不同窖龄窖泥细菌的系统发育多样性分析

Analysis of Bacterial Phylogenetic Diversity of Pit Muds with Different Cellar Ages

提取20、50、150a窖龄窖泥样品总DNA,构建窖泥细菌16S rDNA文库,经过16S rDNA扩增片段的限制性酶切分析(ARDRA)、DNA序列测定和系统发育关系分析,对不同窖龄窖泥细菌群落组成与多样性进行研究。结果表明:398个细菌阳性克隆子分成37个可操作分类单元(OTUs),系统发育分析归为6个门,优势顺序为:厚壁菌门(Firmicute)65.1%、拟杆菌门(Bacteroidetes)21.4%、绿弯菌门(Chloroflexi)4.8%、放线菌门(Actinobacteria)3.8%、未分类细菌(Unclassified Bacteria)3.9%和变形菌门(Proteobacteria)1.0%,窖泥中的大多数克隆子(73.9%)与未培养细菌亲缘关系近。20a窖泥优势菌是Syntrophomonas、Petrimonas、Proteiniphilum,50a和150a龄窖泥优势菌为Natronoanaerobium和Unclassified Clostrida。随着窖龄的增加,窖泥细菌多样性指数呈增加的趋势,从20～50a之间变化幅度较大,50a和150a窖泥的菌群结构最为相似。本研究初步揭示了不同窖龄窖泥细菌系统发育多样性与窖泥老熟老化的关系。

Total DNAs were extracted from pit muds with different cellar ages, 20, 50 a and 150 a, respectively and were used for constructing 16S rDNA clone libraries. The bacterial composition and diversity from the pit muds were investigated by amplified rDNA restriction analysis （ARDRA）, DNA sequencing and phylogenetic analysis. The results showed that 398 clones from the 16S rDNA library fell into 37 operational taxonomic units （OTUs） and 6 phyla including Firmicute （65.1%）, Bacteroidetes （21.4%）, Chloroflexi （4.8%）, Actinobacteria （3.8%）, unclassified bacteria （3.9%） and Proteobacteria （1.0%）. Most of the clones （73.9%） showed more affiliation with the uncultured bacteria. The predominant bacteria from the pit mud with cellar age of 20 years were Syntrophomonas, Petrimonas and Proteiniphilum, while the predominant bacteria from the pit muds with cellar age of 50 a and 150 a were Natronoanaerobium and unclassified Clostrida. The bacterial diversity index of pit mud increased with increasing cellar age, especially when the cellular age increased from 20 to 50 a. The community structure of the pit muds with cellar age of 50 a and 150 a had the highest similarity. This research revealed the relationship between bacterial phylogenetic diversity of pit muds with different cellar ages and the aging of pit mud.