Application of MinION Amplicon Sequencing to Buccal Swab Samples for Improving Resolution and Throughput of Rumen Microbiota Analysis

作者：Hiroto Miura, Masayuki Takeda, Megumi Yamaguchi, Yoshihisa Ohtani, Go Endo, Yasuhisa Masuda, Kaede Ito, Yoshio Nagura, Kunihiro Iwashita, Tomohiro Mitani, Yutaka Suzuki, Yasuo Kobayashi, Satoshi Koike · 发表于：Frontiers in Microbiology · 年份：2022 · DOI：10.3389/fmicb.2022.783058 · 被引用次数：12 · 研究领域：Ruminant Nutrition and Digestive Physiology

The Illumina MiSeq platform has been widely used as a standard method for studying the rumen microbiota. However, the low resolution of taxonomic identification is the only disadvantage of MiSeq amplicon sequencing, as it targets a part of the 16S rRNA gene. In the present study, we performed three experiments to establish a high-resolution and high-throughput rumen microbial profiling approach using a combination of MinION platform and buccal swab sample, which is a proxy for rumen contents. In experiment 1, rumen contents and buccal swab samples were collected simultaneously from cannulated cattle ( n = 6) and used for microbiota analysis using three different analytical workflows: amplicon sequencing of the V3–V4 region of the 16S rRNA gene using MiSeq and amplicon sequencing of near full-length 16S rRNA gene using MinION or PacBio Sequel II. All reads derived from the MinION and PacBio platforms were classified at the species-level. In experiment 2, rumen fluid samples were collected from beef cattle ( n = 28) and used for 16S rRNA gene amplicon sequencing using the MinION platform to evaluate this sequencing platform for rumen microbiota analysis. We confirmed that the MinION platform allowed species-level taxa assignment for the predominant bacterial groups, which were previously identified at the family- and genus-level using the MiSeq platform. In experiment 3, buccal swab samples were collected from beef cattle ( n = 30) and used for 16S rRNA gene amplicon sequencing...