Effects Of Drought On Soil CH₄ Uptake And Underlying Microbial Mechanisms In A Subtropical Forest

Methane(CH₄)is the second largest greenhouse gas after carbon dioxide,the subtle changes in atmospheric CH₄ sinks can cause important feedback on the global greenhouse effect.Forests ecosystems are the largest CH₄ sink in terrestrial ecosystems,and this process is mediated by a specific type of soil functional microorganisms-methanotrophs.A meta-analysis across the world showed that previous studies only focused on the effects of drought on the CH₄ efflux in temperate and tropical forests and its relationship with environmental factors.However,data on subtropical forests were still not available,and little was known about the microbial mechanisms underlying drought effects on CH₄ oxidation in forest soils.Based on existing research,we propose that drought can influence soil CH₄ oxidation in forest soils through two pathways:one is a direct pathway,drought can increase soil aeration,thereby promoting soil CH₄ oxidation capacity;the other is an indirect pathway,drought can influence methanotrophic activity through changing soil soluble carbon and nitrogen contents and the methanotrophic community structure and abundance,then inhibit the soil CH₄oxidation capacity.We selected a long-term extreme drought platform(70%throughfall reduction)in a subtropical forest in Tiantong,Zhejiang Province.A 3-year in situ soil CH₄ efflux observations were conducted via the static chamber approach,and soil samples were collected seasonally to determine soil properties(i.e.,soil moisture contents,pH,extractable carbon and nitrogen and so on).Laboratory incubation were carried out to determine the effects of drought on CH₄ oxidation potential,and metagenomic sequencing as well as real-time quantitative PCR(qPCR)techniques were used to analyze the changes in methanotrophic structures and abundance.By combining the stable isotope techniques(DNA-SIP)with high-throughput sequencing,we detected the changes in active methanotrophic communities under drought.The direct and indirect drought-derived pathways to soil CH₄ efflux in forest ecosystems were finally quantified through structural equation modeling(SEM).The main results are as follows:(1)The long-term in situ observation results showed that the values of soil in situ CH₄ effluxes under drought were negative at most of the sampling time,which represented uptake of atmospheric CH₄.On the whole,Tiantong subtropical forest soil appeared as a sink of atmospheric CH₄,and drought has significantly increased soil CH₄ oxidation capacity in the forest soil.The in situ CH₄ efflux was significantly influenced by soil moisture.(2)Inconsistent with our previous hypothesis,there were no significant differences in soil soluble carbon and nitrogen content such as extractable organic carbon(EOC),microbial biomass carbon(MBC),extractable organic nitrogen(EON),and microbial biomass nitrogen(MBN)content among the treatments,indicating that drought did not affect the CH₄ oxidation capacity by changing soil nutrient contents in the forest soil.(3)Based on the results of metagenomic sequencing analysis,it was found that methanotrophic communities in this area were dominated by type ? methanotrophs(0.072–0.288%vs.0.002–0.045%Type ? methanotrophs).Drought slightly decreased the relative abundance of all type ? methanotrophs,but there were no significant differences among the treatments.The results of laboratory incubation and qPCR analysis showed that drought had no effects on soil CH₄ oxidation potential and methanotrophic abundance(pmo A gene copies).These results indicated that drought did not affect the CH₄ oxidation capacity by changing methanotrophic activity as well as methanotrophic community structure and abundances in the forest soil.(4)Through DNA-SIP combined with 16S rRNA high-throughput sequencing,we detected that the active methanotrophic community in the soil were dominated by Methylosinus(type ? methanotrophs),and drought has slightly decreased its relative abundance in all the heavy fractions of the fractionated DNA gradient,but there were no significant differences among the treatments.SEM analysis showed that soil CH₄oxidation capacity was mainly affected by soil moisture(aeration)and soil CH₄oxidation potentials.Overall,the direct drought-derived pathway contributes about 24%on soil CH₄ oxidation in the forest soils,which exceeded the the effects of indirect drought-derived pathway.In summary,our research results show that Tiantong subtropical forest soils act as a sink of atmospheric CH₄,and drought has significantly increased soil CH₄ oxidation capacity of this forest soil.On the one hand,drought significantly reduced soil moisture and increased soil aeration;on the other hand,drought has no effect on soil soluble carbon and nitrogen content as well as the methanotrophic community and soil CH₄oxidation potential.Therefore,drought mainly promotes soil CH₄ uptake by directly increasing soil aeration.This paper could help us better understand the response of subtropical forest soil CH₄ oxidation capacity to drought,and provide a theoretical basis for the estimation of forest soil CH₄ sink under the scenario of climate change in the future.