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      DNA barcoding and hypopygium shape support delimitation of sympatric Dissomphalus species (Hymenoptera, Bethylidae) from the Atlantic rainforest

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          Abstract

          Dissomphalus is a cosmopolitan genus of Bethylidae and has 269 Neotropical species divided into 32 species-groups, mostly defined by the genital and the tergal process structures. Dissomphalus rectilineus and D. concavatus are sympatric species in the ulceratus species-group. Members of the species-group share many similarities in the morphology of the head, hypopygium, tergal process and genitalia, but may be distinguished by the structure of the hypopygium. Previous studies have found intermediate structures of the hypopygium in the sympatric areas and raised questions about the distinctiveness of these two species. We sequenced 340 bp of the mitochondrial gene cytochrome oxidase I of 29 specimens from Brazil and Paraguay, calculated the genetic divergence among specimens, and recovered the phylogenetic relationships between taxa. In addition, we compared the morphology of the hypopygium to evaluate its use as a species-specific diagnostic character using the genetic divergence values. We recovered three well-supported monophyletic groups (intraclade divergence from 1.3 to 13.4%) and three hypopygium morphologies associated with each clade, two of them associated with D. rectilineus and D. concavatus (as described in the literature); the third one is new, not associated with any known species. The divergence between the D. rectilineus and D. concavatus clades was 19%, while the third clade is divergent from each species by 19–20%. If fully described, the hypopygium shape associated with the COI sequence will represent an extremely promising approach to the diagnosis of Dissomphalus species.

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          Most cited references 29

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          A new versatile primer set targeting a short fragment of the mitochondrial COI region for metabarcoding metazoan diversity: application for characterizing coral reef fish gut contents

          Introduction The PCR-based analysis of homologous genes has become one of the most powerful approaches for species detection and identification, particularly with the recent availability of Next Generation Sequencing platforms (NGS) making it possible to identify species composition from a broad range of environmental samples. Identifying species from these samples relies on the ability to match sequences with reference barcodes for taxonomic identification. Unfortunately, most studies of environmental samples have targeted ribosomal markers, despite the fact that the mitochondrial Cytochrome c Oxidase subunit I gene (COI) is by far the most widely available sequence region in public reference libraries. This is largely because the available versatile (“universal”) COI primers target the 658 barcoding region, whose size is considered too large for many NGS applications. Moreover, traditional barcoding primers are known to be poorly conserved across some taxonomic groups. Results We first design a new PCR primer within the highly variable mitochondrial COI region, the “mlCOIintF” primer. We then show that this newly designed forward primer combined with the “jgHCO2198” reverse primer to target a 313 bp fragment performs well across metazoan diversity, with higher success rates than versatile primer sets traditionally used for DNA barcoding (i.e. LCO1490/HCO2198). Finally, we demonstrate how the shorter COI fragment coupled with an efficient bioinformatics pipeline can be used to characterize species diversity from environmental samples by pyrosequencing. We examine the gut contents of three species of planktivorous and benthivorous coral reef fish (family: Apogonidae and Holocentridae). After the removal of dubious COI sequences, we obtained a total of 334 prey Operational Taxonomic Units (OTUs) belonging to 14 phyla from 16 fish guts. Of these, 52.5% matched a reference barcode (>98% sequence similarity) and an additional 32% could be assigned to a higher taxonomic level using Bayesian assignment. Conclusions The molecular analysis of gut contents targeting the 313 COI fragment using the newly designed mlCOIintF primer in combination with the jgHCO2198 primer offers enormous promise for metazoan metabarcoding studies. We believe that this primer set will be a valuable asset for a range of applications from large-scale biodiversity assessments to food web studies.
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            PATRISTIC: a program for calculating patristic distances and graphically comparing the components of genetic change

            Background Phylogenies are commonly used to analyse the differences between genes, genomes and species. Patristic distances calculated from tree branch lengths describe the amount of genetic change represented by a tree and are commonly compared with other measures of mutation to investigate the substitutional processes or the goodness of fit of a tree to the raw data. Up until now no universal tool has been available for calculating patristic distances and correlating them with other genetic distance measures. Results PATRISTICv1.0 is a java program that calculates patristic distances from large trees in a range of file formats and allows graphical and statistical interpretation of distance matrices calculated by other programs. Conclusion The software overcomes some logistic barriers to analysing signals in sequences. In additional to calculating patristic distances, it provides plots for any combination of matrices, calculates commonly used statistics, allows data such as isolation dates to be entered and reorders matrices with matching species or gene labels. It will be used to analyse rates of mutation and substitutional saturation and the evolution of viruses. It is available at and requires the Java runtime environment.
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              Colour Patterns Do Not Diagnose Species: Quantitative Evaluation of a DNA Barcoded Cryptic Bumblebee Complex

              Cryptic diversity within bumblebees (Bombus) has the potential to undermine crucial conservation efforts designed to reverse the observed decline in many bumblebee species worldwide. Central to such efforts is the ability to correctly recognise and diagnose species. The B. lucorum complex (Bombus lucorum, B. cryptarum and B. magnus) comprises one of the most abundant and important group of wild plant and crop pollinators in northern Europe. Although the workers of these species are notoriously difficult to diagnose morphologically, it has been claimed that queens are readily diagnosable from morphological characters. Here we assess the value of colour-pattern characters in species identification of DNA-barcoded queens from the B. lucorum complex. Three distinct molecular operational taxonomic units were identified each representing one species. However, no uniquely diagnostic colour-pattern character state was found for any of these three molecular units and most colour-pattern characters showed continuous variation among the units. All characters previously deemed to be unique and diagnostic for one species were displayed by specimens molecularly identified as a different species. These results presented here raise questions on the reliability of species determinations in previous studies and highlights the benefits of implementing DNA barcoding prior to ecological, taxonomic and conservation studies of these important key pollinators.
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                Author and article information

                Journal
                Zookeys
                Zookeys
                2
                urn:lsid:arphahub.com:pub:45048D35-BB1D-5CE8-9668-537E44BD4C7E
                urn:lsid:zoobank.org:pub:91BD42D4-90F1-4B45-9350-EEF175B1727A
                ZooKeys
                Pensoft Publishers
                1313-2989
                1313-2970
                2020
                14 August 2020
                : 959
                : 87-97
                Affiliations
                [1 ] Departamento de Ciências Biológicas, Centro de Ciências Humanas e Naturais, Universidade Federal do Espírito Santo, 29.075-910, Vitória, Espírito Santo, Brazil
                Author notes
                Corresponding author: Valéria Fagundes ( valeria.fagundes@ 123456ufes.br )

                Academic editor: Andreas Köhler

                Article
                53737
                10.3897/zookeys.959.53737
                7442767
                Marina Monjardim, Celso O. Azevedo, Valeria Fagundes

                This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                Funding
                Funded by: Fundação Estadual de Amparo à Pesquisa do Estado do Espírito Santo 501100006182 http://doi.org/10.13039/501100006182
                Categories
                Research Article
                Animalia
                Bethylidae
                Hymenoptera
                Insecta
                Molecular Systematics
                Phylogeny
                Taxonomy
                Brazil
                Paraguay
                South America

                Animal science & Zoology

                wasps, new species, molecular systematics, genetic diversity, coi

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