Simultaneous analysis of seven 16S rRNA hypervariable gene regions increases efficiency in marine bacterial diversity detection.

Autor: Leontidou K; Ecology Group, Rheinland-Pfälzische Technische Universität Kaiserslautern-Landau, Kaiserslautern, Germany., Abad-Recio IL; Marine Ecosystems Functioning, AZTI, Marine Research, Basque Research and Technology Alliance, Pasia, Gipuzkoa, Spain., Rubel V; Ecology Group, Rheinland-Pfälzische Technische Universität Kaiserslautern-Landau, Kaiserslautern, Germany., Filker S; Molecular Ecology Group, Rheinland-Pfälzische Technische Universität Kaiserslautern-Landau, Kaiserslautern, Germany., Däumer M; SeqIT, Laboratory for Molecular Diagnostics and Services, Kaiserslautern, Germany., Thielen A; SeqIT, Laboratory for Molecular Diagnostics and Services, Kaiserslautern, Germany., Lanzén A; Marine Ecosystems Functioning, AZTI, Marine Research, Basque Research and Technology Alliance, Pasia, Gipuzkoa, Spain.; IKERBASQUE, Basque Foundation for Science, Bilbao, Bizkaia, Spain., Stoeck T; Ecology Group, Rheinland-Pfälzische Technische Universität Kaiserslautern-Landau, Kaiserslautern, Germany.
Jazyk: angličtina
Zdroj: Environmental microbiology [Environ Microbiol] 2023 Dec; Vol. 25 (12), pp. 3484-3501. Date of Electronic Publication: 2023 Nov 16.
DOI: 10.1111/1462-2920.16530
Abstrakt: Environmental DNA sequencing is the gold standard to reveal microbial community structures. In most applications, a one-fragment PCR approach is applied to amplify a taxonomic marker gene, usually a hypervariable region of the 16S rRNA gene. We used a new reverse complement (RC)-PCR-based assay that amplifies seven out of the nine hypervariable regions of the 16S rRNA gene, to interrogate bacterial communities in sediment samples collected from different coastal marine sites with an impact gradient. In parallel, we employed a traditional one-fragment analysis of the hypervariable V3-V4 region to investigate whether the RC-PCR reveals more of the 'unseen' diversity obtained by the one-fragment approach. As a benchmark for the full deck of diversity, we subjected the samples to PCR-free metagenomic sequencing. None of the two PCR-based approaches recorded the full taxonomic repertoire obtained from the metagenomics datasets. However, the RC-PCR approach detected 2.8 times more bacterial genera compared to the near-saturation sequenced V3-V4 samples. RC-PCR is an ideal compromise between the standard one-fragment approach and metagenomics sequencing and may guide future environmental sequencing studies, in which bacterial diversity is a central subject.
(© 2023 The Authors. Environmental Microbiology published by Applied Microbiology International and John Wiley & Sons Ltd.)
Databáze: MEDLINE
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