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      Detection of avian metapneumovirus subtype A from wild birds in the State of São Paulo, Brazil Translated title: Detecção de metapneumovirus aviário subtipo A em aves silvestres no estado de São Paulo, Brasil

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          Abstract

          ABSTRACT: The present study investigated the circulation of avian metapneumovirus (aMPV) in wild birds in Brazil. To do so, 131 samples from 366 oropharyngeal or cloacal swabs collected from 18 species of birds were tested individually or in pools by RT-PCR. Samples detected by RT-PCR were selected for DNA sequencing. Thirteen (9.9%) samples were detected by the RT-PCR targeting the N gene and four out of 13 samples were sequenced. Sequencing results showed a high identity with the aMPV subtype A. Our results confirm the circulation of the aMPV subtype A in wild birds in Brazil even five years after its last detection.

          Translated abstract

          RESUMO: O presente estudo investigou a circulação de metapneumovírus aviário em aves silvestres no Brasil. Para tanto, 131 amostras de 366 suabes orofaringeanos ou cloacais coletados de 18 espécies de aves foram testadas individualmente ou na forma de pools por RT-PCR. As amostras detectadas por RT-PCR foram selecionadas para sequenciamento. Treze (9,9%) das amostras foram detectadas por RT-PCR tendo o gene N como alvo; destas, quatro foram sequenciadas com sucesso. Resultados do sequenciamento mostraram alta identidade com o aMPV de subtipo A. Nossos resultados confirmam a circulação de aMPV subtipo A em aves silvestres no Brasil mesmo cinco anos após sua última detecção.

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            Nucleotide sequences of the F, L and G protein genes of two non-A/non-B avian pneumoviruses (APV) reveal a novel APV subgroup.

            Sequence analysis was performed of all or part of the genes encoding the fusion (F), polymerase (L) and attachment (G) proteins of two French non-A/non-B avian pneumovirus (APV) isolates (Fr/85/1 and Fr/85/2). The two isolates shared at least 99.7% nt and 99.0% aa sequence identity. Comparison with the F genes from subgroup A, subgroup B or Colorado APVs revealed nt and aa identities of 70.0-80. 5% and 77.6-97.2%, respectively, with the L gene sharing 76.1% nt and 85.3% aa identity with that of a subgroup A isolate. The Fr/85/1 and Fr/85/2 G genes comprised 1185 nt, encoding a protein of 389 aa. Common features with subgroup A and subgroup B G proteins included an amino-terminal membrane anchor, a high serine and threonine content, conservation of cysteine residues and a single extracellular region of highly conserved sequence proposed to be the functional domain involved in virus attachment to cellular receptors. However, the Fr/85/1 and Fr/85/2 G sequences shared at best 56.6% nt and 31.2% aa identity with subgroup A and B APVs, whereas these isolates share 38% aa identity. Phylogenetic analysis of the F, G and L genes of pneumoviruses suggested that isolates Fr/85/1 and Fr/85/2 belong to a previously unrecognized APV subgroup, tentatively named D. G-based oligonucleotide primers were defined for the specific molecular identification of subgroup D. These are the first G protein sequences of non-A/non-B APVs to be determined.
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              A Family-Wide RT-PCR Assay for Detection of Paramyxoviruses and Application to a Large-Scale Surveillance Study

              Family-wide molecular diagnostic assays are valuable tools for initial identification of viruses during outbreaks and to limit costs of surveillance studies. Recent discoveries of paramyxoviruses have called for such assay that is able to detect all known and unknown paramyxoviruses in one round of PCR amplification. We have developed a RT-PCR assay consisting of a single degenerate primer set, able to detect all members of the Paramyxoviridae family including all virus genera within the subfamilies Paramyxovirinae and Pneumovirinae. Primers anneal to domain III of the polymerase gene, with the 3′ end of the reverse primer annealing to the conserved motif GDNQ, which is proposed to be the active site for nucleotide polymerization. The assay was fully optimized and was shown to indeed detect all available paramyxoviruses tested. Clinical specimens from hospitalized patients that tested positive for known paramyxoviruses in conventional assays were also detected with the novel family-wide test. A high-throughput fluorescence-based RT-PCR version of the assay was developed for screening large numbers of specimens. A large number of samples collected from wild birds was tested, resulting in the detection of avian paramyxoviruses type 1 in both barnacle and white-fronted geese, and type 8 in barnacle geese. Avian metapneumovirus type C was found for the first time in Europe in mallards, greylag geese and common gulls. The single round family-wide RT-PCR assay described here is a useful tool for the detection of known and unknown paramyxoviruses, and screening of large sample collections from humans and animals.
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                Author and article information

                Contributors
                Role: ND
                Role: ND
                Role: ND
                Role: ND
                Role: ND
                Role: ND
                Role: ND
                Role: ND
                Journal
                pvb
                Pesquisa Veterinária Brasileira
                Pesq. Vet. Bras.
                Colégio Brasileiro de Patologia Animal - CBPA. Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA) (Rio de Janeiro, RJ, Brazil )
                0100-736X
                1678-5150
                March 2019
                : 39
                : 3
                : 209-213
                Affiliations
                [4] Campinas SP orgnameUniversidade de Campinas orgdiv1Instituto de Biologia orgdiv2Laboratório de Virologia Animal Brazil
                [3] Pirassununga orgnameUniversidade de São Paulo orgdiv1Faculdade de Zootecnia e Engenharia de Alimentos orgdiv2Departamento de Medicina Veterinária Brazil
                [2] São Paulo São Paulo orgnameUniversidade de São Paulo orgdiv1Faculdade de Medicina Veterinária e Ciência Animal orgdiv2Postgraduate Program in Experimental Epidemiology of Zoonoses Brazil
                Article
                S0100-736X2019000300209
                10.1590/1678-5150-pvb-6021
                2f7f9a09-e615-4ce6-86f5-ad67a6141044

                This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

                History
                : 21 September 2018
                : 09 November 2018
                Page count
                Figures: 0, Tables: 0, Equations: 0, References: 33, Pages: 5
                Product

                SciELO Brazil

                Categories
                Wildlife Medicine

                surveillance,aMPV,wildlife animals,birds,viroses,Detecção,metapneumovirus aviário,subtipo A,aves silvestres,São Paulo,Brasil,aves aquáticas,epidemiologia,animais silvestres,Detection,avian metapneumovirus,subtype A,wild birds,Brazil,waterfowl

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