Journal of fish pathology (한국어병학회지)

The Korean Society of Fish Pathology (한국어병학회)
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Immunological characteristics of Edwardsiella tarda grown under iron-restricted condition

철 결핍 조건에서 배양된 Edwardsiella tarda의 면역학적 특성

  • Published : 2006.04.01
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Abstract

The immunogenicity of Edwardsiella tarda was surveyed under two different culture conditions. In SDS-PAGE patterns of the outer membrane proteins (OMPs) extracts of E. tarda, grown under Trypic soy broth (TSB) and TSB supplemented iron chelate 2,2‘-dipyridyl iron-restricted condition, were examined. The results showed that the iron-regulated outer membrane protein (IROMPs) with molecular masses of 68 and 73 kDa were expressed by bacteria grown in iron-chelate TSB.The pathogenicity was examined by intraperitoneal injection with live E. tarda grown under TSB, iron-chelate TSB and iron-supplemented TSB. The result of pathogenicity test showed significantly high mortality in the group of live E. tarda grown under iron-chelate TSB.The effect of formalin killed cell (FKC) of TSB cultured bacteria and 2,2'-dipyridyl FKC (DP-FKC) of cultured bacteria on the iron-chelate TSB on the development of protective immunity in olive flounder was studied. The level of immune response was evaluated with immunized fish at 1, 2, 3 and 4 weeks after immunization. The numbers of specific antibody secreting cells (SASCs) showed significantly increased level at 2 week after immunization in each group. The agglutination titre of immunized fish was significantly high level at 3 weeks after immunization.The level of protection in olive flounder at 1, 2, 3 and 4 weeks after vaccination was examined by intraperitoneal challenge test with live E. tarda.

E. tarda의 독성 비교를 위하여 일반 TSB, iron-chelate인 2,2‘-dipyridyl 첨가 TSB 및 Fe 첨가 TSB의 세 가지 조건에서 E. tarda를 배양하였다. 독성시험의 결과는 iron-chelate 첨가 시험구의 E. tarda에서 가장 낮은 독성을 나타내었다. 그리고 각각의 OMPs와 IROMPs을 분리하여 SDS-PAGE로 항원성을 비교한 결과, iron-chelate 첨가 시험구의 E. tarda IROMPs에서 68, 73 kDa 크기의 분자들이 더 많이 발현된다는 것을 확인할 수 있었다. 두 가지 vaccine 투여구 중에서 2,2‘-dipyridyl 첨가 배지로 배양한 균체로 제작한 DP-FKC 항원 투여구가 응집 항체가가 더 높게 나타났다. 면역반응의 변화를 알기 위한 응집 항체가 조사에서는 FKC 항원과 DP-FKC 항원 투여구에서 모두 대조구보다 높은 값을 나타내었고, 3주 째에 최고치를 나타내었다. ELISPOT을 이용한 항체 생성 세포 수의 검출에서도 응집 항체가와 유사한 pattern을 나타내었는데, 두 가지 vaccine 투여구에서 2주 째에 가장 많은 항체 생성 세포 수를 나타내었다. 공격실험에서는 3주 째에 FKC 항원 투여구가 50%, DP-FKC 항원 투여구에서는 80%의 생존율을 나타내었다. 전체적으로 DP-FKC 항원 투여구의 생존율이 높게 나타났다. 이상의 결과로부터 넙치에 대한 E. tarda의 vaccine으로는 현재 연구되고 있는 FKC vaccine보다 철 결핍 조건에서 배양된 균체의 항원인 DP-FKC vaccine이 동일한 조건으로 사용하였을 때 더 나은 방어력을 가진다는 것을 알 수 있었다.

Keywords

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