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

1994; American Society for Microbiology; Volume: 60; Issue: 4 Linguagem: Inglês

10.1128/aem.60.4.1129-1136.1994

1098-5336

Polly K. Dinsmore, Todd R. Klaenhammer,

Bacterial Genetics and Biotechnology

The abiA gene (formerly hsp ) encodes an abortive phage infection mechanism which inhibits phage DNA replication. To analyze the effects of varying the abiA gene dosage on bacteriophage resistance in Lactococcus lactis , various genetic constructions were made. An IS 946 -based integration vector, pTRK75, was used to integrate a single copy of abiA into the chromosomes of two lactococcal strains, MG1363 and NCK203. In both strains, a single copy of abiA 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 an abiA -specific probe indicated that the integration took place downstream of a promoter causing stronger expression of abiA in this integrant. Three abiA -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-number abiA 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 of abiA significantly affects the phage resistance levels.