Characterization of FosL1, a Plasmid-Encoded Fosfomycin Resistance Protein Identified in Escherichia coli

Abstract

Fosfomycin is gaining renewed interest for treating urinary tract infections. Monitoring fosfomycin resistance is therefore important in order to detect the emergence of novel resistance mechanisms. Here, we used the Rapid Fosfomycin NP test to screen a collection of extended-spectrum-β-lactamase-producing Escherichia coli isolates from Switzerland and found a fosfomycin-resistant isolate in which a novel plasmid-mediated fosfomycin resistance gene, named fosL1, was identified. The FosL1 protein is a putative glutathione S-transferase enzyme conferring high-level resistance to fosfomycin and sharing between 57% to 63% amino acid identity with other FosA-like family members. Genetic analyses showed that the fosL1 gene was embedded in a mobile insertion cassette and had likely been acquired by transposition through a Tn7-related mechanism. In silico analysis over GenBank databases identified the FosL1-encoding gene in addition to another variant (fosL1 and fosL2, respectively) in two Salmonella enterica isolates from the United States. Our study further highlights the necessity of monitoring fosfomycin resistance in Enterobacteriaceae to identify the emergence of novel mechanisms of resistance.

Keywords: Escherichia coli; FosL1; fosfomycin; plasmid.

FIG 1

Phylogenetic tree obtained for all the identified Fos enzymes, including the bacillithiol and glutathione transferases by distance method using a neighbor-joining algorithm (SeaView version 4 software). Branch lengths are drawn to scale and are proportional to the number of amino acid substitutions with 200 bootstrap replications. The distance along the vertical axis has no significance. Percentages of amino acid identity shared between the FosL1 enzyme and the other Fos enzymes are indicated in brackets.

FIG 2

Amino acid sequence comparison between FosL1 and different members of glutathione transferases. Boxes bracket amino acids implicated in Mn²⁺, K⁺, and fosfomycin bond. Hyphens represent the conserved amino acids with the FosL1 sequence and dashes represent the gaps in the amino acid sequence.

FIG 3

Schematic structures of the different mobile insertion cassettes harboring the fosL gene. (A) Genetic structure identified in R249 (this study). (B and C) Genetic structures identified in silico in Salmonella species isolates. Tn7L/R, sequences showing high nucleotide identity with the Tn7 extremities recognized by the transposases of the transposon Tn7; gna, gene encoding a putative acetyltransferase; IS91-like, insertion sequence truncated by the insertion of the fosL1 cassette between nucleotides 150 and 151; pecM, gene encoding a histone acetyltransferase HPA2 and related acetyltransferases (protein ID BBB37423.1); urk, gene encoding a putative uridine kinase; hyp, hypothetical gene.