Impact Of Copper And Sulfadimidine Contamination In Livestock Manure On Greenhouse Gases Emissions And Related Functional Gene Abundance In Paddy Soil

Carbon dioxide（CO₂）,methane（CH₄）and nitrous oxide（N₂O）are the main greenhouse gases contributing to global warming.Among the many sources of emissions,the carbon and nitrogen cycling process of farmland soil is a significant source of greenhouse gases.With the large-scale development of the livestock and poultry breeding,the standard for adding antibiotics and heavy metals in feed additives has has not been keeping up.As a result,the high level of antibiotics and heavy metals in livestock and poultry manure leads to severe contamination in farmland soil.When the soil with different pH values is contaminated,the impact on microbial activity and community structure are different,which in turn affects the circulation of carbon and nitrogen in the soil.Therefore,it is of great significance to study the impact of single and combined contamination of SMZ and Cu on CO₂,CH₄ and N₂O emissions from paddy soil and related functional genes abundance under different pH conditions.In order to investigate the responses to combined contamination of SMZ and Cu on CO₂,CH₄ and N₂O emissions and related functional genes under different pH conditions,SMZ and Cu were selected as the research objects.Different treatments were set up under different pH conditions,including: Cu0+SMZ0,Cu0+SMZ0.1,Cu0+SMZ1,Cu0+SMZ5,Cu100+SMZ0,Cu100+SMZ0.1,Cu100+SMZ1,Cu100+SMZ5,Cu500+SMZ0,Cu500+SMZ0.1,Cu500+SMZ1,Cu500+SMZ5,Cu1500+SMZ0,Cu1500+SMZ0.1,Cu1500+SMZ1 and Cu1500+SMZ5.The number of the treatment code represents Cu or SMZ concentrations（mg/kg dry soil）.As fertilizer source,pig manure was input into soil with Cu and SMZ synchronously.（1）Effect on soil respiration（CO₂）In acidic-environment soil,single SMZ treatment shows low promotion and high inhibition effect on soil CO₂ emission;Cu alone inhibits soil CO₂ emission.The combined pollution of Cu and SMZ has a negative correlation with the concentration of soil CO₂ emission.The effects of single and combined SMZ and Cu pollution on16 S rRNA gene abundance are mainly decreased in the early and middle period and increased in the late period.There is no significant effect on 18 S rRNA gene abundance in the early and middle periods（P>0.05）,but it significantly decreased in the late period（P<0.05）.In neutral-environment soil,single SMZ inhibits soil CO₂ emission,and the low and medium concentrations dose not reach the significant level（P>0.05）.The single Cu treatment shows low and high concentration inhibit the CO₂ emission,but the medium concentration promote the CO₂ emission slightly;The cotamination of Cu and SMZ significantly reduce CO₂（P<0.05）.16 S rRNA: single SMZ low increase and high decrease in the early stage,then increase in the middle stage,and decrease in the middle and high concentration of SMZ in the late stage;Under the contamination of single Cu,the abundance of 16 S rRNA decreases in late stage;The combined pollution of SMZ and Cu decrease in the early stage and increase in the middle and late stage.18 S rRNA: single SMZ decreases in the early stage and increases in the middle stage（P<0.05）,but has no significant effect in the late stage（P>0.05）;Single Cu has no significant effect in the early stage（P>0.05）,but in the middle and late stage of culture,it is low,high and low;In the early stage of combined pollution,the main effect reduces,but in the middle stage it alleviates,and in the late stage it has no significant effect（P>0.05）.（2）Effect on CH₄ emission from soilIn acidic-environment soil,single SMZ treatment have a low promoting and high inhibiting effect on CH₄ emission;The single Cu has a inhibition effect on the CH₄ emission of soil;The combined pollution of Cu and SMZ have a negative effect on CH₄ emission in acid soil.The results shows that the abundance of 16 S rRNA-CH₄ and mcrA genes increase and decrease at low concentration,and then increase and decrease at high and low culture time,resulting in the decrease of pmoA gene abundance;Single Cu reduces the gene abundance of the three microorganisms;In the early stage,Cu and SMZ combined pollution increase 16 S rRNA-CH₄ and mcrA gene abundances mainly.For pmoA gene abundances,low concentration SMZ combined treatment increase pmoA gene abundances,medium concentration SMZ combined treatment decrease the most,while high concentration SMZ combined treatment decrease the degree of gene abundances.In the middle stage,Cu and SMZ combined pollution have no significant effect on gene abundances（P>0.05）.In neutral soil,SMZ has a significant effect on CH₄ emission（P<0.05）;CH₄emission significantly reduces in single Cu treatment;Combined pollution of Cu and SMZ significantly reduce the cumulative emission of CH₄（P<0.05）.Single SMZ pollution has no significant concentration effect on the abundance of the three CH₄ related genes（P>0.05）.The abundance of 16 S rRNA-CH₄ gene followe the alternate effect of decrease-increase-decrease,while the abundance of mcrA and pmoA gene followe the alternate effect of decrease-decrease-increase;Single Cu decreases 16 S rRNA-CH₄ gene abundance,following the rule of increase-decrease-increase for mcrA gene abundance in three culture periods,and has little effect on pmoA gene abundance in the early stage,but decreases significantly in the middle stage（P<0.05）,and increases in the late stage;The results shows that the abundance of the three genes decrease in the early stage,decrease slightly in the middle stage,and increase in the late stage.（3）Effect on N₂O emission from soilIn acid soil,single SMZ treatment have inhibitory effect on N₂O emission;The single Cu has a low promoting and high inhibition effect on N₂O emission;The combined pollution of Cu and SMZ have a negative effect on N₂O emission.The abundance of AOAamoA gene increase significantly in the early and middle stage of SMZ（P<0.05）,and decrease significantly in the late stage（P<0.05）.The abundance of AOBamoA gene increase and decrease alternately in the middle and late stage of culture.The abundance of nirK gene decrease at low and high concentrations.The abundance of nirS gene increase first and then decrease,while the abundance of nosZ gene decrease mainly in the early stage of culture;The results shows that Cu treatment reduce the abundance of five genes;SMZ and Cu combined pollution increase AOAamoA and AOBamoA gene abundances and then decrease them;The nirK,nirS and nosZ genes also show decrease in abundance.In neutral soil,single SMZ promotes N₂O emission,but dose not reach a significant level（P>0.05）;Single copper treatment increase N₂O emission（P<0.05）;The combined pollution of Cu and SMZ promote N₂O emission（P<0.05）.SMZ decreases AOAamoA and AOBamoA gene abundances in the early and middle stages,and then increases them in the late stage.SMZ also decreases nirK gene abundances first and then increases nirS gene abundances first and then decreases AOAamoA and AOBamoA gene abundances in the middle and late stages,decreases nosZ gene abundances in the early stage,and changed to increases in the middle and late stages;The results shows that the abundance of 5 genes is low,high and low,and the abundance of 5 genes decrease before and then increase;After combined pollution,the gene abundances of AOAamoA,AOBamoA,nirK,nirS and nosZ decrease and then increase.