Background

Horizontal gene transfer plays a major role in transferring genetic material between bacteria. The most common type is conjugation, which is considered a significant contributor in the emergence of new antimicrobial-resistant pathogens. This process is highly efficient and depends on the presence of conjugative plasmids. These plasmids provide the necessary genes for their transmission including the transfer (tra) genes involved in the type IV secretion system. Previous work has shown that transfer frequency of a cefotaxime (CTX) resistance plasmid was significantly enhanced when the donor was pre-grown in CTX (1). However, it is still unknown how CTX has this effect.

Objectives

In this study, we aim at identifying genes and regulatory pathways involved in the conjugative spread of resistance plasmids in E. coli exposed to CTX. Furthermore, we intend to identify compounds which prevent plasmid conjugation.

Methods and results

The screening of a transposon mutant library led to the identification of eight genes, five more genes were selected based on their suspected effect on antibiotic-induced conjugation. All genes were deleted using the lambda-red recombinase system. Eleven of these mutants led to reduced antibiotic-induced conjugation frequency compared to the wild-type, while two had the opposite effect. Five mutants, which significantly reduced antibiotic induced conjugation, were complemented to confirm their effect.

A novel protocol was designed to screen a large compound-library for compounds that can inhibit or reduce conjugation. The method provides a rapid and easy way for identifying potential plasmid conjugation inhibitors from a large library.

Summary

These data identify chromosomally encoded genes that contribute to the CTX induced up-regulation of conjugation. By further understanding the regulatory pathway we will be able to identify targets for future new antimicrobial agents.

(1) Liu G, Olsen JE and Thomsen LE (2019) Identification of Genes Essential for Antibiotic-Induced Up-Regulation of Plasmid-Transfer-Genes in Cephalosporin Resistant Escherichia coli. Front. Microbiol. 10:2203. doi: 10.3389/fmicb.2019.02203