沁水煤层气田原位微生物多样性及其在煤生物降解中的作用研究

Microbially enhanced coal bed methane (CBM) not only has the potential to produce new CBM, also could improve the exploitation of CBM via increasing the permeability of coal seam. The essence of the technology is the biodegradation of coal. The key of it is the ability of functional microorganism to produce methane by degrading coal. Thus, that investigating the structure and metabolism of the functional microbial communities in situ would enhance the understanding of the mechanism of coal biodegradation, promote the development of the technology. In this study, the coal and produced water will be sampled from Qinshui CBM field. The bacterial, archaeal and fugal communities in the samples will be analyzed by the culture-independent methods of microbial molecular ecology. The active microbial communities will also be investigated via RNA analysis. Then, the information about the in situ microbial resources in the CBM field will be obtained to explore the microorganism with potential function to degrade coal. Meanwhile, the potential to produce methane by the microbial communities in the coal and produced water will be determined by enrichments. And the intermediates of coal biodegradation will be examined by chromatographic technology. The succession of functional microbial communities will also be determined by the methods of microbial molecular ecology. Then, the metabolic pathway of coal biodegradation and the role of the functional microbial communities will be discussed, to lay a foundation for interpreting the pathway and mechanism of producing methane by the coal biodegradation, and to provide a basis for the application of microbially enhanced CBM in the region with high rank coal.

微生物增产煤层气不仅具有产出新煤层气的资源潜力，而且能够增加煤层渗透性，有利于煤层气的开采。该技术的核心是煤的生物降解，关键在于微生物的降解煤产甲烷能力。研究煤层气田原位微生物群落结构及其降解煤的代谢特征，将有效增强微生物降解煤产甲烷机理的理解，促进微生物增产煤层气技术的发展。本项目拟以沁水煤层气田为研究对象，通过采集产出水及煤样，利用非培养的分子生态学手段，从细菌、古菌和真菌三方面研究煤层气田原位微生物群落结构，同时以RNA表征活性微生物，获取原位微生物资源信息，找寻潜在功能微生物及其相互关系。通过富集培养结合色谱分析技术检测产出水及煤样中功能微生物的产甲烷潜能以及生物降解煤的中间代谢产物，同时利用分子生态学方法研究培养过程中功能微生物的演替，探寻生物降解煤的代谢途径及各功能微生物菌群的作用，为阐明煤的生物降解产甲烷途径和机理奠定基础，为微生物增产煤层气技术在高阶煤区域的应用提供依据。

微生物增产煤层气技术的核心思想是微生物厌氧降解煤产甲烷。通过研究煤层原位微生物群落、分析原位微生物菌群降解煤的代谢特征，能够为阐明微生物厌氧降解煤产甲烷机理奠定基础，从而为找寻技术突破提供指导。项目围绕煤层原位微生物群落结构及其降解煤的代谢特征，选取沁水盆地煤层气产出水和目标煤层煤为样品。研究结果发现，在煤层气产出水和煤中都生存有丰富的真菌、细菌、古菌。其中，真菌主要为Ascomycota和Basidiomycota门，细菌主要为Proteobacteria门，古菌主要是产甲烷菌。在不同煤层气井之间、产出水与煤之间，产甲烷菌无显著差异，都以氢营养型Methanobacterium为主，说明沁水盆地的生物成因气主要是利用氢气和二氧化碳生成；而真菌和细菌群落在属级别差异较大，尽管检测到的真菌和细菌在功能上多以降解大分子化合物或生成简单小分子化合物为主。在实验室模拟产气实验中，产出水中的真菌表现出极为显著的煤降解和产甲烷互作能力，真菌在煤层原位生物煤层气的生成过程中发挥重要作用。在实验室模拟产气过程中，菌群多样性随时间先减小后增大。未添加抗生素的实验组在培养过程中，优势产甲烷菌群不断转变，由氢营养型Methanobacterium到乙酸营养型Methanosaeta再到最后的混合营养型Methanosarcina。由此，产甲烷类型发生了转移。添加抗生素的实验组的优势产甲烷菌群在培养过程中基本保持恒定，都以氢营养型Methanobacterium为主。中间产物方面，整体表现出芳香类减少、小分子化合物增加的趋势。未添加抗生素的实验组内脂肪酸类化合物经历了先升后降的变化过程，在降解中期达到最高值；而芳香类化合物被不断分解。添加抗生素的实验组，脂肪酸在整个过程持续累积，而芳香烃类在早期则全部被降解。官能团方面，添加抗生素后，降解早期的含氧区域发生了截然相反的变化。研究结果揭示了沁水盆地煤层原位真菌、细菌、产甲烷菌菌群结构，明确了产甲烷菌分别与真菌、细菌和真菌互作下的厌氧降解煤产甲烷过程中功能菌群、中间产物、煤官能团的演替规律，为理解微生物增产煤层气的机理、推动其工业应用提供理论指导。