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Metabarcoding meiofauna biodiversity assessment in four beaches of Northern Colombia: effects of sampling protocols and primer choice

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A Correction to this article was published on 12 May 2021

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Abstract

Environmental DNA (eDNA) metabarcoding can enhance understanding of global biodiversity by making it possible to study taxonomic groups that are difficult to sample. However, experimental choices made when generating eDNA data can impact biodiversity surveys and must be carefully considered during study design. Here, we explored the impact of DNA extraction protocol and metabarcode choice on recovery of meiofauna DNA from sand. We extracted DNA from untreated sand and from sand treated with either Ludox or MgCl2 and amplified DNA using the 18S and CO1 metabarcodes. We found differences in species composition and richness both between metabarcodes and among sampling strategies, confirming the sensitivity of the experiments to both parameters. Combining data from multiple barcodes and from multiple extraction protocols increased recovered meiofaunal taxonomic diversity. Future metabarcoding studies and meta-analyses should consider the effects of sampling protocols on biodiversity. Our results also highlight the need to continue to improve existing reference databases of morphological and molecular characterization of meiofauna, in particular of the tropics, which are poorly represented in existing databases.

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Data availability

All raw sequencing data from this study have been deposited in the NCBI Sequence Read Archive (SRA) under BioProject accession number PRJNA694867 (BioSample accessions SAMN17575766– SAMN17575777). All reference databases, taxonomic assignments of Amplicon Sequence Variants, as well as all scripts are available in Zenodo (http://doi.org/10.5281/zenodo.4588923).

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Acknowledgements

Thanks to Maria Victoria Leon, Daniel Giraldo, and Angie Colorado for helping with the collection and processing of samples. This work was supported by a Fulbright visitor research fellowship to Lyda R. Castro and by Fondo Patrimonial de Investigación Fonciencias 2019, Universidad del Magdalena.

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Authors and Affiliations

  1. Grupo de investigación Evolución, Sistemática y Ecología Molecular, Universidad del Magdalena, Carrera 32 # 22-08, Santa Marta DTCH, Colombia

    Lyda R. Castro

  2. Grupo de Investigación en Manejo y Conservación de Fauna, Flora y Ecosistemas Estratégicos Neotropicales MIKU, Universidad del Magdalena, Carrera 32 # 22-08, Hangar B, Santa Marta DTCH, Colombia

    Ana M. Lagos & Sigmer Y. Quiroga

  3. Department of Ecology and Evolutionary Biology, University of California Santa Cruz, Santa Cruz, USA

    Rachel S. Meyer, Beth Shapiro, Sabrina Shirazi & Samuel Cutler

  4. Howard Hughes Medical Institute, University of California Santa Cruz, Santa Cruz, CA, 95064, USA

    Beth Shapiro

Authors
  1. Lyda R. Castro
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  2. Rachel S. Meyer
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  3. Beth Shapiro
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  4. Sabrina Shirazi
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  5. Samuel Cutler
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  6. Ana M. Lagos
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  7. Sigmer Y. Quiroga
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Contributions

All authors contributed to the study conception and design. SYQ and AML collected the material; LRC and SC generated the molecular data; LRC, SC, SS, and RSM contributed to the analysis of the data. All the authors contributed to the interpretation of the results. LRC wrote the first draft of the manuscript and all authors contributed to writing the final version. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Lyda R. Castro.

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The original version of this article was revised: the cited reference for Fais et al. 2020 was incorrect and the name of the used DNA extraction kit has been corrected.

Supplementary Information

Below is the link to the electronic supplementary material.

10750_2021_4576_MOESM1_ESM.xlsx

Supplementary file1 (XLSX 82 kb) Online Resource 1 Taxonomy tables for 18S and CO1 showing observed taxa and their read counts by sample. Read counts are totals after decontamination.

10750_2021_4576_MOESM2_ESM.pdf

Supplementary file2 (PDF 330 kb) Online Resource 2 Barplots of rarefied relative abundance for datasets from 18S and CO1 metabarcode total data as well as datasets filtered to only contain meiofauna. The top 21 taxa are defined. This plot includes local contribution to beta diversity statistics on the proportion of unique taxa in a sample compared to the rest of the samples in its locality

Supplementary file3 (PDF 156 kb) Online Resource 3 Heatmaps of phylum by samples, labeled by sampling type.

10750_2021_4576_MOESM4_ESM.pdf

Supplementary file4 (PDF 233 kb) Online Resource 4 Rarefication curves of 18S and CO1 using total and meiofauna datasets showing the number of taxa observed as a function of sequence depth (number of reads).

10750_2021_4576_MOESM5_ESM.pdf

Supplementary file5 (PDF 174 kb) Online Resource 5 Species accumulation curves obtained with the 18S and CO1 total datasets in four beaches of northern Colombia showing interpolated (sample size-based rarefaction) species richness and extrapolated species richness (dashed lines).

Supplementary file6 (XLSX 22 kb) Online Resource 6 iNEXT asymptotic diversity estimates along with related statistics.

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Supplementary file7 (PDF 283 kb) Online Resource 7 DeSeq2 plots obtained using the meiofauna datasets with the 18S and CO1 metabarcodes. Plot titles indicate the comparison of overrepresented reads.

10750_2021_4576_MOESM8_ESM.xlsx

Supplementary file8 (XLSX 53 kb) Online Resource 8 DESeq2 results showing taxa overrepresented in one sample preparation technique over another (see tab label). Negative values indicate underrepresentation.

10750_2021_4576_MOESM9_ESM.xlsx

Supplementary file9 (XLSX 19 kb) Online Resource 9 Alpha diversity statistics using Shannon, Observed, Chao and Simpson indexes calculated per technique (Sand, MgCl2, Ludox) and Locality (Santa Marta, Rodadero, Monoguaca, Sisiguaca) using 18S and CO1 complete and meiofauna datasets. B. Significant differences among categories were calculated using Kruskal-Wallis and post hoc Dunn test with Benjamini-Hochberg adjustment (adjP < 0.05 considered significant).

10750_2021_4576_MOESM10_ESM.xlsx

Supplementary file10 (XLSX 10 kb) Online Resource 10 Beta Diversity Adonis test results showing significant effects of technique, locality, and their interactions on Jaccard Distance composition.

10750_2021_4576_MOESM11_ESM.xlsx

Supplementary file11 (XLSX 51 kb) Online Resource 11 Blast results obtained from up to 50 sequences from all species found in the area (Table 2), including percentages of identity to each species.

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Castro, L.R., Meyer, R.S., Shapiro, B. et al. Metabarcoding meiofauna biodiversity assessment in four beaches of Northern Colombia: effects of sampling protocols and primer choice. Hydrobiologia 848, 3407–3426 (2021). https://doi.org/10.1007/s10750-021-04576-z

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