Genome-wide identification of novel therapeutic targets for the control of bovine Staphylococcus aureus infection

Supervisors: Ross Fitzgerald, Mark Stevens

Project description:

Staphylococcus aureus is a major cause of bovine mastitis in the global dairy industry. S. aureus mastitis infections are often refractory to treatment, in part due to the rise of antibiotic resistant strains. The development of alternative approaches for treating or preventing major endemic infections such as mastitis is urgently required. We are currently carrying out a reverse vaccinology project supported by a BBSRC-Zoetis LINK award (Sarah Salmon, Zoetis collaborator), and have identified a panel of conserved antigens that are currently being tested for protective efficacy in dairy cows. To complement the ongoing work in order to identify bacterial factors that are important for survival in vivo we will carry out a transposon directed insertion-site sequencing (TraDIS) analysis of bovine S. aureus during both growth in bovine milk and passage with bovine bone marrow-derived macrophages. This approach will allow cross reference and short-listing of antigens identified by the current reverse vaccinology study, and identification of novel targets for vaccine or therapeutic-based interventions. Briefly, the study will involve construction of a transposon (Tn)-mutant library in our reference strain of bovine S. aureus strain Newbold305 which will be cultured in milk or passaged with bovine macrophages and surviving bacteria isolated. High-throughput sequencing specific for transposon insertion sites will facilitate comparison of the original bovine S. aureus input Tn library population with the surviving population after growth/infection. Bioinformatic analysis will allow identification of Tn insertion mutants that are poorly represented in the output population suggesting a key role for those loci in bacterial survival in conditions simulating the in vivo environment. A similar approach will employed using dairy cow challenge model, allowing direct examination of genes required for survival in vivo. The importance of selected genes will be validated by targeted mutagenesis and growth/passage to confirm their importance in survival, and the mechanisms involved will be examined using appropriate functional assays and cellular microbiological analysis in vitro.  Furthermore, selected factors, including any that are identified using both reverse vaccinology and TraDIS may then be taken forward for testing as either vaccine components in experimental challenge studies or as therapeutic targets by screening small molecule libraries. Overall, the study represents a rational method for the identification of novel preventive or therapeutic targets leading to the design of alternative approaches for controlling bovine mastitis. This would have significant benefit for dairy herd health, the economy of the UK and global dairy industry, and for limiting the emergence and spread of antibiotic resistance.

References:

Caitriona M. Guinane, Nouri L. Ben Zakour, Maria A. Tormo-Mas, Lucy A. Weinert, Bethan V. Lowder, Robyn A. Cartwright, Davida S. Smyth, Cyril J. Smyth, Jodi Lindsay, Katherine A. Gould, Adam Witney, Jason Hinds, Jonathan P. Bollback, Andrew Rambaut, José Penadés, and J. Ross Fitzgerald. 2010. Evolutionary genomics of Staphylococcus aureus reveals insights into the origin and molecular basis of ruminant host adaptation. Genome Biol Evol 2:454-66

Chaudhuri RR, Morgan E, Peters SE, Pleasance SJ, Hudson DL, Davies HM, Wang J,van Diemen PM, Buckley AM, Bowen AJ, Pullinger GD, Turner DJ, Langridge GC, Turner AK, Parkhill J, Charles IG, Maskell DJ, Stevens MP. Comprehensiveassignment of roles for Salmonella typhimurium genes in intestinal colonization of food-producing animals. PLoS Genet. 2013 Apr;9(4):e1003456.

 

Other: