海藻糖强化高盐胁迫下异养硝化-好氧反硝化菌群的代谢机制

Mechanism of trehalose-enhanced metabolism of heterotrophic nitrification-aerobic denitrification community under high-salt stress

摘要:

异养硝化-好氧反硝化(heterotrophic nitrification-aerobic denitrification,HN-AD)菌是一类可在高盐环境脱氮的好氧微生物，但其工程应用效果不理想。海藻糖作为相容性溶质，通过参与调节细胞渗透压帮助微生物抵抗高盐胁迫，对提升高盐环境菌群的脱氮效率起重要作用。本研究通过启动膜曝气生物膜反应器(membrane aerobic biofilm reactor,MABR)富集HN-AD菌，设计添加150μmol/L海藻糖的C₁₅₀实验组和未添加海藻糖的C₀对照组，开展了外源性海藻糖对高盐胁迫下HN-AD菌群代谢的强化机制研究。反应器运行性能及群落结构分析结果显示，C₁₅₀组相较C₀组，NH₄⁺-N、总氮(total nitrogen,TN)和化学需氧量(chemical oxygen demand,COD)去除率分别提高29.7%、28.0%和29.1%；以不动杆菌属(Acinetobacter)和假黄褐藻属(Pseudofulvimonas)为优势菌属的耐盐型HN-AD菌群总相对丰度在C₁₅₀组达到66.8%、较C₀组提高了18.2%，添加海藻糖促进高盐环境中耐性型HN-AD菌群富集并强化系统脱氮性能。代谢组学深度分析表明，外源性海藻糖加强脯氨酸合成，提高微生物对高盐胁迫的抵抗能力；通过调节细胞增殖信号通路(cGMP-PKG、PI3K-Akt)、磷脂代谢通路及氨酰基-tRNA合成通路的活性，促使甘油磷脂代谢物磷酸乙醇胺及嘌呤和嘧啶丰度上调，提升细菌聚集能力和细胞增殖，助推微生物在高盐环境生长；同时，添加海藻糖还加快三羧酸循环(tricarboxylic acid cycle,TCA cycle)，为HN-AD菌群的碳、氮代谢提供更多电子供体与能量，进而优化系统脱氮性能。本研究结果为HN-AD菌在高盐高氮废水处理中的运用提供理论指导。

Abstract:

Heterotrophic nitrification-aerobic denitrification (HN-AD) bacteria are aerobic microorganisms that can remove nitrogen under high-salt conditions, but their performance in practical applications are not satisfactory. As a compatible solute, trehalose helps microorganisms to cope with high salt stress by participating in the regulation of cellular osmotic pressure, and plays an important role in promoting the nitrogen removal efficiency of microbial populations in the high-salt environment. We investigated the mechanism of exogenous-trehalose-enhanced metabolism of HN-AD community under high-salt stress by starting up a membrane aerobic biofilm reactor (MABR) to enrich HN-AD bacteria, and designed a C₁₅₀ experimental group with 150 μmol/L trehalose addition and a C₀ control group without trehalose. The reactor performance and the community structure showed that NH₄⁺-N, total nitrogen (TN) and chemical oxygen demand (COD) removal efficiency were increased by 29.7%, 28.0% and 29.1%, respectively. The total relative abundance of salt-tolerant HN-AD bacteria (with Acinetobacter and Pseudofulvimonas as the dominant genus) in the C₁₅₀ group reached 66.8%, an 18.2% increase compared with that of the C₀ group. This demonstrated that trehalose addition promoted the enrichment of salt-tolerant HN-AD bacteria in the high-salt environment to enhance the nitrogen removal performance of the system. In-depth metabolomics analysis showed that the exogenous trehalose was utilized by microorganisms to improve proline synthesis to increase resistance to high-salt stress. By regulating the activity of cell proliferation signaling pathways (cGMP-PKG, PI3K-Akt), phospholipid metabolism pathway and aminoacyl-tRNA synthesis pathway, the abundances of phosphoethanolamine, which was one of the glycerophospholipid metabolites, and purine and pyrimidine were up-regulated to stimulate bacterial aggregation and cell proliferation to promote the growth of HN-AD bacteria in the high-salt environment. Meanwhile, the addition of trehalose accelerated the tricarboxylic acid (TCA) cycle, which might provide more electron donors and energy to the carbon and nitrogen metabolisms of HN-AD bacteria and promote the nitrogen removal performance of the system. These results may facilitate using HN-AD bacteria in the treatment of high-salt and high-nitrogen wastewater.