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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 陳昭倫 | |
dc.contributor.author | Ming-Hui Liao | en |
dc.contributor.author | 廖敏惠 | zh_TW |
dc.date.accessioned | 2021-05-20T20:35:03Z | - |
dc.date.available | 2009-08-05 | |
dc.date.available | 2021-05-20T20:35:03Z | - |
dc.date.copyright | 2008-08-05 | |
dc.date.issued | 2008 | |
dc.date.submitted | 2008-07-30 | |
dc.identifier.citation | Ainsworth, T., Fine, M., Roff, G., and Hoegh-Guldberg, O. (2008) Bacteria are not the
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/9677 | - |
dc.description.abstract | 目前全世界的珊瑚礁都受到破壞,而珊瑚疾病為主要威脅因素之ㄧ。然而受
限於傳統微生物學方法限制,過去對於珊瑚疾病爆發的原因和感染機制仍不清 楚。2006 年,台灣綠島發生大量珊瑚死亡事件,原因為受到如黑色覆蓋物之海綿 (Terpios hoshinota)入侵所引起,簡稱「黑病」。本研究將利用電子顯微鏡方法與 非依賴培養式的微生物分子生物技術,細菌16S rDNA 序列和變性梯度電泳法 (denaturing gradient gel electrophoresis, DGGE),分析T. hoshinota 特有菌相,並比 較受海綿覆蓋之珊瑚(Porites lutea)與未受海綿覆蓋之珊瑚(P. lutea)其菌相變 化情形,期望發現與疾病有關微生物,以利往後病態監測所用。 結果發現,不論珊瑚或海綿,其菌相分布都與周遭海水菌相不同。T. hoshinota 細菌群相專一,主要為具光合作用的單細胞球形藍綠菌,經分子序列及外觀比對, 可能是新種藍綠菌。此藍綠菌數量極大應該是和T. hoshinota 存有某種程度的共生 關係。另外,利用DOTUR、LIBSHUFF 軟體來分析珊瑚P. lutea 的細菌群相之間 的多樣性和組成相似度。結果發現未罹病珊瑚P. lutea 的菌相分佈,主要以 Gammaproteobacteria、Cyanobacteria、unclassified bacteria 為主;而罹病珊瑚P. lutea 的菌相是以Gammaproteobacteria、Alphaproteobacteria 為主,而Cyanobacteria 以 及unclassified bacteria 則大量減少。過去研究發現,珊瑚上有許多共生細菌,當珊 瑚受到環境壓力時,可能改變或抑制原本正常菌叢,導致菌相會以某一類群細菌 為主, 而非專一性的細菌也會伺機而生; 本研究發現罹病珊瑚上 Gammaproteobacteria 大量增加,可能和環境壓力有關(受海綿覆蓋或海水污染), 而Alphaproteobacteria 則可能為伺機生長的細菌,而這樣的改變,影響正常菌叢對 珊瑚的功能,推測可能影響珊瑚健康。由於珊瑚上共生細菌易受環境壓力而改變, 未來若能建立珊瑚共生菌相長期監測資料,應可作為珊瑚的健康指標。 | zh_TW |
dc.description.abstract | Coral diseases are one of the major natural disturbances that threat the survival
of coral reefs worldwide. However, the characterization and mechanism of infection have been difficult in understanding the outbreak of coral diseases due to the limitation of applying classical microbiological assays. In this thesis, the bacteria consortium of the “black disease” caused by a black mat-like sponge, Terpios hoshinota, was characterized by electronic microscopy, denaturing gradient gel electrophoresis (DGGE), and 16S ribosomal DNA library construction. In order to understand the possible pathogenic bacteria in “black disease”, the specificity of T. hoshinota associated bacterial community and the variation between sponge-infected (SI) and non-sponge-infected (NSI) Porites lutea associated bacterial community were investigated in the fringing reef of the Green Island (Lutao), where outbreak of Terpios sponge was first reported in 2006. Result shows that the bacterial communities from seawater, sponge, and coral were specific. T. hoshinota associated bacterial community was specific and the dominant bacteria group was autotrophic Cyanobacteria. The Cyanobacteria associated with T. hoshinota were supposedly a new Cyanobacteria species as demonstrated by 16S rDNA sequence with a unique morphology. Furthermore, the high abundance of cyanobacteria in T. hoshinota may contribute positively to T. iii hoshinota outbreak. On the other hand, the diversity and composition similarity of bacterial community associated with SI and NSI P. lutea were analyzed using DOTUR and LIBSHUFF. In NSI P. lutea, the dominant bacterial groups were Gammaproteobacteria, Cyanobacteria, and unclassified bacteria; however, in SI P. lutea, Gammaproteobacteria and Alphaproteobacteria were major groups and the abundance of Cyanobacteria and unclassified bacteria were low. In previous studies, there were diverse coral holobiont on a coral. The natural coral holobiont changed to dominant bacteria group with abundance of opportunistic bacteria when corals experience environmental stress; similar situation was observed in SI P. lutea. The increase of the abundance of Gammaproteobacteria is probably related to environmental stress (sponge or seawater pollution). In addition, the unexpected emergence of alphaproteobacteria could be opportunistic bacteria because of disturbance of natural bacterial community. Disturbance of coral holobiont possible loss the normal symbiont function for coral and it directly or indirectly supposed to result in coral disease. Coral associated microbes were variable in different environment and may be a good bioindicator for environmental stress; therefore, building up a bioindicator, a long term investigation is apparently essential. | en |
dc.description.provenance | Made available in DSpace on 2021-05-20T20:35:03Z (GMT). No. of bitstreams: 1 ntu-97-R95241211-1.pdf: 5432520 bytes, checksum: 70babccdfc2764657a31c56e3ac6f528 (MD5) Previous issue date: 2008 | en |
dc.description.tableofcontents | 1 Introduction............................................................................................................1
1.1 Coral reef ecosystem degradation and mortality........................................1 1.2 Coral disease ..............................................................................................2 1.2.1 Definition of coral disease ...............................................................3 1.2.2 Microbial diseases of corals.............................................................3 1.2.3 Microbial-induced coral disease: two examples..............................5 1.2.4 The limitation of coral disease research ..........................................8 1.2.5 Coral disease studies in Taiwan .......................................................9 1.2.5.1 Black disease outbreak in Green Island ................................10 1.2.6 Terpios hoshinota, “the coral killer sponge” ................................. 11 1.2.6.1 Biology of Terpios hoshinota................................................13 1.2.7 Symbiotic relationships between sponge and endogenous microbe 14 1.2.7.1 Sponges as hosts of single-celled autotrophs........................17 1.2.7.2 Sponge as hosts of heterotrophic single-celled microbes .....19 1.3 Aim of this thesis .....................................................................................22 2 Materials and Methods.........................................................................................30 2.1 Sampling ..................................................................................................30 2.1.1 Seawater sampling .........................................................................31 2.1.2 Coral sampling...............................................................................31 2.1.3 Sponge sampling............................................................................32 2.2 Electron microscopy ................................................................................32 2.3 Molecular Method for bacterial library construction...............................33 2.3.1 DNA extraction..............................................................................33 2.3.2 Amplification of bacterial 16S ribosomal rRNA by polymer chain reaction 35 2.3.3 Clone library construction .............................................................36 2.3.4 Denaturing gradient gel electrophoresis analysis ..........................38 2.3.5 Recovery of DNA from DGGE gel ...............................................39 2.3.6 Sequencing and phylogenetic analysis ..........................................39 2.3.7 Bacterial group assignment............................................................40 2.3.8 Diversity estimation in DOTUR....................................................40 2.3.9 Sequence library comparison.........................................................41 2.3.10 Phylogenetic analyses....................................................................41 2.3.11 DGGE Cluster analysis..................................................................42 3 Results..................................................................................................................44 3.1 16S rRNA-DGGE analysis.......................................................................44 3.1.1 P. lutea associated bacteria communities changed when T. hoshinota covered .......................................................................44 3.1.2 Gammaproteobacteria were the major group in bacteria communities of seawater and P. lutea.........................................44 3.1.3 T. hoshinota associated bacterial community was specific..........45 3.1.4 Cyanobacteria were the major group in T. hoshinota associated bacterial community....................................................................45 3.2 16S rRNA library construction.................................................................50 3.2.1 Composition analysis of bacterial community.............................50 3.2.1.1 Gammaproteobacteria was the major group in seawater samples..................................................................................50 3.2.1.2 Bacterial communities associated with P. lutea changed when T. hoshinota covered .............................................................50 3.2.1.3 Cyanobacteria were the major group in the sponge .............51 3.2.2 Diversity analysis of bacterial 16S rRNA clone libraries ............58 3.2.2.1 Higher diversity of bacteria community associated with SI Porites lutea (close-to sponge part, PdI) ...............................58 3.2.2.2 Low diversity of bacterial community associated with T. hoshinota...............................................................................59 3.2.3 Comparing different bacterial communities ................................63 3.2.3.1 Different composition between Chai-Kou and Kun-Guan seawater clone libraries.........................................................63 3.2.3.2 Different composition between SI and NSI P. lutea clone libraries .................................................................................63 3.2.3.3 Similar bacterial communities associated with T. hoshinota covered on five different coral species .................................64 3.3 Phylogenetic analysis ...............................................................................68 3.3.1 Bacterial community associated with P. lutea .............................68 3.3.2 Bacterial community associated with T. hoshinota......................70 3.4 Electronic microscopy of T. hoshinota .....................................................79 4 Discussion............................................................................................................84 4.1 The role of bacterial community associated with T. hoshinota................84 4.2 New species of Cyanobacteria associated with T. hoshinota ..................85 4.3 The composition of bacterial community associated P. lutea..................87 4.3.1 The role of Cyanobacteria associated with coral ..........................89 4.3.2 Variation of bacterial community associated with coral................90 4.4 Future research.........................................................................................94 Reference 97 Appendix 107 Table Content Table 1 Standard Systematically Describing Coral Disease/Syndrome......................... 24 Table 2 Coral diseases observed in Taiwan.................................................................... 25 Table 3 Standard systematically description of black disease ....................................... 25 Table 4 The bacteria taxonomy in samples based on 16S rRNA bacteria clone libraries using RDP II analysis....................................................................................... 52 Table 5 Diversity indices for the bacterial communities (Porites lutea and seawater)... 60 Table 6 Diversity indices for the bacterial communities (sponge) ................................. 61 Table 7 Difference in bacteria 16S rRNA clone libraries composition........................... 65 Table 8 Difference in bacteria 16S rRNA clone libraries composition........................... 66 Table 9 16S rRNA gene sequence analysis of bacterial clones from the non-sponge-infected Porites lutea ................................................................... 72 Table 10 16S rRNA gene sequence analysis of bacterial clones from the sponge-infected Porites lutea (far-away sponge, PdH) .............................................................. 73 Table 11 16S rRNA gene sequence analysis of bacterial clones from the sponge-infected Porites lutea (close-to sponge, PdI) ................................................................. 74 Table 12 16S rRNA gene sequence analysis of bacterial clones from Terpios hoshinota (infected Porites lutea) 76 Figure Content Fig. 1 Coral diseases observed in Taiwan ...................................................................... 26 Fig. 2 A large area of coral reef was covered by the blackish mat................................. 26 Fig. 3 Different morphologies of corals were covered by the blackish mat. ................. 27 Fig. 4 Close-up of black disease infecting Porites sp.................................................... 27 Fig. 5 Close-up of sponge crust. .................................................................................... 28 Fig. 6 Portions of tylostyle head (spicule). .................................................................... 28 Fig. 7 Transmission electron microscopy of T. hoshinota. ............................................ 28 Fig. 8 Progress rate of T. hoshinota in Green Island...................................................... 29 Fig. 9 Scars on T. hoshinota suspected by fish bite ....................................................... 29 Fig. 10 News of black disease outbreak in Taiwan........................................................ 29 Fig. 11 P. lutea sampling procedure............................................................................... 43 Fig. 12 T. hoshinota sampling procedure....................................................................... 43 Fig. 13 T. hoshinota covering on four kinds of coral species. ....................................... 43 Fig. 14 Bacterial community composition (according to Fig. 15A 16S rRNA-DGGE gel and cluster diagram)............................................................................................ 46 Fig. 15 Bacterial community composition on Porites lutea, Terpios hoshinota and seawater............................................................................................................... 47 Fig. 16 Bacterial community composition (according to Fig. 17 16S rRNA-DGGE gel and cluster diagram)............................................................................................ 48 Fig. 17 Bacterial community composition in sponge-infected five different coral species ............................................................................................................................. 49 Fig. 18 Pie charts illustrating the diversity of bacterial groups on Porites lutea and seawater............................................................................................................... 55 Fig. 19 Pie charts illustrating the diversity of bacterial groups on Terpios hoshinota.... 56 Fig. 20 Comparison of predominant bacterial community among samples ................... 57 Fig. 21 Rarefaction curves for the 16S rRNA bacteria clone libraries (Porites lutea and seawater). ............................................................................................................ 62 Fig. 22 Rarefaction curves for the 16S rRNA bacteria clone libraries (Terpios hoshinota collected from five coral species) ....................................................................... 62 Fig. 23 Non-metric multidimensional scaling plot (nMDS-2 dimensional) of Porites lutea, Seawater.................................................................................................... 67 Fig. 24 Non-metric multidimensional scaling plot (nMDS-2 dimensional) of Terpios hoshinota............................................................................................................. 67 Fig. 25 Maximum parsimony tree of sponge-associated and black band disease related cyanobacteria based on 16S rRNA. .................................................................... 77 Fig. 26 Neighbor-Joining tree of Porites lutea -associated and seawater related bacteria based on 16S rRNA 78 Fig. 27 Terpios hoshinota infected coral Isopora palifera. Black arrow: close-to coral, white arrow: far-away coral. ............................................................................... 80 Fig. 28 Transmission electron micrographs of Terpios hoshinota .................................. 81 Fig. 29 Transmission electron micrographs of Cyanobacteria....................................... 82 Fig. 30 Transmission electron micrographs of Cyanobacteria membrane ..................... 83 Fig. 31 Transmission electron micrographs of cyanobacterial symbionts in Ircinia variabilis from Marseille. .................................................................................. 96 | |
dc.language.iso | en | |
dc.title | 造礁珊瑚黑病之菌相分佈研究 | zh_TW |
dc.title | Bacterial consortium associated with the“black disease'of reef-building corals | en |
dc.type | Thesis | |
dc.date.schoolyear | 96-2 | |
dc.description.degree | 碩士 | |
dc.contributor.coadvisor | 湯森林 | |
dc.contributor.oralexamcommittee | 王志騰,宋克義,陳文明 | |
dc.subject.keyword | 海綿,藍,綠,菌,珊瑚疾病,珊瑚微生物菌相,非培養式微生物分子技術, | zh_TW |
dc.subject.keyword | sponge,Cyanobacteria,coral disease,coral microbiology,uncultured molecular method, | en |
dc.relation.page | 108 | |
dc.rights.note | 同意授權(全球公開) | |
dc.date.accepted | 2008-07-30 | |
dc.contributor.author-college | 理學院 | zh_TW |
dc.contributor.author-dept | 海洋研究所 | zh_TW |
顯示於系所單位: | 海洋研究所 |
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