title: Studies on Campylobacter fetus subspecies jejuni
Creators: name: Ng, Francis Kee Peng
Publisher: name: University of Glasgow
Year: 1980
Subjects: subject: Microbiology.

description: The work described in this thesis was principally directed at trying to develop a mouse model for human campylobacteriosis and also at defining the antigenic relationships between the human strains. Human diarrhoeal and normal stool specimens were examined for Campylobacter fetus ss, jejuni by culture on plates of Campylobacter selective medium, incubated at 43°C in an atmosphere of 5% O2, 10% CO2 and 85% N2. From 290 specimens of diarrhoeal stool, 11 isolates of Campylobacter were obtained. The organism was not isolated from the stools of 49 normal people. Alkaline peptone water, pH 8.4, was examined as a possible enrichment medium but did not yield additional isolates. For growth in liquid culture, nutrient broth supplemented with yeast extract and cystine and incubated in an atmosphere of 5% O2, 10% CO2 and 85% N2 gave the best cell yields. Bubbling the gas mixture (10% CO2 and 90% N2) through freshly inoculated medium gave no growth. Contrary to what has been reported with Vibrio fetus, the addition of KNO3, MgSO4, CaC12, Na2H2PO4 to nutrient broth did not enhance the growth of C. fetus ss. jejuni. Because of the tendency of Campylobacter colonies to spread on agar, considerable difficulty was experienced in performing colony counts on suspensions of the organism. Success was, however, achieved by using well-dried culture plates and an incubation period of less than 30 hours. Two changes in the culture medium - addition of P-nitrophenylglycerol and extra agar - which have been recommended for colony counts on Proteus strains were not satisfactory with C. fetus ss. jejuni. Freeze-drying was found unsatisfactory for the long term (more than 1 year) preservation of C. fetus ss. jejuni. However, strains could readily be maintained by mixing the broth culture with 15% glycerol and coating glass beads which were stored at -76°C. Mice of the HAM I/CR strain were injected with C. fetus ss. jejuni isolates of human origin by the intraperitoneal and intravenous routes, and were also given the bacteria orally. Although mice died if the dose of organisms was sufficiently large (1010), viable counts on liver and spleen homogenates made at various times indicated that there was little, if any, bacterial multiplication in vivo. However, viable Campylobacter could be recovered from various organs, notably blood, liver, spleen, kidney and gastrointestinal tract. This distribution was irrespective of the route of inoculation. The most consistent recovery of the administered bacteria was from the liver and spleen; persistence in the liver being up to 21 days. No mouse that had been given C. fetus ss. jejuni exhibited signs of diarrhoea. There was a marked age effect of susceptibility of HAM I/CR mice to the lethal effect of C. fetus ss. jejuni. Animals 7 days old were considerably more susceptible than younger (1-and 3-day old) or older (2-, 3-and 5-week old) mice. The virulence of C. fetus ss. jejuni for 7-day old mice could be increased slightly by passaging the bacteria five times through adult mice, with recovery from the spleen. C. fetus ss. jejuni did not produce extracellular toxins, and death of the mice was due to the toxicity of the bacterial cells themselves. Bacteria killed at 56°C had an LD50 of 4.1 X 109 for 7-day old mice, compared with 1.5 x 10 9 of the corresponding live suspension. The toxicity of bacteria killedat 56°C was not diminished by heating at 100°C for 15 min., which suggested that the toxic factor was lipopoly-saccharide and that the lethal effect of live organisms was due to the content of endotoxin. This conclusion was supported by the observation that crude cell envelopes contained the toxic factor while the cytoplasm was non-toxic. (Abstract shortened by ProQuest.)