蓝藻群体颗粒驱动富营养化湖泊反硝化过程的微生物学机制

Eutrophication of the lake water body leads to cyanobacteria blooms and disturbs lake ecosystem functioning, yet cyanobacteria may also have feedback effects on lake N input. Denitrification is an important mechanism of lake N removal, and previous studies mostly focused on denitrification occurring at the sediment-water interface. Based on recent research findings in the country and worldwide, as well as the research progress from our team, we propose that “the interaction mechanisms between cyanobacteria and heterotrophic bacteria within cyanobacteria aggregates could drive denitrification in the water column”. This proposal combines in situ field experiment, laboratory incubation and mesocosm experiment, pure-culture cultivation as well as in situ lake monitoring, and chooses denitrification by cyanobacteria aggregates and cyanobacteria-associated denitrifiers as the breakpoint, to study cyanobacteria-heterotrophic bacteria interactions and the feedback mechanisms to lake N transforming processes. Through N-isotope labeling and membrane sample mass spectrometry analysis, enrichment cultivation experiment, high-throughput sequencing analyses and molecular biological methods, this work will quantify N removal potential by cyanobacteria aggregates, elucidate cyanobacteria-associated denitrifying community structures and dynamics, and further reveal cyanobacteria-denitrifier interactions and regulatory mechanisms by environmental factors. Eventually, results from this work will provide evidences for further understanding of the interactions between cyanobacteria and associated microbes in eutrophic lakes, as well as the role in biogeochemical cycling of N.

湖泊水体富营养化导致蓝藻水华暴发，干扰湖泊生态系统功能，而蓝藻也能够对湖泊氮输入产生反馈作用。反硝化过程是湖泊重要的脱氮机制，以往的研究多关注发生在底泥-水界面的反硝化过程。本课题基于国内外研究进展与本团队最新的研究基础，提出“水柱中蓝藻群体颗粒的藻-菌协作机制能够驱动反硝化过程”这一观点，拟通过现场围隔试验、实验室驯化与微宇宙试验、纯菌培养试验、并结合湖泊原位观测，以蓝藻群体颗粒反硝化作用及藻际反硝化细菌为突破口，研究藻-菌互作关系及其对湖泊氮过程的反馈机制。项目拟通过氮同位素标记与膜进样质谱分析、微生物富集培养、高通量测序技术与分子生物学手段，解析富营养化湖泊蓝藻群体颗粒的脱氮通量、藻际反硝化菌群的结构与动态，并进一步揭示其与蓝藻的互作机制及其受氮素因子的调控机制，为深刻认识富营养化湖泊中蓝藻与微生物的互作关系、及其在氮素生物地球化学循环过程中所起到的作用提供科学依据。

蓝藻群体颗粒是富营养水体中元素地球化学循环的重要介导者。本课题围绕蓝藻群体颗粒驱动的反硝化作用，对其发生的环境条件和通量、藻际反硝化菌群结构与分布格局、及氮水平对藻-菌关系的调控机制开展了一系列研究工作。. 针对蓝藻群体颗粒反硝化的条件和通量问题，首先基于原位水柱实验，揭示了微囊藻浮聚促进了水体黑暗、缺氧环境的形成；在中下层水体中，氮脱除由反硝化作用主导。同时，基于太湖沙渚的月度采样和N15-MIMS法，揭示了蓝藻群体颗粒和太湖底泥的反硝化潜力，前者表现出剧烈的时间差异。. 针对蓝藻群体颗粒中总微生物和反硝化菌群多样性分布格局，首先通过太湖沙渚蓝藻野外调查，基于16S rRNA扩增子测序解析了昼、夜和厌氧下蓝藻群体颗粒中的活跃菌群，发现微生物相对活性主要由丰度水平决定，并与之呈反比。进而基于功能基因扩增子分析，发现nirS、nosZ菌群呈现一定的季节演替模式，而nirK群落较为稳定，以及三种功能基因群落之间较强的相互关系。此外，进一步基于宏基因组手段解析了太湖氮循环微生物的时空分布及其生物和非生物调控因子。. 针对蓝藻群体颗粒藻-菌关系及其调控机制，首先通过微宇宙实验，揭示极度氮限制导致了藻-菌关系由互利向竞争和敌对关系转变。微囊藻大量产生溶菌酶、几丁质酶等，破坏藻际细菌细胞结构的同时，增加对有机氮源的利用；而其自身也须应对高活性氧胁迫。基于太湖原位宏转录组分析进一步揭示了藻-菌关系的季节和昼夜变化模式，发现相较于藻华前期，藻华中后期的藻-菌关系更多偏向于竞争和拮抗关系，且与包括氮在内的养分限制呈现较强的相关性。最后，从微囊藻群体颗粒中分离得到62个反硝化菌株，并基于反硝化纯菌和微囊藻模式菌株的共培养实验，发现氮限制下微囊藻所产生的胞外分泌物可能对反硝化细菌的生长产生显著影响。综上所述，本项目的研究成果综合揭示了微囊藻群体颗粒介导水体反硝化作用的微生物机制及水体氮水平对藻-菌关系的调控机制。