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      Molecular Techniques for Dicistrovirus Detection without RNA Extraction or Purification

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          Abstract

          Dicistroviridae is a new family of small, nonenveloped, and +ssRNA viruses pathogenic to both beneficial arthropods and insect pests as well. Triatoma virus (TrV), a dicistrovirus, is a pathogen of Triatoma infestans (Hemiptera: Reduviidae), one of the main vectors of Chagas disease. In this work, we report a single-step method to identify TrV, a dicistrovirus, isolated from fecal samples of triatomines. The identification method proved to be quite sensitive, even without the extraction and purification of RNA virus.

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          Most cited references10

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          Capsid protein identification and analysis of mature Triatoma virus (TrV) virions and naturally occurring empty particles.

          Triatoma virus (TrV) is a non-enveloped +ssRNA virus belonging to the insect virus family Dicistroviridae. Mass spectrometry (MS) and gel electrophoresis were used to detect the previously elusive capsid protein VP4. Its cleavage sites were established by sequencing the N-terminus of the protein precursor and MS, and its stoichiometry with respect to the other major capsid proteins (VP1-3) was found to be 1:1. We also characterized the polypeptides comprising the naturally occurring non-infectious empty capsids, i.e., RNA-free TrV particles. The empty particles were composed of VP0-VP3 plus at least seven additional polypeptides, which were identified as products of the capsid precursor polyprotein. We conclude that VP4 protein appears as a product of RNA encapsidation, and that defective processing of capsid proteins precludes genome encapsidation. Copyright © 2010 Elsevier Inc. All rights reserved.
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            Triatoma virus pathogenicity in laboratory colonies of Triatoma infestans (Hemiptera:Reduviidae).

            In a survey of wild populations of Triatoma infestans (Klug) in Argentina, 10% were infected with Triatoma virus (TrV). The virus also was detected in a laboratory colony 18 mo after being established, with infection rates up to 100%. Mortality rate was 97.6% in nymphs and the molting process was inhibited, thereby increasing development time. Because the virus was detected in colony nymphs. TrV may be transmitted vertically. However, the higher infection rate in the colony compared with natural populations also indicates other route(s) of transmission.
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              Characterization of Triatoma virus, a picorna-like virus isolated from the triatomine bug Triatoma infestans.

              Some properties of Triatoma virus (TrV), a picorna-like virus recently isolated from Triatoma infestans, have been studied. Electron microscopic observations of purified viral preparations showed the presence of non-enveloped viral particles 30 nm in diameter. The sedimentation coefficient of virus particles was about 165S and the buoyant density in CsCl was 1.39 g/ml. The viral genome was composed of one single-stranded RNA molecule with an Mr of 3 x 10(6). Three major polypeptides with Mr values of 39K, 37K and 33K and a minor one of about 45K were found in the virus particle. TrV particles contain about 35% RNA and 65% protein by weight. These data support the classification of this virus in the family Picornaviridae.
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                Author and article information

                Journal
                Biomed Res Int
                Biomed Res Int
                BMRI
                BioMed Research International
                Hindawi Publishing Corporation
                2314-6133
                2314-6141
                2013
                4 April 2013
                : 2013
                : 218593
                Affiliations
                1Centre for Malaria and Tropical Diseases, Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Rua da Junqueira, 100, 1349-008 Lisboa, Portugal
                2Unidad de Biofísica (UBF), CSIC-UPV/EHU, Barrio Sarriena S/N, Bizkaia, 48940 Leioa, Spain
                3Centro de Estudios Parasitológicos y de Vectores (CEPAVE), CCT-La Plata/CONICET-UNLP, No. 584, 1900 La Plata, Argentina
                Author notes
                *Marcelo Sousa Silva: mssilva@ 123456ihmt.unl.pt

                Academic Editor: Zhirong Sun

                Article
                10.1155/2013/218593
                3654624
                23710438
                348500a2-47b4-404a-b724-cf43e5a29c43
                Copyright © 2013 Jailson F. B. Querido et al.

                This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 10 January 2013
                : 15 March 2013
                : 21 March 2013
                Categories
                Methodology Report

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