Choice of DNA extraction method affects DNA metabarcoding of unsorted invertebrate bulk samples

Majaneva, Markus; Diserud, Ola Håvard; Hajibabaei, Mehrdad; Eagle, Shannon H. C.; Ekrem, Torbjørn

doi:10.3897/mbmg.2.26664

Cited by 46 publications

(59 citation statements)

References 58 publications

(82 reference statements)

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“…As previously described for other types of samples (Deiner, Walser, Mächler, & Altermatt, 2015;Hermans, Buckley, & Lear, 2018;Majaneva, Diserud, Eagle, Hajibabaei, & Ekrem, 2018), the DNA extraction method was one of the main technical factors affecting metabarcoding community estimates from preservative ethanol. Overall, our results showed that column-based DNA extraction methods (TISSUE and SOIL) tended to have lower performance compared to the magnetic-based method (BEAD).…”

Section: Discussionmentioning

confidence: 99%

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

Martins

Galhardo

Filipe

et al. 2019

Molecular Ecology Resources

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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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Section: Discussionmentioning

confidence: 99%

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

Martins

Galhardo

Filipe

et al. 2019

Molecular Ecology Resources

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“…The DNA extraction method itself may affect the quantity and quality of the extracted DNA (Jedlicka, Sharma, & Almeida, ; Majaneva, Diserud, Eagle, Hajibabaei, & Ekrem, ; Oehm et al, ), with obvious downstream consequences. DNA extraction is especially important when targeting metazoans, compared to prokaryotes and micro‐eukaryotes (Dopheide et al, ).…”

Section: Factors Distorting Diet Assessmentsmentioning

confidence: 99%

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Alberdi

Aizpurua

Bohmann

et al. 2018

Molecular Ecology Resources

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195

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The application of high‐throughput sequencing‐based approaches to DNA extracted from environmental samples such as gut contents and faeces has become a popular tool for studying dietary habits of animals. Due to the high resolution and prey detection capacity they provide, both metabarcoding and shotgun sequencing are increasingly used to address ecological questions grounded in dietary relationships. Despite their great promise in this context, recent research has unveiled how a wealth of biological (related to the study system) and technical (related to the methodology) factors can distort the signal of taxonomic composition and diversity. Here, we review these studies in the light of high‐throughput sequencing‐based assessment of trophic interactions. We address how the study design can account for distortion factors, and how acknowledging limitations and biases inherent to sequencing‐based diet analyses are essential for obtaining reliable results, thus drawing appropriate conclusions. Furthermore, we suggest strategies to minimize the effect of distortion factors, measures to increase reproducibility, replicability and comparability of studies, and options to scale up DNA sequencing‐based diet analyses. In doing so, we aim to aid end‐users in designing reliable diet studies by informing them about the complexity and limitations of DNA sequencing‐based diet analyses, and encourage researchers to create and improve tools that will eventually drive this field to its maturity.

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“…The PCR and bioinformatic processing were conducted as described in Majaneva, Diserud, Eagle, Hajibabaei, and Ekrem (2018).…”

Section: Biological Samplingmentioning

confidence: 99%

DNA metabarcoding adds valuable information for management of biodiversity in roadside stormwater ponds

Sun

Majaneva

Sokolova

et al. 2019

Ecology and Evolution

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Stormwater ponds are used to compensate for the adverse effects that road runoff might have on the natural environment. Depending on their design and placement, stormwater ponds can act as both refugia and traps for local biodiversity. To evaluate the impact of stormwater ponds on biodiversity, it is critical to use effective and precise methods for identification of life associated with the water body. DNA metabarcoding has recently become a promising tool for identification and assessment of freshwater biodiversity. Using both morphology and DNA metabarcoding, we analyze species richness and biological composition of samples from 12 stormwater ponds and investigate the impact of pond size and pollution levels in the sediments and water column on the macroinvertebrate community structure. DNA metabarcoding captured and identified more than twice the number of taxa compared to morphological identification. The (dis)similarity of macroinvertebrate community composition in different ponds showed that the ponds appear better separated in the results obtained by DNA metabarcoding, but that the explained variation is higher for the results obtained by morphologically identification, since it provides abundance data. The reliance on morphological methods has limited our perception of the aquatic biodiversity in response to anthropogenic stressors, thereby providing inaccurate information for appropriate design and management of stormwater ponds; these drawbacks can be overcome by DNA metabarcoding. Synthesis and applications. The results indicate that DNA metabarcoding is a useful tool in identifying species, especially Diptera, which are difficult to determine. Application of DNA metabarcoding greatly increases the number of species identified at each sampling site, thereby providing a more accurate information regarding the way the ponds function and how they are affected by management. OPEN PRACTICES This article has earned an Open Data Badge for making publicly available the digitally‐shareable data necessary to reproduce the reported results. The data is available at https://www.ebi.ac.uk/ena/data/view/PRJEB30841.

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Choice of DNA extraction method affects DNA metabarcoding of unsorted invertebrate bulk samples

Cited by 46 publications

References 58 publications

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

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

Promises and pitfalls of using high‐throughput sequencing for diet analysis

DNA metabarcoding adds valuable information for management of biodiversity in roadside stormwater ponds

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