Hybrid read-based and genome-centric investigation of laboratory scale anaerobic digestion using various second and third generation substrates

Biogas production from co-digestion of second and third generation substrates is a promising alternative to replace similar complex polysaccharide containing first generation biomasses. In the present study energy willow (EW), simple water pre-treated chicken manure (TCM) and mixed microalgal-bacterial (MABA) biomass cultivated on wastewater by-product of chicken manure treatment were used. The BMP tests showed that biogas production of co-digestion was ~21% higher compared to the production using EW alone indicating a synergistic effect of co-substrates. During the AD in semi-CSTR, a slow but balanced operation was observed due to the optimized C/N ratio. In order to gain insights into the complex anaerobic decomposing microbiome and its functional network hybrid read-based and genome-centric metagenomics were employed. This hybrid investigation revealed that the major bacterial participants were the representatives of Firmicutes, Actinobacteria, Proteobacteria and Bacteroidetes phyla, while Methanomicrobia and Methanobacteria represented the archaeal constituents of the specific biodegrading community. Several CAZymes were found indicating active complex polysaccharide degradation processes. The bacterial phyla were shown to perform the long chain carbohydrate hydrolysis while methanogenesis were linked to the slow growing members of the complex community. It was concluded that an optimized balance between H2 producers and consumers was critical for the efficient and stable operation of the biogas microbial community utilizing second and third generation feedstock.

以第二代与第三代底物进行共厌氧消化制取沼气，是替代含复合多糖的第一代生物质原料的极具潜力的方案。本研究选用了能源柳（energy willow, EW）、简易水预处理鸡粪（simple water pre-treated chicken manure, TCM）以及以鸡粪处理废水副产物为培养基培养的混合微藻-细菌生物质（mixed microalgal-bacterial biomass, MABA）。生物甲烷潜力（Biochemical Methane Potential, BMP）测试结果显示，共消化体系的沼气产量较单一能源柳体系提升约21%，表明共底物存在协同效应。在半连续搅拌槽式反应器（semi-continuous stirred-tank reactor, semi-CSTR）的厌氧消化（Anaerobic Digestion, AD）过程中，由于优化了碳氮比，体系运行缓慢但保持稳定平衡。为深入解析复杂的厌氧分解微生物组及其功能网络，本研究采用了基于读长与基因组中心相结合的宏基因组学技术。该联合研究揭示，群落中的主要细菌类群隶属于厚壁菌门（Firmicutes）、放线菌门（Actinobacteria）、变形菌门（Proteobacteria）与拟杆菌门（Bacteroidetes）；而产甲烷微菌纲（Methanomicrobia）与产甲烷杆菌纲（Methanobacteria）则构成了该特异性降解群落的古菌组分。研究检测到多种碳水化合物活性酶（Carbohydrate-Active EnZymes, CAZymes），表明体系存在活跃的复合多糖降解过程。细菌类群负责长链碳水化合物的水解过程，而产甲烷作用则与该复杂群落中生长缓慢的类群相关联。本研究最终得出结论：氢气产生菌与消耗菌之间的优化平衡，对于利用第二代与第三代原料的沼气微生物群落实现高效稳定运行至关重要。