Using high-density mutagenesis to identify the genetic requirements for the growth of Escherichia coli

Abstract

Escherichia coli is highly adapted to life within the mammalian gastrointestinal (GI) tract, capable of adapting to multiple environments en route to colonising the intestine. Moreover, the means by which the species copes with changes in the microenvironments of the GI tract strongly influences the nature of E. coli's relationship with the host i.e. whether it exists as a commensal or pathogen. However, the response of E. coli to many of these conditions is complex, often employing a whole-cell response. This necessitates the use of high-throughput approaches in order to fully understand factors the bacterium requires for growth under these conditions. This work outlines the use of transposon-directed insertion site sequencing (TraDIS) to describe genetic requirements for fitness of E. coli K-12 MG1655 during growth in the presence of bile and under anaerobic conditions in the presence of nitrate, representative of bile exposure in the duodenum and conditions of inflammation in the intestine. TraDIS reveals, in detail, genetic requirements for growth under these conditions, revealing new roles for many genes with no prior association with growth under these conditions. This work will, therefore, contribute to future studies of E. coli colonisation of the GI tract by identifying candidate genes required for fitness under these growth conditions.