Identification of host genetic factors modulating β-lactam resistance in <i>Escherichia coli</i> harbouring plasmid-borne β-lactamase through transposon-sequencing

Since β-lactam antibiotics are widely used, emergence of bacteria with resistance to them poses a significant threat to society. In particular, acquisition of genes encoding β-lactamase, an enzyme that degrades β-lactam antibiotics, has been a major contributing factor in the emergence of bacteria that are resistant to β-lactam antibiotics. However, relatively few genetic targets for killing these resistant bacteria have been identified to date. Here, we used a systematic approach called transposon-sequencing (Tn-Seq), to screen the <i>Escherichia coli</i> genome for host genetic factors that, when mutated, affect resistance to ampicillin, one of the β-lactam antibiotics, in a strain carrying a plasmid that encodes β-lactamase. This approach enabled not just the isolation of genes previously known to affect β-lactam resistance, but the additional loci <i>skp</i>, <i>gshA</i>, <i>phoPQ</i> and <i>ypfN</i>. Individual mutations in these genes modestly but consistently affected antibiotic resistance. We have identified that these genes are not only implicated in β-lactam resistance by itself but also play a crucial role in conditions associated with the expression of β-lactamase. <i>GshA</i> and <i>phoPQ</i> appear to contribute to β-lactam resistance by regulating membrane integrity. Notably, the overexpression of the uncharacterized membrane-associated protein, <i>ypfN</i>, has been shown to significantly enhance β-lactam resistance. We applied the genes identified from the screening into <i>Salmonella</i> Typhimurium and <i>Pseudomonas aeruginosa</i> strains, both critical human pathogens with antibiotic resistance, and observed their significant impact on β-lactam resistance. Therefore, these genes can potentially be utilized as therapeutic targets to control the survival of β-lactamase-producing bacteria.