Characterization of wild-type and laboratory attenuated “Brachyspira hampsonii”
This thesis aimed to characterize Western Canadian “Brachyspira hampsonii” clinical isolates and to develop and characterize an attenuated “B. hampsonii” vaccine candidate. Clinical “B. hampsonii” isolates selected from different Western Canadian barns were described biochemically and by sequencing the NADH-oxidase (nox) gene to improve the information available about clinical isolates present in Western Canada. The biochemical profiles of two clade I and eight clade II isolates were compared and variability in some diagnostic biochemical markers (β-glucosidase and hippurate hydrolysis) was detected. Representative isolates of clade I and II “B. hampsonii” were also compared to other swine-associated Brachyspira spp. based on biochemical profiles and morphometric data. Cell dimensions, number of flagella, and select biochemical markers (hippurate hydrolysis, α-glucosidase, and β-glucosidase) were not sufficient to discern “B. hampsonii” from other Brachyspira species. The data highlighted that consistent biochemical characteristics (indole and α-galactosidase negative) of “B. hampsonii” and morphology were not sufficient to differentiate them from other Brachyspira spp. and that nox gene sequencing is a more reliable tool for identification and differentiation of Brachyspira species. Development of a potential vaccine strain was also carried out by serial passage of a virulent “B. hampsonii” clade II strain (passage 13 (P13)) for 100 passages, resulting in strain P113. The virulent P13 and serially passaged P113 strains were phenotypically and genotypically compared to identify any changes in P113. Phenotypes of P13 and P113, including the biochemical profile, β-haemolysis, growth profile, and virulence in a murine model, were compared to identify changes. More extensive β-haemolysis and more rapid growth of P113 over P13 was observed suggesting laboratory adaptation after serial passage. Two experiments were performed to assess the virulence of P113 and P13 in a murine model, which both demonstrated a decrease of P113 virulence, suggesting that partial attenuation was achieved. No difference between the virulent and attenuated strains was detected based on genomic profiling by random amplified polymorphic DNA (RAPD) or whole genome sequence comparison. The results of this thesis demonstrate the complexity of phenotypic identification and discrimination of “B. hampsonii” and other swine-associated Brachyspira species. Serial passage of a virulent “B. hampsonii” resulted in a laboratory adapted, partially attenuated strain that will be attenuated further and used as a potential vaccine. This serially passaged strain also provides opportunities for further studies to identify virulence determinants, which can aid immensely with prevention and treatment of “B. hampsonii”.
Brachyspira hampsonii, Western Canada
Master of Science (M.Sc.)