| CO2,CH4 and N2O are the main greenhouse gases causing global warming.The carbon and nitrogen cycling process in farmland soil contributes significantly to greenhouse gas emissions.Due to the lack of strict control over the heavy metal content of livestock and poultry feed additives in China,the excess rate of heavy metal in livestock and poultry feces applied to farmland is high,which leads to the aggravation of heavy metal pollution in farmland soil.When the farmland soil is polluted by heavy metals,the microbial activities and community structure in the soil will change,which will inevitably affect the soil carbon and nitrogen cycle which is closely related to the microorganisms.Therefore,it is of great practical significance to study the effects of single and compound pollution of typical heavy metals cuprum(Cu)and zinc(Zn)in livestock manure on the emission of CO2,CH4 and N2O in farmland soil and the microbial mechanism under the fertilization condition of livestock manure.In this study,typical heavy metals Cu and Zn in excrements of livestocks were selected as research objects.Through the design of indoor simulated paddy field experiment,domestic pig manure(M),which have not been exposed to heavy metal feed additives,and conventional urea(U),were used as nitrogen source fertilizer to design single pollution with different concentrations of Zn and combined pollution with different concentrations of CuZn.The effects of single Zn and Cu-Zn combined contamination at different concentrations under different fertilizer source conditions on CO2,CH4 and N2O emissions in simulated flooded soil and the related microbial mechanisms were investigated.The simulated submerged chamber culture experiment was divided into two batches:single contamination of Zn and combined contamination of Cu-Zn.The former set a total of 11 treatments,which are:no fertilization and no heavy metal treatment(CK);Using urea as nitrogen fertilizer,Zn stress concentrations were 0、50、500、1500、5000 mg/kg(U-CK、U-Zn50、U-Zn500、U-Zn1500、U-Zn5000);Using pig manure as nitrogen fertilizer,Zn stress concentrations were 0、50、500、1500、5000mg/kg(M-CK、M-Zn50、M-Zn500、M-Zn1500、M-Zn5000).A total of 19 treatments were set for the latter Cu-Zn composite pollution,respectively:no fertilization and no heavy metal treatment(CK);Using urea as nitrogen fertilizer,Cu stress concentration was 0,100,1500 mg/kg and Zn stress concentration was 0,500,5000 mg/kg(U-CK、U-Cu0-Zn500、U-Cu0-Zn5000、U-Cu100-Zn0、U-Cu100-Zn500、U-Cu100-Zn5000、U-Cu1500-Zn0、U-Cul500-Zn500、U-Cu1500-Zn5000);Using pig manure as nitrogen fertilizer,Cu stress concentration was 0,100,1500 mg/kg and Zn stress concentration was 0,500,5000 mg/kg(M-CK、M-Cu0-Zn500、M-Cu0-Zn5000、M-Cu100-Zn0、M-Cu100-Zn500、M-Cu100-Zn5000、M-Cu1500-ZnO、M-Cu1500-Zn500、M-Cu1500-Zn5000).Main research results of this experiment:(1)For soil respiration,Zn single pollution showed low promotion(50mg/kg)and high inhibition(≥500mg/kg).When the concentration was 1500mg/kg,the inhibition effect was strongest,and the CO2 emission of soil was significantly reduced by about 36%compared with the control of the same fertilizer source(P<0.05).However,at 5000mg/kg,the inhibition effect was weakened,and soil CO2 emissions only reduced by about 26%(P<0.05).In the presence of pig manure,the inhibitory effect of Zn can be enhanced,especially at low concentrations(500mg/kg).Under the condition of combined Cu-Zn pollution,it shows inhibition effect,and there is a certain synergistic effect between Cu and Zn.In particular,the high concentration of Cu-Zn(Cu1500-Zn5000)showed an obvious synergistic effect,which could significantly reduce the CO2 emission of soil by about 8-15%on the basis of the single pollution of the same concentration of Cu and Zn(P<0.05).The effect of Zn single contamination on the gene abundance of soil total bacteria 16S rRNA in the early stage of culture(about 3-5d)was low concentration goes up and high concentration goes down.At the late stage of culture,no effect or increasing effect was dominant.In particular,the high concentration of Zn(5000mg/kg)under the coexistence of pig manure showed the most obvious promotion effect,which was 4.4 times higher than the control group(P<0.05).Under the condition of combined Cu-Zn contamination,CO2 emission was positively correlated with the abundance of 16S rRNA gene of total soil bacteria(P<0.05).However,under the single contamination condition of Zn,there was no significant correlation between CO2 emission and the abundance of 16S rRNA gene of total soil bacteria(P>0.05).It indicates that Cu-Zn composite pollution mainly affects the soil CO2 emission by affecting the activity of soil bacteria,while the effect of single Zn on soil respiration is not only soil bacteria,but also other microbial activities.(2)For soil CH4 emission,when the concentration of Zn is 50～5000mg/kg under a single pollution condition,the performance of different soil conditions in the coexistence with urea is different,it has both promoting and inhibiting effects.When coexisting with pig manure,low concentration of Zn promotes methane emission(500mg/kg),while high concentration of Zn inhibits methane emission(≥1500mg/kg).Under the condition of combined pollution of Cu-Zn,no matter what kind of fertilizer source,the combined pollution of Cu-Zn can promote CH4 emission in the form of low concentration Cu-Zn,while high concentration Cu-Zn can inhibit CH4 emission.At low concentration ratio,the role of Cu was dominant.When high concentration of Cu is involved(1500mg/kg),no matter how high or low the concentration of Zn is,it has an inhibitory effect.Especially when coexisting with pig manure,high concentration of Cu and Zn(1500,5000 mg/kg)showed obvious synergism inhibition,which could reduce the CH4 emission of soil by about 6-28%on the basis of single pollution of the same concentration of Cu and Zn(P<0.05).The influence of single contamination with different concentrations of Zn on 16S rRNA-CH4,mcrA and pmoA genes was not obvious.When Cu and Zn co-exist with urea,Cu and Zn have synergistic inhibition effect on the three genes.No matter what the ratio of Cu and Zn concentration was,the stress effect was intensified in the later period of culture.No significant correlation was found between CH4 emission and the abundance of the three genes under either single Zn or Cu-Zn composite contamination conditions(P>0.05),indicating that the influence of Cu and Zn on CH4 emission was not limited to the changes in the abundance of these three genes.(3)For N2O emission,when Zn and urea coexist under a single pollution condition,no matter the concentration is high or low,there is a significant inhibitory effect(P>0.05).The N2O emission treated with high concentration of Zn(5000mg/kg)accounted for only about 1%of the control.When coexisting with pig manure,low concentration of Zn promoted N2O emission,while high concentration of Zn inhibited N2O emission.The inhibition effect was strongest when the concentration was 1500mg/kg.The performance of Zn at the concentration of 5000mg/kg was different in different soil conditions.Under the condition of Cu and Zn combined pollution,when urea was the fertilizer source,Cu-Zn combined pollution showed obvious synergistic inhibition effect on N2O emission.When pig manure was used as the fertilizer source,Cu-Zn had obvious synergistic promoting effect.Especially at low copper(100mg/kg),no matter the ratio of high and low concentrations of Zn,its synergic promotion effect was the most obvious(P<0.05),and its N2O emission was 1.3-2.5 times of the pollution caused by single Cu and Zn at the same concentration.When single contamination of Zn and urea coexist,the inhibition effect on AOB amoA gene is more obvious in the early and late stage of culture.When coexisting with pig manure,AOB amoA gene was promoted in the early stage and inhibited in the later stage.Regardless of the fertilizer source,the single contamination of Zn had a weaker effect on AOA amoA gene than on AOB amoA gene,and the main effect was inhibition.Effects of Zn on soil nirK,nirS and nosZ genes showed that the single contamination of Zn was mainly promoted in the early stage and inhibited in the later stage.The effect of high concentration(5000mg/kg)Zn on promoting and inhibiting the abundance of the five major genes involved in the process of nitrification and denitrification changed with the soil conditions,especially when it coexisted with pig manure to significantly increase the abundance of the five major genes(P<0.05).In addition to the significant increase in the abundance of AOA amoA gene at the late stage of co-culture with pig manure,Cu-Zn composite contamination mainly inhibited AOA amoA and AOB amoA genes involved in the nitrification process.For nirK,nirS and nosZ genes involved in the denitrification process,the co-contamination of Cu-Zn with urea was mainly promoted in the early stage and inhibited in the late stage,while the co-contamination with pig manure was mainly inhibited in the early stage and the late stage.The N2O emission under single and CuZn combined contamination was mainly positively correlated with the abundance of soil AOA amoA,AOB amoA,nirK and nirS genes(P<0.05),indicating that single or combined contamination of Cu and Zn significantly affected the soil nitrification and denitrification related microorganisms,thereby affecting the N2O emission. |
