Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring
Autor(a) principal: | |
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Data de Publicação: | 2018 |
Outros Autores: | , , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/10198/20286 |
Resumo: | DNA 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. |
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Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoringBenthic macroinvertebratesDNA extractionDNA metabarcodingFreshwater bioassessmentPreservative ethanolWater framework directiveDNA 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.This 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.Biblioteca Digital do IPBMartins, Filipa M.S.Galhardo, MafaldaFilipe, Ana FilipaTeixeira, AmílcarPinheiro, PauloPaupério, JoanaAlves, Paulo C.Beja, Pedro2018-01-19T10:00:00Z20192019-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10198/20286engMartins, 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. 863-8761755-098X10.1111/1755-0998.13012info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-11-21T10:46:14Zoai:bibliotecadigital.ipb.pt:10198/20286Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-19T23:11:04.578750Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
title |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
spellingShingle |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring Martins, Filipa M.S. Benthic macroinvertebrates DNA extraction DNA metabarcoding Freshwater bioassessment Preservative ethanol Water framework directive |
title_short |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
title_full |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
title_fullStr |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
title_full_unstemmed |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
title_sort |
Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring |
author |
Martins, Filipa M.S. |
author_facet |
Martins, Filipa M.S. Galhardo, Mafalda Filipe, Ana Filipa Teixeira, Amílcar Pinheiro, Paulo Paupério, Joana Alves, Paulo C. Beja, Pedro |
author_role |
author |
author2 |
Galhardo, Mafalda Filipe, Ana Filipa Teixeira, Amílcar Pinheiro, Paulo Paupério, Joana Alves, Paulo C. Beja, Pedro |
author2_role |
author author author author author author author |
dc.contributor.none.fl_str_mv |
Biblioteca Digital do IPB |
dc.contributor.author.fl_str_mv |
Martins, Filipa M.S. Galhardo, Mafalda Filipe, Ana Filipa Teixeira, Amílcar Pinheiro, Paulo Paupério, Joana Alves, Paulo C. Beja, Pedro |
dc.subject.por.fl_str_mv |
Benthic macroinvertebrates DNA extraction DNA metabarcoding Freshwater bioassessment Preservative ethanol Water framework directive |
topic |
Benthic macroinvertebrates DNA extraction DNA metabarcoding Freshwater bioassessment Preservative ethanol Water framework directive |
description |
DNA 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. |
publishDate |
2018 |
dc.date.none.fl_str_mv |
2018-01-19T10:00:00Z 2019 2019-01-01T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://hdl.handle.net/10198/20286 |
url |
http://hdl.handle.net/10198/20286 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
Martins, 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. 863-876 1755-098X 10.1111/1755-0998.13012 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
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Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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RCAAP |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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