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, pRUM17i48 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 pHTb-like conjugative plasmid, named pHTβ17i48) were investigated by several methods, including PCR mapping and sequencing, S1-PFGE followed by Southern blotting and hybridization, and WGS. Results: Two locations of repApHTβ were detected in both transconjugants, one on a 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 pHTβ17i48 (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 pHTβ17i48 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 pHTβ17i48. The cointegrates could resolve to yield themultiresistance plasmids and a pHTβ17i48 derivative carrying an IS1216 (unlike the pHTβ17i48 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.