Novel plasmid-borne gene qacJ mediates resistance to quaternary ammonium compounds in equine Staphylococcus aureus, Staphylococcus simulans, and Staphylococcus intermedius

Abstract

We identified a novel plasmid-borne gene (designated qacJ) encoding resistance to quaternary ammonium compounds (QACs) in three staphylococcal species associated with chronic infections in four horses. qacJ was located on a 2,650-bp plasmid (designated pNVH01), a new member of the pC194 family of rolling-circle replication plasmids. The 107-amino-acid protein, QacJ, showed similarities to known proteins of the small multidrug resistance family: Smr/QacC (72.5%), QacG (82.6%), and QacH (73.4%). The benzalkonium chloride MIC for a qacJ-containing recombinant was higher than those for otherwise isogenic recombinants expressing Smr, QacG, or QacH. Molecular epidemiological analyses by pulsed-field gel electrophoresis suggested both the clonal spread of a qacJ-harboring Staphylococcus aureus strain and the horizontal transfer of pNVH01 within and between different equine staphylococcal species. The presence of pNVH01 of identical nucleotide sequence in different staphylococcal species suggests that recent transfer has occurred. In three of the horses, a skin preparation containing cetyltrimethylammonium bromide had been used extensively for several years; this might explain the selection of staphylococci harboring the novel QAC resistance gene.

FIG. 1.

Nucleotide sequence of pNVH01 (GenBank accession no. AJ512814) containing two ORFs. The smaller ORF (qacJ; nt 1428 to 1748) encodes a putative protein (QacJ) of 107 aa. An alternative transcription start codon is present at nt 1419 to 1421 and potentially encodes a 110-aa protein. The larger ORF transcribed in the opposite direction (nt 100 to 1104) encodes a putative protein of 334 aa. The putative ribosome binding site (RBS) and the +ori sequence (including the potential replication nick site indicated by a vertical arrow) are underlined. The conserved single-stranded origion (SSO_A; previously known as palA) is boxed. Direct repeat sequences are in boldface.

FIG. 2.

Alignment of different staphylococcal QAC resistance-related proteins. The amino acids that differ from those in Smr are boxed. The completely conserved amino acids in all four proteins are denoted on the line above the alignment. The different SMR proteins consist of four TMSs, TMS1 to TMS4, comprising amino acid residues 3 to 20, 30 to 47, 58 to 74, and 86 to 103, respectively. The only charged TMS amino acid residue is highly conserved glutamate-13 (in boldface).