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Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring

2019Journal article Open access
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dc.contributor.authorMartins, Filipa M.S.
dc.contributor.authorGalhardo, Mafalda
dc.contributor.authorFilipe, Ana Filipa
dc.contributor.authorTeixeira, Amílcar
dc.contributor.authorPinheiro, Paulo
dc.contributor.authorPaupério, Joana
dc.contributor.authorAlves, Paulo C.
dc.contributor.authorBeja, Pedro
dc.date.accessioned2018-02-19T10:00:00Z
dc.date.accessioned2020-01-06T14:10:42Z
dc.date.available2018-01-19T10:00:00Z
dc.date.available2020-01-06T14:10:42Z
dc.date.issued2019
dc.description.abstractDNA metabarcoding can contribute to improving cost‐effectiveness and accuracy of biological assessments of aquatic ecosystems, but significant optimization and standardization efforts are still required to mainstream its application into biomonitoring programmes. In assessments based on freshwater macroinvertebrates, a key challenge is that DNA is often extracted from cleaned, sorted and homogenized bulk samples, which is time‐consuming and may be incompatible with sample preservation requirements of regulatory agencies. Here, we optimize and evaluate metabarcoding procedures based on DNA recovered from 96% ethanol used to preserve field samples and thus including potential PCR inhibitors and nontarget organisms. We sampled macroinvertebrates at five sites and subsampled the preservative ethanol at 1 to 14 days thereafter. DNA was extracted using column‐based enzymatic (TISSUE) or mechanic (SOIL) protocols, or with a new magnetic‐based enzymatic protocol (BEAD), and a 313‐bp COI fragment was amplified. Metabarcoding detected at least 200 macroinvertebrate taxa, including most taxa detected through morphology and for which there was a reference barcode. Better results were obtained with BEAD than SOIL or TISSUE, and with subsamples taken 7–14 than 1–7 days after sampling, in terms of DNA concentration and integrity, taxa diversity and matching between metabarcoding and morphology. Most variation in community composition was explained by differences among sites, with small but significant contributions of subsampling day and extraction method, and negligible contributions of extraction and PCR replication. Our methods enhance reliability of preservative ethanol as a potential source of DNA for macroinvertebrate metabarcoding, with a strong potential application in freshwater biomonitoring.
dc.description.sponsorshipThis article has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 668981 (ERA Chair in Environmental metagenomics, EnvMetaGen) and was supported by EDP‐Biodiversity Chair (EDP/FCT) and the project FRESHING funded by the Portuguese Foundation for Science and Technology (FCT) and COMPETE (PTDC/ AAGMAA/2261/2014—POCI‐01‐0145‐FEDER‐356 016824). FMSM was supported by FCT PhD grant SFRH/BD/104703/2014. PB was supported by EDP‐Biodiversity Chair, MG and JP by the EnvMetaGen, and AFF by FRESHING/FCT. We thank B. Maia‐ Carvalho for his invaluable support on the development of the BEAD protocol, and M. Monaghan for suggestions on DNA quality assessment.
dc.description.versioninfo:eu-repo/semantics/publishedVersionen_EN
dc.identifier.citationMartins, Filipa M.S.; Galhardo, Mafalda; Filipe, Ana F.; Teixeira, Amílcar; Pinheiro, Paulo; Paupério, Joana; Alves, Paulo C.; Beja, Pedro (2019). Have the cake and eat it: Optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring. Molecular Ecology Resources. ISSN 1755-098X. 19, p. 863-876en_EN
dc.identifier.doi10.1111/1755-0998.13012en_EN
dc.identifier.issn1755-098X
dc.identifier.urihttp://hdl.handle.net/10198/20286
dc.language.isoeng
dc.peerreviewedyesen_EN
dc.relation2261/2014
dc.subjectBenthic macroinvertebratesen_EN
dc.subjectDNA extractionen_EN
dc.subjectDNA metabarcodingen_EN
dc.subjectFreshwater bioassessmenten_EN
dc.subjectPreservative ethanolen_EN
dc.subjectWater framework directiveen_EN
dc.titleHave the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoringen_EN
dc.typejournal article
dspace.entity.typePublication
person.familyNameTeixeira
person.givenNameAmilcar
person.identifier2153193
person.identifier.ciencia-id9510-3CF3-0393
person.identifier.orcid0000-0001-5336-1174
person.identifier.scopus-author-id7202385393
rcaap.rightsopenAccessen_EN
rcaap.typearticleen_EN
relation.isAuthorOfPublication4f0753fd-4b4f-46f6-8e2f-20c7f835a32a
relation.isAuthorOfPublication.latestForDiscovery4f0753fd-4b4f-46f6-8e2f-20c7f835a32a

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