腐殖酸对亚硝酸盐依赖型甲烷厌氧氧化过程的生物作用机理研究  

Study on the biological mechanism of humic acid on nitrite-denitrifying anaerobic methane oxidation(nitrite-DAMO)process

作　　者：高冉 金昊 吕娇 徐帆 王如意[1] 楼菊青[1,2] GAO Ran;JIN Hao;LÜJiao;XU Fan;WANG Ruyi;LOU Juqing(School of Environmental Science and Engineering,Zhejiang Gongshang University,Hangzhou 310018;Instrumental Analysis Center of Zhejiang Gongshang University,Hangzhou 310018)

机构地区：[1]浙江工商大学环境科学与工程学院,杭州310018 [2]浙江工商大学分析测试中心,杭州310018

出　　处：《环境科学学报》2024年第5期1-16,共16页Acta Scientiae Circumstantiae

基　　金：浙江省‘尖兵’‘领雁’研发攻关计划(No.2023C03149);浙江工商大学“数字+”学科建设项目(No.SZJ2022B016);浙江省自然科学基金资助项目(No.LY21D030003)。

摘　　要：腐殖酸(Humic acid,HA)是土壤和沉积物中天然有机物的氧化还原产物,其独特的醌官能团可作为电子受体或电子穿梭器,与自然界中的甲烷厌氧氧化(Anaerobic Oxidation of Methane,AOM)过程耦合.反硝化型甲烷厌氧氧化(Denitrifying Anaerobic Methane Oxidation,DAMO)是以硝酸盐/亚硝酸盐作为电子受体的甲烷厌氧氧化与反硝化耦合过程.研究发现,许多自然生境中HA与N-DAMO系统共存.本研究以亚硝酸盐依赖型甲烷厌氧氧化(N-DAMO)系统为研究对象,考察750 mg·L^(-1)(促进)和2000 mg·L^(-1)(抑制)的HA对该系统的生物作用机理.研究结果表明,当外源腐殖酸存在时,系统中挥发性脂肪酸(Volatile Fatty Acid,VFA)含量明显降低,并促进了胞外聚合物(Extracellular polymers,EPS)的分泌.在高浓度HA影响下,细胞形态会发生变化.腐殖酸的加入导致N-DAMO系统菌群结构也发生了变化,OTU指数和Shannon指数分析结果显示促进组的生物多样性较空白组显著增加,而抑制组的生物多样性则较空白组显著降低,但优势菌种始终是Proteobacteria和Chloroflexi,并且Chloroflexi和Firmicutes丰度有所增加.宏基组测序结果发现促进组的氮代谢基因(NirK、NirS和NOD)丰度高于对照组,腐殖酸的加入会导致NO歧化作用的增强,与N_(2)O还原相关的基因(NosZ)受到了外源腐殖酸的抑制,且浓度越高越显著,导致了N_(2)O的累积.促进组和抑制组的甲烷代谢基因(pmoABC、mdhA和FDH)丰度均有提高,基因丰度的改变影响了N-DAMO系统反应代谢途径.Humic acid(HA)is a redox product of natural organic matter in soils and sediments.Its unique quinone functional group can act as an electron acceptor or electron shuttle to couple with the anaerobic oxidation of methane(AOM)process in nature.Denitrifying Anaerobic Methane Oxidation(DAMO)is a coupled process of methane anaerobic oxidation and denitrification using nitrate/nitrite as electron acceptors.HA was found to coexist with nitrite dependent anaerobic oxidation of methane(N-DAMO)process in many natural habitats.This study focused on the N-DAMO system,and investigated the biological mechanisms of 750 mg·L^(-1)(promoted)and 2000 mg·L^(-1)(inhibited)HA on the N-DAMO process.The results showed that when exposed to exogenous humic acid,the VFA content in the system was markedly reduced and the secretion of extracellular polymers(EPS)was promoted.Under the influence of high concentrations of HA,cell morphology changes.The addition of humic acid also resulted in changes in the microbial community structure of the N-DAMO system.The result of the OTU number and the Shannon index showed that the biodiversity of the promoted group increased significantly compared to the control,while the biodiversity of the suppressed group decreased significantly.Proteobacteria and Chloroflexi were always the dominating strains in all experimental groups.Meanwhile,the abundance of Chloroflexi and Firmicutes has increased.The metagenome sequencing results showed that the abundance of nitrogen metabolism genes(NirK,NirS,and NOD)in the promotion group was higher than that in the control.The addition of humic acid can lead to an enhancement of NO disproportionation.The NosZ genes involved in nitrogen monoxide(N_(2)O)reduction were inhibited by exogenous humic acid,leading to the accumulation of N_(2)O.The higher the concentration of humic acid,the more remarkable the inhibition was.The abundance of methane metabolism genes(pmoABC,mdhA,and FDH)in the promotion group and the inhibition group were both increased.The changes in gene abundan

关 键 词：N-DAMO 腐殖酸 细胞形态 群落结构 基因丰度 

分 类 号：X172[环境科学与工程—环境科学]