基于多组学的产酸发酵菌群纤维素代谢网络分析

Carboxylate platform based on undefined mixed culture-based fermentation technology is a new strategy for the production of biofuels and biobased products from lignocellulosic biomass. However, the mixed culture is undefined and the mechanisms of cellulose degradation and carboxylate production are not clear. To reveal the cellulose metabolism of carboxylate-producing mixed culture is of importance in promoting the development and application of mixed culture-based fermentation technology. With the pointcut of how mixed culture to response pH stress via regulating and controlling of metabolic networks, multi-omics analysis is employed to reveal the cellulose metabolism in a carboxylate-producing microbial community. Metagenomic and metatranscriptomic sequencing technologies will be used to characterize the microbial community structure and to determine the expression activities of key genes related to cellulose metabolism. The cellulose metabolic network of the microbial community will be predicted based on metagenomic and metatranscriptomic data. Metabolomic study will be performed to determine the major metabolite changes in the microbial community induced by pH stress, and to discover the metabolic pathways related to the response to pH stress. Association analysis of bacteria spectrum, gene expression profile and metabolic phenotypes will be performed to reveal the cellulose metabolism of carboxylate-producing microbial community, which offers theoretical support for the process optimization of mixed culture-based process. In consideration of general characteristics in acidogenic fermentation, the findings in this proposal will have reference meaning to perform similar investigation and research in anaerobic digestion and anaerobic biotreatment of wastewater.

基于混合菌群发酵技术的羧酸平台是木质纤维素生物质转化生物能源与生物基产品的新策略，产酸发酵菌群的纤维素代谢网络研究对于推动混合菌群发酵技术的发展与应用具有重要的科学意义。本项目以菌群如何通过代谢网络运行与调控以响应pH变化为切入点，运用多组学技术扩展探索产酸发酵菌群的纤维素代谢网络及其调控机制。采用宏基因组和宏转录组技术监测pH调控过程中混合菌群的遗传多样性，比较特定功能菌群及关键基因转录变化，预测菌群纤维素代谢网络及其响应pH变化的调控策略。采用代谢组学技术监测混合菌群代谢组变化，比较并鉴定其中的关键代谢物，通过与宏基因组学和宏转录组学研究结果的关联分析与交叉验证，综合解析混合菌群的纤维素代谢网络，为混合菌群发酵工艺的过程优化与控制提供理论支持。鉴于产酸发酵过程的共性，本项目研究结果亦对厌氧消化以及废水厌氧生物处理等研究与应用具有借鉴意义。

本项目首先以纤维素降解产酸发酵混合菌群的pH响应机制为切入点，运用高通量测序技术研究了pH调控下的纤维素降解产酸发酵混合菌群演替规律。研究结果表明：纤维素发酵系统中pH为4.0时，优势菌为Pseudomonas、Clostridium sensu stricto、Lactobacillus，发酵类型表现为典型的乙醇型发酵；当pH为5.0时，功能菌主要有Clostridium sensu stricto、Phascolarctobacterium，呈现出典型的混合酸发酵；当pH为6.0时，优势菌为Clostridium sensu stricto、Levilinea、Longilinea，表现出典型的丁酸型发酵；而当pH为pH7.0时，功能菌主要有Levilinea、Longilinea Phascolarctobacterium，呈现出典型的丙酸型发酵。在次基础上，采用宏基因组和转录组技术解析了pH调控过程中混合菌群的遗传多样性及其代谢多样性，构建了产酸发酵混合菌群的纤维素代谢网络。研究结果表明：纤维素降解产酸发酵代谢存在着显著的多样性，其多样性与pH值的调控有正相关。研究结果表明： pH为4.0时，纤维素产酸发酵系统中主要代谢产物为乙酸和乙醇；pH为5.0时，纤维素产酸发酵系统中主要代谢产物为乙酸、丙酸和丁酸。pH为6.0时，纤维素产酸发酵系统中主要代谢产物为乙酸和丁酸。而pH为7.0时，纤维素产酸发酵系统中主要代谢产物为乙酸和丙酸。最后，研究了中性DES介导的双螺旋挤压生物质预处理技术，DES介导的挤压预处理工艺作为一种高效的生物质预处理技术具有较大的应用潜力。总之，本项目的研究结果为混合菌群发酵工艺的过程优化与控制提供理论支持，为实现纤维素的高效综合利用和资源化提供了新技术。