Clonal differences in Staphylococcus aureus bacteraemia-associated mortality.

The bacterium Staphylococcus aureus is a major human pathogen for which the emergence of antibiotic resistance is a global public health concern. Infection severity, and in particular bacteraemia-associated mortality, has been attributed to several host-related factors, such as age and the presence of comorbidities. The role of the bacterium in infection severity is less well understood, as it is complicated by the multifaceted nature of bacterial virulence, which has so far prevented a robust mapping between genotype, phenotype and infection outcome. To investigate the role of bacterial factors in contributing to bacteraemia-associated mortality, we phenotyped a collection of sequenced clinical S. aureus isolates from patients with bloodstream infections, representing two globally important clonal types, CC22 and CC30. By adopting a genome-wide association study approach we identified and functionally verified several genetic loci that affect the expression of cytolytic toxicity and biofilm formation. By analysing the pooled data comprising bacterial genotype and phenotype together with clinical metadata within a machine-learning framework, we found significant clonal differences in the determinants most predictive of poor infection outcome. Whereas elevated cytolytic toxicity in combination with low levels of biofilm formation was predictive of an increased risk of mortality in infections by strains of a CC22 background, these virulence-specific factors had little influence on mortality rates associated with CC30 infections. Our results therefore suggest that different clones may have adopted different strategies to overcome host responses and cause severe pathology. Our study further demonstrates the use of a combined genomics and data analytic approach to enhance our understanding of bacterial pathogenesis at the individual level, which will be an important step towards personalized medicine and infectious disease management

Development of an efficient technique for gene deletion and allelic exchange in Geobacillus spp.

Abstract Background Geobacillus thermoglucosidasius is a thermophilic, natural ethanol producer and a potential candidate for commercial bioethanol production. Previously, G. thermoglucosidasius has been genetically modified to create an industrially-relevant ethanol production strain. However, creating chromosomal integrations and deletions in Geobacillus spp. is laborious. Here we describe a new technique to create marker-less mutations in Geobacillus utilising a novel homologous recombination process. Results Our technique incorporates counter-selection using β-glucosidase and the synthetic substrate X-Glu, in combination with a two-step homologous recombination process where the first step is a selectable double-crossover event that deletes the target gene. We demonstrate how we have utilised this technique to delete two components of the proteinaceous shell of the Geobacillus propanediol-utilization microcompartment, and simultaneously introduce an oxygen-sensitive promoter in front of the remaining shell-component genes and confirm its functional incorporation. Conclusion The selectable deletion of the target gene in the first step of our process prevents re-creation of wild-type which can occur in most homologous recombination techniques, circumventing the need for PCR screening to identify mutants. Our new technique therefore offers a faster, more efficient method of creating mutants in Geobacillus

Additional file 2: of Epistasis analysis uncovers hidden antibiotic resistance-associated fitness costs hampering the evolution of MRSA

Table S3. Protein profile differences between SH1000 (mupS) and MY40 (mupR). Proteins highlighted in yellow are also differentially produced by exposure to mupirocin. (XLSX 32 kb