基于16S rRNA的高通量测序和DGGE法对比新鲜与4℃贮藏腐败的凡纳滨对虾微生物菌群变化

Comparative Analysis of Microbial Communities of Fresh and Spoiled Pacific White Shrimp Stored at 4 ℃ by 16s rRNA-based Pyrosequencing and DGGE

以新鲜和4℃条件下贮藏4 d腐败后的凡纳滨对虾为研究对象,分别采用高通量测序和变性梯度凝胶电泳(DGGE)指纹图谱技术研究对虾微生物菌群组成。结果显示:微生物在门、纲、目、科和属的分类水平上,高通量测序技术检测出新鲜对虾和腐败对虾分别有18和19门、33和46纲、60和89目、93和150科、191和304属;而DGGE法对新鲜对虾和腐败对虾均仅检测到1门、1纲,分别检测到4和5目,3和4科,6和7属。高通量测序检测到新鲜和腐败凡纳滨对虾微生物的4个优势门类群分别为变形菌门(87.54%和91.17%)、拟杆菌门(8.93%和1.05%)、厚壁菌门(1.01%和4.54%)和梭杆菌门(1.01%和0.2%),而DGGE法检测到的门水平优势类群仅变形菌门,表明高通量测序法与DGGE法在反映样本微生物群落规模上具有较大差异。通过DGGE法和高通量测序法发现贮藏期间腐败凡纳滨对虾中弧菌属微生物的相对丰度较新鲜对虾明显降低,而以腐败希瓦氏菌和波罗的海希瓦氏菌为主的希瓦氏菌属微生物相对丰度上升,说明这两种方法在反映优势菌群演替变化规律上结果一致,其中,高通量测序法检测灵敏度和精度更高。

The bacterial community of fresh and spoiled Pacific white shrimp was studied by pyrosequencing and Denaturing gradient gel electrophoresis（DGGE） technology. The results of pyrosequencing showed that there were 18 and19 phyla, 33 and 46 class, 60 and 89 order, 93 and 150 family and 191 and 304 genus in fresh and spoiled Pacific white shrimp, respectively. However, only 1 phyla, 1 class, 4 and 5 order, 3 and 4 family and 6 and 7 genus were detected by DGGE in fresh and spoiled Pacific white shrimp, respectively. The results of pyrosequencing showed that the fresh and spoiled shrimp were predominant by Proteobacteria（87.54% and 91.17%）, Bacteroidetes（8.93% and 1.05%）,Firmicutes（1.01% and 4.54%） and Fusobacteria（1.01% and 0.2%）, while only Proteobacteria was detected by DGGE.Both DGGE and pyrosequencing found that the richness of Vibrio decreased during storage when the spoiled shrimp compared with the fresh, and the richness of Shewanella such as Shewanella putrefaciens and Shewanella baltica increased.This indicated that both of the two methods got some similar results, but pyrosequencing displayed higher sensitivity and accuracy.