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초록갈파래로부터 분리한 세균의 군집 분석 및 신종 미생물의 동정

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Alternative Title
Bacterial community analysis and identification of a novel bacterium isolated from Umbraulva japonica
Abstract
This study sought to carry out bacterial community analysis and examine the identification of a novel bacterium isolated from the surface of Umbraulva japonica, which is a marine algae collected in Jeju Island. A total of 79 strains were isolated from U. japonica and cultivated using Marine agar, 1/10 Marine agar, R2A. The 16S rRNA gene sequence of the isolated bacteria were analyzed using NCBI and EzbioCloud. Based on the result, the major groups were classified into 4 phyla as Proteobacteria (α-proteobacteria, β-proteobacteria, γ -proteobacteria) (74.7%), Bacteroidetes (20.3%), Actinobacteria (2.5%) and Firmicute (2.5%) along with 7 classes, 13 order, 17 families and 31 genera. The phylogenetic analysis showed 3 strains were showed similarity of <97% with the previously identified strains, and could be reported as a new genus or species. Therefore, it is considered that additional experiments should be conducted together with the standard strain. The novel strain, UJT9T, was identified as an orange colony, gram-negative, rod-shaped, non-motility and aerobic bacterium. Strain UJT9T grew at a temperature range of 15-35℃ (optimum, 20-25℃), a pH range 5.5-8.0 (optimum, pH 6.0-7.0) and in the presence 1.0-3.0% (w/v) NaCl (optimum, 1.0-2.0%). The 16S rRNA gene sequence- based phylogenetic analysis showed that strain UJT9T belong to the Nonlabens and was most closely related to Nonlabens arenilitoris M-M3T, Nonlabens ulvanivorans PLRT with a sequence similarity of 97.7%, 97.6%, respectively. Strain UJT9T contained menaquinones type 6 (MK-6) as the major quinones and Fatty acids (>5%) were iso-C15:0 (20.1%), iso-C16:0 3-OH (14.1%), iso-C16:0 (10.2%), iso-17:0 3-OH (7.8%), anteiso-C15:0 (6.3%), iso-C16:1 H (5.6%). The major polar lipids were phosphatidylenthanolamine (PE), two unidentified lipids and two unidentified amino lipids. The DNA G+C content of strain UJT9T was determined to be 38.50 mol%. The DNA-DNA hybridization values for strain between N. arenilitoris M-M3T and N. ul vanivorans PLRT were 67.42 ± 1.41%, 48.21 ± 4.53%, respectively. On the basis of the data presented, strain UJT9T is considered to represented a novel species of the genus Nonlabens. The type strain is Nonlabens sp. UJT9T (=KACC 19635T = JCM 32766T). The antibacterial activity of 79 strains including the novel bacterium was evaluated. Strain UJT7, UJT20 and UJR17 showed antibacterial activity against Vibrio vulnificus, while UJR17 showed antibacterial activity against Streptococcus parauberis. UJT7 and UJT20, UJR17 have been identified as Bacillus sp. and Pseudomonas sp. and it can be safely presumed that it is beneficial to carry out additional experiments for various applications.
Author(s)
김지현
Issued Date
2019
Awarded Date
2019. 2
Type
Dissertation
URI
http://dcoll.jejunu.ac.kr/common/orgView/000000008865
Alternative Author(s)
Kim, Ji Hyun
Affiliation
제주대학교 대학원
Department
대학원 해양생명과학과
Advisor
허문수
Table Of Contents
목 차 ⅰ
List of Tables iv
List of Figures vi
Abstract vii
Ⅰ. 서 론 1
Ⅱ. 재료 및 방법 4
2.1. 시료채집 4
2.2. 미생물의 분리, 배양 및 보관 4
2.3. 염기서열 분석 7
2.3.1. Genomic DNA 추출 및 유전자 증폭 7
2.3.2. 염기서열 및 계통학적 분석 8
2.4. 신종 미생물 UJT9T의 동정 9
2.4.1. 계통분류학적 분석 9
2.4.1.1. Genomic DNA 및 계통학적 분석 9
2.4.1.2. 표준균주 10
2.4.1.3. DNA-DNA hybridization 11
2.4.2. 형태학적 특성 12
2.4.2.1. 주사전자현미경(SEM, Scanning Electron Microscopy) 12
2.4.2.2. 그람염색 및 운동성 12
2.4.3. 생리학적 특성 13
2.4.4. 생화학적 특성 15
2.4.4.1. CatalaseOxidase test 15
2.4.4.2. 가수분해(Hydrolysis) test 15
2.4.4.3. API 20 NE 및 ZYM 16
2.4.5. 화학 분류학적 특성 17
2.4.5.1. 지방산(Fatty acid) 17
2.4.5.2. 퀴논(Quinones) 18
2.4.5.3. 극성지방(Polar lipids) 19
2.4.5.4. DNA G+C content 20
2.4.6. 항생제 시험(Antibiotic test) 21
2.5. 항균 활성 탐색 22
Ⅲ. 결 과 24
3.1. 염기서열 및 계통학적 분석 24
3.2. 세균 군집 분석 31
3.3. 신종 미생물 UJT9T의 동정 34
3.3.1. 계통분류학적 분석 34
3.3.2. 형태학적 특성 36
3.3.3. 생리학적 특성 37
3.3.4. 생화학적 특성 38
3.3.5. 화학 분류학적 특성 40
3.3.6. 항생제 시험(Antibiotic test) 44
3.4. 항균 활성 탐색 46
Ⅳ. 고 찰 47
Ⅴ. 요 약 52
Ⅵ. 참고 문헌 54
Ⅶ. 감사의 글 64
Degree
Master
Publisher
제주대학교 대학원
Citation
김지현. (2019). 초록갈파래로부터 분리한 세균의 군집 분석 및 신종 미생물의 동정
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