Staphylococcal tetracycline-MLSB resistance plasmid pSTE2 is the product of an RSA-mediated in vivo recombination

Objectives: The complete nucleotide sequence of the 6913 bp plasmid pSTE2 from Staphylococcus lentus, which mediates inducible resistance to tetracyclines, macrolides and lincosamides, was determined. The plasmid was analysed for potential reading frames and structural features to gain insight into its development from potential ancestor plasmids. Methods: Plasmid pSTE2 was transformed into Staphylococcus aureus RN4220. Suitable restriction fragments were cloned into E. coli plasmid vectors and sequenced. In vitro susceptibility testing was performed to confirm the resistance phenotype mediated by this plasmid. Results: Plasmid pSTE2 consisted of two parts, each of which corresponded closely to previously identified staphylococcal plasmids. The initial 4439 bp represented a pT181-analogous tet(K)-carrying tetracycline resistance plasmid, whereas the remaining 2474 bp represented a pPV141-related erm(C)-carrying macrolide-lincosamide-streptogramin B resistance plasmid. Both putative parental plasmids harboured the staphylococcal recombination site A (RSA) and the pT181-like plasmid also carried the recombinase gene pre whose product acts at RSA. Analysis of the junctions of the pT181-like and the pPV141-like homologous parts in pSTE2 suggested that plasmid pSTE2 developed from pT181- and pPV141-like ancestor plasmids by cointegrate formation at RSA. Conclusion: Plasmid pSTE2 is the first completely sequenced plasmid from S. lentus and represents the product of an in vivo derived RSA-mediated recombination between two compatible plasmids.