Assessment of vancomycin resistance transfer among enterococci of clinical importance in milk matrix

Dissemination of vancomycin resistance in enterococci has been associated with horizontal transfer of mobile genetic elements. Aim of the study was to evaluate if milk matrix is a suitable environment to support transferability of vancomycin resistance (vanA) gene from clinical vancomycin-resistant Enterococcus faecium to vancomycin-sensitive Enterococcus faecalis. Enterococci strains were firstly screened for the presence of cpd (inducible sex pheromone determinant) gene, vanA and tetL genes (vancomycin and tetracycline resistance markers, respectively) and the gelE (extracellular metalloendopeptidase) gene to define the mating pairs. Based on these selection markers, we investigated the transferability of eight plasmid-borne vanA harbored by E. faecium (vanA+, cpd-, tetL- and gelE-) into two E. faecalis (vanA-, cpd+, tetL + and gelE+) recipient strains in milk matrix. The strains were mated in a 1:1 ratio in 7% reconstituted milk and incubated at 37 °C. Transconjugants emerged from all 16 matings within 2 h of incubation and were evidenced by dual antibiotic resistance (vancomycin and tetracycline). The vancomycin-resistance of trasconjugants was maintained even after ten subsequent passages on nonselective medium. Transconjugants were positive for vanA, tetL and gelE genes. This study indicates milk matrix as suitable environment to support gene exchange between Enterococcus species.