Partial characterization of the genetic basis for sucrose metabolism and nisin production in Streptococcus lactis

Abstract

We attempted to identify the genetic loci for sucrose-fermenting ability (Suc+), nisin-producing ability (Nip'), and nisin resistance (Nisr) in certain strains of Streptococcus lactis. To obtain genetic evidence linking the Suc+ Nip' Nisr phenotype to a distinct plasmid, both conjugal transferand transformation were attempted. A conjugation procedure modified to protect the recipients against the inhibitory action of nisin allowed the conjugal transfer of the Suc" Nip+ Nisr marker from three Suc+ Nip' Nisr donors to various recipients. The frequency of transfer ranged from 1.7 x 10-4 to 5.6 x 10-8 per input donor, depending on the mating pair. However, no additional plasmid DNA was apparent in these transconjugants. Transformation of S. lactis LM0230 to the Suc+ Nip' Nisr phenotype by using the plasmid pool of S. lactis ATCC 11454 was not'achieved, even though other plasmids present in the pool were successfully transferred. Howevet, two results imply the involvement of plasmid DNA in coding for the Suc+ Nip' Nisr phenotype. The Suc+ Nip' Nisr marker was capable of conjugal transfer to a recipient deficient in host-mediated homologous recombination (Rec-), and the Suc+ Nip' Nisr marker exhibited bilateral plasmid incompatibility with a number of lactose plasmids found in S. lactis. Although our results indicate that the Suc+ Nip' Nisr phenotype is plasmid encoded, no physical evidence linking this phenotype to a distinct plasmid was obtained.

Genetics of bacteriocins produced by lactic acid bacteria

“…The close genetic linkage of Nip + Imm ~ Suc has been observed consistently in conjugal matings where transfer of Suc + is accompanied by cotransfer of Nip ~ Imm + [96,97,99,103]. The transconjugants did not acquire new plasmids and the phenotypes were genetically stable, even in cases where an unstable Nis + Imm ~ Suc + plasmid was implicated in the conjugal donor [96][97][98].…”

“…It was recognized in 1974 [95] that Nip + and Imm + were genetically unstable in Lactococcus prompting speculation that nisin biosynthesis was plasmid encoded. Genetic instability of Nip + Imm + was also accompanied by irreversible loss of Suc +, indicating that the three phenotypes were genetically linked [96][97][98][99]. Such genetic behavior in lactococci is highly characteristic of plasmid-encoded functions.…”

“…Nip ~ Imm + can be conjugally transferrcd within Lactococcus species using a variety of different mating techniques that minimize the lethal effects of the Nip + donor against a nisinsensitive recipient (reviewed in [7]). The approaches have included the following: (i) conducting matings in the presence of ~-chymotrypsin [99]; (ii) washing donor cells to remove residual nisin immediately before mixing with recipicm cells on membranes or directly on agar [96,103]; and (iii) donor cells arc mated during log-phasc, when little nisin is produccd [97].…”

“…However, this event was not confirmed in later attempts to repeat the conjugation experiment and the original Nip + Imm* Leuconostoc transconjugants were lost (Sandine, personal communication). It is significant in the studies of Steele and McKay [99] and Broadbent and Kondo [1(/3] that Nip + Imm + Suc + could bc transfcrred into L. lactis subsp, cremoris strains, where their fermentative characteristics and phage resistance are oftcn more acceptable for industrial starter cultures. 52 The improved frequencies of Nip' transfer through the direct plating method [103] may provide expanded opportunities to create nisin-producing starters for applications in the dairy industry.…”

Looking backward and forward at the practical applications of genetic researches on lactic acid bacteria

“…However, no plasmid DNA was found in the transconjugates, a situation also noted by Gasson [132]. Steel and McKay [135] also were unable to obtain physical evidence that nisin production was plasmid linked. For diplococcin production, the data are unambiguous; Davey showed that this activity could be conjugally transferred from S. cremoris 346 donor to two plasmid-free S. cremoris recipients at a frequency of 10 -1 per donor.…”