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

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dc.contributor.authorMartins, Filipa M.S.
dc.contributor.authorGalhardo, Mafalda
dc.contributor.authorFilipe, Ana F.
dc.contributor.authorTeixeira, Amilcar
dc.contributor.authorPinheiro, Paulo
dc.contributor.authorPaupério, Joana
dc.contributor.authorAlves, Paulo C.
dc.contributor.authorBeja, Pedro
dc.date.accessioned2019-07-02T11:27:32Z
dc.date.available2019-07-02T11:27:32Z
dc.date.issued2019
dc.descriptionResource Articlept_PT
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 biomonitoringpt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.citationMol Ecol Resour. 2019;19:863–876pt_PT
dc.identifier.doi10.1111/1755-0998.13012pt_PT
dc.identifier.urihttp://hdl.handle.net/10400.5/18086
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherJohn Wiley and Sonspt_PT
dc.relationPTDC/ AAGMAA/2261/2014–POCI-1- 145-FEDER-356016824pt_PT
dc.relationSFRH/ BD/104703/2014pt_PT
dc.relationHorizon 2020 Framework Programme, Grant/Award Number: 668981pt_PT
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/pt_PT
dc.subjectbenthic macroinvertebratespt_PT
dc.subjectDNA extractionpt_PT
dc.subjectDNA metabarcodingpt_PT
dc.subjectfreshwater bioassessmentpt_PT
dc.subjectpreservative ethanolpt_PT
dc.subjectWater Framework Directivept_PT
dc.titleHave the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoringpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.citation.titleMolecular Ecology Resourcespt_PT
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT

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