Characterization of the Tn916 conjugative transposon in a food borne strain of Lactobacillus paracasei

Food-borne antibiotic-resistant lactic acid bacteria have received growing attention in the past few years. We have recently identified tetracycline-resistant Lactobacillus paracasei in samples of milk and natural whey starter cultures employed in the manufacturing process of a typical Italian fermented dairy product, Mozzarella di Bufala Campana. In the present study, we have characterized at the molecular level the genetic context of tetracycline resistance determinants in these natural strains, which we have identified as tet(M). This gene was present in 21 independent isolates, whose fingerprinting profiles were distributed into eight different repetitive extragenic palindromic groups by cluster analysis. We provide evidence that the gene is associated with the broad-host, conjugative transposon Tn916, which had never before been described to occur in L. paracasei. PCR analysis of four independent isolates by use of specifically designed primer pairs detected the presence of a circular intermediate form of the transposon, carrying a coupling sequence (GGCAAA) located between the two termini of Tn916. This novel coupling sequence conferred low conjugation frequency in mating experiments with the recipient strain JH2-2 of Enterococcus faecalis.