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      A Formal Re-Description of the Cockroach Hebardina concinna Anchored on DNA Barcodes Confirms Wing Polymorphism and Identifies Morphological Characters for Field Identification

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          Abstract

          Background

          Hebardina concinna is a domestic pest and potential vector of pathogens throughout East and Southeast Asia, yet identification of this species has been difficult due to a lack of diagnostic morphological characters, and to uncertainty in the relationship between macroptyrous (long-winged) and brachypterous (small-winged) morphotypes. In insects male genital structures are typically species-specific and are frequently used to identify species. However, male genital structures in H. concinna had not previously been described, in part due to difficulty in identifying conspecifics.

          Methods/Principal Findings

          We collected 15 putative H. concinna individuals, from Chinese populations, of both wing morphotypes and both sexes and then generated mitochondrial COI (the standard barcode region) and COII sequences from five of these individuals. These confirmed that both morphotypes of both sexes are the same species. We then dissected male genitalia and compared genital structures from macropterous and brachypterous individuals, which we showed to be identical, and present here for the first time a detailed description of H. concinna male genital structures. We also present a complete re-description of the morphological characters of this species, including both wing morphs.

          Conclusions/Significance

          This work describes a practical application of DNA barcoding to confirm that putatively polymorphic insects are conspecific and then to identify species-specific characters that can be used in the field to identify individuals and to obviate the delay and cost of returning samples to a laboratory for DNA sequencing.

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          Most cited references8

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          DNA Barcoding the Geometrid Fauna of Bavaria (Lepidoptera): Successes, Surprises, and Questions

          Background The State of Bavaria is involved in a research program that will lead to the construction of a DNA barcode library for all animal species within its territorial boundaries. The present study provides a comprehensive DNA barcode library for the Geometridae, one of the most diverse of insect families. Methodology/Principal Findings This study reports DNA barcodes for 400 Bavarian geometrid species, 98 per cent of the known fauna, and approximately one per cent of all Bavarian animal species. Although 98.5% of these species possess diagnostic barcode sequences in Bavaria, records from neighbouring countries suggest that species-level resolution may be compromised in up to 3.5% of cases. All taxa which apparently share barcodes are discussed in detail. One case of modest divergence (1.4%) revealed a species overlooked by the current taxonomic system: Eupithecia goossensiata Mabille, 1869 stat.n. is raised from synonymy with Eupithecia absinthiata (Clerck, 1759) to species rank. Deep intraspecific sequence divergences (>2%) were detected in 20 traditionally recognized species. Conclusions/Significance The study emphasizes the effectiveness of DNA barcoding as a tool for monitoring biodiversity. Open access is provided to a data set that includes records for 1,395 geometrid specimens (331 species) from Bavaria, with 69 additional species from neighbouring regions. Taxa with deep intraspecific sequence divergences are undergoing more detailed analysis to ascertain if they represent cases of cryptic diversity.
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            Design and applicability of DNA arrays and DNA barcodes in biodiversity monitoring

            Background The rapid and accurate identification of species is a critical component of large-scale biodiversity monitoring programs. DNA arrays (micro and macro) and DNA barcodes are two molecular approaches that have recently garnered much attention. Here, we compare these two platforms for identification of an important group, the mammals. Results Our analyses, based on the two commonly used mitochondrial genes cytochrome c oxidase I (the standard DNA barcode for animal species) and cytochrome b (a common species-level marker), suggest that both arrays and barcodes are capable of discriminating mammalian species with high accuracy. We used three different datasets of mammalian species, comprising different sampling strategies. For DNA arrays we designed three probes for each species to address intraspecific variation. As for DNA barcoding, our analyses show that both cytochrome c oxidase I and cytochrome b genes, and even smaller fragments of them (mini-barcodes) can successfully discriminate species in a wide variety of specimens. Conclusion This study showed that DNA arrays and DNA barcodes are valuable molecular methods for biodiversity monitoring programs. Both approaches were capable of discriminating among mammalian species in our test assemblages. However, because designing DNA arrays require advance knowledge of target sequences, the use of this approach could be limited in large scale monitoring programs where unknown haplotypes might be encountered. DNA barcodes, by contrast, are sequencing-based and therefore could provide more flexibility in large-scale studies.
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              An evaluation of sampling effects on multiple DNA barcoding methods leads to an integrative approach for delimiting species: a case study of the North American tarantula genus Aphonopelma (Araneae, Mygalomorphae, Theraphosidae).

              The North American tarantula genus Aphonopelma provides one of the greatest challenges to species delimitation and downstream identification in spiders because traditional morphological characters appear ineffective for evaluating limits of intra- and interspecific variation in the group. We evaluated the efficacy of numerous molecular-based approaches to species delimitation within Aphonopelma based upon the most extensive sampling of theraphosids to date, while also investigating the sensitivity of randomized taxon sampling on the reproducibility of species boundaries. Mitochondrial DNA (cytochrome c oxidase subunit I) sequences were sampled from 682 specimens spanning the genetic, taxonomic, and geographic breadth of the genus within the United States. The effects of random taxon sampling compared traditional Neighbor-Joining with three modern quantitative species delimitation approaches (ABGD, P ID(Liberal), and GMYC). Our findings reveal remarkable consistency and congruence across various approaches and sampling regimes, while highlighting highly divergent outcomes in GMYC. Our investigation allowed us to integrate methodologies into an efficient, consistent, and more effective general methodological workflow for estimating species boundaries within the mygalomorph spider genus Aphonopelma. Taken alone, these approaches are not particularly useful - especially in the absence of prior knowledge of the focal taxa. Only through the incorporation of multiple lines of evidence, employed in a hypothesis-testing framework, can the identification and delimitation of confident species boundaries be determined. A key point in studying closely related species, and perhaps one of the most important aspects of DNA barcoding, is to combine a sampling strategy that broadly identifies the extent of genetic diversity across the distributions of the species of interest and incorporates previous knowledge into the "species equation" (morphology, molecules, and natural history). Copyright © 2013 Elsevier Inc. All rights reserved.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2014
                18 September 2014
                : 9
                : 9
                : e106789
                Affiliations
                [1 ]Zhongshan Entry-Exit Inspection and Quarantine Bureau Technology Center, Zhongshan, Guangdong, China
                [2 ]Guangdong Entry-Exit Inspection and Quarantine Bureau Technology Center, Guangzhou, Guangdong, China
                [3 ]The EMBL-European Bioinformatics Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, United Kingdom
                Institute of Zoology, Chinese Academy of Sciences, China
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: QY DQ. Performed the experiments: QY KW DQ JH DL XW JC. Analyzed the data: QY KW DQ JH JC CEC. Contributed reagents/materials/analysis tools: CEC. Contributed to the writing of the manuscript: QY KW CEC.

                Article
                PONE-D-14-14512
                10.1371/journal.pone.0106789
                4169431
                25232993
                e7e9b7a5-2188-41f8-880e-bdbf110d5602
                Copyright @ 2014

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

                History
                : 2 April 2014
                : 2 August 2014
                Page count
                Pages: 12
                Funding
                This work was financially supported by the National Science and Technology support program “2012BAK11B05 ( http://program.most.gov.cn/)”, AQSIQ (Administration of Quality Supervision, Inspection and Quarantine) support program “2012IK223 ( http://zjkygl.aqsiq.gov.cn)”, GDCIQ (Guangdong Entry-Exit Inspection and Quarantine Bureau) support program “2013GDK39 ( http://www.gdciq.gov.cn)” and “2013GDK36 ( http://www.gdciq.gov.cn)” and Zhongshan City support program “20123A298 ( http://www.zskj.gov.cn/).” The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and life sciences
                Evolutionary biology
                Evolutionary systematics
                Molecular systematics
                DNA barcoding
                Organisms
                Animals
                Invertebrates
                Arthropoda
                Insects
                Cockroaches
                Taxonomy
                Custom metadata
                The authors confirm that all data underlying the findings are fully available without restriction. All the new amplified sequences for Hebardina concinna and other species are available from Genbank ( http://www.ncbi.nlm.nih.gov/nuccore/) (KF640073, KF640074, KF640076, KF640077, KF640075, KF876003, KF876004, KF876006, KF876007, KF876005, KF640067, KF640069, KF640071, KF640072, KF640066, KF876000, KF640067, KF876001, KF640068, KF876002). Data are also available from the Zhongshan Entry-Exit Inspection and Quarantine Bureau study whose authors may be contacted at 2 Zhongshan 6 Road, Zhongshan 528403, Guangdong, China, or by email yueqy@ 123456zs.gdciq.gov.cn .

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