Molecular Characterization of Three Small Isometric-Headed Bacteriophages Which Vary in Their Sensitivity to the Lactococcal Phage Resistance Plasmid pTR2030

Abstract

Lactococcus lactis LMA12-4 is a pTR2030 transconjugant that has been used as an industrial starter culture because of its resistance to phages predominant in cheese plants. Plasmid pTR2030 interferes with susceptible phages in this host strain via two mechanisms, restriction and modification (R/M) and abortive infection (Hsp). After prolonged use of LMA12-4 transconjugants in the industry, two different bacteriophages, designated nck202.phi48 (phi48) and nck202.phi50 (phi50), were isolated which could produce plaques on LMA12-4 containing pTR2030. In this study, these two phages were characterized and compared with a third phage, nck202.phi31 (phi31), which is susceptible to both the R/M and Hsp activities encoded by pTR2030. Phage phi48 was not susceptible to inhibition by Hsp, whereas phi50 was unaffected by either the R/M or Hsp mechanisms. All three were small isometric-headed phages, but small differences were noted between the phages in the structural details of the tail base plate, susceptibility to chloroform treatment, and requirements for calcium infectivity. The phage genomes were all between 29.9 and 31.9 kb in length. Phages phi31 and phi48 harbored cohesive ends, whereas the phage phi50 genome was circularly permuted, terminally redundant, and carried a putative packaging initiation site. DNA-DNA hybridization experiments conducted between the phages revealed a common region in phi48 and phi50 that may correlate with the resistance of the two phages to the Hsp-abortive infection induced by pTR2030. Phage phi50 also harbored DNA sequences that shared homology to pTR2030 in the region where R/M activities have been localized on the plasmid. Molecular characterization of the three phages localized regions within the genomes of the pTR2030-resistant phages that may be responsible for circumventing plasmid-encoded Hsp and R/M defense mechanisms in lactococci.