[目的]非共生固氮是农田生态系统中氮素输入的重要途径之一,探明非共生固氮速率、固氮微生物丰度及其群落结构对施肥量的响应,为农业可持续生产提供施肥参考.[方法]试验地点位于东北黑土区,该地块自2012年起一直实行免耕及秸秆还田.田间试验设置6个施氮量处理:0、120、150、180、210、240kg/hm2,分别记作N0、N120、N150、N180、N210、N240.玉米收获后,采集0-5、5-20 cm 土层土壤样品,测定土壤pH和速效氮、磷、钾含量;采用乙炔还原法、定量PCR和高通量测序技术,分析土壤非共生固氮速率、固氮微生物丰度、群落结构多样性.[结果]不同施氮量下,土壤非共生固氮速率为C2H4 0.26～0.63 nmol/(g·d),N150处理0-5和5-20 cm 土层土壤非共生固氮速率均最高,分别为C2H4 0.53和0.63 nmol/(g·d);固氮酶基因nifH丰度范围为5.34× 106～15.64× 106 copies/g,0-5和5-20 cm 土层土壤nifH基因丰度的最大值也为N150处理,分别为15.64×106和8.74×106copies/g.主成分分析(PCoA)结果显示,不同施氮量之间固氮微生物群落结构存在显著差异.各施氮量处理土壤固氮菌群属水平以 unclassified_o_Burkholderiales、unclassified_k_norank_d_Bacteria、Skermanella、Methylocystis为优势菌属.Spearman相关性分析结果表明,非共生固氮速率与土壤速效钾含量呈显著正相关,与pH呈负相关;nifH基因拷贝数与土壤速效钾含量呈显著正相关,与施氮量呈负相关.冗余分析显示,nifH群落结构与土壤pH及NH4+-N、NO3-N、有机碳、全氮、有效磷、速效钾含量呈显著相关.[结论]在长期免耕和秸秆全量还田条件下,施用N150kg/hm2有利于增加土壤非共生固氮速率和固氮微生物丰度,丰富群落结构多样性,是该地区较为适宜的施氮量.
Optimal nitrogen application rate enhances the asymbiotic nitrogen fixation rate and nitrogen-fixing bacteria abundance in black soil of Northeast China
[Objectives]Asymbiotic nitrogen fixation is one of the important sources of nitrogen input in farmland ecosystems.Investigating the responses of asymbiotic nitrogen fixation rate,abundance of nitrogen-fixing microorganisms and diversity of community structure to nitrogen fertilizer application rates will provide fertilization reference for sustainable agricultural production.[Methods]The experiment was conducted in black soil area of Northeast China,the test field was under conservation tillage and straw return since 2012.Six N application rate treatments were setup,0(N0),120(N120),150(N150),180(N180),210(N210),and 240(N240)kg/hm2.At harvest of maize,0-5 cm and 5-20 cm soil samples were collected,conventional chemical methods were used to determine soil pH and available N,P and K contents,and acetylene reduction method,quantitative PCR,and high-throughput sequencing technology were used to analyze the soil asymbiotic nitrogen fixation rate,diazotroph community abundance and structure diversity.[Results]Under the six N rate treatments,the asymbiotic nitrogen fixation rate ranged from C2H4 0.26-0.63 nmol/(g·d)in both 0-5 cm and 5-20 cm soil layers,and N150 treatment exhibited the maximum asymbiotic nitrogen fixation rate,C2H4 0.53 nmol/(g·d)in 0-5 cm layer and C2H4 0.63 nmol/(g·d)in 5-10 cm layer.The nifH gene abundance ranged from 5.34× 106 to 15.64× 106 copies/g,and N150 treatment was recorded the maximum nifH gene abundance as well,they were 15.64×106 and 8.74× 106 copies/g in the two aforementioned layers.Principal component analysis(PCoA)revealed notable differences among fertilization rates regarding the composition of soil diazotroph community.The dominant bacterial genera at species level were unclassified_o_Burkholderiales,unclassified_k_norank_d_Bacteria,Skermanella,and Methylocystis in all the six treatments.Spearman correlation analysis between environmental factors and indicators for soil diazotroph community showed that asymbiotic nitrogen fixation was positively correlated with available K and negatively correlated with pH.nifH gene copy number was positively correlated with AK and negatively correlated with fertilizer application rates.Redundancy analysis indicated that nifH community structure had a significant correlation with soil pH,NH4+-N,NO3-N,soil organic C,total N,available P,and available K.[Conclusions]Application of N 150 kg/hm2 is beneficial for increasing the asymbiotic nitrogen fixation and nitrogen-fixing microbial abundance,enriching the community structure diversity,so is thought an optimal fertilizer rate in black soil farmland under long-term no-tillage and full-crop return.
万方数据
维普
