覆盖作物对猕猴桃园土壤氨氧化微生物丰度和群落结构的影响

  【目的】  果园行间播种覆盖作物有利于改善土壤理化性状，改变氮循环过程中的氨氧化微生物群落结构。因此，我们分析不同覆盖作物下猕猴桃园土壤氨氧化微生物的群落结构和基因丰度，以探究其与土壤硝化作用的关系。  【方法】  选择湖北十堰的一个5年生猕猴桃果园进行了种植多年生覆盖作物试验，供试覆盖作物有白三叶草 (Trifolium repens L，W)、毛苕子 (Vicia villosa Roth，V)、黑麦草 (Lolium perenne L，R) 和高羊茅 (Festuca elata Keng ex E. Alexeev，F)。设置单种 (W)、两种混种 (WR)、3种混种 (WRV)、4种混种 (WRVF) 和清耕对照 (CK) 共5种试验处理。在覆盖作物种植后的第2年，取0—20 cm土层样品，利用荧光定量PCR (fluorescence quantitative PCR)、限制性末端片段长度多态性 (terminal restriction fragment length polymorphism, T-RFLP) 以及克隆测序技术，测定了土壤中氨氧化古菌 (ammonia-oxidizing archaea, AOA) 与氨氧化细菌 (ammonia-oxidizing bacteria, AOB) 基因丰度、群落多样性及组成。  【结果】  4种覆盖作物处理的果园地上部生物量没有显著差异。除两种混种处理显著提高了土壤含水率、铵态氮和硝态氮含量和降低了土壤pH外，其他3个处理的土壤理化指标之间也没有显著差异。与清耕对照相比，单种处理的AOA-amoA基因丰度显著提高了2.85%，单种和两种混种处理的AOB-amoA基因丰度分别显著提高了4.95%、6.42%。不同 T-RFs 所代表的AOA在不同种植模式中所占比例发生变化，但AOA群落结构未发生明显改变；不同T-RFs所代表的AOB在不同种植模式下发生改变，其AOB群落结构可能发生变化。两种混种处理显著降低了AOA的多样性指数，两种混种和4种混种处理显著降低了AOA均匀度指数，而3种混种和4种混种处理显著提高了AOB的多样性指数。4个处理土壤中，AOA的优势菌属为Nitrososphaera (44.2%) 和Nitrosotalea (45.8%)，AOB的优势菌属为Nitrosospira (96.98%) 和Nitrosomonas (3.02%)。Pearson相关分析和冗余 (RDA) 分析表明，土壤pH、含水率、铵态氮和硝态氮含量是影响氨氧化微生物群落结构及丰度的主要环境因子。  【结论】  土壤pH、含水率、铵态氮和硝态氮含量是影响氨氧化微生物群落结构及丰度的主要环境因子。白三叶草和黑麦草混种模式 (WR处理) 可显著提升土壤的水分含量、铵态氮和硝态氮含量，降低土壤pH，但是不同覆盖作物对土壤中AOA和AOB的群落结构尚未显现出显著影响。

Effects of cover crops on gene abundance and community structure of soil ammonia-oxidizing microorganism in a kiwifruit orchard

Objectives]Planting of cover crops between fruit tree rows can improve soil physicochemical properties,and change the structure of ammonia-oxidizing microorganism community in the process of the nitrogen cycle. Here, we analyzed the effects of different cover crop treatments on soil ammonia-oxidizingmicroorganisms in a kiwifruit orchard, and examined its relationship with soil nitrification.Methods]Soilsamples were collected from a five-year kiwifruit orchard where cover crops experiment was set up including fourkinds of cover crops white clover (Trifolium repens L, W), hairy vetch (Vicia Villosa Roth, V), ryegrass (Loliumperenne L, R) and tall fescue (Festuca elata Keng ex E. Alexeev, F)]. The treatments included single cover crop(W), two cover crops (WR), three cover crops (WRV), four cover crops (WRVF) and cleaning tillage (CK). QPCR,T-RFLP and cloning and sequencing were employed to analyze the changes in gene abundance, communitydiversity and structure of AOA and AOB of the different cover crop treatments.Results]There was nosignificant difference in the above-ground biomass of the four cover crop treatments. However, soil water content,ammonium and nitrate nitrogen (P < 0.05) increased while soil pH decreased in the WR treatment. Similarly, therewas no significant difference in soil physicochemical properties among the other three cover crop treatments.Compared with CK, the abundance of the AOA-amoA gene in W (P < 0.05) was increased by 2.85%, and theabundance of the AOB-amoA gene in W and WR was significantly increased by 4.95% and 6.42%, respectively.The proportion of T-RFs representing AOA varied among the treatmenta but did not cause significant changes inAOA community structure. AOB represented by T-RFs changed, and the AOB community structure mightchange. AOA diversity index significantly decreased in WR, AOA evenness index decreased in WR and WRVF,while AOB diversity index increased in WRV and WRVF. Under the four treatments, the dominant soil bacteriaof the AOA group were Nitrososphaera (44.2%) and Nitrosotalea (45.8%), and those of AOB group wereNitrosospira (96.98%) and Nitrosomonas (3.02%). Pearson correlation analysis and redundancy analysis showedthat soil pH, water content, ammonium nitrogen and nitrate nitrogen were the main factors affecting soil ammoniaoxidizingmicroorganism abundance and community structure.Conclusions]Soil pH, soil water, ammonium and nitrate nitrogen content are the main environmental factors affecting the community structure and abundanceof ammonia-oxidizing microorganisms. White clover and ryegrass mixed cropping system significantlyincreases soil moisture content, ammonium and nitrate nitrogen content but decreases soil pH. However, thedifferences in soil properties lead to increased diversity of microorganism, but not sufficient enough to cause avariation in the microbial structure.