ÇİĞDEM SEZER, Abamüslüm GÜVEN

Investigation of bacteriocin production capability of lactic acid bacteria ısolated from foods

Bu çalışmada, gıdalardan bakteriyosinojenik laktik asit bakterilerinin (LAB) izolasyonu amaçlanmıştır. Gıdalardan izole edilen toplam 12700 LAB’nin antimikrobiyel etkisi test edilmiştir. Bu izolatlardan sadece 601 adedi çalışmada kullanılan indikatör suşlara karşı antimikrobiyel etki göstermiştir. LAB’nin ürettiği laktik asitin ve hidrojen peroksitin güçlü antimikrobiyel etkisi göz önüne alındığında bu etkinin ortadan kaldırılması için katalaz ilavesiyle birlikte ortam nötrlendiğinde antimikrobiyel etkisini sürdüren sadece 35 izolat olduğu belirlenmiştir. İzolatların etkisi proteolitik enzimlerin varlığında kaybolmuştur. Üretilen antimikrobiyel etkili metabolitlerin gıda üretim ve muhafaza sıcaklıklarında ve geniş pH (pH 2-12) aralığında aktivitelerini sürdürdükleri gözlenmiştir. Kaşar peynirinden izole edilen Lactococcus lactis ssp. lactis suşunun ürettiği bakteriyosin kısmi olarak saflaştırılarak tanımlanmaya çalışılmıştır. İzole edilen bakteriyosinojenik suşların Gram-negatif ve Gram-pozitif indikator mikroorganizmalara karşı oldukça güçlü antimikrobiyel etkiye sahip olduğu belirlenmiştir. Bakteriyosinojenik LAB ve/veya onların bakteriyosinleri gıda kaynaklı patojenlerden özellikle Listeria monocytogenes’in inhibisyonu için doğal bir katkı maddesi olarak kullanılabilir.

Anahtar Kelimeler:

katalaz, antimikrobiyal özellikler, bakteriyosin, laktik asit bakterileri, Listeria monocytogenes

Gıdalardan izole edilen laktik asit bakterilerilerinin bakteriyosin üretme yeteneklerinin araştırılması

The aim of this study was to isolate bacteriocin-producing lactic acid bacteria (LAB) from foods. A total of 12700 LAB were isolated from foods and screened for their bacteriocin production. Among them, 601 isolate showed a clear antimicrobial activity against indicator strains. After neutralization 35 of the cultures retained active. The antimicrobial activities of strains were lost after treatment with proteases. The antimicrobial metabolites were heat stable and were active pH range of 2-12. The bacteriocin which produced by Lactococcus lactis ssp. lactis isolated from kashar samples was partially purified by ammonium sulphate precipitation, dialysis, following ion-exchange chromatography. It was also found that this bacteriocin was cationic. The apparent molecular mass of partially purified bacteriocin, as indicated by activity detection after Tricine-SDS-PAGE, was 2.4 kDa, 4 kDa or 15 kDa. In this study antimicrobial substance of LAB from food showed strong antibacterial activity against Gram-negative and Gram-positive indicator microorganisms. Those were also found that these bacteriocins were stable at temperatures used in food manufacturing and storage. Bacteriosinogenic LAB or bacteriocins might be useful as a natural preservative for inhibiting food-borne pathogens in particular Listeria monocytogenes in foods.

Keywords:

catalase, antimicrobial properties, bacteriocins, lactic acid bacteria, Listeria monocytogenes,

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Daeschel MA: Antimicrobial substance from lactic acid bacteria for use as food preservatives. Food Technol, 43, 164-166, 1989.
Jack RW, Tagg JR, Ray B: Bacteriocins of Gram-positive bacteria. Microbiol Rev, 59 (2): 171-200, 1995.
Holzapfel WH, Geisen R, Schillinger U: Biological preservation of foods with reference to protective cultures, bacteriocins and food-grade enzymes. Review Int J Food Microbiol, 24 (3): 343-62, 1995.
Abee T, Krockel L, Hill C: Bacteriocins: modes of action and potentials in food preservation and control of food poisoning. Int J Food Microbiol, 28, 169-185, 1995. Marshall WM, Cole WM: Methods for making kefir and fermented milks based on kefir. J Dairy Res, 52, 451-456,1985.
Vanos V, Cox L: Rapid routine method for the detection of lactic acid bacteria among competitive flora. Food Microbiol, 3, 223-234, 1986.
Kandler O, Weiss N: Regular, Nonsporing Gram positive rods. Genus Lactobacillus. In, Sneath PHA, Mair NS, Sharpe ME, Holt JG (Eds): Bergey’s Manual of Systematic Bacteriology. Vol. 2, Williams and Wilkins, Baltimore M D, 1208-1234, 1986.
Schillinger U, Lücke FK: Antibacterial activity of Lactobacillus sake isolated from meat. Appl Environ Microbiol, 55 (8): 1901-1906, 1989.
Lewus CB, Montville TJ: Detection of bacteriocins produced by lactic acid bacteria. J Microbiol Methods, 13, 145-150, 1991.
Campos CA, Rodriquez O, Calo-Mata P, Prado M, Barros-Velazquez J: Preliminary characterization of bacteriocins from Lactococcus lactis, Enterococcus faecium and Enterococcus mundtii strains isolated from turbot. Food Res Intern, 39 (3): 356-364, 2006.
Vignolo GM, Suriani F, Holgado APR, Oliver G: Antibacterial activity of Lactobacillus strains isolated from dry fermented sausages. J Appl Bacteriol, 75, 34-349, 1993.
Bromberg R, Moreno I, Zaganini CL, Delboni RR, Olivera J: Isolation of bacteriocin-producing lactic acid bacteria from meat and meat products and its spectrum of inhibitory activity. Braz J Microbiol, 35, 137-144, 2004.
Deraz SF, Karlsson EN, Hedstrom M, Andersson MM, Mattiasson B: Purification and characterisation of acidocin D20079, a bacteriocin produced by Lactobacillus acidophilus DSM 20079. J Biotechnol, 117 (4): 343-354, 2005.
Equchi T, Kamınaka K, Shıma J, Kawamoto S, Mori K, Choi SH, Doi K, Ohmomo S, Ogata S: Isolation and
characterization of enterocin SE-K4 produced by thermophilic enterococci, Enterococcus faecalis K–4. Biosci Biotechnol Biochem, 65 (2): 247-253, 2001.
Schillinger U, Lücke FK: Identification of lactobacilli from meat and meat products. Food Microbiol, 4,199-208, 1987.
Carr FJ, Chill D, Miada N: The lactic acid bacteria: A literature survey. Crit Rev Microbiol, 28 (4): 281-370, 2002.
Hammes WP, Weiss N, Holzapfel W: The Genera Lactobacillus and Carnobacterium. In, Balows A, Truper HG, Dworkin M, Harder W, Schleifer KH (Eds): The Prokaryotes. 2nd ed., Springer-Verlag, New York, Vol.2, pp.1535-1594, 1992.
Budde BB, Hornbaek T, Jacobsen T, Barkholt V, Koch AG: Leuconostoc carnosum 4010 has the potential for use as a protective culture for vacuum-packed meats: Culture isolation, bacteriocin identification, and meat application experiments. Int J Food Microbiol, 83 (2): 171-84, 2003.
Yamato M, Ozaki K, Ota F: Partial purification and characterization of the bacteriocin produced by Lactobacillus acidophilus YIT 0154. Microbiol Res,158, 169-172, 2003.
Kanatani K, Oshimura M, Sano K: Isolation and characterization of acidocin A and cloning of the bacteriocin gene from Lactobacillus acidophilus. Appl Environ Microbiol, 61 (3): 1061-1067, 1995.
Schagger H: Tricine-SDS-PAGE. Protocol. Nature Protocols, 1, 16-22, 2006.
Blum H, Beier H, Gross HJ: Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis, 8 (2): 93-99, 1987.
Delves- Broughton J: Nisin as a food preservative. Food Australia, 57 (12): 525-527, 2005.
Tahiri I, Desbiens M, Benech R, Kheadr E, Lacroix C, Thibault S, Ouellet D, Fliss I: Purification, characterization and amino acid sequencing of divergicin M35: A novel class IIa bacteriocin produced by Carnobacterium divergens M35. Int J Food Microbiol, 97 (2): 123-36, 2004.
Todorov SD, Dicks LMT: Screening for bacteriocinproducing lactic acid bacteria from boza, a traditional cereal beverage from Bulgaria comparison of the bacteriocins. Process Biochemistry, 41, 11-19, 2006.
Nissen-Meyer J, Holo H, Havarstein LS, Sletten K, Nes IF: A novel Lactococcal bacteriocin whose activity depends on the complementary action of two peptides. J Bacteriol, 174 (17): 5686-5692, 1992.
Piard JC, Muriana PM, Desmazeaud MJ, Klaenhammer TR: Purification and partial characterization of lactacin 481, a lanthionine-containing bacteriocin produced by Lactococcus lactis subsp. lactis CNRZ 481. Appl Environ Microbiol, 58 (1): 279-284, 1992.
Klaenhammer TR: Genetics of bacteriocins produced by lactic acid bacteria. FEMS Microbiol Rev, 12, 39-86, 1993.