In vivo survival and growth of bacterial mutants quantified by next-generation sequencing

Screening pools of random transposon mutants is a powerful approach to link genotype with phenotype. TraDIS (transposon-directed insertion site-sequencing) is a high-throughput method that exploits next-generation sequencing to allow the genomic location of each transposon insertion site to be determined in a massively parallel manner. Here we develop the method to allow quantitative evaluation of the relative abundance of each mutant before and after an in vivo selective screen. The approach is applied to Salmonella enterica serovar Typhimurium mutants screened by infection of BALB/c mice, a widely used model of systemic typhoid fever. Comparison of the TraDIS results with those obtained from a study using a microarray-based approach to quantify transposon insertion frequency (TMDH, transposon-mediated differential hybridization) supports and extends the conclusions of that study. We show that TraDIS offers significant advantages over existing approaches as the method is quantitative, fast and highly cost-effective, opening up the possibility that TraDIS technology can be applied to a range of functional genomics projects.