Widespread Fosfomycin Resistance in Gram-Negative Bacteria Attributable to the Chromosomal <i>fosA</i> Gene

Abstract

Fosfomycin is a decades-old antibiotic which is being revisited because of its perceived activity against many extensively drug-resistant Gram-negative pathogens. FosA proteins are Mn²⁺ and K⁺-dependent glutathione <i>S</i>-transferases which confer fosfomycin resistance in Gram-negative bacteria by conjugation of glutathione to the antibiotic. Plasmid-borne <i>fosA</i> variants have been reported in fosfomycin-resistant <i>Escherichia coli</i> strains. However, the prevalence and distribution of <i>fosA</i> in other Gram-negative bacteria are not known. We systematically surveyed the presence of <i>fosA</i> in Gram-negative bacteria in over 18,000 published genomes from 18 Gram-negative species and investigated their contribution to fosfomycin resistance. We show that FosA homologues are present in the majority of genomes in some species (e.g., <i>Klebsiella</i> spp., <i>Enterobacter</i> spp., <i>Serratia marcescens</i>, and <i>Pseudomonas aeruginosa</i>), whereas they are largely absent in others (e.g., <i>E. coli</i>, <i>Acinetobacter baumannii</i>, and <i>Burkholderia cepacia</i>). FosA proteins in different bacterial pathogens are highly divergent, but key amino acid residues in the active site are conserved. Chromosomal <i>fosA</i> genes conferred high-level fosfomycin resistance when expressed in <i>E. coli</i>, and deletion of chromosomal <i>fosA</i> in <i>S. marcescens</i> eliminated fosfomycin resistance. Our results indicate that FosA is encoded by clinically relevant Gram-negative species and contributes to intrinsic fosfomycin resistance.<b>IMPORTANCE</b> There is a critical need to identify alternate approaches to treat infections caused by extensively drug-resistant (XDR) Gram-negative bacteria. Fosfomycin is an old antibiotic which is routinely used for the treatment of urinary tract infections, although there is substantial interest in expanding its use to systemic infections caused by XDR Gram-negative bacteria. In this study, we show that <i>fosA</i> genes, which encode dimeric Mn²⁺- and K⁺-dependent glutathione <i>S</i>-transferase, are widely distributed in the genomes of Gram-negative bacteria-particularly those belonging to the family <i>Enterobacteriaceae</i>-and confer fosfomycin resistance. This finding suggests that chromosomally located <i>fosA</i> genes represent a vast reservoir of fosfomycin resistance determinants that may be transferred to <i>E. coli</i> Furthermore, they suggest that inhibition of FosA activity may provide a viable strategy to potentiate the activity of fosfomycin against XDR Gram-negative bacteria.