ArticleMicrobial ecology2013
Effects of OTU clustering and PCR artifacts on microbial diversity estimates.
Article in Microbial ecology, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 22 papers.
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Who cites it
22 citing papers in PubMed.
- Multi-Omic Assessment of Microbial Communities and Their Polyketide Biosynthetic Potential Across Abyssal Sediments.Environmental microbiology · 2026Article
- Comparison of the full-length sequence and sub-regions of 16S rRNA gene for skin microbiome profiling.mSystems · 2024Article
- Small molecule in situ resin capture provides a compound first approach to natural product discovery.Nature communications · 2024Article
- An independent evaluation in a CRC patient cohort of microbiome 16S rRNA sequence analysis methods: OTU clustering, DADA2, and Deblur.Frontiers in microbiology · 2023Article
- Fecal Microbiota Transplantation as New Therapeutic Avenue for Human Diseases.Journal of clinical medicine · 2022Review
- Microbial diversity in tropical marine sediments assessed using culture-dependent and culture-independent techniques.Environmental microbiology · 2021Article
- Toward Standards in Clinical Microbiota Studies: Comparison of Three DNA Extraction Methods and Two Bioinformatic Pipelines.mSystems · 2020Article
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- Issues and current standards of controls in microbiome research.FEMS microbiology ecology · 2019Article
- Detection of Natural Products and Their Producers in Ocean Sediments.Applied and environmental microbiology · 2019Article
- The impact of OTU sequence similarity threshold on diatom-based bioassessment: A case study of the rivers of Mayotte (France, Indian Ocean).Ecology and evolution · 2019Article
- Quality control of the traditional Chinese medicine Ruyi jinhuang powder based on high-throughput sequencing and real-time PCR.Scientific reports · 2018Article
- The Madness of Microbiome: Attempting To Find Consensus "Best Practice" for 16S Microbiome Studies.Applied and environmental microbiology · 2018Review
- Effects of Actinomycete Secondary Metabolites on Sediment Microbial Communities.Applied and environmental microbiology · 2017Article
- Metagenomic Analysis of Silage.Journal of visualized experiments : JoVE · 2017Article
- Systematic improvement of amplicon marker gene methods for increased accuracy in microbiome studies.Nature biotechnology · 2016Article
- Article
- Ecology and exploration of the rare biosphere.Nature reviews. Microbiology · 2015Review
- Metagenomics: tools and insights for analyzing next-generation sequencing data derived from biodiversity studies.Bioinformatics and biology insights · 2015Review
Corrections and comments
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Authors and funding
4 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Next-generation sequencing has increased the coverage of microbial diversity surveys by orders of magnitude, but differentiating artifacts from rare environmental sequences remains a challenge. Clustering 16S rRNA sequences into operational taxonomic units (OTUs) organizes sequence data into groups of 97 % identity, helping to reduce data volumes and avoid analyzing sequencing artifacts by grouping them with real sequences. Here, we analyze sequence abundance distributions across environmental samples and show that 16S rRNA sequences of >99 % identity can represent functionally distinct microorganisms, rendering OTU clustering problematic when the goal is an accurate analysis of organism distribution. Strict postsequencing quality control (QC) filters eliminated the most prevalent artifacts without clustering. Further experiments proved that DNA polymerase errors in polymerase chain reaction (PCR) generate a significant number of substitution errors, most of which pass QC filters. Based on our findings, we recommend minimizing the number of PCR cycles in DNA library preparation and applying strict postsequencing QC filters to reduce the most prevalent artifacts while maintaining a high level of accuracy in diversity estimates. We further recommend correlating rare and abundant sequences across environmental samples, rather than clustering into OTUs, to identify remaining sequence artifacts without losing the resolution afforded by high-throughput sequencing.
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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.