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采用PCR-RFLP法分析了灌溉猪沼液、牛沼液和施用化肥的土壤细菌多样性的季节性变化。通过直接提取土壤DNA,扩增细菌16S rDNA基因片段,建立克隆文库,用限制性内切酶AfaⅠ和HindⅢ进行酶切,得到了96～123种酶切类型。采用多样性测度对试验结果进行统计分析。结果表明,细菌多样性指数H′、Ds、Dg、dMa、R2和E变化规律为:施用猪沼液处理组>施用牛沼液处理组>对照组(P<0.05),Simpson指数λ则表现为对照组>牛沼液组>猪沼液组(P<0.05),细菌群落的均匀度指数Jgi和种间相遇机率的PIE指数均为猪沼液组>牛沼液组>对照组(P<0.05);细菌多样性指数H′、dMa、R和λ的变异系数在2.65%～18.87%之间,尤以Simpson指数λ变化最为敏感,处理间差异最大。最终得到如下结论:处理以及季节对细菌多样性的影响极显著,不同处理土壤细菌多样性为猪沼液组>牛沼液组>对照组,春季细菌多样性高于秋季;实施种养结合循环利用模式3年后,土壤细菌多样性有不同程度的提高。
PCR-RFLP method was used to analyze the seasonal variation of bacterial diversity in the soil of swine biogas slurry, biogas slurry and fertilizer application. Through direct extraction of soil DNA and amplification of bacterial 16S rDNA gene fragment, a cloning library was constructed and digested with restriction endonucleases Afa Ⅰ and Hind Ⅲ to obtain 96-123 digested fragments. The test results were statistically analyzed using diversity measures. The results showed that the change rules of bacterial diversity index H ’, Ds, Dg, dMa, R2 and E were as follows: Swine biogas slurry treatment group> Niunuma liquid treatment group> control group (P <0.05) (P <0.05). The evenness index (Jgi) of bacterial community and PIE index of inter-species probability of control group were all higher than that of control group (P <0.05) The coefficient of variation of bacterial diversity index H ’, dMa, R and λ was between 2.65% and 18.87%, especially the change of Simpson index λ was the most sensitive and the difference among treatments was the largest. Finally, the following conclusions are drawn: the effects of treatment and season on bacterial diversity are extremely significant. The bacterial diversity of different treatments is that of Swine Biogas Liquid Group> Niunuma Liquid Group> control group, with higher bacterial diversity in spring than in autumn; Three years after the model, soil bacterial diversity increased to varying degrees.