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      The use of citizen science in fish eDNA metabarcoding for evaluating regional biodiversity in a coastal marine region: A pilot study

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      Metabarcoding and Metagenomics
      Pensoft Publishers

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          Abstract

          To test the feasibility of a citizen science program for fish eDNA metabarcoding in coastal marine environments, we recruited six groups of voluntary citizens for a science education course at a natural history museum. We held a seminar on eDNA and a workshop for seawater sampling and on-site filtration using syringes and filter cartridges for the participants. After that, they selected single survey sites following the guidelines for conducting a safe field trip. They performed seawater sampling and on-site filtration at these sites during their summer holidays. The six selected sites unexpectedly included diverse coastal habitats within a 40 km radius, located at temperate latitudes in central Japan (~35°N). After the field trips, they returned filtered cartridges to the museum, and we extracted eDNA from the filters. We performed fish eDNA metabarcoding, along with data analysis. Consequently, we identified 140 fish species across 66 families and 118 genera from the six samples, with species richness ranging from 14 to 66. Despite its limited sample size, such a diverse taxonomic range of fish species exhibited spatial biodiversity patterns within the region, which are consistent with species distribution. These include north-south and urbanization gradients of species richness, geographic structure of the fish communities, and varying salinity preferences of the component species. This case study demonstrates the potential of fish eDNA metabarcoding as an educational and scientific tool to raise public awareness and perform large-scale citizen science initiatives encompassing regional, national, or global fauna.

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          MAFFT Multiple Sequence Alignment Software Version 7: Improvements in Performance and Usability

          We report a major update of the MAFFT multiple sequence alignment program. This version has several new features, including options for adding unaligned sequences into an existing alignment, adjustment of direction in nucleotide alignment, constrained alignment and parallel processing, which were implemented after the previous major update. This report shows actual examples to explain how these features work, alone and in combination. Some examples incorrectly aligned by MAFFT are also shown to clarify its limitations. We discuss how to avoid misalignments, and our ongoing efforts to overcome such limitations.
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            Search and clustering orders of magnitude faster than BLAST.

            Biological sequence data is accumulating rapidly, motivating the development of improved high-throughput methods for sequence classification. UBLAST and USEARCH are new algorithms enabling sensitive local and global search of large sequence databases at exceptionally high speeds. They are often orders of magnitude faster than BLAST in practical applications, though sensitivity to distant protein relationships is lower. UCLUST is a new clustering method that exploits USEARCH to assign sequences to clusters. UCLUST offers several advantages over the widely used program CD-HIT, including higher speed, lower memory use, improved sensitivity, clustering at lower identities and classification of much larger datasets. Binaries are available at no charge for non-commercial use at http://www.drive5.com/usearch.
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              Exact sequence variants should replace operational taxonomic units in marker-gene data analysis

              Recent advances have made it possible to analyze high-throughput marker-gene sequencing data without resorting to the customary construction of molecular operational taxonomic units (OTUs): clusters of sequencing reads that differ by less than a fixed dissimilarity threshold. New methods control errors sufficiently such that amplicon sequence variants (ASVs) can be resolved exactly, down to the level of single-nucleotide differences over the sequenced gene region. The benefits of finer resolution are immediately apparent, and arguments for ASV methods have focused on their improved resolution. Less obvious, but we believe more important, are the broad benefits that derive from the status of ASVs as consistent labels with intrinsic biological meaning identified independently from a reference database. Here we discuss how these features grant ASVs the combined advantages of closed-reference OTUs—including computational costs that scale linearly with study size, simple merging between independently processed data sets, and forward prediction—and of de novo OTUs—including accurate measurement of diversity and applicability to communities lacking deep coverage in reference databases. We argue that the improvements in reusability, reproducibility and comprehensiveness are sufficiently great that ASVs should replace OTUs as the standard unit of marker-gene analysis and reporting.
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                Author and article information

                Contributors
                Journal
                Metabarcoding and Metagenomics
                MBMG
                Pensoft Publishers
                2534-9708
                May 23 2022
                May 23 2022
                : 6
                Article
                10.3897/mbmg.6.80444
                d51c1505-693c-42e5-8496-b9145afdc3af
                © 2022

                http://creativecommons.org/licenses/by/4.0/

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