Low-concentration chlorine disinfection promotes conjugative transfer from an ESBL-producing Escherichia coli donor within and across Gram types

Abstract

To elucidate the effects of chlorine disinfection on conjugative transfer of antibiotic resistance genes, conjugation experiments were conducted using an ESBL-producing Escherichia coli T413 as the donor strain and E. coli NK5449 and Bacillus cereus X524 as recipient strains, representing intra- and inter-Gram conjugation models, respectively. Conjugative transfer frequency was quantified under different chlorine concentrations by exposing either the donor strain or the recipient strain to chlorine prior to conjugation, and the underlying mechanisms were investigated through physiological and molecular analyses. The results demonstrated that low-concentration chlorine disinfection at 0.5 mg/L promoted conjugative transfer regardless of whether the donor or the recipient was chlorinated, and in both intra- and inter-Gram conjugation models, with the highest increase of up to 14.3-fold observed. Low-concentration chlorine induced oxidative stress in donor cells, as reflected by elevated ROS levels and upregulation of genes recA, rpoS, and sodA. This stress enhanced cell membrane permeability via upregulation of outer membrane protein genes (ompA, ompC, and ompF), facilitating plasmid release, while also upregulating conjugative transfer-related genes (trbC and flgC) to promote conjugation apparatus formation. In recipient bacteria, chlorine increased membrane permeability, thereby enhancing plasmid uptake. Together, these gene expression changes and cellular responses contributed to the enhanced conjugative transfer. These mechanistic insights highlight the potential risk of low-concentration chlorine disinfection in inadvertently driving the spread of antibiotic resistance.

Graphical abstract

Keywords

Low-concentration chlorine disinfection / Conjugative transfer / ESBL-producing Escherichia coli / Oxidative stress / Cell membrane permeability / Intra- and inter-Gram conjugation models

Highlight

	● Low-concentration chlorine disinfection promotes conjugative transfer.
	● Promotion occurs from an ESBL-producing E. coli donor within and across Gram types.
	● Chlorine induces oxidative stress and increases membrane permeability.
	● Low-concentration chlorine upregulates expression of conjugation-related genes.

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