ArticleInternational microbiology : the official journal of the Spanish Society for Microbiology2024
A new spike-in-based method for quantitative metabarcoding of soil fungi and bacteria.
Article in International microbiology : the official journal of the Spanish Society for Microbiology, 2024. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Who cites it
7 citing papers in PubMed.
- Limitations of Common Molecular Markers in Fungal Biodiversity Analysis and the Benefits of Their Synergistic Use.Molecular ecology resources · 2026Article
- Quantitative metagenomics using a portable protocol.Applied and environmental microbiology · 2026Article
- From Sampling to Identification of Arbuscular Mycorrhizal Fungi Through Next Generation Sequencing.Methods in molecular biology (Clifton, N.J.) · 2026Article
- Year-Round Quantification, Structure and Dynamics of Epibacterial Communities From Diverse Macroalgae Reveal a Persistent Core Microbiota and Strong Host Specificities.Environmental microbiology reports · 2025Article
- The quest for environmental analytical microbiology: absolute quantitative microbiome using cellular internal standards.Microbiome · 2025Review
- Advancing time-since-interval estimation for clandestine graves: a forensic ecogenomics perspective into burial and translocation timelines using massively parallel sequencing.Frontiers in microbiology · 2025Review
- Assessment of absolute abundance in mother-infant gut microbiome using marine-sourced bacterial DNA spike-in and comparison with conventional quantification methods.Microbiome research reports · 2025Article
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Authors and funding
1 author.
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Abstract
Metabarcoding is a powerful tool to characterize biodiversity in biological samples. The interpretation of taxonomic profiles from metabarcoding data has been hindered by their compositional nature. Several strategies have been proposed to transform compositional data into quantitative data, but they have intrinsic limitations. Here, I propose a workflow based on bacterial and fungal cellular internal standards (spike-ins) for absolute quantification of the microbiota in soil samples. These standards were added to the samples before DNA extraction in amounts estimated after qPCRs, to target around 1-2% coverage in the sequencing run. In bacteria, proportions of spike-in reads in the sequencing run were very similar (< 2-fold change) to those predicted by the qPCR assessment, and for fungi they differed up to 40-fold. The low variation among replicates highlights the reproducibility of the method. Estimates based on multiple bacterial spike-ins were highly correlated (r = 0.99). Procrustes analysis evidenced significant biological effects on the community composition when normalizing compositional data. A protocol based on qPCR estimation of input amounts of cellular spikes is proposed as a cheap and reliable strategy for quantitative metabarcoding of biological samples.
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Registered trials
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