Benchmark of 16S rRNA gene amplicon sequencing using Japanese gut microbiome data from the V1–V2 and V3–V4 primer sets

BMC Genomics

Satoshi Watanabe¹  Yu Sawai¹  Yuki Midorikawa¹  Ryuichi Kubo¹  Natsuko O. Shinozaki¹  Aya K. Takeda¹  Nicolas Jung²  Shoichiro Kameoka³  Daisuke Motooka⁴  Shota Nakamura⁵ 

[1] Cykinso, Inc. Shibuya, Tokyo, Japan;Department of Infection Metagenomics, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;Department of Infection Metagenomics, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;Cykinso, Inc. Shibuya, Tokyo, Japan;Department of Infection Metagenomics, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;Next-Generation Sequencing Core Facility, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;Integrated Frontier Research for Medical Science Division, Institute for Open and Transdisciplinary Research Initiatives, Osaka University, Suita, Osaka, Japan;Department of Infection Metagenomics, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;Next-Generation Sequencing Core Facility, Genome Information Research Center, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;Integrated Frontier Research for Medical Science Division, Institute for Open and Transdisciplinary Research Initiatives, Osaka University, Suita, Osaka, Japan;Laboratory of Pathogen Detection and Identification, International Research Center for Infectious Diseases, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan;

关键词: Microbiota;    16S rRNA;    Next-generation sequencing;   

DOI  :  10.1186/s12864-021-07746-4

来源: Springer

【 摘 要 】

Background16S rRNA gene amplicon sequencing (16S analysis) is widely used to analyze microbiota with next-generation sequencing technologies. Here, we compared fecal 16S analysis data from 192 Japanese volunteers using the modified V1–V2 (V12) and the standard V3–V4 primer (V34) sets to optimize the gut microbiota analysis protocol.ResultsQIIME1 and QIIME2 analysis revealed a higher number of unclassified representative sequences in the V34 data than in the V12 data. The comparison of bacterial composition demonstrated that at the phylum level, Actinobacteria and Verrucomicrobia were detected at higher levels with V34 than with V12. Among these phyla, we observed higher relative compositions of Bifidobacterium and Akkermansia with V34. To estimate the actual abundance, we performed quantitative real-time polymerase chain reaction (qPCR) assays for Akkermansia and Bifidobacterium. We found that the abundance of Akkermansia as detected by qPCR was close to that in V12 data, but was markedly lower than that in V34 data. The abundance of Bifidobacterium detected by qPCR was higher than that in V12 and V34 data.ConclusionsThese results indicate that the bacterial composition derived from the V34 region might differ from the actual abundance for specific gut bacteria. We conclude that the use of the modified V12 primer set is more desirable in the 16S analysis of the Japanese gut microbiota.

【 授权许可】

CC BY