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어병균 Vibrio anguillarum 생육 저해 인자를 생산하는 Bacillus amyloliquefaciens H41의 분리

Isolation of Bacillus amyloliquefaciens H41 Producing Growth Inhibition Factor against Vibrio anguillarum

  • 발행 : 2006.07.31

초록

양식 산업의 문제점인 어병균을 방제하기 위한 생물 전구체의 개발을 목적으로 해산어의 내장 추출물에서 어병균인 Vibrio anguillarum의 생육을 저해하는 새로운 균주를 분리, 동정하여 Bacillus amyloliquefaciens H41으로 명명하고 생육특성 및 생육저해인자 생성 조건을 검토하였다. 분리 균의 생육 최적 조건은 기본배지로 1% peptone, 질소원으로 1.5% yeast extract, 탄소원으로 1% sucrose, 0.5% NaCl, 금속이온으로는 0.05% $MgSO_4{\cdot}7H_20$이었으며 pH는 7.0-8.0, 온도는 $28-35^{\circ}C$에서 20시간의 진탕배양이었다. V. anguillarum 생육 저해물질은 B. amyloliquefaciens H41의 배양 상등 액에서 볼 수 있었는데 paper-disk 법으로 투명환의 생성여부로 확인하였다. 분리 균에 의한 생육저해 물질 생산 최적 조건은 1% peptone, 1.5% yeast extract, 1% NaCl, 0.05% $MgSO_4{\cdot}7H_20$, 1%의 sucrose이었으며 최적 pH는 7.5, 최적온도는 $35^{\circ}C$이었다. 분리 균은 최적 배양조건하에서 저해물질을 배양 후 16시간부터 24시간대에 가장 높은 활성을 나타내었다. 저해 활성반응은 분리 균에서는 저해 활성을 나타내었으나 B. amyloliquefaciens KCTC1724 표준균주에는 저해활성을 나타나지 않는 대조를 보였다. 또한 분리 균에 의해 생산된 저해물질은 V. anguillarum 에는 생육 억제 효과를 나타내었으나 다른 인체 병원성 Vibrio에는 전혀 영향을 미치지 않는 것으로 나타나 균주 선택성이 특이적이었다.

To investigate the possible use of probiont in fish farming industry, a bacterium with inhibitory effect against Vibrio anguillarum was isolated from gastrointestinal tract of the marine fish of yellow tail. It was identified to be Bacillus amyloliquefaciens H41 based on biochemical and physiological characterization. The optimal growth conditions of the isolated strain were 1% peptone, 1.5% yeast extract, 1% sucrose, 0.5% NaCl, 0.05% $MgSO_4{\cdot}7H_20$, pH 7.0-8.0, and 20 hr of incubation between $28-35^{\circ}C$ under aeration. The culture supernatant of the isolated strain showed inhibition activity against V. anguillarum. Inhibition activity was cleared by forming a clear zone by a paper-disk method. The maximal production of growth inhibition factor was induced by cultivation under 1% peptone, 1.5% yeast extract, 1% sucrose, 1% NaCl, 0.05% $MgSO_4{\cdot}7H_20$, pH 7.5 and at $35^{\circ}C$. The highest growth inhibition factor production was observed after 16-24 hr cultivation under aeration. The culture supernatant of the isolated strain showed inhibition activity whereas no inhibition activity was shown from the standard B. amyloliquefaciens KCTC 1724 strain. The growth inhibition affected only against V. anguillarum among other pathogenic Vibrios tested here.

키워드

참고문헌

  1. Austin, B., L. F. Stuckey, P. A. W. Robertson, I. Effendi and D. R. Griffith. 1995. A probiotic strain of Vibrio alginolyticus effective in reducing diseases caused by Aeromonas salmonicida, Vibrio anguillarum and Vibrio ordalli. J. Fish Dis. 18, 93-96 https://doi.org/10.1111/j.1365-2761.1995.tb01271.x
  2. Bly, J. E., S. M. A. Quiniou, M. A. Lawson and L. W. Clem. 1997. Inhibition of Saprolegnia pathogenic for fish by Pseudomona fluorescens. J. Fish Dis. 20, 35-40 https://doi.org/10.1046/j.1365-2761.1997.d01-104.x
  3. Cross, J. H., M. H. Schiewe and S. Falkaw. 1977. Evidence for plasmid contribution to the virulence of the fish pathogen Vibrio anguillarum. Infect. Immun. 27, 897-902
  4. Dopazo, C. P., Lemos, M. L., Lodeiros, C. Bolinches, J. L. Barja and A. Torranzo. 1988. Inhibitory activity of antibiotic- producing marine bacteria against fish pathogens. J. Appl. Bacteriol. 65, 97-101 https://doi.org/10.1111/j.1365-2672.1988.tb01497.x
  5. Febregas, J., A. Munoz, A. Otero, J. L. Barja and M. Romaris. 1991. A preliminary study on antimicrobial activities of some bacteria isolates from marine environment. Nippon Suisan Gakkaishi 57, 1377-1382 https://doi.org/10.2331/suisan.57.1377
  6. Gram, L., J. Melchiorsen, B. Spanggard, I. Huber and T. F. Nielsen. 1999. Inhibition of Vibrio anguillarum by Pseudomonas flourecence AH2, a possible probiotic treatment of fish. Appl. Environ. Microbiol. 65(3), 969-973
  7. Horne M. T. and A. Baxendale. 1983. The adhesion of Vibrio anguillarum to host tissues and its role in pathogenesis. Journal of Fish Diseases 6, 461-471 https://doi.org/10.1111/j.1365-2761.1983.tb00100.x
  8. Horne M. T., M. Tatner, S. Mcderment and C. Agius. 1982. Vaccination of rainbow trout, Richardson, at low temperatures and the long-term persistence of protection. Journal of Fish Diseases 5, 343-345 https://doi.org/10.1111/j.1365-2761.1982.tb00490.x
  9. Itoh, S., Yoshimizu, M. and Ezura, Y. 1997. Disinfectant effects of low level of total residual oxidants in artificial seawater on fish pathogenic microorganisms. pp. 92-102, Nippon. Suisan Gakkaishi 63, Japan
  10. Kim. E. and Aoki, T. 1993. Drug resistance and broad geographical distribution of identical R plasmid of Pasteurella pisicida isolated from cultured yellowtuil in Japan. Microbiol. Immunol. 37, 103-109 https://doi.org/10.1111/j.1348-0421.1993.tb03186.x
  11. Kim, Y. H. 1998. Purification of hemolysin from Vibrio anguillarum isolated from fish. J. Life Science 8(5), 598-603
  12. Larsen, J. L. and H. B. Rasmussen. 1988. Study of Vibrio anguillarum strains from different sources with emphasis on ecological and pathological properties. Appl. Environ. Microbiol. 54(9), 2264-2267
  13. Lemos, M. L., A. E. Toranzo, and J. L. Barja. 1985. Antibiotic activity of epiphytic bacteria isolated from intertidal seaweeds. Microb. Ecol. 11, 149-163 https://doi.org/10.1007/BF02010487
  14. Nakamura, A., Takahashi, K. G. and, K. Mori. 1999. Vibriostatic bacteria isolated from rearing seawater of oyster blood stock: potentially as biocontrol agents for vibriosis in oyster larvae. Fish Pathol. 34, 139-144 https://doi.org/10.3147/jsfp.34.139
  15. Olsson, J. C., A. Westerdahl, P. L. Conway, and S. Kjelleberg. 1992. Intestinal colonization potential of turbot( Scophthalmus maximus) and dab(Limanda limanda)-associated bacteria with inhibitory effects against Vibrio anguillarum. Appl. Environ. Microbiol. 58(2), 551-556
  16. Spanggaard, B., I. huber, J. Nielson, E. B. Sick, C. B. Pipper, T. Martinussen, W. J. Slierendrecht and L. Gram. 2001. The probiotic potential against vibriosis of the indigenous microflora of rainbow trout. Environ. Microbiol. 3(12), 755-765 https://doi.org/10.1046/j.1462-2920.2001.00240.x
  17. Westerdahl, A., J. C. Olsson, S. Kjelleberg and P. L. Conway. 1991. Isolation and characterization of turbot (Scophthalmus maximus) associated bacteria with inhibitory effects against Vibrio anguillarum. Appl. Environ. Microbiol. 57, 2223-2228