Abstract
A specific method to identify nisin-producing strains was developed based on Nisin-Controlled gene Expression (NICE) vector pSec:Nuc. The plasmid pSec:Nuc was transformed into non-nisin-producing strain Lactococcus lactis NZ9000, a host commonly used for the NICE system. The generating strain L. lactis NZ9000/pSec:Nuc could sense extracellular inducer nisin and efficiently secrete a reporter protein Nuc, the staphylococcal nuclease (Nuc) into the medium. Instead of using purified nisin, the culture supernatants of nisin-producing strains were also used as inducers. Therefore, the NICE system could be used to identify nisin-producing strains. With this principle, 4 among 56 lactococci strains isolated from raw milk were identified as nisin producers. The results were further confirmed by polymerase chain reaction amplification with their genomic DNA as templates, and nucleotide sequencing revealed that three of them produced nisin A, and the others produced nisin Z. Those results made it possible to isolate and identify nisin-producing strains specifically and rapidly using NICE system.
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References
Klaenhammer TR (1993) Genetics of bacteriocins produced by lactic acid bacteria. FEMS Microbiol Rev 12:39–85
Nes IF, Diep DB, Håvarstein LS, Brurberg MB, Eijsink V, Holo H (1996) Biosynthesis of bacteriocins in lactic acid bacteria. Antonie Van Leeuwenhoek 70:113–128
Tagg JR, Dajani AS, Wannamaker LW (1976) Bacteriocins of gram-positive bacteria. Bacteriol Rev 40:722–756
Gross E, Morell JL (1971) The structure of nisin. J Am Chem Soc 93:4634–4635
Mulders JW, Boerrigter IJ, Rollema HS, Siezen RJ, de Vos WM (1991) Identification and characterization of the lantibiotic nisin Z, a natural nisin variant. Eur J Biochem 201:581–584
Zendo T, Fukao M, Ueda K, Higuchi T, Nakayama J, Sonomoto K (2003) Identification of the lantibiotic nisin Q, a new natural nisin variant produced by Lactococcus lactis 61–14 isolated from a river in Japan. Biosci Biotechnol Biochem 67:1616–1619
Wirawan RE, Klesse NA, Jack RW, Tagg JR (2006) Molecular and genetic characterization of a novel nisin variant produced by Streptococcus uberis. Appl Environ Microbiol 72:1148–1156
Delves-Broughton J, Blackburn P, Evans RJ, Hugenholtz J (1996) Applications of the bacteriocin, nisin. Antonie Van Leeuwenhoek 69:193–202
Cheigh CI, Park H, Choi HJ, Pyun YR (2005) Enhanced nisin production by increasing genes involved in nisin Z biosynthesis in Lactococcus lactis subsp. lactis A164. Biotechnol Lett 27:155–160
Rollema HS, Kuipers OP, Both P, de Vos WM, Siezen RJ (1995) Improvement of solubility and stability of the antimicrobial peptide nisin by protein engineering. Appl Environ Microbiol 61:2873–2878
Beasley SS, Saris PE (2004) Nisin-producing Lactococcus lactis strains isolated from human milk. Appl Environ Microbiol 70:5051–5053
Cai Y, Ng LK, Farber JM (1997) Isolation and characterization of nisin-producing Lactococcus lactis subsp. lactis from bean-sprouts. J Appl Microbiol 83:499–507
Noonpakdee W, Santivarangkna C, Jumriangrit P, Sonomoto K, Panyim S (2003) Isolation of nisin-producing Lactococcus lactis WNC 20 strain from nham, a traditional Thai fermented sausage. Int J Food Microbiol 81:137–145
Tramer J, Fowler JJ (1964) Estimation of nisin in foods. J Sci Food Agric 15:522–528
Wolf CE, Gibbons WR (1996) Improved method for quantification of the bacteriocin nisin. J Appl Bacteriol 80:453–457
Martínez MI, Rodríguez E, Medina M, Hernández PE, Rodríguez JM (1998) Detection of specific bacteriocin-producing lactic acid bacteria by colony hybridization. J Appl Microbiol 84:1099–1103
Sonia G, Rodríguez E, Gaya P, Medina M, Nuñez M (2001) PCR detection of the structural genes of nisin Z and lacticin 481 in Lactococcus lactis subsp. lactis INIA 415, a strain isolated from raw milk Manchego cheese. Biotechnol Lett 23:85–89
de Ruyter PG, Kuipers OP, Beerthuyzen MM, Alen-Boerrigter I, de Vos WM (1996) Functional analysis of promoters in the nisin gene cluster of Lactococcus lactis. J Bacteriol 178:3434–3439
Ribeiro LA, Azevedo V, Le Loir Y, Oliveira SC, Dieye Y, Piard JC, Gruss A, Langella P (2002) Production and targeting of the Brucella abortus antigen L7/L12 in Lactococcus lactis: a first step towards food-grade live vaccines against brucellosis. Appl Environ Microbiol 68:910–916
Kuipers OP, Beerthuyzen MM, Siezen RJ, de Vos WM (1993) Characterization of the nisin gene cluster nisABTCIPR of Lactococcus lactis. Requirement of expression of the nisA and nisI genes for development of immunity. Eur J Biochem 216:281–291
Kuipers OP, de Ruyter PGGA, Kleerebezem M, de Vos WM (1998) Quorum sensing-controlled gene expression in lactic acid bacteria. J Biotechnol 64(7):15–21
Holo H, Nes IF (1989) High-frequency transformation, by electroporation, of Lactococcus lactis subsp. cremoris grown with glycine in osmotically stabilized media. Appl Environ Microbiol 55:3119–3123
Engelke G, Gutowski-Eckel Z, Hammelmann M, Entian KD (1992) Biosynthesis of the lantibiotic nisin: genomic organization and membrane localization of the NisB protein. Appl Environ Microbiol 58:3730–3743
Rodríguez JM, Cintas LM, Casaus P, Horn N, Dodd HM, Hernández PE, Gasson MJ (1995) Isolation of nisin-producing Lactococcus lactis strains from dry fermented sausages. J Appl Bacteriol 78:109–115
Mitra S, Chakrabartty PK, Biswas SR (2007) Production of nisin Z by Lactococcus lactis isolated from Dahi. Appl Biochem Biotechnol 143:41–53
Kuipers OP, Beerthuyzen MM, de Ruyter PGGA, Luesink EJ, de Vos WM (1995) Autoregulation of nisin biosynthesis in Lactococcus lactis by signal transduction. J Biol Chem 270:27299–27304
Mierau I, Kleerebezem M (2005) 10 years of the nisin-controlled gene expression system (NICE) in Lactococcus lactis. Appl Microbiol Biotechnol 68:705–717
Park SH, Itoh K, Kikuchi E, Niwa H, Fujisawa T (2003) Identification and characteristics of nisin Z-producing Lactococcus lactis subsp. lactis isolated from Kimchi. Curr Microbiol 46:385–388
Acknowledgements
We thank Mr N. Galleron (Institut National de la Recherche Agronomique, France) to kindly gave us the plasmid pSec:Nuc and Lactococcus lactis NZ9000. This work was supported by the National Science Foundation of China (NSFC project number 30570043) and 863 Hi-Tech Research and Development Program of China (project number 2006AA10Z344, 2006AA10Z321).
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Hu, S., Kong, J., Kong, W. et al. Identification of Nisin-Producing Strains by Nisin-Controlled Gene Expression System. Curr Microbiol 58, 604–608 (2009). https://doi.org/10.1007/s00284-009-9378-y
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DOI: https://doi.org/10.1007/s00284-009-9378-y