不同生育期苹果园土壤氨氧化微生物丰度研究

Abundance of ammonia oxidizers in apple orchard soil at different growth stages

【目的】大量施氮引起的土壤酸化问题已严重制约苹果的安全生产。氨氧化微生物驱动的土壤硝化作用是土壤氮素循环的重要环节,探析苹果园土壤中氨氧化微生物氨单加氧酶基因(amo A)丰度与土壤硝化强度(Potential Nitrification,PN)和土壤理化性质的相关性,有助于评价土壤氨氧化微生物类群在苹果园土壤生态系统中的作用。【方法】本研究以辽宁省丹东东港地区‘寒富’苹果园土壤为研究对象,采用实时荧光定量PCR(Real-time PCR)技术,检测苹果树不同生育时期,4月28日(萌芽期)、7月24日(新梢停长期)、10月23日(落叶期)的两个苹果园(分别编号为D1和D2)的土壤理化性质、土壤硝化强度、氨氧化古菌(Ammonia-oxidizing archaea,AOA)和氨氧化细菌(Ammonia-oxidizing bacteria,AOB)的amo A基因丰度,并分析了氨氧化微生物丰度与土壤理化性质和土壤硝化强度之间的关系。【结果】不同生育时期‘寒富’苹果园土壤理化因子差异较大。所有供试土壤的硝态氮(NO-3-N)、速效磷(AP)和速效钾(AK)含量在4月份最高,铵态氮(NH+4-N)含量在7月最高,NO-3-N、NH+4-N、AP和AK含量在10月多为最低。且所有供试土壤p H值均在4.25 6.09之间。同一生育时期内,D2土壤p H均显著高于D1土壤,但其NO-3-N和NH+4-N含量则不同程度地低于D1土壤。不同采样时期的果园土壤硝化强度随季节变化表现出先降后增的趋势,除7月D2土壤硝化强度显著高于D1土壤外,4月与10月D2土壤硝化强度均显著低于D1土壤。尽管不同采样时期的土壤AOA与AOB丰度随生育期而各异,所有供试土壤中AOA丰度均显著高于AOB丰度。同一时期内,D2土壤AOA和AOB丰度均显著高于D1土壤。尽管土壤p H、NO-3-N与AOA、AOB均表现出显著相关性,土壤PN仅与AOA丰度明显正相关。【结论】长期施肥导致苹果园土壤p H值降低,p H值的改变是影响AOA与AOB丰度的重要因子,果园土壤的硝化过程主要由AOA来完成,土壤硝化强度与季节变化引起的温度和土壤环境因子等的改变密切相关。苹果园无机氮肥混合有机肥的施入,同时结合自然生草、人工刈割等管理制度,在一定程度上可改变土壤氮素的含量与种类,减缓土壤酸化。

【Objectives】Soil acidification caused by over application of N fertilizers has posed a great challenge for apple security production. Nitrification is a central process of soil nitrogen cycle,which is driven by ammoniaoxidizing microorganisms,including ammonia-oxidizing bacteria（ AOB） and ammonia-oxidizing archaea（ AOA）.Studying the correlation of the abundance of gene fragment amo A with the potential nitrification（ PN）,as well as the soil physical and chemical properties, will be of help to objective evaluation of soil ammonia-oxidizingmicroorganisms in soil ecological system.【Methods】Two apple orchards in east of Liaoning Province were used for investigation,in both of the orchards,apple cultivar was ‘hanfu＇. In one orchard,the weeds had been removed every year（ D1） and in the other the weeds had been cut and returned to soil（ D2）. Soil samples were collected on April 28th（budding period）,July 24th（young shoot-stop growing period） and October 23th（deciduous period）.Soil PN and properties were determined,and the abundance of AOA and AOB was measured by real-time PCR using amo A gene copies. 【Results 】 The highest contents of NO-3-N,available P and readily available K all appeared in April and those of NH＋4-N in July,the lowest contents of NO-3-N,NH＋4-N,available P and readily available K were in October in both orchard soils mostly. The p H of all soil samples was in the range from 4. 25 to6. 09. The soil p H in D2 was significantly higher than in D1 in the same growth stages,while the NO-3-N and NH＋4-N contents in D2 were lower than in D1 in different degrees. Soil samples collected in April had the highest PN rates and those collected in July had the lowest rates. Compared to D2,the PN values of D1 groups were significantly higher in April and October,while there were opposite results in July. Although the AOA and AOB abundances changed with the growth stages,the abundances of AOA were significantly higher than the abundances of AOB in all tested soils. In the same period,the AOA and AOB abundances of D2 soils were significantly higher than those of D1 soils,which might be related to soil p H. The correlation analysis showed that there were significant correlations between soil p H,NO-3-N and AOA and AOB,but the soil PN was only positively correlated with the AOA abundance.【Conclusions】The long-term fertilization leads to low soil p H,which is an important factor affecting the abundances of AOA and AOB. The nitrification process in apple orchard soils is mainly related to AOA,and closely related to soil temperature and environmental factors. Fertilization and the returning of mowed weeds affect soil nitrogen contents and forms,slow down the acidification process of orchard soils.