Anaerobic Oxidation Of Methane Cooperatively Driven By Nitrite And Ferric Hydroxide

Methane is one of the most important greenhouse gases,and biotechnology is the most economical and effective way to reduce methane emissions.Nitrate/nitrite reduction coupled with methane anaerobic oxidation plays an important role in reducing methane emissions in freshwater ecosystems,but its engineering application is limited due to its extremely slow growth of microorganisms.In this study,a reactor system（fresh water system）with nitrite/iron hydroxide coexistence coupled with methane anaerobic oxidation was constructed,and nitrite coupled with methane anaerobic oxidation system was used as a control to track and monitor the anaerobic reaction that had been running for three years.Analyzed the characteristics of nitrite/ferric hydroxide synergistic coupling of methane anaerobic oxidation;combined high-throughput metagenomic sequencing,bacterial and archaeal diversity sequencing,functional genes（differences in the abundance of related enzymes）and network analysis and other methods analyze the microbial diversity and functional microbial metabolic pathways,and explored the process and mechanisms of the anaerobic oxidation of methane driven by microbes under the coexistence of NO₂^-/Fe³⁺.The research achieved the following results:（1）The coexistence of iron hydroxide and nitrite promoted the denitrification-coupled anaerobic oxidation process of methane:the consumption of methane,nitrite nitrogen,and ammonia nitrogen was consumed faster in the synergistic system,although the concentration of Fe（Ⅱ）and Fe（Ⅲ）was consumed in dynamic equilibrium and higher than the control system.It showed that iron promoted the N-DAMO process and not only participated in the reaction in the synergistic system,but also may be a potential catalyst for the N-DAMO process.（2）The microbial community structure of bacteria and archaea in the iron hydroxide synergistic system had different changes.The bacterial diversity of the synergistic coupling system was lower than that of the control system,while the diversity of archaea was higher than that of the control system.It showed that Fe（OH）₃was more selective for bacteria,and bacteria may be the main functional microorganisms in the cooperative coupling system.（3）The four environmental factors of methane,nitrite,Fe（Ⅲ）,and Fe（Ⅱ）were positively correlated with each other.The degree of influence of bacterial community structure by environmental factors was as follows:Nitrate>Fe（Ⅱ）>methane>Fe（Ⅲ）;the degree of influence of the archaeal community structure by environmental factors was as follows:methane>nitrite>Fe（Ⅲ）>Fe（Ⅱ）.It showed that the conversion of methane and Fe（Ⅲ）may be related to archaea,and the conversion of nitrite and Fe（Ⅱ）may be related to bacteria.（4）Compared with the control system,the key genes（pmo-amo,mdh,mxa,nar,nir,nor）in the process of oxidation of methane to carbon dioxide and reduction of nitrite to nitrogen（pmo-amo,mdh,mxa,nar,nir,nor）were up-regulated in the coordinated coupling system.In the control system,the abundance of key genes for ammonia nitrogen production and transformation was up-regulated（CPS1,gin A,cyn T,cyn S）,and key genes related to siderophore metabolism were up-regulated（HEPH,PPOX,hem H）.It indicated that the addition of iron hydroxide in the synergistic system may contribute to the conversion of ammonia nitrogen,and iron as an essential life activity factor provided support for the life activities of functional microorganisms in the synergistic coupling system.Hyphomicrobium and Methylocystis had higher species contributions in methane metabolism,nitrogen metabolism,and porphyrin metabolism,and the key genes were annotated to these two types of bacteria,indicating that they were potentially functional microorganisms in a synergistic system;however,they were more effective than the control system.There were differences in gene abundance in the metabolic process,indicating that the synergy of ferric hydroxide and nitrite will affect the metabolic trend of functional microorganisms.