Phenotypic and genetic characteristics of Vibrio ichthyoenteri isolated from the olive flounder, Paralichthys olivaceus of culturing size

미성어 양식 넙치, Paralichthys olivaceus에서 분리한 Vibrio icthyoenteri의 표현형 및 유전형적 특성

  • Published : 2006.08.01


From 2002 to 2004, various vibrios were isolated from the olive flounder, Paralichthys olivaceus of culturing size with disease signs. During this survey, it was known that the high proportion of Vibrio ichthyoenteri was occupied among the isolated vibrios. Generally, V. ichthyoenteri is well known as the pathogen of bacterial enteritis of olive flounder larvae. The aim of the present study was the compare the characteristics of two groups of V. ichthyoenteri, culturing sized olive flounder, and larvae of olive flounder showing the intestinal necrosis. The research was focused on the physiology, biochemistry, genetics in the two bacterial groups. The physiological and biochemical characteristics of the tested strains were very similar. The intergenic spacer (IGS) region between the 16S and 23S rRNA genes of 21 isolated strains and 3 reference strains, V. ichthyoenteri, were investigated by PCR fragment length typing and DNA sequencing. After the isolated strains were identified as V. ichthyoenteri, not only phenotypic characteristics of the isolated and reference strains but also homology of 16S-23S IGS of all isolated strains and reference strains as 99.1~100%. The V. ichthyoenteri showed 4 specific 16S-23S patterns and contained no-tRNA, tRNAGlu(TTC) , tRNAIle(GAT) tRNAAla(TGC) type .

2002년에서 2004년에 걸쳐, 우리 나라의 넙치 양식장에서 질병의 증상이 보이는 중간 육성 단계의 넙치를 대상으로 다수의 Vibrio속 세균을 분리 동정하였으며 그 중에서 V. ichthyoenteri균이 높은 비율을 차지하고 있었다. 일반적으로 V. ichthyoenteri는 넙치의 자어기 장관백탁증의 원인균으로 잘 알려져 있으며, 미성어의 병어로부터 분리되었다는 점에서 여러 크기의 넙치 및 종묘 생산에 미치는 영향을 알고자 하였다. 본 연구는 미성어 넙치에서 분리된 균주그룹과 장관백탁증에 걸린 넙치 자어에서 분리한 V. ichthyoenteri 참조 균주 두 그룹간의 생화학적 및 생리학적 성상, 유전학적 특성을 비교 분석함으로서 분리 균주를 V. ichthyoenteri로 동정하고, 이들의 어병학적 특성을 조사하였다.실험 결과 시험 균주와 참조 균주간의 생화학적 특성과 생리학적 성상이 거의 동일한 것으로 나타났다.본 연구에서 분리 균주와 참조 균주의 16S-23S rRNA intergenic space (IGS) region을 cloning하여 염기 서열을 분석한결과 3개의 참조 균주와 19개의 분리 균주들은 V. ichthyoenteri 참조 균주와 99.1 ~ 100% 동일하였으며 V. ichthyoenteri는 3개 이상의 PCR products를 보였다. 이들 각각에 대한 tRNA type을 분석한 결과, no-tRNA, tRNAGlu(TTC) type 그리고 tRNAIle(GAT)와 tRNAAla(TGC) 를 coding하는 3 가지 tRNA type을 밝혔다.



  1. Alsian, M. and Blanch, A. R.: A set of keys for biochemical identification of environmental Vibrio species. J. Appl. Bacteriol., 76: 79-85, 1994
  2. Amann, R. I., Ludwing, W. and Schleifer, K. H.: Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev. 59: 143-169, 1995
  3. Barry, T., Colleran, G., Glennon, M., Dunican, L. K. and Gannon, F.: The 16S/23S ribosomal spacer region as a target for DNA probes to identify eubacteria. PCR Methods and Applications. 1: 51-56, 1991
  4. Brant, T. N., Lee, J. V., West, P. A. and Colwell, R. P.: Numerical classification of species of vibrio and related genera. J. Appl. Bacteriol., 61: 437-467, 1986
  5. Chen, D., Hanna, P. J., Altmann, K., Smith, A., Moon, P. and Hammond, L. S.: Development of monoclonal antibodies that identify vibrio species commonly isolated from infections of humans, fish and shellfish. Appl. Environ. Microbiol., 58: 3694-3700, 1992
  6. Chun, J. A. H. and Colwell, R. R.: Analysis of 16S/23S rRNA intergenic Spacer Regions of Vibrio cholerae and Vibrio mimicus. Appl. Environ. Microbiol., 65: 2202-2208, 1999
  7. Diggles, B. K., Carson, J., Hine, P.M., Hickman, R. W. and Tait, M. J.: Vibrio species associated with mortalities in hatchery-reared turbot (Colistium mudipinns) and brill (C. guntheri) in New York, pp. 1-36, 2000
  8. Gurtler, V. and Stanisich, V. A.: New approaches to typing and identification of bacteria using the 16S-23S rDAN spacer region. Microbiology, 142: 3-6, 1996
  9. Ishimaru, K., Akagawa-Matsushita, M. and Muroga, K.: Vibrio ichthyoenteri sp. nov., a pathogen of Japanese flounder (Paralichthys olivaceus) larvae. Int. J. Syst. Bacteriol., 46: 155-159, 1996
  10. Kim, D. H., Han, H. J., Kim, S. M., Lee, D. C. and Park, S. I.: Bacterial enteritis and the development of the larval digestive tract in olive flounder, Paralichthys olivaceus (Temminck & Schlegel). J. Fish Dis., 27: 497-505, 2004
  11. Kim, S.M., Won, K. M., Woo, S. H., Li, H., Kim, E. J., CHoi, K. J., Cho, M. Y., Kim, M. S. and Park, S. I.: Vibrio isolated from diseased marine culturing fishes in Korea. J. Fish pathol., 18: 133-145, 2005
  12. Krawiec, S. and Riley, M.: Organization of the bacterial chromosome. Microbiol. Rev., 54: 502-539, 1990
  13. Maeda, T., Takada, N., Furushita, M. and Shiba T.: Structural variation in the 16S-23S rRNA intergenic spacers of Vibrio parahaemolyticus. FEMS Microbiol. Lett., 192: 73-77, 2000
  14. Maeda, T., Matsuo, Y., Furushita, M. and Shiba T.: Seasonal dynamics in a coastal Vibrio community examined by a rapid clustering method based on 16S rDNA. Fisheries Sci., 69: 385-394, 2003
  15. Masumura, K., Yasunobu, H., Okada, N. and Muroga, K.: Isolation of a Vibrio sp. the causative bacterium of intestinal necrosis of Japanese flounder larvae. Fish Pathol., 24: 135-141, 1989
  16. Moon, Y. G., Park, G. T., Son, H. J., Lee, S. H., Lee, J. M. and Heo, M. S.: Rapid detection of the pathogenic of bacterial white enteritis of larval and juvenile stages in olive flounder (Paralichthys olivaceus). J. Fish Pathol., 17: 159-169, 2004
  17. Murata, O.: Infectious intestinal necrosis in flounder. Fish Pathol., 22: 59-61, 1987
  18. Muroga, K.: Viral and bacterial diseases of marine fish and shellfish in Japanese hatcheries. Aquaculture, 202: 23-44, 2001
  19. Muroga, K., Higashi, M. and Keitoku, H.: The isolation of intestinal microflora of farmed red seabream (Pagrus major) and black seabream (Acanthopagrus schlegeli) at larval and juvenile stages. Aquaculture, 65: 79-88, 1987
  20. Muroga, K., Yasunobu H., Okada, N. and Masumura, K.: Bacterial enteritis of cultured flounder Paralichthys olivaceus larvae. Dis. Aquat. Org., 9: 121-125, 1990
  21. Nishioka, T., Furusawa, T. and Mizuta, Y.: Diseases occurring in marine fish and shellfish hatcheries in Japan (1989-1994). Suisan Zoshoku, 45: 285-290, 1997
  22. Osorio, C. and Toranzo, A.: DNA-based diagnostics in sea farming, p. 253-310. In M. Fingerman and R. Nagahushanam (ed.), Seafood safely and human health. Science Publishers, Inc., Enfield, N. H, 2002
  23. Simon, K. Y., Wang, H. Z., Sheran, H. W., Rudolf, S. S. and Richard, Y. C.: Analysis of 16s-23s rDNA intergenic spacers (IGSS) of marine vibrios for species-specific signature DNA sequences. Marine pollution Bulletin, 44: 412-420, 2002
  24. 이정백, 노섬, 송춘복: 넙치 Paralichthys olivaceus 자어에서 분리한 장관백탁증의 원인균인 Vibrio sp. (INFL group)의 생물학적 및 생화학적 특성. 한국어병학회지, 8: 99-109, 1995