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构建微域培养结合梯度凝胶电泳(DGGE)、Illumina MiSeq高通量测序、生物信息学分析等分子生态学技术,以不施肥土壤为对照(CK),研究长期施化肥(NPK)和有机肥(OM)对酸性旱地土壤硝化活性及自养硝化微生物群落的影响,并认知其与土壤理化因子间的关系.结果表明,施化肥和有机肥显著提高土壤有机碳和无机氮含量,施有机肥提高土壤pH和总氮含量、降低C/N;供试土壤自养硝化作用占据主导(73.60%~85.32%),施肥显著提升土壤自养硝化活性,且施有机肥提升效果更为明显;微域培养后,OM土壤氨氧化古菌(AOA)和细菌(AOB)amoA基因绝对丰度及16S rRNA基因相对丰度显著上升,而CK和NPK土壤仅AOA相对丰度显著上升,即3种土壤AOA均有明显活性(主要类群为Nitrososphaera,>99.30%),而AOB仅在OM土壤有活性(主要类群为Nitrosospira,>99.99%),另外还发现OM土壤中亚硝酸盐氧化细菌(NOB)有较强活性(主要类群为Nitrospira,>96.69%);逐步回归分析显示自养硝化活性显著受总氮含量影响,AOA和AOB amoA基因丰度分别受有机碳含量和pH影响,Nitrososphaera相对丰度与NO_3~--N含量显著正相关,而Nitrosospira和Nitrospira相对丰度则与C/N显著负相关.可见,长期施肥后土壤总氮含量的提升显著刺激自养硝化活性;以Nitrososphaera为主的AOA在酸性旱地土壤硝化作用中发挥了重要作用,施有机肥土壤pH上升及C/N下降刺激了Nitrosospira(AOB)生长,从而改变了酸性旱地土壤中活跃的自养硝化微生物类群.
To study the effects of long-term chemical fertilizers (NPK) and organic fertilizers (CK) on the development of micro-culture in combination with gradient gel electrophoresis (DGGE), Illumina MiSeq high throughput sequencing and bioinformatics analysis, (OM) on soil nitrification activity and autotrophic nitrifying microbial communities in acidic dry land and its relationship with soil physico-chemical factors were studied.The results showed that applying chemical fertilizers and organic fertilizers significantly increased soil organic carbon and inorganic nitrogen content, Fertilizer increased the soil pH and total nitrogen content and decreased C / N. The autotrophic nitrification played a dominant role (73.60% -85.32%) in the soil. Fertilization significantly increased soil autotrophic nitrification activity, After micro-region cultivation, the absolute abundance of OM and AOB amoA gene and the relative abundance of 16S rRNA gene in OM soil increased significantly, while the relative abundance of AOA in CK and NPK increased significantly Soil AOA had obvious activity (the major group was Nitrososphaera,> 99.30%), whereas AOB was only active on OM soil (Nitrosospira, the major group was> 99.99%). In addition, nitrite oxidizing bacteria (NOB) Strong activity (major group Nitros pira,> 96.69%). The stepwise regression analysis showed that the autotrophic nitrification activity was significantly affected by the total nitrogen content. The abundance of AOA and AOB amoA genes were affected by organic carbon content and pH respectively. The relative abundance of Nitrososphaera and NO_3 ~ While the relative abundance of Nitrosospira and Nitrospira was significantly negatively correlated with C / N. It can be seen that the increase of total nitrogen content in soil after long-term fertilization significantly stimulated the autotrophic nitrification activity; AOA mainly in Nitrososphaera was under nitrification in acid dry soil Played an important role. The increase of organic fertilizer soil pH and the decrease of C / N stimulated the growth of Nitrosospira (AOB), thus changing the active autotrophic nitrifying microorganism group in acid dry soil.