pHTβ-promoted mobilization of non-conjugative resistance plasmids from Enterococcus faecium to Enterococcus faecalis

Objectives: To analyse the recombination events associated with conjugal mobilization of two multiresistance plasmids, pRUM(17i48) and pLAG (formerly named pDO1-like), from Enterococcus faecium 17i48 to Enterococcus faecalis JH2-2. Methods: The plasmids from two E. faecalis transconjugants (JH-4T, tetracycline resistant, and JH-8E, erythromycin resistant) and from the E. faeciumdonor (also carrying a pHT beta-like conjugative plasmid, named pHT beta(17i48)) were investigated by several methods, including PCR mapping and sequencing, S1-PFGE followed by Southern blotting and hybridization, and WGS. Results: Two locations of repA(pHT beta) were detected in both transconjugants, one on a similar to 50 kb plasmid (as in the donor) and the other on plasmids of larger sizes. In JH-4T, WGS disclosed an 88.6 kb plasmid resulting from the recombination of pHT beta(17i48) (similar to 50kb) and a new plasmid, named pLAG (35.3 kb), carrying the tet(M), tet(L), lsa(E), lnu(B), spw and aadE resistance genes. In JH-8E, a 75 kb plasmid resulting from the recombination of pHTb17i48 and pRUM17i48 was observed. In both cases, the cointegrates were apparently derived from replicative transposition of an IS1216 present in each of the multiresistance plasmids into pHT beta(17i48). The cointegrates could resolve to yield themultiresistance plasmids and a pHT beta(17i48) derivative carrying an IS1216 (unlike the pHT beta(17i48) of the donor). Conclusions: Our results completed the characterization of the multiresistance plasmids carried by the E. faecium 17i48, confirming the role of pHT plasmids in the mobilization of non-conjugative antibiotic resistance elements among enterococci. Results also revealed that mobilization to E. faecalis was associated with the generation of cointegrate plasmids promoted by IS1216-mediated transposition.