Mutants constructed in selected candidate virulence genes in S. suis strains
In the PIGSs project we will determine which S. suis are conditionally essential for survival and growth in the ecological niches within the host mainly using data from (1). Transcriptomics under in vivo mimicking conditions, (2). TnSeq mutagenesis under in vivo mimicking conditions and (3). Genomics data from the collection of clinical isolates sequenced during the initial phases of the project.
The transcriptomics data under in vivo mimicking conditions, revealed no new candidate virulence candidates not already described in the literature (apart from genes encoding hypothetical proteins).
Using transposon insertion sequencing (TnSeq) we identified a total of 116 transposon mutants that were attenuated in growth under in vivo mimicking conditions: growth in peroxide (mimicking oxidative stress resulting from host innate defences), growth without transition metals (mimicking very low abundance of free iron at mucosal surfaces), or growth in serum (mimicking invasive infections). It is known that mutants generated by insertional tranposon mutagenesis can be unstable. Therefore we generated deletion mutants from a rationally selected subset of these genes. In total, 17 genes are selected for targeted mutagenesis.
Genetic approaches are also being used to investigate the function of genes which are highly conserved in the genomes of clinical isolates but much less conserved in non-disease associated strains.