sacA and nisA genes are not always linked in Lactococcus lactis subsp. lactis strains (original) (raw)
Related papers
Novel sucrose transposons from plant strains of Lactococcus lactis
FEMS Microbiology Letters, 2000
Lactococcus lactis strains isolated from vegetable products transferred the ability to ferment sucrose in conjugation experiments with the recipient strain L. lactis MG1614. Nisin production and sucrose fermentation were transferred together from two strains, but transfer also occurred from several other strains which did not produce nisin. Pulsed-field gel electrophoresis analysis showed that all transconjugants had acquired large chromosomal insertions at two main sites. Nisin^sucrose transconjugants had gained inserts of 70 kb, while those that fermented sucrose without nisin production contained inserts of between 50 and 110 kb. Transconjugants from one donor had acquired a separate insertion of 55 kb which correlated with enhanced bacteriophage resistance, but contained neither nisin nor sucrose fermentation genes. ß
Fems Microbiology Letters, 1996
Sixty-four lactococcal strains isolated from natural whey starters were screened for the presence of the nisin structural gene by polymerase chain reaction. Seven of them showed a specific PCR product of 320 bp; only two produced antagonistic activity and were resistant to nisin. Southern blots of SnzaI-digested DNA from PCR-positive strains hybridized with a nisA probe displayed a location of the gene on different SmaI fragments. Among PCR-positive strains. nisin producers showed specific transcript after reverse transcriptase-PCR, as well as some non-nisin-producing strains. The RT-PCR product could not be shown in one non-nisin-producing PCR-positive strain.
Genes & Genomics, 2011
Nisin is an antimicrobial peptide produced by Lactococcus lactis. It has a long history of safe use, mainly in food production. This bacteriocin has been studied from many aspects of genetics, biosynthesis, immunity, regulation and mode of action. The strain Lac. lactis M78 has already been described in previous studies as a good nisin A producer with equally good potential to be used in food production. The main objective of the present study was to determine the complete nucleic acid sequence of the nisin A gene cluster from this strain. This is the first time that all 11 genes that form the nisin A gene cluster were sequenced. The obtained sequence (GenBank: HM219853) was compared to other known nucleic acid sequences of bacteriocin nisin. The results of the comparison showed certain differences in sequences that might influence the structure and function of proteins involved in nisin biosynthesis, immunity and regulation.
Gene-Specific PCR Amplification of Technologically Important Lactococcal Genes
Biotechnology & Biotechnological Equipment, 2012
A set of gene-specific PCR techniques were used to characterize lactococcal cultures from the LBB collection. Initially the species identification of lactococci was confirmed by targeting the glutamate decarboxylase gene (gadB). PCR amplification of the genes for membrane proteinase (prtP) and citrate permease (citP) was used to select strains which grow rapidly in milk and/or ferment citrate. The prtP + and citP + phenotype was confirmed using differentiating microbiological media. It was proved that the citP + phenotype was always associated with strains of L. lactis ssp. lactis biovar. diacetylactis-an important aroma forming variety of dairy lactococci. In all diacetylactis strains the citP gene was localized on a 8.2 kbp plasmid. The lactococcal cultures were also tested for the presence of nisin synthesis (nisA/Z) genes. Potentially lysogenic lactococcal cultures which carry in their genome the gene of prophage integrase (int) were detected. From the tested strains nearly one third were int-positive. The thermal induction of prophage activity was demonstrated for the int + strain L. lactis ssp. lactis LBB.C1/6.
Transfer of Sucrose-Fermenting Ability and Nisin Production Phenotype among Lactic Streptococci
Applied and environmental microbiology, 1985
Transfer of sucrose fermentation ability, nisin production, and nisin resistance from Streptococcus lactis to S. lactis and Streptococcus lactis subsp. diacetylactis occurred between cells immobilized on nitrocellulose filters in the presence of DNase. Transconjugants were able to act as donors to transfer the Suc-Nis phenotype in subsequent mating. No changes in sensitivity to lytic phage c2 were noted in S. lactis transconjugants. However, temperature-independent restriction of lytic phage 18-16 was noted in transconjugants of S. lactis subsp. diacetylactis 18-16. Adsorption studies with phage-resistant transconjugants showed that resistance was not due to lack of adsorption by the lytic phage. Physical evidence for the presence of introduced plasmid DNA was not found in lysates of transconjugants.
Applied and Environmental Microbiology
Conjugation was used to construct nisin-producing Lactococcus lactis subsp. cremoris strains. Recipients were obtained by electroporation of L. lactis subsp. cremoris strains with the drug resistance plasmid pGK13 or pGB301. A method, direct-plate conjugation, was developed in which donor and recipient cells were concentrated and then combined directly on selective media. This method facilitated transfer of the nisin-sucrose (Nip' Suc+) phenotype from the donor strain, L. lactis subsp. lactis 11454, to three L. lactis subsp. cremoris recipient strains. Nip' Suc+ L. lactis subsp. cremoris transconjugants were obtained at frequencies which ranged from 10-7 to 10-8 per donor CFU. DNA-DNA hybridization to transconjugant DNAs, performed with an oligonucleotide probe synthesized to detect the nisin precursor gene, showed that this gene was transferred during conjugation but was not associated with detectable plasmid DNA. Further investigation indicated that L. lactis subsp. cremoris Nip' Suc+ transconjugants retained the recipient strain phenotype with respect to bacteriophage resistance and acid production in milk. Results suggested that it would be feasible to construct nisin-producing L. lactis subsp. cremoris strains for application as mixed and multiple starter systems. Additionally, the direct-plate conjugation method required less time than ifiter or milk agar matings and may also be useful for investigations of conjugal mechanisms in these organisms.
Journal of Applied Microbiology, 2009
Aims: This paper describes optimization of electrotransformation of Mu transposition complexes into Lactococcus lactis cells and identification of genes affecting nisin production. Methods and Results: The highest transformation efficiency, 1AE1 • 10 2 transformants lg)1 of input transposon DNA, was achieved when cells were grown to an OD 600 of 0AE5 in the presence of 1AE5% of glycine and treated with 20 lg ml)1 ampicillin for 60 min. Three insertions affecting nisin production, which were identified at nisB, fhuR, and rpiA genes, were screened from a library of 2000 erythromycin-resistant transformants using a nisin bioassay method. NisB is part of the nisin biosynthetic machinery, explaining the loss of nisin production in nisB mutant. FhuR is a transcription regulator involved in sulphur acquisition. Inactivation of fhuR presumably results in a low cellular cystein level, which affects nisin biosynthesis that involves utilization of cystein. RpiA is involved in pentose phosphate pathway and carbon fixation. The rpiA mutant showed reduction in nisin production and slow growth rate. Conclusions: The results showed that Mu transposition complex mutagenesis can be used to identify genes in L. lactis. Three genes involved in nisin production were identified. Significance and Impact of the Study: Expanding the Mu transposition-based mutagenesis to Lactococci adds a new tool for studies of industrially important bacteria.
Biotechnology Letters, 2001
A Lactococcus strain with strong antimicrobial activity was isolated from raw milk Manchego cheese during a survey on the production of bacteriocins by lactic acid bacteria present in raw milk cheeses. It was identified as Lactococcus lactis subsp. lactis, phenotypically by its morphological and physiological characteristics and genotypically by a PCR technique. When tested for tolerance to known bacteriocins produced by lactococci, it was shown to be resistant to nisin A and nisin Z. The presence of genes encoding nisin and lacticin 481 was revealed by PCR techniques with specific probes. Sequences of the respective PCR amplified fragments matched sequences reported for nisin Z and lacticin 481.