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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 references 10

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          MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods.

          Comparative analysis of molecular sequence data is essential for reconstructing the evolutionary histories of species and inferring the nature and extent of selective forces shaping the evolution of genes and species. Here, we announce the release of Molecular Evolutionary Genetics Analysis version 5 (MEGA5), which is a user-friendly software for mining online databases, building sequence alignments and phylogenetic trees, and using methods of evolutionary bioinformatics in basic biology, biomedicine, and evolution. The newest addition in MEGA5 is a collection of maximum likelihood (ML) analyses for inferring evolutionary trees, selecting best-fit substitution models (nucleotide or amino acid), inferring ancestral states and sequences (along with probabilities), and estimating evolutionary rates site-by-site. In computer simulation analyses, ML tree inference algorithms in MEGA5 compared favorably with other software packages in terms of computational efficiency and the accuracy of the estimates of phylogenetic trees, substitution parameters, and rate variation among sites. The MEGA user interface has now been enhanced to be activity driven to make it easier for the use of both beginners and experienced scientists. This version of MEGA is intended for the Windows platform, and it has been configured for effective use on Mac OS X and Linux desktops. It is available free of charge from http://www.megasoftware.net.
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            DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates.

            We describe "universal" DNA primers for polymerase chain reaction (PCR) amplification of a 710-bp fragment of the mitochondrial cytochrome c oxidase subunit I gene (COI) from 11 invertebrate phyla: Echinodermata, Mollusca, Annelida, Pogonophora, Arthropoda, Nemertinea, Echiura, Sipuncula, Platyhelminthes, Tardigrada, and Coelenterata, as well as the putative phylum Vestimentifera. Preliminary comparisons revealed that these COI primers generate informative sequences for phylogenetic analyses at the species and higher taxonomic levels.
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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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                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
                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

                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.

                Counts
                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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