Dataset Information

Assessing insect biodiversity with automatic light traps in Brazil: Pearls and pitfalls of metabarcoding samples in preservative ethanol.

ABSTRACT: Automated species identification based on data produced with metabarcoding offers an alternative for assessing biodiversity of bulk insect samples obtained with traps. We used a standard two-step PCR approach to amplify a 313 bp fragment of the barcoding region of the mitochondrial COI gene. The PCR products were sequenced on an Illumina MiSeq platform, and the OTUs production and taxonomic identifications were performed with a customized pipeline and database. The DNA used in the PCR procedures was extracted directly from the preservative ethanol of bulk insect samples obtained with automatic light traps in 12 sampling areas located in different biomes of Brazil, during wet and dry seasons. Agricultural field and forest edge habitats were collected for all sampling areas. A total of 119 insect OTUs and nine additional OTUs assigned to other arthropod taxa were obtained at a ?97% sequence similarity level. The alpha and beta diversity analyses comparing biomes, habitats, and seasons were mostly inconclusive, except for a significant difference in beta diversity between biomes. In this study, we were able to metabarcode and HTS adult insects from their preservative medium. Notwithstanding, our results underrepresent the true magnitude of insect diversity expected from samples obtained with automatic light traps in Brazil. Although biological and technical factors might have impacted our results, measures to optimize and standardize eDNA HTS should be in place to improve taxonomic coverage of samples of unknown diversity and stored in suboptimal conditions, which is the case of most eDNA samples.

SUBMITTER: Zenker MM

PROVIDER: S-EPMC7069332 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Assessing insect biodiversity with automatic light traps in Brazil: Pearls and pitfalls of metabarcoding samples in preservative ethanol.

Zenker Mauricio M MM Specht Alexandre A Fonseca Vera G VG

Ecology and evolution 20200225 5

Automated species identification based on data produced with metabarcoding offers an alternative for assessing biodiversity of bulk insect samples obtained with traps. We used a standard two-step PCR approach to amplify a 313 bp fragment of the barcoding region of the mitochondrial COI gene. The PCR products were sequenced on an Illumina MiSeq platform, and the OTUs production and taxonomic identifications were performed with a customized pipeline and database. The DNA used in the PCR procedures ...[more]

PMID: 32184986

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Project description:BackgroundCharacterizing biodiversity in a habitat or in targeted taxonomically or socioeconomically important groups remains a challenge. Standard DNA-based biodiversity identification tools such as DNA barcoding coupled with high-throughput Next-Generation Sequencing (NGS) technologies are rapidly changing the landscape of biodiversity analysis by targeting various habitats and a wide array of organisms. However, effective use of these technological advances requires optimized protocols and benchmarking against traditional tools. Here we investigate the use of commonly used preservative ethanol as a non-destructive and inexpensive source of DNA for NGS biodiversity analysis of benthic macroinvertebrates. We used the preservative ethanol added to field collected organisms (live sorted bulk benthic samples) as a source of community DNA for NGS environmental barcoding. We directly compare this approach with a DNA barcode library generated using Sanger sequencing of all individuals separated from abenthic sample as well as with NGS environmental barcoding of DNA extracted from mixed/homogenized tissue specimens of the same benthic sample. We also evaluate a multiplex PCR strategy, as compared to commonly used single amplicon workflow, using three newly designed primer sets targeting a wide array of benthic macroinvertebrate taxa.ResultsOur results indicate the effectiveness of ethanol-based DNA in providing sequence information from 87% of taxa identified individually from mixture as compared to 89% in conventional tissue extracted DNA. Missing taxa in both DNA sources were from species with the lowest abundance (e.g. 1 individual) in the benthic mixture. Interestingly, we achieved 100% detection for taxa represented with more than 1% individuals in the mixture in both sources of DNA. Our multiplex amplification regime increased the detection as compared to any single primer set indicating the usefulness of using multiple primer sets in initial amplification of target genes.ConclusionsAlthough NGS approaches have significantly increased the potential of using DNA information in biodiversity analysis, robust methods are needed to provide reliable data and alleviate sample-processing bottlenecks. Here we coupled non-destructive DNA access and a multiplex PCR approach in NGS environmental barcoding for effective data generation from benthic live-sorted samples collected in bulk and preserved in ethanol. Our study provides a possible solution to sampling and vouchering challenges in using benthic samples through next-generation environmental barcoding and facilitates wider utility of DNA information, especially species-specific DNA barcodes, in ecological and environmental studies and real-world applications such as biomonitoring programs.

Dataset Information

Assessing insect biodiversity with automatic light traps in Brazil: Pearls and pitfalls of metabarcoding samples in preservative ethanol.

Publications

Assessing insect biodiversity with automatic light traps in Brazil: Pearls and pitfalls of metabarcoding samples in preservative ethanol.

Similar Datasets

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