Taxonomic Assessment of Rumen Microbiota Using Total RNA and Targeted Amplicon Sequencing Approaches.
Taxonomic Assessment of Rumen Microbiota Using Total RNA and Targeted Amplicon Sequencing Approaches.
复制标题
DOI：
10.3389/fmicb.2016.00987
复制
发表时间：
2016
影响因子：
5.2
通讯作者：
Guan le L
中科院分区：
文献类型：
--
作者：
Li F;Henderson G;Sun X;Cox F;Janssen PH;Guan le L
活性胃肠道微生物群的分类表征对于检测不同条件下微生物群落和功能的变化至关重要。本研究旨在利用总RNA测序技术鉴定和量化潜在活性的瘤胃微生物群，并将该方法的结果与广泛使用的靶向RNA/DNA扩增子测序技术进行比较。从5只肉牛瘤胃食糜样品中分离总RNA进行Illumina配对端测序（RNA-seq），并对细菌和古细菌的部分16S rRNA/rDNA进行454焦磷酸测序（RNA/DNA扩增子-seq）。RNA-seq、RNA Amplicon-seq和DNA Amplicon-seq数据集的分类评估使用内部开发的管道进行。检测到的主要微生物类群在三个数据集中是共同的，在所有数据集中共有7个细菌门，15个细菌科和5个古细菌分类群。在每个数据集中还检测到独特的微生物分类群。Elusimicrobia和Verrucomicrobia门；Desulfovibrionaceae， elusimicroaceae和Sphaerochaetaceae科；和Methanobrevibacter woesei仅在RNA- seq和RNA Amplicon-seq数据集中检测到，而Streptococcaceae仅在DNA Amplicon-seq数据集中检测到。此外，4个细菌门、8个细菌科和1个古细菌分类群的相对丰度在3个数据集之间存在差异。这是第一个比较RNA-seq和RNA/DNA扩增子-seq数据集之间瘤胃微生物群分析结果的研究。我们的结果说明了这些方法在定性和定量地表征同一样品的微生物群方面的差异，因此在比较数据时必须谨慎。
Taxonomic characterization of active gastrointestinal microbiota is essential to detect shifts in microbial communities and functions under various conditions. This study aimed to identify and quantify potentially active rumen microbiota using total RNA sequencing and to compare the outcomes of this approach with the widely used targeted RNA/DNA amplicon sequencing technique. Total RNA isolated from rumen digesta samples from five beef steers was subjected to Illumina paired-end sequencing (RNA-seq), and bacterial and archaeal amplicons of partial 16S rRNA/rDNA were subjected to 454 pyrosequencing (RNA/DNA Amplicon-seq). Taxonomic assessments of the RNA-seq, RNA Amplicon-seq, and DNA Amplicon-seq datasets were performed using a pipeline developed in house. The detected major microbial phylotypes were common among the three datasets, with seven bacterial phyla, fifteen bacterial families, and five archaeal taxa commonly identified across all datasets. There were also unique microbial taxa detected in each dataset. Elusimicrobia and Verrucomicrobia phyla; Desulfovibrionaceae, Elusimicrobiaceae, and Sphaerochaetaceae families; and Methanobrevibacter woesei were only detected in the RNA-Seq and RNA Amplicon-seq datasets, whereas Streptococcaceae was only detected in the DNA Amplicon-seq dataset. In addition, the relative abundances of four bacterial phyla, eight bacterial families and one archaeal taxon were different among the three datasets. This is the first study to compare the outcomes of rumen microbiota profiling between RNA-seq and RNA/DNA Amplicon-seq datasets. Our results illustrate the differences between these methods in characterizing microbiota both qualitatively and quantitatively for the same sample, and so caution must be exercised when comparing data.