In the food chain, foods with high moisture, such as some cheeses, are the main route of antimicrobial resistance dissemination between different bacteria. Listeria monocytogenes is an important agent of foodborne diseases that can display antimicrobial resistance, while Enterococci are frequently found in cheeses and can receive resistance genes from other bacteria. In this study, Minas Frescal cheese, was used as model system to evaluate horizontal transfer of the tetM gene from L. monocytogenes to Enterococcus faecalis. The analysis confirmed that both donor and recipient strains were maintained in the food model with similar growth kinetics, during the shelf life of 21 days with storage at 4 °C. Transconjugant cells were detected on the second day, and were increasing during the 21 days evaluated. Our results suggest that the Minas Frescal cheese provides a suitable environment for horizontal transfer of conjugative plasmids from foodborne L. monocytogenes to E. faecalis.