采用限制性片段长度多态性(restriction fragment length polymorphism,RFLP)方法,研究了巢湖西半湖北岸湖滨带蓝藻堆积处不同深度层次的细菌多样性。利用直接法提取了蓝藻堆积处3个不同深度层次样品(A层0～2 cm、B层2～10 cm、C层10～20 cm)中的细菌总DNA,并进一步构建各层细菌的16S rDNA克隆文库。结果表明,蓝藻堆积处各层细菌的Margalef丰富度指数(d_Ma)、Shannon-Wiener指数(H)和Simpson指数(D)均伴随深度的增加而减小。系统发育树分析显示,各层的细菌优势类群主要归属于厚壁菌门(Firmicutes)和变形菌门(Proteobacteria);且随深度层次的增加,归属于变形菌门的细菌比例减少,而归属于厚壁菌门的细菌比例增加;在A层样品中存在少量归属于蓝藻门(Cyanobacteria)、放线菌门(Actinobacteria)、浮霉菌门(Planctomycetes)、螺旋体门(Spirochaetes)类群的细菌;在C层样品中也存在少量归属于拟杆菌门(Bacteroidetes)类群的细菌。研究结果在一定程度上揭示了蓝藻堆积腐烂过程中细菌多样性和种类组成的变化。

Cyanobacterial bloom has been a hot issue and the microbial community is one of the important functional groups in the decomposition process of cyanobacteria accumulation.In this study,restriction fragment length polymorphism(RFLP)was applied to analyze the bacterial diversity of different depths in the cyanobacterial accumulation of lakefront at northern shore of west region of Chaohu Lake.The total DNA of the microorganisms from three different layers of cyanobacterial accumulation(Layer A:0-2 cm,Layer B:2-10 cm,Layer C:10-20 cm)were extracted by the direct method.The DNA extracted was amplified using bacterial universal primers 27F and 1492R.PCR products were ligated into the pMD-18T Vector and ligations were transformed into Escherichia coli DH5α to construct 16S rDNA clone library of microbes in the cyanobacterial accumulation.120 positive clones each layer from the library were screened and their 16S rDNA fragments were reamplified.The fragments were digested with HhaⅠ and RsaⅠ respectively and their fingerprints were analyzed.The results showed that the layers from top to bottom include 51,32,27 restriction endonnuclease types and the coverage(C- value)of the clone library were 80.8%, 87.5%, 89.2%, respectively.Margalef indices(d_Ma), Shannon-Wiener indices(H)and Simpson indices(D)all decreased with the depth increasing.Phylogenetic analysis suggested that the dominant bacteria in different layers of cyanobacterial accumulation belonged to Firmicutes and Proteobacteria.With the depth increasing,the species of Proteobacteria decreased,but those of Firmicutes increased.Some bacterial species in Layer A also belonged to Cyanobacteria,Actinobacteria,Planctomycete,and Spirochaetes.And some in Layer C belonged to Bacteroidetes.The results revealed the changes of bacterial diversity and species composition during the decomposition of cyanobacteria accumulation.This study on the bacterial diversity not only provided microbiology information for understanding the decomposition process of cyanobacteria accumulation,but also supplied the theoretical basis for using the resources of unknown bacteria in the cyanobacteria accumulation.

国家“九七三”重点基础研究发展计划项目(2008CB418004)