DNA和cDNA水平对比研究施肥对稻田土壤细菌多样性的影响

Effects of Fertilization on Soil Microbial Abundance and Community Structure at DNA and cDNA Levels in Paddy Soils

依托长沙农业环境观测研究站不同施肥方式长期定位试验,直接从土壤中抽提总DNA和RNA,应用T-RFLP和RTPCR技术在DNA和c DNA水平对比研究施肥对亚热带稻田土壤细菌丰度和群落结构的影响.结果表明,施肥和秸秆还田显著改变了土壤细菌群落组成(DNA水平)和转录组成(c DNA水平)结构,且不同处理中DNA水平的T-RFLP图谱与cDNA水平相差很大.施肥和秸秆还田降低了土壤中存在的细菌多样性,对土壤中转录的细菌多样性没有影响.土壤16S rRNA基因丰度(DNA水平)平均是其转录丰度(c DNA水平)的337倍,与不施氮肥处理(N0)相比,平衡施肥(NPK)没有改变土壤细菌的数量,在平衡施肥的基础上进行秸秆还田增加了土壤细菌的数量,但高量与低量秸秆还田没有显著差异.RDA分析表明,稻田土壤中铵态氮含量是调控各处理土壤中存在及转录的细菌群落的关键因子.总体而言,不管在DNA还是c DNA水平,研究施肥对土壤细菌丰度的影响均是可行并可信的;但只有在c DNA水平开展研究,才能有效发现细菌群落对环境的适应性.

Fertilizer applications have important effects on soil microbial abundance and community structure. In this study,total soil microbial DNA and RNA were directly extracted from paddy soils of N0（ control treatment,no nitrogen fertilizer）,NPK（ balanced fertilization）,NPK ＋ LS（ balanced fertilization with additional 3. 0 t·hm- 2rice straw incorporation） and NPK ＋ HS（ balanced fertilization with additional 6. 0 t·hm- 2rice straw incorporation） treatments in a long-term fertilization experiment of double rice cropping system in Changsha County,Hunan Province. Soil bacteria community structures were evaluated by analyzing the 16 S rRNA gene fragments at DNA and cDNA levels with Terminal Restriction Fragment Length Polymorphism（ T-RFLP） and quantitative PCR techniques. Balancing fertilization with chemical fertilizers and rice straw incorporation significantly changed the composition of bulk（ DNA-based） and potentially active（ mRNA-based） soil bacterial community as shown in T-RFLP profiles,and also reduced the bulk soil microbial diversity,but not the potentially active ones,as compared with the control treatment. The DNA-based abundance of16 S rRNA gene was on average 377 times as many as the m-RNA based population size. Compared to N0,balanced fertilization with rice straw incorporation（ NPK ＋ LS and NPK ＋ HS） increased the bulk and active copy numbers of 16 S rRNA gene,but not for balanced fertilization（ NPK）. The abundance and microbial community structure were not significantly different between the NPK ＋ LS and NPK ＋ HS treatments. Redundancy analysis（ RDA） showed that soil ammonium was the key environmental factor determining the bulk and active soil microbial community structure among the treatments. In conclusion,the effect of fertilization on soil microbial abundance and community structure could be indicated at both DNA and cDNA levels; the cDNA information could better reflect the adaptability of bacterial community to the environmental stress.