Phage Resistance in a Phage-Insensitive Strain of Streptococcus lactis : Temperature-Dependent Phage Development and Host-Controlled Phage Replication

Abstract

Streptococcus lactis ME2 is a dairy starter strain that is insensitive to a variety of phage, including 418. The efficiency of plating of 418 on ME2 and Ni could be increased from <1 x 10-9 to 5.0 x 10-2 and from 7.6 x 10-7 to 2.1 x 10-2, respectively, when the host strains were subcultured at 40°C before plating the phage and the phage assay plates were incubated at 40°C. Host-dependent replication was demonstrated in Ni at 30°C and in Ni and ME2 at 40°C, suggesting the operation of a temperature-sensitive restriction and modification system in ME2 and Ni. The increased sensitivity of ME2 and Ni to 418 at 40°C was also demonstrated by lysis of broth cultures and increased plaque size. ME2 grown at 40°C showed an increased ability to adsorb 418, indicating a second target for temperature-dependent phage sensitivity in ME2. Challenge of Ni with a 418 preparation that had been previously modified for growth on Ni indicated that at 40°C phage development was characterized by a shorter latent period and larger burst size than at 30°C. The evidence presented suggests that the high degree of phage insensitivity expressed by ME2 consists of a variety of temperature-sensitive mechanisms, including (i) the prevention of phage adsorption, (ii) hostcontrolled restriction of phage, and (iii) suppression of phage development. At 30°C these factors appear to act cooperatively to prevent the successful emergence of lytic phage active against S. lactis ME2.

Plasmid-directed mechanisms for bacteriophage defense in lactic streptococci

“…These mechanisms include plasmid-induced interference with phage adsorption, restriction and modification of phage, and a heat-sensitive mechanism that suppresses the burst size of infecting phage (Hsp +). The mechanisms acted cooperatively to confer a phage-insensitive state to S. lactis ME2 [36]. However, similar to pNP40, phage resistance at each of the three levels was inoperative following growth of the culture at 40°C.…”

“…In the prototype phage-insensitive strain, S. lactis ME2 [28,36], curing of pME0030 resulted in increased phage adsorption of four heterologous phages. In one case, the degree of phage adsorption correlated with the plaquing efficiencies for phage 18 on S. lactis ME2 (EOP < 10 -9 at 40% adsorption) and the pME0030-cured variant, N1 (EOP = 10 -7 at 99.7% adsorption).…”

“…Detailed phenotypic and genetic examination of the prototype phage-insensitive strain, S. lactis ME2, revealed that at least three independent mechanisms operate to confer resistance to virulent phage [28][29][30]36]. These mechanisms include plasmid-induced interference with phage adsorption, restriction and modification of phage, and a heat-sensitive mechanism that suppresses the burst size of infecting phage (Hsp +).…”

“…It has been suggested that collective expression of different mechanisms of phage defense is the basis of the phage-insensitive state in certain lactic streptococci [2,36]. In this light, genetic programs for development of phage-insensitive strains should emphasize gene and plasmid combinations that provide expression of multiple mechanisms of phage defense.…”

Bacteriophage and bacteriophage resistance in lactic acid bacteria

“…Watanabe et al [76] have suggested that two types of adsorption can exist; irreversible adsorption which can lead to infection, and reversible adsorption, which does not. It is impossible to distinguish between the two forms of adsorption on the basis of standard adsorption assays, it may 94 be that a new definition is required for insensitivity due to the failure of phage to initiate an infection by injecting its DNA after adsorption. "Injection blocking' is one possible phenotypic designation.…”