Antimicrobial susceptibility of bifidobacteria from probiotic milk products and determination of the genetic basis of tetracycline resistance in Enterococcus species after in vitro conjugation with Bifidobacterium animalis subsp lactis

The study aimed at investigating the antimicrobial resistance (AMR) profiles of bifidobacteria from fermented milk products sold in the UK that are claimed to have beneficial effects on humans, and assess the potential transferability of AMR to other bacteria. Isolates of different Bifidobacterium species were screened for their susceptibility to 24 antimicrobials and the presence of AMR genes [tet(M), tet(L), tet(S), tet(Q), tet(K), tet(0), tet(W), aph(3)-I, ant(2)-I, strA, strB, andA, andE, erm(A), enn(B), and erm(C)]. The ability of Bifidobacterium animalis subsp. lactis isolates to transfer the tet(W) gene to Enterococcus faecalis JH2-2 and Enterococcus faecium BM4105 was investigated by conjugation. Potential E. faecalis transconjugants (PETs) were analysed by whole genome sequencing (WGS) for the presence of tet(W), other DNA fragments from the donor and single-nucleotide polymorphisms (SNPs). A high level of resistance to aminoglycosides and a moderate level of resistance to tetracycline were observed, while susceptibility to macrolides, vancomycin, chloramphenicol and beta-lactams was determined. Only the tet (W) gene was detected in all B. animalis subsp. lactis isolates. Some PETs exhibited a significant tetracycline MIC increase from 1 to up to 16 mg/L. WGS did not detect tet(W) in the PETs, but SNPs in the rpsJ gene encoding the ribosomal protein S10, which is part of the 30S ribosomal subunit and contains a proposed tetracycline binding site. Substitutions (Tyr-58 -> Asp, Tyr-58 -> Cys, Asp-60 -> Tyr, Tyr-58 -> Ser, Ser-61 -> Tyr, Ala-54 -> Glu, Asp 60 -> Tyr) and an insertion of Thr between Thr55 and His56 were detected. No E. faecium potential transconjugants were recovered. B. animalis subsp. lactis isolates did not transfer the tet(W) gene to the Enterococcus species. The majority of the PETs had mutations within or near the ribosomal protein SIO vertex loop suggesting that mutations in the rpsJ gene confer tetracycline resistance to E. faecalis JH2-2.