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<p>The overall goal of this application is to understand the bacterial factors and host immunity that influence the virulence, persistence, and dominance of Campylobacter jejuni clone SA in sheep. The specific objectives of this project are </p>
<p>1) to identify the bacterial factors that are important for systemic infection of C. jejuni clone SA, </p>
<p>2) to determine the mechanisms responsible for the emergence of C. jejuni clone SA in sheep abortion, and </p>
<p>3) to evaluate protective immunity induced by commercial and experimental vaccines against C. jejuni clone SA. This is a 5-year-long research project. </p>
<p>Specific Objectives 1 and 2 will be conducted during the entire project period, while Specific Objective 3 will be carried out during the last 4 years. Once completed, the planned work will identify the bacterial factors contributing to systemic infection of C. jejuni, which will not only significantly advance our understanding of Campylobacter pathogenesis, but also identify potential candidates for developing anti-Campylobacter vaccines. It will also discover the adaptive genetic changes associated with the evolution of C. jejuni clone SA. This finding will provide important insights into the evolutionary trajectory of this pathogenic clone and facilitate the identification of intervention strategies to control the emergence of new pathogenic variants. Additionally, the work will ascertain if commercially available vaccines are protective against this newly emerged virulent clone and if the vaccines induce strain-specific protection. Together, these outputs will significantly advance our understanding of Campylobacter pathogenesis, adaptive mechanisms, and protective immunity. The discoveries will also benefit sheep producers by identifying appropriate immunization strategies. Control of C. jejuni in ruminants will also impact food safety as this hypervirulent clone is transmitted to humans via raw milk and other routes.</p>
<p>NON-TECHNICAL SUMMARY: <br/>Campylobacter jejuni, a Gram-negative bacterium, is a major cause of foodborne illnesses in humans and septic abortion in sheep around the world, including the United States. A ruminant C. jejuni clone (named clone SA) has become the predominant cause of sheep abortion and is associated with outbreaks (related to raw milk) and sporadic cases of foodborne gastroenteritis in humans. Despite the increasing importance of clone SA in animal production and public health, little is known about its disease-causing mechanisms and the factors that contribute to its emergence and dominance. There is also very little information on protective immunity against this pathogen. Addressing these knowledge gaps is critically needed for developing practical and effective strategies to mitigate this emerging zoonotic threat. Our central hypothesis is that through
evolution Campylobacter jejuni clone SA has acquired a unique ability to cause systemic infection and evade immunity generated by non-homologous vaccines. Specific objectives of this study are: 1) to identify bacterial factors that are important for systemic infection caused by clone SA; 2) to determine the mechanisms involved in emergence and persistence of clone SA in sheep abortion, and 3) to evaluate the protective immunity of commercial and experimental vaccines against sheep abortion. We expect that these studies will identify previously unrecognized bacterial factors involved in systemic disease, reveal the evolutionary mechanisms for dominance of clone SA in sheep abortion, and develop new vaccines to protect against sheep abortion. The findings will significantly benefit sheep producers by identifying appropriate strategies to control sheep abortion as it incurs a major economic
loss. In addition, the work will positively impact public health because C. jejuni clone SA is a zoonotic pathogen and its control in ruminants will reduce its transmission to the human hosts. Thus the proposed studies truly serve ?Dual Purpose with Dual Benefit?, improving both animal and human health.
<p>APPROACH: <br/>For Specific Objective 1, two complementary approaches will be used to identify bacterial factors involved in systemic infection: targeted mutation of genes and systemic screening of the entire genome by transposon mutagenesis coupled with high-throughput deep sequencing technology. These mutants will be examined in infection models using guinea pigs and sheep to determine attenuation of virulence in causing systemic infection. For genome-wide identification of genes involved in systemic infection, a saturated library of transposon mutants (input pool) of clone SA will be inoculated into guinea pigs and sheep and Campylobacter will be isolated from different tissues (output pools). Each output population will be compared with the input population by deep sequencing (using Illumina sequencing platforms) to identify mutants which are negatively selected
during systemic infection. These experiments will allow us to identify bacterial factors that are important for systemic infection, which are potential targets for disease control. For Specific Objective 2, a historical collection of C. jejuni isolates from sheep abortion will be analyzed using whole genome sequencing. Bioinformatics analyses will be performed to identify the genomic changes associated with emergence of clone SA. The genomics findings will also be compared with the results from experimental studies to identify links between specific genomic features and evolution of virulence. In addition, animal experiments using lambs will be performed to assess the within-host and between-host fitness and transmission of clone SA isolates derived from different time periods. These experiments will allow us to understand how C. jejuni clone SA became predominant and hypervirulent.For
Specific Objective 3, a series of laboratory immunization experiments using ewes will be conducted to evaluate the efficacy of commercially available vaccines and a clone SA-specific bacterin. Groups of ewes will be immunized with these vaccines and then challenged with this hypervirulent clone. At necropsy, various tissues will be collected for culturing Campylobacter and for histopathology. Protection will be assessed using clinical signs, bacteremia, fecal and bile colonization, and histopathological lesions in various tissues. These experiments will yield important information on immunization strategies, which will facilitate the control of sheep abortion in the United States.