CHARACTERISTICS OF TN5307 EXCHANGE AND INTERGENERIC TRANSFER OF GENESASSOCIATED WITH NISIN PRODUCTION

Transfer of the Lactococcus lactis 11454 nisin-sucrose conjugative tra nsposon, Tn5307,was investigated to develop a methodology for conjugat ion of this element to other lactic acid bacteria. Tn5307 ex change wa s sensitive to temperature and pH but was not affected by protease or amylase treatments to donor cells. Moreover, conjugation studies demon strated that the direct-plate method could be employed to rapidly iden tify LM2301 transconjugants able to transfer Tn5307 at least ten times more efficiently than 11454. Intergeneric transfer of nisin and sucro se genes between L. lactis and a dairy Enterococcus sp. was also inves tigated. Erythromycin-resistant Enter ococcus sp. recipients were deve loped by electro-transformation with pGK13 or by conjugal introduction of the broad-host-range plasmid pAM beta 1. Matings between L. lactis 11454 and an Enter ococcus sp. recipient that contained pAM beta 1 yi elded sucrose-positive, nisin-immune transconjugants at a frequency of 2.3 x 10(-9) transconjugants per donor cfu. Agar-overlay assays for n isin production revealed that enterococcal transconjugants did not pro duce the bacteriocin, but DNA . DNA hybridization with a nisA-specific probe demonstrated that these bacteria had acquired the nisin structu ral gene.