X. METHODS IN CAMPYLOBACTER
449
C. K antigen
Wiidik and Hlidar (1955) demonstrated that the capsular antigen may
inhibit 0 agglutination of living or formolised C. fetus suspensions in
homologous 0 serum. The presence of K antigen in C. fetus in this investigation is detected by (1) K agglutination with diluted K serum performed
in the slide agglutination test, (2) the K agglutination with diluted K serum
in the tube agglutination test, (3) the swelling reaction which is usual for
pneumococci, and (4) by heating the cultures at 120°C for 2 h. It seems
that this surface antigen appears as a layer of different thickness in various
strains and that different strains vary only quantitatively in their K antigen
content.
VII. PHAGES OF C. FETUS
C. fetus is lysogenic and carries temperate bacteriophage (Bryner et uZ.,
1968; Firehammer and Border, 1968).
A. Maintenance of phages
The fluid thioglycollate medium is useful for maintenance and enrichment of lysogenic strains of C. fetus. The method is described by Bryner
et aZ. (1970) : Broth for shaken cultures and phage dilution blanks is BruceZZu
broth (Albimi) supplemented with 3 g sodium succinate and 0.1 g cysteine/
litre (pH 6-9). BruceZZa agar (Albimi) is the nutrient base for double agar
layer cultures; the soft top layer contains per litre of water: agar (Colab
Ion agar no. 2) 5 g; NH4CI 1 g; KHzP04 6.5 g; NazHP04 3.5 g; sodium
succinate 3 g; glucose 1 g. The agar cultures are incubated as described.
B. Phage isolation (Bryner et al., 1970)
For isolation of lysogenic strains of C. fetus, each test strain is inoculated
in a separate tube containing 9 ml of thioglycollate medium and then 1 ml
of the host culture is added to each tube. The mixed cultures are incubated
for three days at 36°C and then centrifuged at 2500 x g for 20 min to remove
cells from unadsorbed phage in the supernatant fluid. This fluid is transferred to separate 100 ml flasks containing 5 x 109 log phase host cells in
20 ml supplemented Albimi broth (as described above) and incubated for
5 h. One millilitre samples from each flask are plated in overlay agar to
test for plaque formation.
Bryner et aZ. (1970) isolated 22 bacteriophages from 38 strains of C.
fetus and characterised the phage V-45 according to its stability in broth,
growth kinetics, morphology, and physicochemical properties. The minimum latent period was 135 min, rise time was 75 min, and average burst
size 35 plaque forming units per infected cell.
449
C. K antigen
Wiidik and Hlidar (1955) demonstrated that the capsular antigen may
inhibit 0 agglutination of living or formolised C. fetus suspensions in
homologous 0 serum. The presence of K antigen in C. fetus in this investigation is detected by (1) K agglutination with diluted K serum performed
in the slide agglutination test, (2) the K agglutination with diluted K serum
in the tube agglutination test, (3) the swelling reaction which is usual for
pneumococci, and (4) by heating the cultures at 120°C for 2 h. It seems
that this surface antigen appears as a layer of different thickness in various
strains and that different strains vary only quantitatively in their K antigen
content.
VII. PHAGES OF C. FETUS
C. fetus is lysogenic and carries temperate bacteriophage (Bryner et uZ.,
1968; Firehammer and Border, 1968).
A. Maintenance of phages
The fluid thioglycollate medium is useful for maintenance and enrichment of lysogenic strains of C. fetus. The method is described by Bryner
et aZ. (1970) : Broth for shaken cultures and phage dilution blanks is BruceZZu
broth (Albimi) supplemented with 3 g sodium succinate and 0.1 g cysteine/
litre (pH 6-9). BruceZZa agar (Albimi) is the nutrient base for double agar
layer cultures; the soft top layer contains per litre of water: agar (Colab
Ion agar no. 2) 5 g; NH4CI 1 g; KHzP04 6.5 g; NazHP04 3.5 g; sodium
succinate 3 g; glucose 1 g. The agar cultures are incubated as described.
B. Phage isolation (Bryner et al., 1970)
For isolation of lysogenic strains of C. fetus, each test strain is inoculated
in a separate tube containing 9 ml of thioglycollate medium and then 1 ml
of the host culture is added to each tube. The mixed cultures are incubated
for three days at 36°C and then centrifuged at 2500 x g for 20 min to remove
cells from unadsorbed phage in the supernatant fluid. This fluid is transferred to separate 100 ml flasks containing 5 x 109 log phase host cells in
20 ml supplemented Albimi broth (as described above) and incubated for
5 h. One millilitre samples from each flask are plated in overlay agar to
test for plaque formation.
Bryner et aZ. (1970) isolated 22 bacteriophages from 38 strains of C.
fetus and characterised the phage V-45 according to its stability in broth,
growth kinetics, morphology, and physicochemical properties. The minimum latent period was 135 min, rise time was 75 min, and average burst
size 35 plaque forming units per infected cell.