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      Comparative transcriptome analyses reveal the genetic basis underlying the immune function of three amphibians’ skin

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          Abstract

          Skin as the first barrier against external invasions plays an essential role for the survival of amphibians on land. Understanding the genetic basis of skin function is significant in revealing the mechanisms underlying immunity of amphibians. In this study, we de novo sequenced and comparatively analyzed skin transcriptomes from three different amphibian species, Andrias davidianus, Bufo gargarizans, and Rana nigromaculata Hallowell. Functional classification of unigenes in each amphibian showed high accordance, with the most represented GO terms and KEGG pathways related to basic biological processes, such as binding and metabolism and immune system. As for the unigenes, GO and KEGG distributions of conserved orthologs in each species were similar, with the predominantly enriched pathways including RNA polymerase, nucleotide metabolism, and defense. The positively selected orthologs in each amphibian were also similar, which were primarily involved in stimulus response, cell metabolic, membrane, and catalytic activity. Furthermore, a total of 50 antimicrobial peptides from 26 different categories were identified in the three amphibians, and one of these showed high efficiency in inhibiting the growth of different bacteria. Our understanding of innate immune function of amphibian skin has increased basis on the immune-related unigenes, pathways, and antimicrobial peptides in amphibians.

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          Most cited references22

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          The genome of the Western clawed frog Xenopus tropicalis.

          The western clawed frog Xenopus tropicalis is an important model for vertebrate development that combines experimental advantages of the African clawed frog Xenopus laevis with more tractable genetics. Here we present a draft genome sequence assembly of X. tropicalis. This genome encodes more than 20,000 protein-coding genes, including orthologs of at least 1700 human disease genes. Over 1 million expressed sequence tags validated the annotation. More than one-third of the genome consists of transposable elements, with unusually prevalent DNA transposons. Like that of other tetrapods, the genome of X. tropicalis contains gene deserts enriched for conserved noncoding elements. The genome exhibits substantial shared synteny with human and chicken over major parts of large chromosomes, broken by lineage-specific chromosome fusions and fissions, mainly in the mammalian lineage.
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            The natural history of amphibian skin secretions, their normal functioning and potential medical applications.

            Amphibians occupy a wide range of habitat types from arid deserts to deep freshwater lakes; they may spend most of their life underground or high in cloud forest canopy. Some are found north of the Arctic Circle and can tolerate freezing conditions, while others have evolved a range of adaptations to avoid desiccation in some of the hotter areas of the world. The skin plays key roles in the everyday survival of amphibians and their ability to exploit a wide range of habitats and ecological conditions. The normal functions of the skin are surveyed and Eisner's biorational approach to chemical prospecting--seeking clues from an animal's behaviour and its interactions with its environment to reveal the presence of chemical compounds with potential medical or veterinary applications--is applied to amphibians. The biology and natural history of amphibian skin, its glands and their secretions are briefly reviewed. Four categories of compounds are found in the granular or poison glands, these are: biogenic amines, bufodienolides (bufogenins), alkaloids and steroids, peptides and proteins. Toads, particularly members of the genus Bufo, are identified as a particularly convenient and useful source of granular gland secretions. The potential medical-pharmaceutical significance of products derived from amphibian skin secretions is discussed. The need for a humane approach to this work is noted.
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              Benchmarking next-generation transcriptome sequencing for functional and evolutionary genomics.

              Next-generation sequencing has opened the door to genomic analysis of nonmodel organisms. Technologies generating long-sequence reads (200-400 bp) are increasingly used in evolutionary studies of nonmodel organisms, but the short-sequence reads (30-50 bp) that can be produced at lower cost are thought to be of limited utility for de novo sequencing applications. Here, we tested this assumption by short-read sequencing the transcriptomes of the tropical disease vectors Aedes aegypti and Anopheles gambiae, for which complete genome sequences are available. Comparison of our results to the reference genomes allowed us to accurately evaluate the quantity, quality, and functional and evolutionary information content of our "test" data. We produced more than 0.7 billion nucleotides of sequenced data per species that assembled into more than 21,000 test contigs larger than 100 bp per species and covered approximately 27% of the Aedes reference transcriptome. Remarkably, the substitution error rate in the test contigs was approximately 0.25% per site, with very few indels or assembly errors. Test contigs of both species were enriched for genes involved in energy production and protein synthesis and underrepresented in genes involved in transcription and differentiation. Ortholog prediction using the test contigs was accurate across hundreds of millions of years of evolution. Our results demonstrate the considerable utility of short-read transcriptome sequencing for genomic studies of nonmodel organisms and suggest an approach for assessing the information content of next-generation data for evolutionary studies.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data curationRole: Funding acquisitionRole: Writing – original draft
                Role: ConceptualizationRole: Data curationRole: Funding acquisitionRole: Writing – original draft
                Role: Funding acquisitionRole: Project administration
                Role: Formal analysis
                Role: Formal analysisRole: Investigation
                Role: ConceptualizationRole: SupervisionRole: Writing – review & editing
                Role: Investigation
                Role: Investigation
                Role: Funding acquisitionRole: ResourcesRole: Writing – review & editing
                Role: Funding acquisitionRole: ResourcesRole: SupervisionRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                21 December 2017
                2017
                : 12
                : 12
                : e0190023
                Affiliations
                [1 ] Chongqing Research Center of Conservation and Development on Rare and Endangered Aquatic Resources, Chongqing University of Arts and Sciences, Yongchuan, Chongqing, China
                [2 ] Chongqing Key Laboratory of Kinase Modulators as Innovative Medicine, Yongchuan, Chongqing, China
                [3 ] Chongqing Engineering Laboratory of Targeted and Innovative Therapeutics, Yongchuan, Chongqing, China
                The National Orchid Conservation Center of China; The Orchid Conservation & Research Center of Shenzhen, CHINA
                Author notes

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

                Author information
                http://orcid.org/0000-0003-2397-5081
                Article
                PONE-D-17-09798
                10.1371/journal.pone.0190023
                5739465
                29267366
                7f2f0fa7-90f7-4fdc-9304-9ded70c76d37
                © 2017 Fan et al

                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
                : 12 March 2017
                : 6 December 2017
                Page count
                Figures: 4, Tables: 4, Pages: 14
                Funding
                Funded by: Natural Science Foundation of Chongqing Municipal Science and Technology Commission
                Award ID: cstc2015jcyjBX0013
                Funded by: Natural Science Foundation of Chongqing Municipal Science and Technology Commission
                Award ID: cstc2014jcyjA80035
                Funded by: Science and Technology Research Project of Chongqing Municipal Education Commission
                Award ID: KJ1601109
                Funded by: Science and Technology Research Project of Chongqing Municipal Education Commission
                Award ID: KJ1601107
                Funded by: Natural Science Foundation of Chongqing Yongchuan Municipal Science and Technology Commission
                Award ID: Ycstc, 2015nc1001
                Funded by: Foundation for High-level Talents of Chongqing University of Art and Sciences
                Award ID: R2014LX07
                This work was supported by the Natural Science Foundation of Chongqing Municipal Science and Technology Commission (Grant No. cstc2015jcyjBX0013, cstc2014jcyjA80035), http://www.cstc.gov.cn/, Chongqing Science and Technology Commission; the Science and Technology Research Project of Chongqing Municipal Education Commission (Grant No. KJ1601109, KJ1601107), http://www.cstc.gov.cn/, Chongqing Science and Technology Commission; the Natural Science Foundation of Chongqing Yongchuan Municipal Science and Technology Commission (Grant No. Ycstc, 2015nc1001), http://www.cstc.gov.cn/, Chongqing Science and Technology Commission; and the Foundation for High-level Talents of Chongqing University of Art and Sciences (No.R2014LX07), http://edf.cqu.edu.cn/, Chongqing University Education Foundation.
                Categories
                Research Article
                Biology and Life Sciences
                Organisms
                Eukaryota
                Animals
                Vertebrates
                Amphibians
                Biology and Life Sciences
                Computational Biology
                Genome Analysis
                Transcriptome Analysis
                Biology and Life Sciences
                Genetics
                Genomics
                Genome Analysis
                Transcriptome Analysis
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Immunology
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                Immunology
                Immune Response
                Medicine and Health Sciences
                Immunology
                Immune Response
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                Genetics
                Genomics
                Animal Genomics
                Amphibian Genomics
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
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                Taxonomy
                Evolutionary Systematics
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                Computer and Information Sciences
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                Taxonomy
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                Biology and Life Sciences
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