생명과학회지 (Journal of Life Science)

  • 제28권12호
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  • Pages.1477-1488
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  • 2018
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  • 1225-9918(pISSN)
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  • 2287-3406(eISSN)
한국생명과학회 (Korean Society of Life Science)
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16S rRNA 염기서열 분석을 통한 오분자기(Sulculus diversicolor supertexta)내 미생물 군집 조사 및 인체유해 질병세균에 대한 항균활성 모니터링

Investigation of Microbial Communities in Sulculus diversicolor supertexta Through 16S rRNA Sequencing and Antibacterial Monitoring of Harmful Strains

  • 김민선 (국립수산과학원 제주수산연구소) ;
  • 이승종 (국립수산과학원 제주수산연구소) ;
  • 허문수 (제주대학교 해양과학대학 수산생명의학과)
  • Kim, Min-Seon (Jeju Fisheries Research Institute, National Institute of Fisheries Science) ;
  • Lee, Seung-Jong (Jeju Fisheries Research Institute, National Institute of Fisheries Science) ;
  • Heo, Moon-Soo (Major of Aquatic Life Medicine, Faculty of Marine Biomedical Sciences, Jeju National University)
  • 투고 : 2018.08.07
  • 심사 : 2018.10.22
  • 발행 : 2018.12.30
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초록

본 연구는 제주 연안에서 채집한 오분자기(Sulculus diversicolor supertexta)를 구성하는 미생물 군집의 다양성을 알아보기 위하여 근육, 장, 생식소 각 부위별로 조사하였다. 배지로 1차 순수 분리한 결과 근육은 MA, 장 1% BHIA, 생식소 1% TSA에서 각각 최대 군락 계수가 나타났다. 16S rRNA sequence로 표준 균주와 비교 유사도 분석 결과 총 190개의 순수 colony가 분리되었다. NBLAST program 분석 결과 크게 5문 25과 39속 71종으로 나타났다. 표준 균주와 상동성은 91-100%를 나타냈다. 오분자기 내 미생물 군집은 크게 Probacteria (Gamma-proteobacteria, Alpha-proteobacteria) 48%, Actinobacteria 32.5%, Firmicutes 16.9%, Bacteroide 1.3%, Deinococcus-thermus 1.3%로 나타났다. 근육, 장, 생식소 모든 부위에서 Moraxellaceae과 Psychrobacter cibarius가 우점하였다. 근육, 장, 생식소 모든 부위에서 Alteromonadaceae, Enterobacteriaceae, Pasturellaceae, Moraxellaceae, Rhodobacteraceae, Geminicoccaceae, Dietziaceae, Intrasporangiaceae, Microbacteriaceae, Micrococcaceae, Micromonosporaceae, Streptomycetaceae, Aerococcaceae, Bacillaceae, Paenibacillaceae, Planococcaceae, Staphylcoccaceae가 공통적으로 분리되었으며, 장에서 Flavobacteriaceae, Corynebacteriaceae, Yesiniaceae, Vibrionaceae, Hahellaceae, Pseudomonadaceae가 추가 분리되었다. 분리 균주로부터 인체 유해 질병 세균에 대한 항균활성 모니터링 결과 Sterptomyces albus (96%)가 4균주 모두 항균활성을 보였고 Agrococcus baldri (99%), Psychrobacter nivimaris (99%)가 E. coli, E. aerogens에 대한 항균활성을 나타냈다. 그 외 상동성이 낮은 일부 균주가 분리되어 신균주 실험을 비롯한 항균활성물질 정제 등 추가 실험이 필요한 것으로 사료된다. 본 실험은 오분자기 미생물 군집의 다양성과 유전학적 자원을 확보하는데 의의를 두며, 응용 미생물의 개발 가능성에 있어 기초 자료를 제공하고자 하였다.

This study investigated the muscles, intestines, and gonads of Sulculus diversicolor supertexta to examine the diversity of microbial communities within examples collected from the Jeju Coast. Using different media, initial pure isolation in MA, 1% BHIA, and 1% TSA indicated that the muscles, intestines, and gonads supported more communities, respectively. In analysis of relative similarity with 16s rRNA sequencing, 190 pure colonies were isolated, and further analysis with NBLAST identified 71 species, 39 genera, 25 families, and five phyla. Homogeny with the reference strain was 91-100%. Microbial communities in S. supertexta consisted of gamma and alpha Proteobacteria (48%), Actinobacteria (32.5%), Firmicutes (16.9%), Deinococcus-Thermus (1.3%), and Bacteroides (1.3%). In all tissue, Psychrobacter cibarius in Moraxellaceae was dominant. Alteromonadaceae, Enterobacteriaceae, Pasturellaceae, Moraxellaceae, Rhodobacteraceae, Geminicoccaceae, Dietziaceae, Intrasporangiaceae, Microbacteriaceae, Micrococcaceae, Micromonosporaceae, Streptomycetaceae, Aerococcaceae, Bacillaceae, Paenibacillaceae, Planococcaceae, and Staphylcoccaceae were commonly isolated across all tissues, and Flavobacteriaceae, Corynebacteriaceae, Yesiniaceae, Vibrionaceae, Hahellaceae, Pseudomonadaceae were also identified from the intestines. In microbial monitoring of four harmful bacteria, Streptomyces albus (96%) showed antibacterial activity against all four strains, and Agrococcus baldri (99%) and Psychrobacter nivimaris (99%) presented against E. Coli and E. aerogens. In addition, some strains with low homogeny were isolated and further experiments are therefore required, for example to refine the antimicrobial substances including new strain investigations. These additional experiments would aim to establish generic resources for the microbial communities in S. Supertexta and provide basic data for applied microbiological research.

키워드

SMGHBM_2018_v28n12_1477_f0001.png 이미지

Fig. 1. Neighbour-joining phylogenetic tree based on 16S rRNA gene sequences showing the position of microorganisms strains from each organs (muscle, intestine, gonads) of Sulculus diversicolor supertexta. Bootstrap percentage (from 1,000 replications) >50% are shown at brach points.

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Fig. 2. Distribution of each organs bacteria in Sulculus diversicolor supertexta. (a; muscle, b; Intestine, c; gonads.)

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Fig. 3. Antimicrobial monitoring of human harmful disease by cross- streak method of Microorganisms isolated from Sulculus diversicolor supertexta (harmful disease of human body: gram negative; Enterobacter aerogens KCTC 2190, Escherichia coli KCTC 1682, gram postitive: Streptococcus mutans KCCM 40105, Enterococcus faecalis KCTC 5290), (tested organisms; A;P. nivimaris (B-12), b; A. bibardi (T-25), S. albus (M-62)

Table. 1. Microbial communities isolated from each organ of Sulculus diversicolor supertexta

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Table 2. Analysis of 16S rRNA sequence of microorganisms isolated from muscle, intestines and gonads of Sulculus diversicolor supertexta. (A group) Proteobacteria, (B group) Actinobacteria, (C group) Firmicutes, Bacteroide, Deinococcus-thermus

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Table 2. Continued

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