Detection of avian reovirus (ARV) by ELISA based on recombinant σB, σC and σNS full-length proteins and protein fragments.

Introduction. Avian reovirus (ARV) is associated with arthritis/tenosynovitis and malabsorption syndrome in chickens. The σC and σB proteins, both exposed to the virus capsid, are highly immunogenic and could form the basis for diagnostic devices designed to assess the immunological status of the flock.Gap Statement. Commercial ARV ELISAs cannot distinguish between vaccinated and infected animals and might not detect circulating ARV strains.Aim. We aimed to develop a customized test to detect the circulating field ARV strains as well as distinguish between vaccinated and unvaccinated animals.Methodology. We developed ELISA assays based on recombinant (r) σB, σC and the nonstructural protein σNS and tested them using antisera of vaccinated and unvaccinated chickens as well as negative controls. Fragments of σB and σC proteins were also used to study regions that could be further exploited in diagnostic tests.Results. Vaccinated and unvaccinated birds were positive by commercial ELISA, with no difference in optical density values. In contrast, samples of unvaccinated animals showed lower absorbance in the rσB and rσC ELISA tests and higher absorbance in the rσNS ELISA test than the vaccinated animals. Negative control samples were negative in all tests. Fragmentation of σB and σC proteins showed that some regions can differentiate between vaccinated and unvaccinated animals. For example, σB amino acids 128-179 (σB-F4) and σC amino acids 121-165 (σC-F4) exhibited 85 and 95% positivity among samples of vaccinated animals but only 5% and zero positivity among samples of unvaccinated animals, respectively.Conclusion. These data suggest that unvaccinated birds might have been exposed to field strains of ARV. The reduction in absorbance in the recombinant tests possibly reflects an increased specificity of our test since unvaccinated samples showed less cross-reactivity with the vaccine proteins immobilized on ELISAs. The discrepant results obtained with the protein fragment tests between vaccinated and unvaccinated animals are discussed in light of the diversity between ARV strains.

A Fijivirus Major Viroplasm Protein Shows RNA-Stimulated ATPase Activity by Adopting Pentameric and Hexameric Assemblies of Dimers.

Fijiviruses replicate and package their genomes within viroplasms in a process involving RNA-RNA and RNA-protein interactions. Here, we demonstrate that the 24 C-terminal residues (C-arm) of the P9-1 major viroplasm protein of the mal de Río Cuarto virus (MRCV) are required for its multimerization and the formation of viroplasm-like structures. Using an integrative structural approach, the C-arm was found to be dispensable for P9-1 dimer assembly but essential for the formation of pentamers and hexamers of dimers (decamers and dodecamers), which favored RNA binding. Although both P9-1 and P9-1ΔC-arm catalyzed ATP with similar activities, an RNA-stimulated ATPase activity was only detected in the full-length protein, indicating a C-arm-mediated interaction between the ATP catalytic site and the allosteric RNA binding sites in the (do)decameric assemblies. A stronger preference to bind phosphate moieties in the decamer was predicted, suggesting that the allosteric modulation of ATPase activity by RNA is favored in this structural conformation. Our work reveals the structural versatility of a fijivirus major viroplasm protein and provides clues to its mechanism of action. IMPORTANCE The mal de Río Cuarto virus (MRCV) causes an important maize disease in Argentina. MRCV replicates in several species of Gramineae plants and planthopper vectors. The viral factories, also called viroplasms, have been studied in detail in animal reovirids. This work reveals that a major viroplasm protein of MRCV forms previously unidentified structural arrangements and provides evidence that it may simultaneously adopt two distinct quaternary assemblies. Furthermore, our work uncovers an allosteric communication between the ATP and RNA binding sites that is favored in the multimeric arrangements. Our results contribute to the understanding of plant reovirids viroplasm structure and function and pave the way for the design of antiviral strategies for disease control.

High prevalence of CsRV2 in cultured Callinectes danae: Potential impacts on soft-shell crab production in Brazil.

Crabs can be infected by a variety of pathogenic micro-organisms but the most damaging are viruses. Naturally-occurring Callinectes sapidus reovirus 1 (CsRV1) is thought to contribute to mortality of Callinectes sapidus in soft crab culture in the USA. In Brazil, soft crabs are frequently produced using Callinectes danae, which suffers a similar rate of mortality in culture as C. sapidus. This study investigated whether CsRV1 could be detected in healthy or dead Callinectes danae from Paraná, Brazil and kept in captivity, we also evaluated the relationship between viral infection, and biochemical and behavioral parameters. C. danae from Paranaguá Bay were kept in a recirculation system for 14 days and subjected to weekly biochemical analyses and a reflex action mortality predictors (RAMP) test. RT-qPCR assays for CsRV1 were negative for all samples. However, electrophoretic analysis of extracted RNA from some crabs showed a pattern of 12 dsRNA bands that indicated intense infection by a reovirus with a genome organization different from CsRV1. The banding pattern was indistinguishable from a putative novel reovirus detected in C. sapidus in Rio Grande do Sul, Brazil, provisionally called CsRV2. The prevalence of dsRNA of CsRV2 showed no significant difference between crabs that died and survived. Interestingly, the presence of CsRV2 dsRNA was correlated with a significant reduction in glycogen concentration in hepatopancreas and a decrease in reflex action. The results obtained in this study are an early glimpse of the occurrence of reoviruses in C. danae and their potential effects in soft-shell crab systems in Brazil.

Evaluation of Avian Reovirus S1133 Vaccine Strain in Neonatal Broiler Chickens in Gastrointestinal Integrity and Performance in a Large-Scale Commercial Field Trial.

Avian reovirus (ARV) is the principal cause of several diseases. The vaccination of breeders allows for the control of viral arthritis and delivery of maternal-derived antibodies to the progeny. The vaccination of broiler chickens with ARV strain S1133 is used to prevent viral arthritis. However, the post-vaccination enteric effects have not been well-characterized. The purpose of this study was to evaluate the effect of vaccination with the S1133 strain on the weight gain and feed conversion of broiler chickens and to characterize the gastric, enteric, and pancreatic lesions that the strain could induce. A total of 672,000 chickens were divided into two groups: a group vaccinated with ARV strain S1133 (S1133ARV) and a control group (not vaccinated). Upon histological analysis, the vaccine group showed less proventricular glandular tissue and atrophy of the pancreas and duodenal villi, as well as having a lower average daily profit. The conclusion based on the results of this investigation is that neonatal vaccination with S1133ARV causes atrophy of the pancreatic acini, proventricular glands, and intestinal villi, leading to an increased diameter of the glandular lumen and atrophy of the enteric villous, as well as weight loss, in broiler chickens.

Emerging new avian reovirus variants from cases of enteric disorders and arthritis/tenosynovitis in Brazilian poultry flocks.

1. The objective of this study was to characterise circulating Brazilian avian reovirus (ARV) strains by genetic analysis of the σC protein encoded by segment 1 of the viral genome and compare these with those of viral strains used for immunising commercial poultry.2. The analysis detected the presence of ARV genomes by quantitative reverse transcriptase PCR (RT-qPCR) in the enteric samples and the joint tissues (JT) of birds with signs of viral arthritis/tenosynovitis. Nucleotide sequencing used 16 strains (three commercial vaccines, 10 from enteric tissues and three from JT). The results indicated high variability in the amino acid sequences of 13 wild strains, showing between 40% and 75% similarity compared with the vaccine strains (S1133 and 2177).3. The sequences were grouped into three well-defined clusters in a phylogenetic tree, two of these clusters together with previous Brazilian σC ARV sequences, and one cluster (VII) that was novel for Brazilian strains. Antigenic analysis showed that there were amino acids within putative epitopes located on the surface of the receptor-binding region of the σC protein with a high degree of variability.4. The study confirmed the presence of ARV genetic variants circulating in commercial birds in Brazil, and according to the antigenic prediction, the possibility of antigenic variants appears to be high.

Real-time RT-qPCR assay to detect sequences in the Piscine orthoreovirus-1 genome segment S1 associated with heart and skeletal muscle inflammation in Atlantic salmon.

During the last decade, Piscine orthoreovirus was identified as the main causative agent of heart and skeletal muscle inflammation (HSMI) in Atlantic Salmon, Norway. A recent study showed that PRV-1 sequences from salmonid collected in North Atlantic Pacific Coast (NAPC) grouped separately from the Norwegian sequences found in Atlantic Salmon diagnosed with HSMI. Currently, the routine assay used to screen for PRV-1 in NAPC water and worldwide cannot differentiate between the two groups of PRV-1. Therefore, this study aimed at developing a real-time polymerase chain reaction (RT-qPCR) assay to target the PRV-1 genome segments specific for variants associated with HSMI. The assay was optimized and tested against 71 tissue samples collected from different regions including Norway, Chile and both coast of Canada and different hosts farmed Atlantic Salmon, wild Coho Salmon and escaped Atlantic Salmon collected in British Columbia, West Coast of Canada. This assay has the potential to be used for screening salmonids and non-salmonids that may carry PRV-1 potentially causing HSMI.

Molecular detection of Mycoplasma synoviae and avian reovirus infection in arthritis and tenosynovitis lesions of broiler and breeder chickens in Santa Catarina State, Brazil.

Infectious arthritis or tenosynovitis in broiler and breeder chickens results in major loss of productivity because of reduced growth and downgrading at processing plants. The most common causative agents of avian infectious arthritis are the bacterium Mycoplasma synoviae and avian reoviruses (ARVs) (family Reoviridae, genus Orthoreovirus). In this study, we evaluated the occurrence of these two pathogens in arthritis or tenosynovitis lesions of broilers and breeder flocks in southern Brazil using molecular detection. Tissue sections from tibiotarsal joints with visible lesions from 719 broilers and 505 breeders were analysed using pathogen-specific polymerase chain reaction (PCR) assays. In breeders, 41.2% (n = 296) of lesions were positive for M. synoviae, 26.4% (n = 190) were positive for ARV, while co-infection was present in 12.2% (n = 88) of the samples. In broilers, 20.8% (n = 105) of lesions were positive for M. synoviae, 11.9% (n = 60) for ARV and 7.7% (n = 39) of these cases were positive for both pathogens. Post-mortem examination revealed lesions with varying degrees of gross pathological severity. Histopathological examination showed intense, diffuse lymphohistiocytic inflammatory infiltrates with heterophil accumulation, primarily in the synovial capsule and digital flexor tendon, in all samples. Improved strategies for early detection and control of these major avian pathogens are highly desirable for preventing the spread of infection and reducing economic losses in the poultry industry.

Detection of RNA-Dependent RNA Polymerase of Hubei Reo-Like Virus 7 by Next-Generation Sequencing in Aedes aegypti and Culex quinquefasciatus Mosquitoes from Brazil.

Advancements in next-generation sequencing and bioinformatics have expanded our knowledge of the diversity of viruses (pathogens and non-pathogens) harbored by mosquitoes. Hubei reo-like virus 7 (HRLV 7) was recently detected by the virome analysis of fecal samples from migratory birds in Australia. We now report the detection of RNA-dependent RNA polymerase sequences of HRLV 7 in pools of Aedes aegypti and Culex quinquefasciatus mosquitoes species from the Brazilian Amazon forest. Phylogenetic inferences indicated that all HRLV 7 strains fall within the same independent clade. In addition, HRLV 7 shared a close ancestral lineage with the Dinovernavirus genus of the Reoviridae family. Our findings indicate that HRLV 7 is present in two species of mosquitoes.

Reovirus: Friend and Foe.

PURPOSE OF REVIEW: Mammalian orthoreovirus (reovirus) is a powerful tool for studying viral replication and pathogenesis. Most reovirus infections are subclinical, however recent work has catapulted reovirus into the clinical spotlight. RECENT FINDINGS: Owing to its capacity to kill cancer cells more efficiently than normal cells, reovirus is under development as a therapeutic for a variety of cancers. New efforts have focused on genetically engineering reovirus to increase its oncolytic capacity, and determining how reovirus potentiates immunotherapy. Other recent studies highlight a potential role for reovirus in celiac disease (CeD). Using mouse models of CeD, reovirus caused loss of oral tolerance to dietary antigens, opening the possibility that reovirus could trigger CeD in humans. SUMMARY: We will focus on new developments in reovirus oncolysis and studies suggesting a role for reovirus as a trigger for celiac disease (CeD) that make reovirus a potential friend and foe to human health.

Distribution of IFITM3 in yellow-feathered broilers and inhibition of Avian Reovirus multiplication by IFITM3

This study was carried to express the interferon-induced transmembrane protein 3 (IFITM3) in vitro and examine its function in inhibition of avian reovirus (ARV) replication. The recombinant prokaryotic vector expressing yellow-feathered broiler IFITM3 was successfully constructed, and the recombinant protein was expressed in competent Escherichia coli BL21 cells. New Zealand white rabbits were immunized with the purified recombinant protein to prepare a polyclonal antibody, with a titer of 1:128,000. Immunohistochemistry, reverse transcription-PCR, and real-time fluorescence quantitative PCR showed that IFITM3 was distributed in the yellow-feathered broiler immune organs, and the expression of IFITM3 in bursa of Fabricius was more than in spleen and thymus. It was found that in the thymus, spleen and bursa of Fabricius the mRNA expression levels of IFN and IFITM3 were significantly induced after ARV infection. And it was also certified in the chicken embryo fibroblasts (CEFs) which infected with ARV. Then the IFN was added into the cell culture medium before CEFs were infected with ARV. The results indicated that the mRNA of IFITM3 expression was significantly increased and ARV multiplication was significantly inhibited. And when the expression of IFITM3 was knocked down by siRNA-IFITM3, the expression of IFITM3 was significantly reduced, but the ARV multiplication was significantly increased, which indicated that IFITM3 protein could inhibit the ARV replication.(AU)

Distribution of IFITM3 in yellow-feathered broilers and inhibition of Avian Reovirus multiplication by IFITM3

This study was carried to express the interferon-induced transmembrane protein 3 (IFITM3) in vitro and examine its function in inhibition of avian reovirus (ARV) replication. The recombinant prokaryotic vector expressing yellow-feathered broiler IFITM3 was successfully constructed, and the recombinant protein was expressed in competent Escherichia coli BL21 cells. New Zealand white rabbits were immunized with the purified recombinant protein to prepare a polyclonal antibody, with a titer of 1:128,000. Immunohistochemistry, reverse transcription-PCR, and real-time fluorescence quantitative PCR showed that IFITM3 was distributed in the yellow-feathered broiler immune organs, and the expression of IFITM3 in bursa of Fabricius was more than in spleen and thymus. It was found that in the thymus, spleen and bursa of Fabricius the mRNA expression levels of IFN and IFITM3 were significantly induced after ARV infection. And it was also certified in the chicken embryo fibroblasts (CEFs) which infected with ARV. Then the IFN was added into the cell culture medium before CEFs were infected with ARV. The results indicated that the mRNA of IFITM3 expression was significantly increased and ARV multiplication was significantly inhibited. And when the expression of IFITM3 was knocked down by siRNA-IFITM3, the expression of IFITM3 was significantly reduced, but the ARV multiplication was significantly increased, which indicated that IFITM3 protein could inhibit the ARV replication.

Enteric Virus Diversity Examined by Molecular Methods in Brazilian Poultry Flocks.

Enteric viruses play an important role in the Brazilian poultry industry due to the economic impact of resulting low yields of broilers, layers, and breeders. The most common enteric viruses affecting commercial flocks in Brazil include Fowl Adenovirus of group I (FAdV-I), Chicken Parvovirus (ChPV), Chicken Astrovirus (CAstV), Avian Nephritis Virus (ANV), Infectious Bronchitis Virus (IBV), Avian Reovirus (AReo), and Avian Rotavirus (ARtV). The aim of this study was to identify single and multiple infections using data obtained from 270 samples from eleven Brazilian states, corresponding to the period between 2010 and 2017. This was accompanied by an analysis of the relationship between the age of birds, clinical signs, and geographical distribution, using Polymerase Chain Reaction (PCR) and Reverse Transcription-PCR (RT-PCR) techniques. Twenty-five profiles of virus combinations were detected. Single infections were encountered in 86.3% of samples, and multiple infections were present in the remaining 13.7%. Both single and multiple infections affected all kinds of commercial chickens with digestive problems, stunting syndrome, decreases in egg and meat production, increased mortality, and respiratory signs. FAdV-I, ChPV, CAstV, ANV, and ARtV were mostly detected in young broilers, in contrast with IBV, which was detected in hens from one to greater than 51 weeks of age. These results exhibit the complexity of enteric diseases and the still poorly understood role of each pathogen as a unique etiological agent.

Investigation of enteric viruses in the feces of neotropical migratory birds captured on the coast of the State of Pará, Brazil

Migratory birds can become long-distance vectors for a wide range of microorganisms and can cause human disease, being the Brazilian coast a gateway for northern migratory birds. These animals are considered natural reservoirs of different viruses that cause important diseases, being relevant research of viral pathogens in migratory birds to epidemiology surveillance. The objective of the study was to investigate the presence of avian rotavirus (AvRV), avian reovirus (ARV) and picobirnavirus (PBV) in Neotropical migratory birds captured on the coast of Brazil. A total of 23 individual fecal samples of the migratory birds species Calidris pusilla (20 birds), Numenius phaeopus (1 bird) and Charadrius semipalmatus (2 birds) were collected. Fecal suspensions were prepared from the collected samples for subsequent extraction of double-stranded RNA (dsRNA), which was subjected to polyacrylamide gel electrophoresis (PAGE) and reverse transcription polymerase chain reaction (RT-PCR). The electrophoretic profiles were not detected by PAGE, and the amplification for the studied viruses PBV, ARV and AvRV (specie D, gene VP6 and NSP4) were negative. Positivity for AvRVD, VP7 gene was of 4.35% (1/23) for the migratory bird Calidris pusilla. After sequencing and building the tree of phylogenetic relationships avian Rotavirus Group D identified in this study was phylogenetically related and grouped into one branch, together to previously reported AvRVD from Brazil in chicken flocks with 99.8% nucleotide and 100% amino acid similarities.(AU)

Investigation of enteric viruses in the feces of neotropical migratory birds captured on the coast of the State of Pará, Brazil

Migratory birds can become long-distance vectors for a wide range of microorganisms and can cause human disease, being the Brazilian coast a gateway for northern migratory birds. These animals are considered natural reservoirs of different viruses that cause important diseases, being relevant research of viral pathogens in migratory birds to epidemiology surveillance. The objective of the study was to investigate the presence of avian rotavirus (AvRV), avian reovirus (ARV) and picobirnavirus (PBV) in Neotropical migratory birds captured on the coast of Brazil. A total of 23 individual fecal samples of the migratory birds species Calidris pusilla (20 birds), Numenius phaeopus (1 bird) and Charadrius semipalmatus (2 birds) were collected. Fecal suspensions were prepared from the collected samples for subsequent extraction of double-stranded RNA (dsRNA), which was subjected to polyacrylamide gel electrophoresis (PAGE) and reverse transcription polymerase chain reaction (RT-PCR). The electrophoretic profiles were not detected by PAGE, and the amplification for the studied viruses PBV, ARV and AvRV (specie D, gene VP6 and NSP4) were negative. Positivity for AvRVD, VP7 gene was of 4.35% (1/23) for the migratory bird Calidris pusilla. After sequencing and building the tree of phylogenetic relationships avian Rotavirus Group D identified in this study was phylogenetically related and grouped into one branch, together to previously reported AvRVD from Brazil in chicken flocks with 99.8% nucleotide and 100% amino acid similarities.

Avian reovirus infection: an overview / Reovirose aviária: um panorama

Poultry production is an activity of great importance in Brazilian economy, both due to the domestic consumption and the large amount of chicken meat exportation. Poultry activity modernization allowed the creation of animals in high density facilities, however, it facilitates the rapid dissemination of pathogens, which reduces the productivity rates. This review aims to highlight the avian reovirus, an important agent of arthritis in birds that has a worldwide distribution. The affected birds present a reduction in weight gain due to movement difficulties. In addition to arthritis, the virus may be related to a variety of pathological conditions, such as enteric and respiratory disorders, Hepatitis and myocarditis. The main prevention and control measure is the flock vaccination. Nevertheless, due to the avian reovirus great genetic variability, the vaccine may not be effective against circulating strains. This article aims to overview the virus biology, its variability and classification, and the infection pathology and diagnosis.(AU)

A avicultura é um setor de grande importância na economia brasileira tanto pelo aumento do consumo interno quanto pelo crescimento na exportação de carne de frango. A modernização da atividade avícola permitiu a criação adensada de animais, facilitando, no entanto, a rápida disseminação de patógenos que reduzem os índices de produtividade dos plantéis. Nesta revisão, é destacado o reovírus aviário, importante agente de artrite em aves que apresenta distribuição mundial. As aves acometidas apresentam redução no ganho de peso devido à dificuldade de locomoção. Além da artrite, o vírus pode estar relacionado a uma variedade de condições patológicas, como distúrbios entéricos e respiratórios, hepatite e miocardite. A principal forma de prevenção e controle é a vacinação do plantel. No entanto, devido à grande variabilidade genética do reovírus aviário, a vacina utilizada pode não ser eficiente contra estirpes que circulam no campo. O artigo traz uma visão geral sobre a biologia do vírus, sua variabilidade e propostas de classificação dos isolados, patologia da doença e diagnóstico da infecção.(AU)

Avian reovirus infection: an overview / Reovirose aviária: um panorama

Poultry production is an activity of great importance in Brazilian economy, both due to the domestic consumption and the large amount of chicken meat exportation. Poultry activity modernization allowed the creation of animals in high density facilities, however, it facilitates the rapid dissemination of pathogens, which reduces the productivity rates. This review aims to highlight the avian reovirus, an important agent of arthritis in birds that has a worldwide distribution. The affected birds present a reduction in weight gain due to movement difficulties. In addition to arthritis, the virus may be related to a variety of pathological conditions, such as enteric and respiratory disorders, Hepatitis and myocarditis. The main prevention and control measure is the flock vaccination. Nevertheless, due to the avian reovirus great genetic variability, the vaccine may not be effective against circulating strains. This article aims to overview the virus biology, its variability and classification, and the infection pathology and diagnosis.

A avicultura é um setor de grande importância na economia brasileira tanto pelo aumento do consumo interno quanto pelo crescimento na exportação de carne de frango. A modernização da atividade avícola permitiu a criação adensada de animais, facilitando, no entanto, a rápida disseminação de patógenos que reduzem os índices de produtividade dos plantéis. Nesta revisão, é destacado o reovírus aviário, importante agente de artrite em aves que apresenta distribuição mundial. As aves acometidas apresentam redução no ganho de peso devido à dificuldade de locomoção. Além da artrite, o vírus pode estar relacionado a uma variedade de condições patológicas, como distúrbios entéricos e respiratórios, hepatite e miocardite. A principal forma de prevenção e controle é a vacinação do plantel. No entanto, devido à grande variabilidade genética do reovírus aviário, a vacina utilizada pode não ser eficiente contra estirpes que circulam no campo. O artigo traz uma visão geral sobre a biologia do vírus, sua variabilidade e propostas de classificação dos isolados, patologia da doença e diagnóstico da infecção.

Unraveling complex viral infections in cassava (Manihot esculenta Crantz) from Colombia.

In the Americas, different disease symptoms have been reported in cassava including leaf mosaics, vein clearings, mottles, ring spots, leaf distortions and undeveloped and deformed storage roots. Some viruses have been identified and associated with these symptoms while others have been reported in symptomless plants or latent infections. We observed that reoviruses associated with severe root symptoms (RS) of Cassava Frogskin Disease (CFSD) are not associated with leaf symptoms (LS) observed in the cassava indicator plant 'Secundina'. Neither were these LS associated with the previously characterized Cassava common mosaic virus, Cassava virus X, Cassava vein mosaic virus or phytoplasma, suggesting the presence of additional pathogens. In order to explain LS observed in cassava we used a combination of biological, serological and molecular tests. Here, we report three newly described viruses belonging to the families Secoviridae, Alphaflexiviridae and Luteoviridae found in cassava plants showing severe RS associated with CFSD. All tested plants were infected by a mix of viruses that induced distinct LS in 'Secundina'. Out of the three newly described viruses, a member of family Secoviridae could experimentally induce LS in single infection. Our results confirm the common occurrence of complex viral infections in cassava field-collected since the 1980s.

The occurrence of orthoreovirus, rotavirus and chicken anemia virus in chickens of the poultry industry in Minas Gerais, Brazil / Ocorrência de orthoreovirus, rotavirus e vírus da anemia das galinhas em frangos de corte da indústria avícola de Minas Gerais

Fifty-four fecal samples taken from broiler chickens from 1 to 45 days of age, and of pullets from 10 to 13 weeks of age, original from eight different poultry regions in the state of Minas Gerais, Brazil, were collected from March 2008 to January 2010 for avian Orthoreovirus (ARV) and avian Rotavirus (AvRV) analyses. For the assay of ARV, RNA was immediately extracted (Trizolâ) and transcribed into cDNA for assaying in a nested-PCR with ARV-specific primers. For AvRV, polyacrylamide gel electrophoresis (PAGE) was performed with RNA extracts obtained by phenol-chloroform extraction. CAV was additionally investigated through a nested-PCR of thymus and spleen. Results found 5.55% positive for ARV and 9.25% for AvRV. Also, CAV and ARV genomes were detected in co-infection, in a highly prostrated and claudicating chicken flock. No ARV or AvRV infections were detected in pullets. Material of a clinically affected flock was inoculated into SPF embryos, resulting in embryonic hemorrhage, whitish foci in the chorio-allantoic membrane and death. Sequencing of ARV amplicons and isolate cDNA grouped local strains with the ARV S1133 strain, historically used in live vaccines, suggesting the continued circulation of this vaccine virus strain in intensive poultry regions. Detection rates for ARV and AvRV, as well as the presence of CAV, were additionally indicative of failing biosecurity strategies for the intensive poultry regions examined.

Avaliou-se a ocorrência de Orthoreovirus (ARV) e Rotavirus (AvRV) aviários na avicultura industrial de Minas Gerais. Foram colhidas cinquenta e quatro amostras de fezes de frangos de corte entre um e 45 dias e de frangas de postura de 10 a 13 semanas de idade. Para análise de ARV, o RNA foi imediatamente extraído (Trizol), transcrito em cDNA e avaliado em uma PCR com oligonucleotídeos iniciadores específicos para ARV. Para a investigação de AvRV, os extratos de RNA foram obtidos por fenol-clorofórmio e submetidos à eletroforese em gel de poliacrilamida. Todas as amostras foram também avaliadas para o DNA do vírus da anemia das galinhas (CAV) em uma nested-PCR específica. Em frangos de corte, a positividade encontrada para ARV foi de 5,55% e para AvRV de 9,25%. CAV foi detectado em coinfecção em um plantel com refugagem, claudicação e prostração. Nenhuma amostra de poedeiras foi positiva para ARV ou AvRV. Material de plantel com sinais clínicos foi purificado e inoculado em ovos SPF embrionados, sendo obtidas lesões hemorrágicas e focos brancos na membrana cório-alantóide. O sequenciamento dos produtos de PCR e de embrião agrupou os isolados de ARV com a estirpe S1133, historicamente usada como vacina viva. Os resultados sugerem a continuada circulação da infecção por estirpes assemelhadas a ARV S1133 nas regiões de avicultura industrial. Os índices de detecção de ARV, AvRV e CAV indicam que a intensificação nas regiões produtoras tem resultado em falhas de biosseguridade.

The occurrence of Orthoreovirus, Rotavirus and chicken anemia virus in chickens of the poultry industry in Minas Gerais, Brazil / Ocorrência de Orthoreovirus, Rotavirus e vírus da anemia das galinhas em frangos de corte da indústria avícola de Minas Gerais

Fifty-four fecal samples taken from broiler chickens from 1 to 45 days of age, and of pullets from 10 to 13 weeks of age, original from eight different poultry regions in the state of Minas Gerais, Brazil, were collected from March 2008 to January 2010 for avian Orthoreovirus (ARV) and avian Rotavirus (AvRV) analyses. For the assay of ARV, RNA was immediately extracted (Trizolâ) and transcribed into cDNA for assaying in a nested-PCR with ARV-specific primers. For AvRV, polyacrylamide gel electrophoresis (PAGE) was performed with RNA extracts obtained by phenol-chloroform extraction. CAV was additionally investigated through a nested-PCR of thymus and spleen. Results found 5.55% positive for ARV and 9.25% for AvRV. Also, CAV and ARV genomes were detected in co-infection, in a highly prostrated and claudicating chicken flock. No ARV or AvRV infections were detected in pullets. Material of a clinically affected flock was inoculated into SPF embryos, resulting in embryonic hemorrhage, whitish foci in the chorio-allantoic membrane and death. Sequencing of ARV amplicons and isolate cDNA grouped local strains with the ARV S1133 strain, historically used in live vaccines, suggesting the continued circulation of this vaccine virus strain in intensive poultry regions. Detection rates for ARV and AvRV, as well as the presence of CAV, were additionally indicative of failing biosecurity strategies for the intensive poultry regions examined.(AU)

Avaliou-se a ocorrência de Orthoreovirus (ARV) e Rotavirus (AvRV) aviários na avicultura industrial de Minas Gerais. Foram colhidas cinquenta e quatro amostras de fezes de frangos de corte entre um e 45 dias e de frangas de postura de 10 a 13 semanas de idade. Para análise de ARV, o RNA foi imediatamente extraído (Trizol), transcrito em cDNA e avaliado em uma PCR com oligonucleotídeos iniciadores específicos para ARV. Para a investigação de AvRV, os extratos de RNA foram obtidos por fenol-clorofórmio e submetidos à eletroforese em gel de poliacrilamida. Todas as amostras foram também avaliadas para o DNA do vírus da anemia das galinhas (CAV) em uma nested-PCR específica. Em frangos de corte, a positividade encontrada para ARV foi de 5,55% e para AvRV de 9,25%. CAV foi detectado em coinfecção em um plantel com refugagem, claudicação e prostração. Nenhuma amostra de poedeiras foi positiva para ARV ou AvRV. Material de plantel com sinais clínicos foi purificado e inoculado em ovos SPF embrionados, sendo obtidas lesões hemorrágicas e focos brancos na membrana cório-alantóide. O sequenciamento dos produtos de PCR e de embrião agrupou os isolados de ARV com a estirpe S1133, historicamente usada como vacina viva. Os resultados sugerem a continuada circulação da infecção por estirpes assemelhadas a ARV S1133 nas regiões de avicultura industrial. Os índices de detecção de ARV, AvRV e CAV indicam que a intensificação nas regiões produtoras tem resultado em falhas de biosseguridade.(AU)

Anticorpos contra vírus de galinha em cracídeos / Detection of antibodies against commercial poultry viruses in cracids

Os cracídeos são Galliformes silvestres das Américas. Com o objetivo de investigar a presença de anticorpos contra vírus de galinhas em cracídeos, foram coletadas 51 amostras de soro de 10 diferentes espécies dessas aves. Esses animais eram mantidos em criatórios conservacionistas e zoológicos nos Municípios de Santa Maria, Soledade, Passo Fundo, Sapucaia, Gravataí, Viamão e Três Coroas, Estado do Rio Grande do Sul, Brasil. Anticorpos neutralizantes foram detectados em 5,9 por cento (3/51) do total de amostras testadas contra o vírus da bronquite infecciosa das galinhas, 15,7 por cento (8/51) contra o reovírus aviário e 35,3 por cento (18/51) contra o vírus da doença infecciosa da bolsa. Todas as amostras foram negativas para o vírus da bouba aviária no teste de IDGA. A detecção de anticorpos para vírus de aves comerciais sugere que os cracídeos podem ser susceptíveis à infecção por esses vírus.

The cracids are wild Galliformes native from the Americas. Fifty one serum samples were collected from individuals of 10 different species of cracids in order to obtain information regarding to the antibody status of different viruses. These birds were kept in shelters and zoos localized in Santa Maria, Soledade, Passo Fundo, Sapucaia, Gravataí, Viamão and Três Coroas counties, in the Rio Grande do Sul State, Brazil. Neutralizing antibodies were detected in the individuals serum from different species specific referring to infectious bronchitis virus in 5.9 percent (3/51) of the samples, to avian reovirus in 15.7 percent (8/51) and, to infectious bursal disease virus in 35.3 percent (18/51). All samples were negative for fowlpox virus, as measured by IDGA test. The detection of commercial poultry viruses antibodies suggests that cracids could be susceptible to infection by those viruses.