Comparison of different microbiome analysis pipelines to validate their reproducibility of gastric mucosal microbiome composition
Lehr K, Oosterlinck B, Then CK, Gemmell MR, Gedgaudas R, Bornschein J, Kupcinskas J, Smet A, Hold G, Link A (2025)
Publication Type: Journal article
Publication year: 2025
Journal
Book Volume: 10
Journal Issue: 2
DOI: 10.1128/msystems.01358-24
Abstract
Microbiome analysis has become a crucial tool for basic and translational research due to its potential for translation into clinical practice. However, there is ongoing controversy regarding the comparability of different bioinformatic analysis platforms and a lack of recognized standards, which might have an impact on the translational potential of results. This study investigates how the performance of different microbiome analysis platforms impacts the final results of mucosal microbiome signatures. Across five independent research groups, we compared three distinct and frequently used microbiome analysis bioinformatic packages (DADA2, MOTHUR, and QIIME2) on the same subset of fastQ files. The source data set encompassed 16S rRNA gene raw sequencing data (V1–V2) from gastric biopsy samples of clinically well-defined gastric cancer (GC) patients (n = 40; with and without Helicobacter pylori [H. pylori] infection) and controls (n = 39, with and without H. pylori infection). Independent of the applied protocol, H. pylori status, microbial diversity and relative bacterial abundance were reproducible across all platforms, although differences in performance were detected. Furthermore, alignment of the filtered sequences to the old and new taxonomic databases (i.e., Ribosomal Database Project, Greengenes, and SILVA) had only a limited impact on the taxonomic assignment and thus on global analytical outcomes. Taken together, our results clearly demonstrate that different microbiome analysis approaches from independent expert groups generate comparable results when applied to the same data set. This is crucial for interpreting respective studies and underscores the broader applicability of microbiome analysis in clinical research, provided that robust pipelines are utilized and thoroughly documented to ensure reproducibility.
Involved external institutions
Otto-von-Guericke-Universität Magdeburg
Germany (DE)
University of Antwerp / Universiteit Antwerpen
Belgium (BE)
University of Oxford
United Kingdom (GB)
The University of Liverpool
United Kingdom (GB)
Lithuanian University of Health Sciences / Lietuvos sveikatos mokslų universitetas (LSMU)
Lithuania (LT)
MRC Weatherall Institute Of Molecular Medicine
United Kingdom (GB)
University of New South Wales (UNSW)
Australia (AU)
Germany (DE)
University of Antwerp / Universiteit Antwerpen
Belgium (BE)
University of Oxford
United Kingdom (GB)
The University of Liverpool
United Kingdom (GB)
Lithuanian University of Health Sciences / Lietuvos sveikatos mokslų universitetas (LSMU)
Lithuania (LT)
MRC Weatherall Institute Of Molecular Medicine
United Kingdom (GB)
University of New South Wales (UNSW)
Australia (AU)
How to cite
APA:
Lehr, K., Oosterlinck, B., Then, C.K., Gemmell, M.R., Gedgaudas, R., Bornschein, J.,... Link, A. (2025). Comparison of different microbiome analysis pipelines to validate their reproducibility of gastric mucosal microbiome composition. mSystems, 10(2). https://doi.org/10.1128/msystems.01358-24
MLA:
Lehr, Konrad, et al. "Comparison of different microbiome analysis pipelines to validate their reproducibility of gastric mucosal microbiome composition." mSystems 10.2 (2025).
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