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TheabiA gene (formerlyhsp ) encodes an abortive phage infection mechanism which inhibits phage DNA replication. To analyze the effects of varying theabiA gene dosage on bacteriophage resistance inLactococcus lactis , various genetic constructions were made. An IS946 -based integration vector, pTRK75, was used to integrate a single copy ofabiA into the chromosomes of two lactococcal strains, MG1363 and NCK203. In both strains, a single copy ofabiA did not confer any significant phage resistance on the host except for one of the MG1363 integrants, NCK625, which exhibited a slightly higher level of resistance to phages sk1 and p2. Hybridization of the total cellular RNA from NCK625 to anabiA -specific probe indicated that the integration took place downstream of a promoter causing stronger expression ofabiA in this integrant. ThreeabiA -containing plasmids of various copy numbers were introduced into both strains, and the recombinants were evaluated for resistance to phages c2, p2, sk1, and φ31. Plasmid pTRK18 has a copy number of approximately six (cn = 6) and caused a decreased plaque size for all phages evaluated. Integration of pTRK75 into a native plasmid of NCK203 generated pTRK362 (cn = 13), which caused a reduced efficiency of plaquing (EOP = 10-2 ) and reduced plaque size. A high-copy-numberabiA plasmid (pTRK363), based on the pAMβ1 origin of replication, was also constructed (cn = 100). Plasmid pTRK363 caused a significant reduction in EOP (10-4 to 10-8 ) and plaque size for all phages tested, although in some cases, this plasmid caused the evolution of AbiA-resistant phage derivatives. Altering the gene dosage or expression level ofabiA significantly affects the phage resistance levels.

applied and environmental microbiology

Phenotypic Consequences of Altering the Copy Number of <i>abiA</i> , a Gene Responsible for Aborting Bacteriophage Infections in <i>Lactococcus lactis</i>

Phenotypic Consequences of Altering the Copy Number of abiA , a Gene Responsible for Aborting Bacteriophage Infections in Lactococcus lactis