应用PCR-DGGE技术比较绵羊瘤胃、网胃、瓣胃和皱胃菌群的多样性

A Comparison Study on Bacteria Diversity of Rumen,Reticulum,Omasum and Abomasum from Sheep Using PCR-DGGE Analysis

本研究采用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术结合DGGE图谱条带的克隆和测序比较了绵羊瘤胃、网胃、瓣胃和皱胃菌群的多样性,同时对菌群进行了聚类分析和主成分分析。结果表明:绵羊瘤胃、网胃、瓣胃和皱胃内容物DGGE图谱的平均条带数分别为18、13、16和15条;瘤胃和瓣胃内菌群的多样性指数、均匀度和丰富度均较高,分别为2.83、0.79、17.00和2.82、0.79、16.80,而网胃和皱胃内容物较低,分别为2.52、0.70、12.60和2.73、0.76、15.40;聚类分析结果显示不同胃的内容物菌群具有一定差异,但不同动物个体相同胃的内容物相似性系数均高于0.63;PCR-DGGE图谱中测序的条带大多数归为拟杆菌门(Bacteroidetes)和厚壁菌门(Firmicutes),优势菌群为厚壁菌门(Firmicutes)细菌、拟杆菌门(Bacteroidetes)细菌、未培养瘤胃细菌(uncultured rumen bacterium)、未培养细菌(uncultured bacterium)和韦荣球菌科(Veillonellaceae)细菌,特异性细菌为不动杆菌属(Acinetobacter sp.)细菌和瘤胃球菌属(Ruminococcaceae)细菌。结果提示,绵羊4个胃中均含有种类丰富和数量巨大的细菌,且随着消化道部位由前往后的顺序,菌群的多样性呈现先高后降低再升高的趋势。

In order to compare the bacterial flora diversity of rumen, reticulum, omasum and abomasum from sheep, polymerase chain reactiondenaturing gradient gel electrophoresis （ PCR-DGGE） analysis compounded with cloning and sequencing of bands in DGGE profiles were used, and cluster analysis and principal constituent analysis （ PCA） of bacterial flora were carried out. The results showed as follows： the average number of DGGE bands of contents in rumen, reticulum, omasum and abomasum was 18, 13, 16 and 15, respectively;Shannon diversity index, evenness and richness of bacteria in rumen and omasum were higher, which were 2.83, 0.79 and 17.00, and 2.82, 0.79 and 16.80, respectively, however, those of reticulum and abomasum were lower, which were 2.52, 0.70 and 12.60, and 2.73, 0.76 and 15.40, respectively; clustering analysis results suggested that there were some differences in bacterial flora among the contents in different stomachs, but the similarity coefficients from the same segment of stomach of different animals were all above 0.63;most of the sequenced bands belonged to Bacteroidetes and Firmicutes, and Bacteroidetes, Firmicutes, uncultured rumen bacterium, uncultured bacterium and Veillonellaceae were predominant in stomach, while the special bacteria were Acinetobacter sp. and Ruminococcaceae bacteria. In conclusion, there are rich kinds and large a-mount of bacteria in the four stomachs of sheep. With rearward moving of position of intestinal tract, the diversity increased firstly and then decreased and finally increased.