54
IV. STOEV
B. Serotyping
The method of Atkinson as modified by Nikolov (1964) was used to
produce an acetic acid extract from cells. The method involves the use of
dilute acetic acid held in a thermostat during extraction. Group specific
A and B antisera were produced by immunisation of rabbits.
The microprecipitation method was carried out by Nikolov’s modification of Mansi’s method (Nikolov, 1964). Both the agar base and the
specific antisera were stored at 6 6 ° C after the addition of merthiolate
(1 in 10 000).
C. Bacteriophage
One to two drops of serogroup A specific Erysipelothrix bacteriophage
(107 plaque-forming units per ml) were added to young (1 to 2 h incubation) broth cultures of the strains to be studied. Lytic action of the phage
was determined after 18-24 h cultivation at 37°C.
D. Haemadsorption
The ability of strains to adsorb erythrocytes was determined by the
examination of colonies grown on the surface of agar. Broth cultures were
inoculated on to serum agar (pH 7.8) in Petri dishes. After cultivation for
24,48,72 and 96 h at 37°C the agar surface was flooded with a 2% chicken
erythrocyte suspension using a dropper and applying the reagent with
great care at the edge of the dish. The flooded plate was allowed to stand
for 15-20min at room temperature after which the erythrocytes were
gently decanted and the agar suface carefully washed with physiological
saline, again using a dropper. Washing was carried out by slow shaking
and rotary movement of the dishes. Sharp or careless movements can lead
to the separation of Erysipelothrix colonies from the surface of the agar.
The agar was then dried in an inverted po.sition for 5-10 min and the
colonies observed under a microscope at a magnification of 80 diameters.
Erythrocytes for the test were prepared from poultry, rabbit, sheep and
.guinea pigs and they were carefully washed several times with physiological
saline.
In order to check the specificity of haemadsorption, a section of the
agar surface carrying Erysipelothrix colonies was overlayered with specific
anti-Erysipelothrix serum derived from pigs and destined for therapeutic
use. Colonies were exposed to the serum for 60 min at room temperature,
after which time it was carefully poured off and the whole surface of the
agar covered with erythrocyte suspension and the haemadsorption characteristics of both the treated and untreated colonies studied by the methods
described above.
IV. STOEV
B. Serotyping
The method of Atkinson as modified by Nikolov (1964) was used to
produce an acetic acid extract from cells. The method involves the use of
dilute acetic acid held in a thermostat during extraction. Group specific
A and B antisera were produced by immunisation of rabbits.
The microprecipitation method was carried out by Nikolov’s modification of Mansi’s method (Nikolov, 1964). Both the agar base and the
specific antisera were stored at 6 6 ° C after the addition of merthiolate
(1 in 10 000).
C. Bacteriophage
One to two drops of serogroup A specific Erysipelothrix bacteriophage
(107 plaque-forming units per ml) were added to young (1 to 2 h incubation) broth cultures of the strains to be studied. Lytic action of the phage
was determined after 18-24 h cultivation at 37°C.
D. Haemadsorption
The ability of strains to adsorb erythrocytes was determined by the
examination of colonies grown on the surface of agar. Broth cultures were
inoculated on to serum agar (pH 7.8) in Petri dishes. After cultivation for
24,48,72 and 96 h at 37°C the agar surface was flooded with a 2% chicken
erythrocyte suspension using a dropper and applying the reagent with
great care at the edge of the dish. The flooded plate was allowed to stand
for 15-20min at room temperature after which the erythrocytes were
gently decanted and the agar suface carefully washed with physiological
saline, again using a dropper. Washing was carried out by slow shaking
and rotary movement of the dishes. Sharp or careless movements can lead
to the separation of Erysipelothrix colonies from the surface of the agar.
The agar was then dried in an inverted po.sition for 5-10 min and the
colonies observed under a microscope at a magnification of 80 diameters.
Erythrocytes for the test were prepared from poultry, rabbit, sheep and
.guinea pigs and they were carefully washed several times with physiological
saline.
In order to check the specificity of haemadsorption, a section of the
agar surface carrying Erysipelothrix colonies was overlayered with specific
anti-Erysipelothrix serum derived from pigs and destined for therapeutic
use. Colonies were exposed to the serum for 60 min at room temperature,
after which time it was carefully poured off and the whole surface of the
agar covered with erythrocyte suspension and the haemadsorption characteristics of both the treated and untreated colonies studied by the methods
described above.