本文關(guān)鍵詞：云南沼氣發(fā)酵生態(tài)系統(tǒng)的原核生物群落時(shí)空動(dòng)態(tài)研究　出處：《云南大學(xué)》2016年博士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 沼氣發(fā)酵系統(tǒng) 原核生物群落 時(shí)空動(dòng)態(tài) 高通量測序技術(shù) DGGE 微生物多樣性

【摘要】：沼氣工程的推廣應(yīng)用在保護(hù)生態(tài)環(huán)境和減少溫室氣體排放等方面具有重要的意義。在農(nóng)村地區(qū)發(fā)展戶用沼氣工程能夠改進(jìn)農(nóng)村能源使用途徑,治理農(nóng)業(yè)面源污染和促進(jìn)農(nóng)業(yè)生態(tài)的良性循環(huán)。沼氣發(fā)酵生態(tài)系統(tǒng)中大量的原核生物與系統(tǒng)的高效穩(wěn)定運(yùn)行有著密切關(guān)系。沼氣發(fā)酵過程中的工藝參數(shù)和原料來源等環(huán)境因子從多方面影響著發(fā)酵系統(tǒng)中原核生物的數(shù)量、群落結(jié)構(gòu)和群落功能,而原核生物的代謝活動(dòng)反過來又會(huì)影響沼氣發(fā)酵生態(tài)系統(tǒng)的穩(wěn)定運(yùn)行。因此,沼氣發(fā)酵系統(tǒng)中原核生物的群落結(jié)構(gòu)組成對(duì)產(chǎn)氣效率起著十分重要的作用。目前,云南農(nóng)村戶用沼氣池?cái)?shù)量多,但沼氣池在管理和使用過程中仍然存在許多問題,如沼氣池產(chǎn)氣不穩(wěn)定、產(chǎn)氣量少、冬季停用現(xiàn)象。針對(duì)云南農(nóng)村戶用沼氣池的問題,目前的研究主要從工程角度出發(fā),多集中在沼氣池的工藝參數(shù)優(yōu)化、保溫措施和活性污泥馴化等方面研究上,但是對(duì)云南農(nóng)村戶用沼氣發(fā)酵生態(tài)系統(tǒng)的重要生物因素—原核生物及其與環(huán)境因子相關(guān)性的研究幾乎無人涉及。因此,對(duì)云南農(nóng)村戶用沼氣發(fā)酵生態(tài)系統(tǒng)中原核生物的群落結(jié)構(gòu)和功能、原核生物與環(huán)境因子相互關(guān)系的研究十分有意義。通過生態(tài)因子綜合分析,揭示沼氣發(fā)酵系統(tǒng)中原核生物群落的演替規(guī)律等基本生態(tài)學(xué)問題,能夠?yàn)樵O(shè)計(jì)具有應(yīng)用前景的高效沼氣發(fā)酵系統(tǒng),活性污泥馴化,發(fā)酵條件提供最優(yōu)監(jiān)控方案和規(guī)�；瘧�(yīng)用奠定基礎(chǔ)。本研究是從云南4個(gè)不同氣候類型與地理區(qū)域的代表性農(nóng)村戶用沼氣發(fā)酵池中采集活性污泥,用Illumina高通量測序技術(shù),系統(tǒng)地分析云南不同氣候類群與地理區(qū)域的農(nóng)村戶用沼氣池中原核生物的類群組成、群落結(jié)構(gòu)和群落功能,并闡述其與環(huán)境中非生物因子間的相關(guān)性,進(jìn)而探討不同氣候區(qū)域的沼氣池中原核生物群落空間動(dòng)態(tài)(地理格局)分布。其次,運(yùn)用DGGE和Illumina高通量測序技術(shù),研究模擬云南熱帶氣候區(qū)域的農(nóng)村戶用沼氣發(fā)酵過程中原核生物群落的時(shí)間動(dòng)態(tài)變化。深入分析整個(gè)過程中原核生物群落結(jié)構(gòu),探討原核生物群落在時(shí)間上的動(dòng)態(tài)演替規(guī)律。研究結(jié)果顯示,云南農(nóng)村戶用沼氣發(fā)酵生態(tài)系統(tǒng)中存在豐富的原核生物類群。在不同氣候類型與地理區(qū)域的沼氣發(fā)酵生態(tài)系統(tǒng)中,原核生物群落結(jié)構(gòu)存在較大差異。在模擬沼氣發(fā)酵過程中,不同的原核生物類群的動(dòng)態(tài)變化規(guī)律不一樣。1.云南農(nóng)村戶用沼氣發(fā)酵生態(tài)系統(tǒng)中存在豐富的原核生物類群。Illumina高通量測序結(jié)果顯示,云南4個(gè)不同氣候區(qū)域的農(nóng)村戶用沼氣生態(tài)系統(tǒng)中存在著豐富的細(xì)菌和古菌類群。從10個(gè)沼氣池中平均獲得61,000多條有效序列,聚類分析發(fā)現(xiàn)發(fā)酵系統(tǒng)中有16個(gè)已知門類群：泉古菌門(Crenarchaeota)、廣古菌門(Euryarchaeota)、擬桿菌門(Bacteroidetes)、酸桿菌門(Acidobacteria)、放線菌門(Actinobacteria)、綠菌門(Chlorobi)、纖維桿菌門(Fibrobacteres)、綠彎菌門(Chloroflexi)、厚壁菌門(Firmicutes)、芽單胞菌門(Gemmatimonadetes)、變形菌門(Proteobacteria)、螺旋體門(Spirochaetes)、Synergistetes、熱袍菌門(Thermotogae)、疣微菌門(Verrucomicrobia)和Caldithrix等和5個(gè)候選門類群。其中,厚壁菌門(Firmicutes)、廣古菌門(Euryarchaeota)和擬桿菌門(Bacteroidetes)在該生態(tài)系統(tǒng)中占著絕對(duì)的優(yōu)勢地位。2.PCA結(jié)果顯示,不同氣候區(qū)域的沼氣發(fā)酵系統(tǒng)中原核生物的群落結(jié)構(gòu)存在差異,同一氣候區(qū)域的原核生物群落結(jié)構(gòu)相近。厚壁菌門(Firmicutes)所占的比例范圍為22-55%,其中它在北溫帶的XGP2和XGP3比例最高,分別為54.6%和41.1%。廣古菌門(Euryarchaeota)的豐度變化范圍為4.1%到46%,同樣差異明顯。與厚壁菌門(Firmicutes)豐度相反,廣古菌門(Euryarchaeota)在XGP2和XGP3中比例最低,僅為8.5%和4.1%。擬桿菌門(Bacteroidetes),在各個(gè)樣點(diǎn)中的豐度存在差異,為10-27%。原核生物的群落結(jié)構(gòu)與沼氣發(fā)酵系統(tǒng)的產(chǎn)氣量存在一定的相關(guān)性。3.應(yīng)用DGGE指紋圖譜技術(shù)研究模擬沼氣發(fā)酵過程,結(jié)果表明,系統(tǒng)中的細(xì)菌和古菌類群具有很高的多樣性,細(xì)菌類群的數(shù)量高于古菌類群。對(duì)原核生物的動(dòng)態(tài)變化進(jìn)行聚類分析,發(fā)現(xiàn)整個(gè)發(fā)酵過程中細(xì)菌和古菌的群落結(jié)構(gòu)不斷在變化；細(xì)菌類群的豐度變化明顯大于古菌類群。整個(gè)發(fā)酵過程中細(xì)菌和古菌的群落結(jié)構(gòu)變化規(guī)律可以分為3個(gè)階段,但細(xì)菌類群的變化階段明顯早與古菌類群的變化。4. Illumina高通量測序結(jié)果表明,模擬沼氣發(fā)酵過程中涉及的原核生物類群主要?dú)w屬于19個(gè)門一級(jí)的系統(tǒng)發(fā)育類群：泉古菌門(Crenarchaeota)、廣古菌門(Euryarchaeota)、厚壁菌門(Firmicutes)、擬桿菌門(Bacteroidetes)、變形菌門(Proteobacteria)、螺旋體門(Spirochaetes)、Synergistetes、綠彎菌門(Chloroflexi)、酸桿菌門(Acidobacteria)、放線菌門(Actinobacteria)、纖維桿菌門(Fibrobacteres)、綠菌門(Chlorobi)、疣微菌門(Verrucomicrobia)、Armatimonadetes、Caldithrix、 Lentisphaerae、熱袍菌門(Thermotogae)和OP9、OP11等。廣古菌門(Euryarchaeota)、厚壁菌門(Firmicutes)、擬桿菌門(Bacteroidetes)和變形菌門(Proteobacteria)同樣是沼氣發(fā)酵系統(tǒng)中的主要優(yōu)勢類群。在模擬沼氣發(fā)酵過程中,沼氣發(fā)酵系統(tǒng)的原核生物變化規(guī)律同樣分為3個(gè)階段,不同類群的原核生物變化趨勢不一致。廣古菌門(Euryarchaeota)在發(fā)酵過程中的豐度變化趨勢為增加,相對(duì)豐度由發(fā)酵初期的32.4%到發(fā)酵末期的60.5%。厚壁菌門(Firmicutes)相對(duì)豐度由發(fā)酵初期的19.2%到發(fā)酵末期的6.4%,與廣古菌門(Euryarchaeota)的變化趨勢相反。擬桿菌門(Bacteroidetes),相對(duì)豐度由發(fā)酵初期的15.7%到發(fā)酵末期的5.9%,變化趨勢同樣是減少。一些細(xì)菌類群的相對(duì)豐度變化與產(chǎn)氣量變化相一致,與產(chǎn)氣量存在一定的相關(guān)性。5.云南不同氣候類型與地理區(qū)域的農(nóng)村戶用沼氣發(fā)酵系統(tǒng)中,原核生物的群落分布與環(huán)境因子(發(fā)酵條件)存在著顯著的相關(guān)性。RDA分析結(jié)果顯示,沼氣發(fā)酵系統(tǒng)中的總固體(TS)和化學(xué)需氧量(COD)是影響細(xì)菌類群地理空間分布的最主要非生物因子,而溫度是影響古菌屬群落分布的主要環(huán)境因子。而在模擬沼氣發(fā)酵過程中,TS和COD同樣是影響沼氣發(fā)酵細(xì)菌類群時(shí)間動(dòng)態(tài)變化的主要非生物因子;恒溫條件下,主要產(chǎn)甲烷古菌類群受到發(fā)酵條件的影響不大。
[Abstract]:It has important significance to promote the application of Biogas Engineering in protecting ecological environment and reduce greenhouse gas emissions. The development of household biogas project can improve the rural energy use way in rural areas, management of agricultural non-point source pollution and promote agricultural ecological circulation. There is a close relationship between the efficient and stable operation of methane generator systems with a large number of prokaryotes and yeast in the ecological system. The process parameters and sources of raw materials and other environmental factors in the process of biogas fermentation from many aspects affect the number of fermentation system of prokaryotic, community structure and function, and metabolic activity of prokaryotic organisms will in turn affect the stable operation of Biogas fermentation ecosystem. Therefore, the community structure of the biogas fermentation system of Central Plains nuclear biological composition plays an important role in gas production efficiency. At present, the number of household biogas digesters in rural areas of Yunnan, but the tube There are still many problems and the use of the process, such as methane stable gas production, gas consumption, winter stop phenomenon. For biogas plants in rural areas of Yunnan, the current study mainly from the engineering point of view, optimization of process parameters and more concentrated in the digester, research on thermal insulation measures and activated sludge etc. However, research on important biological factors in Yunnan Prokaryotae rural household biogas fermentation ecosystem and its correlation with environmental factors almost nobody involved. Therefore, the community structure and function of biogas fermentation ecosystem of prokaryotic households in rural areas of Yunnan, the relationship between prokaryotes and environmental factors is of great significance. Through comprehensive analysis ecological factors, reveal the biogas fermentation system of prokaryotic community succession and other basic ecological problems, efficient biogas to design with application prospect Gas fermentation system, activated sludge, fermentation conditions to provide optimal solutions and lay the foundation for monitoring large-scale applications. This study is the active sludge collected from Yunnan biogas 4 different types of climate and geographical areas representative of rural household fermentation tank, using Illumina high-throughput sequencing technology, system analysis of Yunnan climate and different groups geographical area of rural household biogas digesters of prokaryotic species, community structure and function, and expounds the relationship between the non biological factors and the environment, and to explore the different climatic regions of biogas pool of prokaryotic community spatial dynamic (geographical pattern) distribution. Secondly, using DGGE and Illumina high-throughput sequencing technology dynamic research on Simulation of Yunnan tropical climate region of the rural household biogas fermentation process of prokaryotic organisms. In-depth analysis of the whole process of prokaryotic biological community To investigate the structure, dynamic succession at the time of the prokaryotic community. The results showed that the presence of prokaryote taxa rich Yunnan rural household biogas fermentation ecosystem. Biogas fermentation ecosystem in different types of climate and geographical area, there is a big difference between the prokaryotic community structure. In the simulation process of biogas fermentation, high.Illumina groups of prokaryotes the sequencing results showed the presence of abundant amount of dynamic changes of different groups are not the same as the prokaryote.1. Yunnan rural household biogas fermentation ecosystem, there are bacteria and archaea rich groups of Yunnan in 4 different climate regions of the rural household biogas system. The average obtained more than 61000 effective sequences from 10 digesters in the cluster analysis showed that the fermentation system there are 16 known categories: Crenarchaeota group (Crenarchaeota), Euryarchaeota (Euryarcha Eota), Bacteroidetes (Bacteroidetes), acidobacteria (Acidobacteria), actinobacteria (Actinobacteria), green (Chlorobi), bacterial phylum fibrobacteres (Fibrobacteres), Chloroflexi (Chloroflexi), Firmicutes (Firmicutes), gemmatimonadetes (Gemmatimonadetes). Proteobacteria (Proteobacteria), spiral gate (Spirochaetes), Synergistetes, thermotogae (Thermotogae), verrucomicrobia (Verrucomicrobia and Caldithrix) and 5 candidate door groups. Among them, the Firmicutes (Firmicutes), Euryarchaeota (Euryarchaeota) and Bacteroidetes (Bacteroidetes in the ecological system) dominating the.2.PCA absolute that community structure in different climate regions of the biogas fermentation system of prokaryotic differences, the same group of prokaryotes in climatic regions fall. Similar structures of Firmicutes (Firmicutes) the proportion of the range of 22-55%, in which it North temperate XGP2 and XGP3 highest proportion, respectively 54.6% and 41.1%. Euryarchaeota (Euryarchaeota) abundance ranged from 4.1% to 46%, the same difference. And Firmicutes (Firmicutes) abundance in Euryarchaeota (Euryarchaeota) in XGP2 and XGP3 in the lowest rate, only 8.5% and 4.1%. (Bacteroidetes), Bacteroidetes abundance in each sample point of difference for community structure and gas production of biogas fermentation system 10-27%. in prokaryotes have certain correlation.3. DGGE fingerprint technique of simulated biogas fermentation process, the results show that the system of bacteria and archaea, bacteria with high diversity the number of groups is higher than that of bacteria, archaea. The dynamic changes of the prokaryotic community structure found by clustering analysis of bacteria and archaea in the whole fermentation process of the continuous changes in the abundance of bacterial groups significantly; More than archaea. Community structure changes of bacteria and archaea in the whole fermentation process can be divided into 3 stages, but the change was earlier stage groups of bacteria and archaea. The changes of.4. Illumina high-throughput sequencing results showed that prokaryotic organisms belonging mainly involved in the process of biogas fermentation system simulation in 19 a development group: Crenarchaeota (Crenarchaeota), Euryarchaeota (Euryarchaeota), (Firmicutes) the Firmicutes, Bacteroidetes and Proteobacteria (Bacteroidetes) (Proteobacteria), spiral gate (Spirochaetes), Synergistetes, Chloroflexi (Chloroflexi), acid bacillus the door (Acidobacteria), actinobacteria (Actinobacteria), fibrobacteres (Fibrobacteres), green (Chlorobi), bacterial phylum verrucomicrobia (Verrucomicrobia), Armatimonadetes, Caldithrix, Lentisphaerae, thermotogae (Thermotogae) and OP9, OP11 and so on. The Archaea The door (Euryarchaeota), (Firmicutes) the Firmicutes, Bacteroidetes and Proteobacteria (Bacteroidetes) (Proteobacteria) is also the dominant group biogas fermentation system. In the simulation process of biogas fermentation, prokaryotes variation of biogas fermentation system of the same is divided into 3 stages, the change trend of different groups of prokaryotes inconsistent. Euryarchaeota (Euryarchaeota) abundance change trend in the process of fermentation was increased by 60.5%., the relative abundance of Firmicutes to 32.4% at the end of fermentation at the initial stage of fermentation of the relative abundance (Firmicutes) from 19.2% to 6.4% at the end of fermentation fermentation stage, and Euryarchaeota (Euryarchaeota) changes on the contrary. The Bacteroidetes (Bacteroidetes), the relative abundance of from 15.7% to 5.9% at the end of fermentation fermentation stage, the same trend is reduced. Some bacterial taxa relative abundance changes and the changes of the gas production. The correlation between.5. and Yunnan in different climate types with a certain geographic region and gas production of rural household biogas fermentation system, community distribution and environmental factors in prokaryotes (fermentation) there is a significant correlation between.RDA analysis results showed that the total solids in the biogas fermentation system (TS) and chemical oxygen demand (COD) is the influence of geographical space distribution of bacterial taxa most major abiotic factors, while temperature is the main environmental factors affecting the community distribution of ancient bacteria. And in the simulation of methane fermentation process, TS and COD are also major abiotic factors affecting the change of population dynamic time fine biogas fermentation fungus; under the condition of constant temperature, mainly produced little effect methane Archaea by fermentation conditions.

【學(xué)位授予單位】：云南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：S216.4

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