Evaluation of different 16S rRNA gene hypervariable regions and reference databases for profiling engineered microbiota structure and functional guilds in a swine wastewater treatment plant.

IF 3.6 3区生物学 Q1 BIOLOGY

Interface Focus Pub Date : 2023-06-09 eCollection Date: 2023-08-06 DOI:10.1098/rsfs.2023.0012

Limin Lin, Feng Ju

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引用次数: 0

Abstract

High-throughput 16S rRNA gene amplicon sequencing technology is widely applied for environmental microbiota structure analysis to derive knowledge that informs microbiome-based surveillance and oriented bioengineering. However, it remains elusive how the selection of 16S rRNA gene hypervariable regions and reference databases affects microbiota diversity and structure profiling. This study systematically evaluated the fitness of different frequently used reference databases (i.e. SILVA 138 SSU, GTDB bact120_r207, Greengenes 13_5 and MiDAS 4.8) and primers of 16S rRNA gene in microbiota profiling of anaerobic digestion and activated sludge collected from a full-scale swine wastewater treatment plant (WWTP). The comparative results showed that MiDAS 4.8 achieved the highest levels of taxonomic diversity and species-level assignment rate. For whichever sample groups, microbiota richness captured by different primers decreased in the following order: V4 > V4-V5 > V3-V4 > V6-V8/V1-V3. Using primer-bias-free metagenomic data results as the judging standard, V4 region also best characterized microbiota structure and well represented typical functional guilds (e.g. methanogens, ammonium oxidizers and denitrifiers), while V6-V8 regions largely overestimated the archaeal methanogens (mainly Methanosarcina) by over 30 times. Therefore, MiDAS 4.8 database and V4 region are recommended for best simultaneous analysis of bacterial and archaeal community diversity and structure of the examined swine WWTP.

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评估不同的 16S rRNA 基因超变区和参考数据库，以分析猪废水处理厂中的工程微生物群结构和功能区。

高通量 16S rRNA 基因扩增片段测序技术被广泛应用于环境微生物区系结构分析，为基于微生物的监测和定向生物工程提供信息。然而，16S rRNA 基因高变异区和参考数据库的选择如何影响微生物群的多样性和结构分析仍是一个未知数。本研究系统地评估了不同常用参考数据库（即 SILVA 138 SSU、GTDB bact120_r207、Greengenes 13_5 和 MiDAS 4.8）和 16S rRNA 基因引物在从大型猪场污水处理厂（WWTP）收集的厌氧消化和活性污泥微生物区系分析中的适用性。比较结果表明，MiDAS 4.8 的分类多样性和物种水平赋值率最高。无论哪组样品，不同引物捕获的微生物群丰富度按以下顺序下降：v4 > v4-v5 > v3-v4 > v6-v8/v1-v3。以无引物偏倚的元基因组数据结果为评判标准，V4 区域也能最好地描述微生物群结构，并能很好地代表典型的功能区（如甲烷菌、氨氧化菌和反硝化菌），而 V6-V8 区域则在很大程度上高估了古甲烷菌（主要是 Methanosarcina），高估幅度超过 30 倍。因此，推荐使用 MiDAS 4.8 数据库和 V4 区域来同时分析猪污水处理厂的细菌和古细菌群落多样性和结构。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Interface Focus BIOLOGY-

CiteScore

9.20

自引率

0.00%

发文量

审稿时长

6-12 weeks

期刊介绍： Each Interface Focus themed issue is devoted to a particular subject at the interface of the physical and life sciences. Formed of high-quality articles, they aim to facilitate cross-disciplinary research across this traditional divide by acting as a forum accessible to all. Topics may be newly emerging areas of research or dynamic aspects of more established fields. Organisers of each Interface Focus are strongly encouraged to contextualise the journal within their chosen subject.