Korean Journal of Microbiology (미생물학회지)

The Microbiological Society of Korea (한국미생물학회)
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Biodiversity and Isolation of Gut Microbes from Digestive Organs of Harmonia axyridis

무당벌레 소화기관으로부터 장내세균의 분리 및 계통학적 다양성

  • Kim, Ki-Kwang (Department of Microbial & Nanomaterials, Mokwon University) ;
  • Han, Song-Ih (Department of Microbial & Nanomaterials, Mokwon University) ;
  • Moon, Chung-Won (Department of Applied Biology, Chungnam University) ;
  • Yu, Yong-Man (Department of Applied Biology, Chungnam University) ;
  • Whang, Kyung-Sook (Department of Microbial & Nanomaterials, Mokwon University)
  • 김기광 (목원대학교 미생물나노소재학과) ;
  • 한송이 (목원대학교 미생물나노소재학과) ;
  • 문청원 (충남대학교 응용생물학과) ;
  • 유용만 (충남대학교 응용생물학과) ;
  • 황경숙 (목원대학교 미생물나노소재학과)
  • Received : 2011.01.26
  • Accepted : 2011.03.23
  • Published : 2011.03.31
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Abstract

Bacterial density distributions of gut microbes in the digestive organs of Harmonia axyridis collected from three different sources (JK, CK, and CJ) were $6.0{\times}10^4$ CFU/gut under aerobic culture condition and $8.0{\times}10^6$ CFU/gut under anaerobic culture condition. Seven colony types were observed under aerobic condition and three types of similarity were detected under anaerobic condition. In total, 116 strains, including 34 strains under aerobic condition, were isolated from the digestive organs of H. axyridis. Based on the analysis of the 16S rRNA gene sequence, aerobic gut microbes were assigned to the Proteobacteria, Actinobacteria, Firmicutes, and Deinococcus-Thermus. A large number of isolates belonged to the genus Bacillus and Staphylococcus of the Firmicutes commonly found in H. axyridis from different sites. Anaerobic gut microbes were found to be similar according to colony morphological, phylogenetic analysis using ARDRA. Eighty-two anaerobic gut microbes were clustered into 17 different ARDRA types according to HaeIII. Representative anaerobic gut microbes in each ARDRA group were divided into five species of ${\gamma}$-Proteobacteria based on 16S rRNA gene sequence analysis; Hafnia alvei, Enterobacter ludwigii, Enterobacter kobei, Pseudomonas oryzihabitans and Pseudomonas koreensis. Phylogenetic analysis indicated that about 70% of the isolates belonged to ${\gamma}$-Proteobacteria, suggesting predominance of gut microbes.

각 지역에서 수집한 무당벌레(JK, CK, CJ)의 소화기관을 채취하여 장내세균의 밀도를 조사한 결과, 호기배양의 경우 $6.0{\times}10^4$ CFU/gut, 혐기배양 결과 $8.0{\times}10^6$ CFU/gut로 계수되었다. 호기적 조건에서 배양된 세균 집락은 총 7가지 형태로 분류되었으며, 혐기적 조건에서 배양된 집락은 총 3가지 형태로 유사한 특징을 나타내었다. 무당벌레 각 소화기관으로부터 호기성세균 34균주와 혐기성세균 82균주, 총 116균주의 장내세균을 순수분리하였다. 호기성 세균 34균주를 대상으로 16S rRNA 유전자 염기서열을 해석한 결과, ${\alpha}$-Proteobacteria (3균주), ${\gamma}$-Proteobacteria (2 균주), Firmicutes (24 균주), Actinobacteria (4 균주) 그리고 Deinococcus-Thermus (1균주) 계통군으로 분류되었다. Firmicutes 계통군의 Bacillus thuringiensis와 Staphylococcus 속의 다양한 종은 JK, CK 및 CI 무당벌레 소화기관에서 모두 공통적으로 분리되었다. 형태적으로 유사한 혐기세균의 16S rRNA-ARDRA 패턴양상을 분석하여 유사도 70%에서 비교한 결과, 17개 ARDRA group으로 분류되었다. 각 ARDRA group에 속하는 대표 혐기성세균의 16S rRNA 유전자 염기서열을 해석한 결과, 무당벌레 소화기관에서 분리된 모든 혐기성 장내세균은 ${\gamma}$-Proteobacteria 계통군에 속하는 것으로 나타났으며 Hafnia alvei, Enterobacter ludwigii, Enterobacter kobei, Pseudomonas oryzihabitans 그리고 Pseudomonas koreensis와 높은 유연관계를 갖는 것으로 확인되었다. 무당벌레 소화기관으로부터 분리된 전체 장내세균의 약 70%가 ${\gamma}$-Proteobacteria 계통군에 속하였으며, 23%가 Firmicutes 계통군으로 무당벌레 소화기관 내 주요 계통군임이 확인되었다.

Keywords

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