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Molecular characterisation of Australian Coxiella burnetii isolates

Vincent, Gemma (2014) Molecular characterisation of Australian Coxiella burnetii isolates. PhD thesis, Murdoch University.

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The intracellular bacterium Coxiella burnetii is the causative agent of Q fever, a human zoonotic disease with acute and chronic forms, which is most commonly associated with exposure to infected animals such as sheep, goats and cattle. Q fever occurs worldwide and is endemic in Australia with around 300 cases confirmed by laboratory testing each year. Although the first cases of the disease were first recognised in Australia in the 1930s, at the start of this study little was known about the molecular epidemiology of C. burnetii in this country. The work presented in this thesis provides the first extensive molecular analysis of the strains of the bacterium causing Q fever in Australia.

The small, pre-existing collection of seven Australian C. burnetii isolates was expanded to a larger collection comprising 43 isolates. The majority of the new isolates were obtained from serum samples taken from acute Q fever patients during the early stage of their disease in a two year period from 2010-2012. This demonstrated that the organisms remained viable in these specimens despite the absence of host cells, thus acute serum is a valuable source of C. burnetii. Attempted isolations of C. burnetii from kangaroo faeces and an Australian wombat tick were unsuccessful but bacterial DNA was obtained from these samples for further characterisation.

Several genotyping methods were used to characterise the Australian C. burnetii isolate collection at the molecular level. All the human isolates were found to contain the plasmid QpRS and were negative for the acute disease antigen A gene. Single nucleotide polymorphism typing also failed to discriminate between the human isolates but demonstrated that C. burnetii representing three different genotypes was present in the kangaroo faecal samples. Discrimination between the human isolates was only achieved using an extended panel of PCRs targeting the repetitive insertion sequence element IS1111 and multi-locus VNTR analysis. Both methods identified 14 genotypes, most of which were novel compared to the genotypes identified in characterised strains of C. burnetii from other countries and in combination the two methods determined 24 genotypes, providing an even greater discriminatory power. Many of the genotyping targets were not amplified from the bacterial DNA in the wombat tick leading to the conclusion that the organism present was a Coxiella species other than C.burnetii.

Overall, results showed that the Australian C. burnetii isolates are genetically closely related and unique to this country. The evaluation of different genotyping methods enabled the development of a set of guidelines that will reduce the cost and workload required to characterise new Australian isolates of this important pathogen.

Publication Type: Thesis (PhD)
Murdoch Affiliation: School of Veterinary and Life Sciences
Supervisor: Stenos, John, Graves, Stephen and Fenwick, Stan
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