical tools such as gas-liquid chromatography have enhanced
the identification of many anaerobic bacteria by detecting specific organic
acids produced by the fermentation of defined carbohydrates. This technology is described in detail by Holdeman and Moore (1972).
IV. NATURE AND MODE O F ACTION OF CLOSTRIDIAL
BACTERIOCINS
A. Bacteriocins of C. perf ringens
1. Incidence of bacteriocinogenicity in C. perfringens
Although the first observation of bacteriocin-like activity in this species
was made by Smith in 1959 while studying lysogeny, the first major screen-
3
biotic sensitivity testing, and defined volumes of the different bacteriocins
are dropped on to sectors of the seeded plate. The pattern of inhibition
which develops after subsequent growth of the lawn defines the bacteriocin
type of the organism.
There are factors in favour of each method and further detail may be
found in the literature (Shannon, 1957; MacPherson and Gillies, 1969).
111. GROWTH AND IDENTIFICATION O F
CLOSTRIDIA
Many excellent references may be found on the growth and identification
of the genus Clostridium (Dowell and Hawkins, 1974; Holdeman and
Moore, 1972; Smith, 1975; Smith and Holdeman, 1968; Willis, 1969).
In most laboratories, cooked meat or thioglycollate-containing broths and
the Gaspak anaerobic jar (Bioquest, B.B.L.) are used in cultivation. I t is
beyond the scope of this Chapter to discuss the many variations in growth
techniques and identification procedures.
The clostridia are usually Gram-positive, spore-forming, catalasenegative anaerobic bacteria. Some species are much more aerotolerant than
others, but anaerobic techniques must be applied to assure the isolation of
any member of the genus. C. perfringens is often reluctant to produce
spores and is also non-motile. I n addition, a double zone of haemolysis
surrounds the smooth, entire-edged colonies of this organism on human
or sheep blood agar plates; the inner zone of beta haemolysis is produced
by the theta toxin while the outer weaker zone is caused by the alpha toxin.
Some isolates, however, may lack the theta toxin or produce rough, irregular
colonies which will confuse identification. The organism can be specifically
identified by its saccharolytic activity on various sugars and neutralisation
of its toxins with specific antisera. Some clostridia may only be specifically
identified by toxin neutralisation tests, e.g. C. botulism and C. tetani.
Newer analytical tools such as gas-liquid chromatography have enhanced
the identification of many anaerobic bacteria by detecting specific organic
acids produced by the fermentation of defined carbohydrates. This technology is described in detail by Holdeman and Moore (1972).
IV. NATURE AND MODE O F ACTION OF CLOSTRIDIAL
BACTERIOCINS
A. Bacteriocins of C. perf ringens
1. Incidence of bacteriocinogenicity in C. perfringens
Although the first observation of bacteriocin-like activity in this species
was made by Smith in 1959 while studying lysogeny, the first major screen-