ArticlemSphere2021
Primer, Pipelines, Parameters: Issues in 16S rRNA Gene Sequencing.
Article in mSphere, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 263 papers, 4 of them syntheses that pooled it.
What it found
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.
Who cites it
263 citing papers in PubMed, 4 syntheses or guidelines pooled it, 436 citations in OpenAlex.
- Standardized 16S Meta-Analysis Reveals Consistent Host-Associated Patterns in Laminariales Epiphytic Microbiomes.MicrobiologyOpen · 2026Pooled it
- Current Methods in Synovial Fluid Microbiota Characterization: A Systematic Review.International journal of molecular sciences · 2025Pooled it
- Meta-analysis of oral microbiome reveals sex-based diversity in biofilms during periodontitis.JCI insight · 2024Pooled it
- The effects of time-restricted eating and Ramadan fasting on gut microbiota composition: a systematic review of human and animal studies.Nutrition reviews · 2024Pooled it
- Nasal microbiome and the effect of nasal decolonization with a novel povidone-iodine antiseptic solution: a prospective and randomized clinical trial.Scientific reports · 2024Trial
- Early risk stratification of late-onset sepsis in very preterm infants by intestinal microbiota profiling: a multicenter case-control validation study.Gut microbes · 2026Article
- Systematic quantification and removal of host DNA contamination in 16S rRNA gene sequencing.Quantitative biology (Beijing, China) · 2026Article
- High-yield gamma-aminobutyric acid synthesis by wild-type Levilactobacillus brevis strains isolated from traditional Iranian Kashk.International microbiology : the official journal of the Spanish Society for Microbiology · 2026Article
- Urbanization-Associated Microbial Diversity Loss and Its Impact on Elderly Health: Integrating Environmental and Host Microbiome Dynamics.Environmental microbiology · 2026Article
- SURE-Pipe: a pipeline to compare genomes and extract shared and unique regions.NAR genomics and bioinformatics · 2026Article
- A scalable and cost-effective nanopore workflow for 16S rRNA microbiome profiling validated in a zebrafish gut model.Microbial genomics · 2026Article
- Single-cell detection and quantification of the microbiota by MicFLY.Cell host & microbe · 2026Article
- Elucidating the Impact of Bacterial Composition and Diversity on Skin Homeostasis Using a Human 3D In Vitro Skin Model.Microorganisms · 2026Article
- Voluntary wheel running modulates murine gut microbiome during hyperammonemic stress.Microbiology spectrum · 2026Article
- Innovative technologies and future perspectives in tumor microbiomics.Science China. Life sciences · 2026Review
- Article
- A comparative analysis of gut microbiome in dogs using short- and long-reads of 16S rRNA sequences reveals workflow-dependent biases.Veterinary research communications · 2026Article
- Comparison of a long-read amplicon sequencing approach to short-read amplicons for microbiome analysis.Microbiology spectrum · 2026Article
- Commercial washing and storage over shelf life impact bacterial communities more than the fungal communities on baby spinach.Microbiology spectrum · 2026Article
- Comparison of library preparation protocols and bioinformatic pipelines in high-throughput 16S rRNA gene sequencing.BMC microbiology · 2026Article
203 more citing papers are in PubMed but not listed here.
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
8 authors at 3 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Short-amplicon 16S rRNA gene sequencing is currently the method of choice for studies investigating microbiomes. However, comparative studies on differences in procedures are scarce. We sequenced human stool samples and mock communities with increasing complexity using a variety of commonly used protocols. Short amplicons targeting different variable regions (V-regions) or ranges thereof (V1-V2, V1-V3, V3-V4, V4, V4-V5, V6-V8, and V7-V9) were investigated for differences in the composition outcome due to primer choices. Next, the influence of clustering (operational taxonomic units [OTUs], zero-radius OTUs [zOTUs], and amplicon sequence variants [ASVs]), different databases (GreenGenes, the Ribosomal Database Project, Silva, the genomic-based 16S rRNA Database, and The All-Species Living Tree), and bioinformatic settings on taxonomic assignment were also investigated. We present a systematic comparison across all typically used V-regions using well-established primers. While it is known that the primer choice has a significant influence on the resulting microbial composition, we show that microbial profiles generated using different primer pairs need independent validation of performance. Further, comparing data sets across V-regions using different databases might be misleading due to differences in nomenclature (e.g.,
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What OpenQuestion holds
Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.