Identification of a novel tetracycline resistance gene, tet(63), located on a multiresistance plasmid from Staphylococcus aureus

Objectives

To identify and characterize a novel tetracycline resistance gene on a multiresistance plasmid from Staphylococcus aureus SA01 of chicken origin.

Methods

MICs were determined by broth microdilution according to CLSI recommendations. The whole genome sequence of S. aureus SA01 was determined via Illumina HiSeq and Oxford Nanopore platforms followed by a hybrid assembly. The new tet gene was cloned and expressed in S. aureus. The functionality of the corresponding protein as an efflux pump was tested by efflux pump inhibition assays.

Results

A novel tetracycline resistance gene, tet(63), was identified on a plasmid in S. aureus SA01. The cloned tet(63) gene was functionally expressed in S. aureus and shown to confer resistance to tetracycline and doxycycline, and a slightly elevated MIC of minocycline. The tet(63) gene encodes a 459 amino acid efflux protein of the major facilitator superfamily that consists of 14 predicted transmembrane helices. The results of efflux pump inhibitor assays confirmed the function of Tet(63) as an efflux protein. The deduced amino acid sequence of the Tet(63) protein exhibited 73.0% identity to the tetracycline efflux protein Tet(K). The plasmid pSA01-tet, on which tet(63) was located, had a size of 25664 bp and also carried the resistance genes aadD, aacA-aphD and erm(C).

Conclusions

A novel tetracycline resistance gene, tet(63), was identified in S. aureus. Its location on a multiresistance plasmid might support the co-selection of tet(63) under the selective pressure imposed by the use of macrolides, lincosamides and aminoglycosides.