Pathological and molecular findings of avian avulavirus type 1 outbreak in pigeons (Columba livia) of southern Brazil / Achados patológicos e moleculares da infecção por avulavirus aviário tipo 1 em pombos (Columba livia) do sul do Brasil

The Newcastle disease, caused by avian avulavirus type 1 strains (APMV-1) is an important avian disease involved into high rates of mortality and economic losses. Several outbreaks have been reported over the last 30 years in Columbiformes in different parts of the world, caused by a adapted variant strain of AAvV-1, called pigeon paramyxovirus type 1 (PPMV-1). A high mortality associated with an outbreak was analyzed in free-living pigeons (Columba livia) in a public square in Porto Alegre in Southern Brazil. A total of 24 pigeons moribund or freshly dead, within five weeks interval were submitted to necropsy, histopathological, immunohistochemical (anti-Newcastle), and RT-PCR followed by sequencing of the amplification products analysis. They presented neurological signs, non-suppurative encephalitis and encephalomyelitis, and mononuclear inflammatory infiltrate in different organs. Immunohistochemical analysis in nine pigeons tissue showed that anti-Newcastle was expressed in brain, kidney, liver and pancreas. The RT-PCR test for the M protein of Newcastle disease virus was positive in six pigeons. The differential diagnosis of Influenza, West Nile, Mycoplasma gallisepticum and Mycoplasma synoviae in all pigeons presented negative results. The sequence of amino acids in the cleavage site region of the F protein was 112RRQKRF117 classifying the strain as virulent. The phylogenetic analysis classified this virus strain into Class II and VI genotype.(AU)

A doença de Newcastle, causada por cepas de avulavirus aviário tipo 1 (AAvV-1), é uma doença de aves importante por causar altos índices de mortalidade e perdas econômicas. Vários surtos têm sido relatados ao longo de 30 anos em aves da ordem Columbiformes, em diferentes partes do mundo, causados por uma cepa variante específica de AAvV-1, denominada Pigeon paramyxovirus tipo 1 (PPMV-1). Foi analisado um surto de mortalidade em pombos domésticos (Columba livia), provenientes de uma praça pública em Porto Alegre, no Sul do Brasil. Vinte e quatro aves moribundas ou mortas foram submetidas, no intervalo de cinco semanas, ao exame de necropsia, exame histopatológico, imuno-histoquímico anti-Newcastle, RT-PCR e sequenciamento. Apresentaram sinais neurológicos, encefalite e encefalomielite não supurativas, além de infiltrado inflamatório mononuclear em diversos órgãos. Nove aves demonstraram exame imuno-histoquímico positivo em órgãos como cérebro, rim, fígado e pâncreas. Seis aves foram positivas no exame de RT-PCR para a proteína M do vírus da Doença de Newcastle. Nos exames de diagnósticos diferenciais de Influenza, West Nile, Mycoplasma gallisepticum e Mycoplasma synoviae, todas as aves testadas foram negativas. A sequência dos aminoácidos na região do sítio de clivagem da proteína foi 112RRQKRF117, classificando a cepa como virulenta. De acordo com a análise filogenética o vírus identificado foi classificado como pertencente à classe II e ao genótipo VI.(AU)

Biological Property of Recombinant Hemagglutinin-Neuraminidase Protein of Avian Paramyxovirus Type 6 Expressed by Recombinant Baculovirus

Hemagglutination inhibition (HI) test employing whole virus antigen is a prescribed serological test for serotyping, diagnosis and surveillance for avian paramyxoviruses (APMVs). For use as alternative to the virus antigen, hemagglutinin-neuraminidase (HN) protein gene of the wild duck isolate APMV-6/WB12-163FS of APMV serotype 6 (APMV-6) was amplified, cloned and expressed in Spodoptera frugiperda insect cells. The HN gene of 1,842 bps in length showed nucleotide and amino acid homology of 93.4% and 97.1%, respectively with that of APMV-6 prototype strain. Putative sialic acid binding motif and potential N-linked glycosylation sites were conserved. In Western blot analysis, the expressed protein had a molecular mass of 66 kDa and reacted specifically with antiserum to APMV-6. In addition, the recombinant HN protein showed biological properties such as hemagglutination (HA) and elution. The recombinant HN protein produced from infected cells showed high HA titers (approximately 2(13) HA unit/ml). The HA activity of the recombinant HN protein was inhibited by antisera to APMV-6. In cross HA inhibition test, the recombinant HN protein had the highest titers with antisera to homologous APMV serotype, although there was weak cross reaction with some of antisera to other APMV serotypes. Our results indicated that recombinant APMV-6 HN protein would have the potential as alternative to the APMV-6 antigen in HI assays.

Characterization and comparison of the pathogenicity of viscerotropic velogenic Newcastle disease virus isolates in Korea / 대한수의학회지

A total of 18 Newcastle disease virus (NDV) isolates that were recovered from 1949 through 1997 were characterized and pathotyped. All viruses were highly virulent as determined by intracerebral pathogenicity indices > or = 1.81 in day-old. These pathotypes are typical for viscerotropic velogenic NDV (VVNDV) pathotype viruses. Some differences were observed for the chicken red blood cell elution rate and thermostability of the hemagglutinin at 56degrees C. Three antigenic groups were identified by a hemagglutination-inhibition assay using NDV monoclonal antibodies. And the predominant gross lesions were as follows: discharge from the nasal cavity, tracheal mucus, petechial hemorrhage in the heart fat, kidney urates and hemorrhage with or without necrosis in the gastrointestinal tract. Severe hemorrhagic or necrotic lesions were also noted in the lymphoid organs and were localized primarily in the spleen and cecal tonsil. However, differences in the occurrence and frequency of the gross lesions were observed between the virus strains. Among them, NDV strains that induced neurological symptoms belonged only to genotype VI. This strain had spread throughout Korea during the late 1980s to the 1990s, which suggests that specific VVNDVs genotypes might result in neurological symptoms.

Research of Viral Agent in Free-living Pigeon Feces (Columba livia) in the City of São Paulo, SP, Brazil, for Transmission Electron Microscopy / Investigación de Agentes Virales en Heces de Palomas de Vida Libre (Columba livia) en la Ciudad de São Paulo, SP, Brasil, Através de Miscroscopía Electrónica de Transmisión

The pigeon feces are vehicle of diseases both for humans and other animal species. In these birds, the most important viral diseases of the digestive tract are transmitted by the paramyxovirus, adenovirus and coronavirus. Avian paramyxoviruses have been isolated from a variety of species of free living and domestic birds worldwide, with several symptoms and clinical signs and economic losses. Paramyxoviruses belong to the Paramyxoviridae family and Avulovirus genus that includes nine serotypes (APMV 1 to 9). Avian adenoviruses belong to the Adenoviridae family and Aviadenovirus genus. In pigeons, cause classical adenovirosis and necrotizing hepatitis. The respiratory and enteric tracts are common targets of coronavirus. They belong to the Coronaviridae family and to 3a and 3c groups. In this study, we described the presence of viral agents in free-living pigeon feces (Columba livia) from the city of São Paulo, Brazil. The feces were processed by negative staining technique (rapid preparation) for transmission electron microscopy. In this technique paramyxoviruses particles, pleomorphic, roughly spherical or filamentous, measuring 100 to 500 nm of diameter containing an envelope covered by spikes, with characteristic helical herring-bone-like nucleocapsid, measuring 15 to 20 nm in diameter, were visualized in 45 (79 percent) out of 57 feces samples. In 2 (3.5 percent) samples, paramyxovirus and adenovirus particles were simultaneously visualized. Adenovirus particles were isometric, spherical, characterized as "complete "or" empty ", measuring between 70 and 90 nm in diameter. Paramyxovirus and coronavirus particles were detected in 3 (5.2 percent) samples. Coronaviruses were pleomorphic with a diameter of 75-160 nm containing a solar corona-shaped envelope, with projections of approximately 20 nm of diameter. Seven (12.3 percent) samples were negative for viral particles.

Los heces de las palomas constituyen vehículos de enfermedades importantes, tanto para el Hombre como para otras especies animales. En estas aves, las enfermedades virales más importantes del tracto digestivo son transmitidas por los paramixovirus, adenovirus y coronavirus. Paramixovirus aviario, en todo el mundo, ha sido aislado de una variedad de especies de vida libre y de aves domésticas, que causan variados síntomas y señales clínicas con pérdidas económicas. El Paramixovirus pertenece a la familia Paramyxoviridae y al género Avulavirus que incluye nueve serotipos (APMV 1 a 9). Adenovirus aviario pertenece al género de la familia Adenoviridae y género Aviadenovirus. En las palomas, causan la adenovirosis clásica y la hepatitis necrotizante. El tracto respiratorio y entérico son los albos comunes de los coronavirus. Ellos pertenecen a la familia Coronaviridae y a los grupos 3a y 3c. En este trabajo, se describe la presencia de agentes virales en las heces de palomas de vida libre (Columba livia) en la ciudad de São Paulo, Brasil. Las heces se procesaron para la técnica de microscopía electrónica de transmisión, a través de la técnica de contrastación negativa (preparación rápida). A través de esta técnica fueron visualizadas las partículas de paramixovirus, pleomórficas, más o menos esféricas o filamentosas, de 100 a 500 nm de diámetro que contiene un envoltorio cubierto con espículas, con nucleocapside con características helicoidales, midiendo de 15 a 20 nm de diámetro en 45 (79 por ciento) de 57 muestras. En 2 (3,5 por ciento) muestras, fueron observadas simultáneamente partículas de paramixovirus y de adenovirus. Las partículas de adenovirus eran isométricas, esféricas, caracterizadas como “completa " o “vacía ", midiendo entre 70 y 90 nm de diámetro. Fueron analizadas en tres muestras (5,2 por ciento) las partículas de paramixovirus y coronavirus. Los coronavirus son pleomórficos, con un diámetro de 75 a 160 nm, que contiene un capa en forma de corona solar con proyecciones de aproximadamente 20 nm de diámetro. Siete (12,3 por ciento) muestras fueron negativas para las partículas virales.

Prevalence of newcastle disease virus in broiler chickens (gallus gallus) in Brazil

This study was carried out during 2002/2003, aiming to determine the prevalence of virulent Newcastle disease virus strains (NDV) in Brazilian commercial poultry farms. Clinical samples were obtained from the Southeastern, Southern and Central-Western regions, which comprise the main area of the Brazilian poultry production. Serum samples and tracheal and cloacal swabs of 23,745 broiler chickens from 1,583 flocks, including both vaccinated chickens and those with no vaccination information, were tested for NDV using a diagnostic ELISA kit. The seropositivity was 39.1 percent, and the isolation percentage by flock varied from 1.0 to 7.6 percent, and by region from 6.5 to 58.4 percent. Higher isolation rates (74.3-83.3 percent) were obtained after three passages in embryonated chicken eggs. All isolates preliminarily identified as NDV were characterized as nonpathogenic strains, as their Intracerebral Pathogenicity Index (ICPI) was below 0.7. Based on results of this study, Brazil can claim a virulent NDV-free status for commercial flocks.

A survey for maintenance of virulent newcastle disease virus-free area in poultry production in Brazil

In 2003, Brazil was recognized as a pathogenic Newcastle Disease Virus (NDV) strain-free country for commercial poultry. This research was conducted in Brazil between December 2003 and March 2005 to verify the maintenance of this virulent NDV-free status. Serum samples from 5,455 flocks for commercial poultry farms were collected, comprising 81,825 broiler chickens. The farms were located in nine states of the country, grouped in three geographic regions. Serological evidence of NDV infection was detected in 28.8 percent of the surveyed farms. However, all fifteen viruses isolated and identified as Newcastle Disease Virus (NDV) were characterized as nonpathogenic strains, based on the Intracerebral Pathogenicity Index. These results showed that Brazil preserves the virulent NDV-free status for commercial flocks.

Preparation and identification of the monoclonal antibody specific to hemagglutinin of avian paramyxovirus type 2 / 病毒学报

A hybridoma cell line 1G4A7 secreting monoclonal antibody (McAb) specific to hemagglutinin of avian paramyxovirus type 2 (APMV-2) was developed by fusing the spleen cells of APMV-2 immunized BAlb/c mice with SP2/0 myeloma cells. The immunoglobulin subclass of 1G4A7 was IgG1 with light chain kappa and the affinity constant against APMV-2 was 1.02 X 10(10). Identified by HI and indirect ELISA, the McAb titers in ascities were 10 log 2 and 1 : 10(6) respectively. The McAb did not cross react with the common avian viruses, showing good specificity. There existed obvious differences in antigenitic relationship among APMV-2 viruses analyzed by HI and indirect ELISA using McAb 1G4A7.

Resposta imune conferida pelas amostras La Sota e VG/GA do vírus da doença de Newcastle / Immune response conferred by La Sota and VG/GA strains of the Newcastle Disease Virus

Estudou-se a resposta imune de aves infectadas experimentalmente com amostras lentogênicas do vírus da doença de Newcastle (VDN), incluindo uma amostra com tropismo para o sistema respiratório e outra com tropismo entérico. Foram utilizadas as vias ocular, nasal, água de bebida e aspersão para a inoculação das amostras virais. A pesquisa foi realizada em 1.600 aves de linhagem para corte, divididas em oito grupos, de acordo com a amostra do vírus e a via de inoculação. Dez aves de cada grupo foram sacrificadas semanalmente. A resposta imune foi avaliada pela determinaçäo de anticorpos por ELISA e inibição da hemaglutinação. Os grupos inoculados com a amostra La Sota exibiram níveis mais elevados de anticorpos anti-VDN, superiores aos dos grupos inoculados com a amostra VG/GA. A resposta anamnéstica foi mais baixa para a amostra VG/GA inoculada pela água de bebida, e os níveis mais elevados de anticorpos circulantes, no pré-desafio, näo impediram a infecçäo pelo vírus