APPLICATION AND EVALUATION OF THE PHAGE RESISTANCE-ENCODING AND BACTERIOCIN-ENCODING PLASMID PMRC01 FOR THE IMPROVEMENT OF DAIRY STARTER CULTURES

The conjugative 63-kb lactococcal plasmid pMRC01 encodes bacteriophage resistance and production of and immunity to a novel broad-spectrum b acteriocin, designated lacticin 3147 (M. P. Ryan, M. C. Rea, C. Hill, and R. P. Ross, Appl. Environ. Microbiol, 62:612-619, 1996), The phage resistance is an abortive infection mechanism which targets the phage -lytic cycle at a point after phage DNA replication. By using the gene tic determinants for bacteriocin immunity encoded on the plasmid as a selectable marker, pMRC01 was transferred into a variety of lactococca l starter cultures to improve their phage resistance properties, Selec tion of resulting transconjugants was performed directly on solid medi a containing the bacteriocin. Since the starters exhibited no spontane ous resistance to the bacteriocin as a selective agent, this allowed t he assessment of the transfer of the naturally occurring plasmid into a range of dairy starter cultures, Results demonstrate that efficient transfer of the plasmid was dependent on the particular recipient stra in chosen, and while high-frequency transfer (10(-3) per donor) of the entire plasmid to some strains was observed, the plasmid could not be conjugated into a number of starters, In this study, transconjugants for a number of lactococcal starter cultures which are phage resistant and bacteriocin producing have been generated, This bacteriocin-produ cing phenotype allows for control of nonstarter flora in food fermenta tions, and the phage resistance property protects the starter cultures in industry. The 63-kb plasmid was also successfully transferred into Lactococcus lactis MG1614 cells via electroporation.