Identification of a Virulent Newcastle Disease Virus Strain Isolated from Pigeons (Columbia livia) in Northeastern Brazil Using Next-Generation Genome Sequencing.

Newcastle disease virus (NDV), also known as avian paramyxoviruses 1 (APMV-1) is among the most important viruses infecting avian species. Given its widespread circulation, there is a high risk for the reintroduction of virulent strains into the domestic poultry industry, making the surveillance of wild and domestic birds a crucial process to appropriately respond to novel outbreaks. In the present study, we investigated an outbreak characterized by the identification of sick pigeons in a large municipality in Northeastern Brazil in 2018. The affected pigeons presented neurological signs, including motor incoordination, torticollis, and lethargy. Moribund birds were collected, and through a detailed histopathological analysis we identified severe lymphoplasmacytic meningoencephalitis with perivascular cuffs and gliosis in the central nervous system, and lymphoplasmacytic inflammation in the liver, kidney, and intestine. A total of five pigeons tested positive for NDV, as assessed by rRT-PCR targeted to the M gene. Laboratory virus isolation on Vero E6 cells confirmed infection, after the recovery of infectious NVD from brain and kidney tissues. We next characterized the isolated NDV/pigeon/PE-Brazil/MP003/2018 by next-generation sequencing (NGS). Phylogenetic analysis grouped the virus with other NDV class II isolates from subgenotype VI.2.1.2, including two previous NDV isolates from Brazil in 2014 and 2019. The diversity of aminoacid residues at the fusion F protein cleavage site was analyzed identifying the motif RRQKR↓F, typical of virulent strains. Our results all highlight the importance of virus surveillance in wild and domestic birds, especially given the risk of zoonotic NDV.

An Outbreak in Pigeons Caused by the Subgenotype VI.2.1.2 of Newcastle Disease Virus in Brazil.

Newcastle disease virus (NDV) can infect over 250 bird species with variable pathogenicity; it can also infect humans in rare cases. The present study investigated an outbreak in feral pigeons in São Paulo city, Brazil, in 2019. Affected birds displayed neurological signs, and hemorrhages were observed in different tissues. Histopathology changes with infiltration of mononuclear inflammatory cells were also found in the brain, kidney, proventriculus, heart, and spleen. NDV staining was detected by immunohistochemistry. Twenty-seven out of thirty-four tested samples (swabs and tissues) were positive for Newcastle disease virus by RT-qPCR test, targeting the M gene. One isolate, obtained from a pool of positive swab samples, was characterized by the intracerebral pathogenicity index (ICPI) and the hemagglutination inhibition (HI) tests. This isolate had an ICPI of 0.99, confirming a virulent NDV strain. The monoclonal antibody 617/161, which recognizes a distinct epitope in pigeon NDV strains, inhibited the isolate with an HI titer of 512. A complete genome of NDV was obtained using next-generation sequencing. Phylogenetic analysis based on the complete CDS F gene grouped the detected isolate with other viruses from subgenotype VI.2.1.2, class II, including one previously reported in Southern Brazil in 2014. This study reports a comprehensive characterization of the subgenotype VI.2.1.2, which seems to have been circulating in Brazilian urban areas since 2014. Due to the zoonotic risk of NDV, virus surveillance in feral pigeons should also be systematically performed in urban areas.

Isolation and genetic characterization of virulent strains of avian paramyxovirus-1 from multiple avian species in Azad Jammu and Kashmir 2017-2018.

Despite intensive vaccination, endemicity of Avian paramyxoviruses-1 (APMV-1) is a significant problem in developing countries in Africa, Middle East, and Asia. Given the importance of APMV-1 in poultry and multiple non-poultry avian species, it is important to continue surveillance programs, routine monitoring and characterization of field isolates in the region where viruses are endemic. The purpose of this study was to pathotyped and genetically characterized 21 APMV-1s isolated from multiple avian species reared in different regions of Azad Jammu and Kashmir (AJK). Phylogenetic analysis based on complete fusion (F) gene sequences showed that 17 APMV-1 isolates obtained from commercial poultry and backyard birds belonged to sub-genotype VIIi. Though, one pigeon-origin APMV-1 isolate was clustered in sub-genotype VIg and three in recently designated new sub-genotype VIm of genotype VI. The pigeon-origin isolates had the following two motifs 113-RKKR↓F-117 and 113-RQRR↓F-117, while all other isolates had the polybasic amino acid sequence 113-RQKR↓F-117 at the F-cleavage site, which is characteristic of virulent APMV-1 strains. These results are consistent with the five viruses that had intracerebral pathogenicity indices (ICPIs) of between 1.50 and 1.73, corresponding to a velogenic pathotype. The APMV-1s isolated from commercial poultry and backyard birds in this study showed low nucleotide distance (0.3-0.9%) and genetically closely related (> 97%) to viruses repeatedly isolated (2011-2017) from multiple avian species in other states of Pakistan. Strengthened surveillance programs in both commercial poultry and backyard flocks are needed to better assess the commercial-backyard bird interface and form a basis for evidence-based measures to limit and prevent APMV-1 transmission.

Characterization of Colombian serotype 1 avian paramyxoviruses, 2008-2010.

Newcastle disease (ND) still remains one of the most important diseases affecting domestic poultry in Colombia. Here, for the first time, we report on the molecular characterization of 12 virulent and 12 avirulent or lentogenic avian paramyxovirus type 1 (APMV-1) strains that were isolated from commercial, backyard, and game poultry in Colombia from 2008 to 2010. The 12 virulent isolates had a fusion (F) protein cleavage site with basic amino acids at positions 113, 115, and 116 and a phenylalanine at position 117 (112RRQKR*F117), characteristic of virulent strains. The remaining 12 isolates had the F protein cleavage sites 112GKQGR*L117 or 112GRQGR*L117 typical of avirulent or lentogenic APMV-1 strains. Phylogenetic analysis of full-length F genes of all isolates was performed, and based on the recently proposed criteria for classification of APMV-1 strains, the 24 Colombian isolates were found to belong to class II viruses and clustered into four different genotypes. Ten virulent isolates clustered with genotype VII (sub-genotype VIId), seven lentogenic strains within genotype II, five lentogenic strains with genotype I (sub-genotype Ia), and two virulent isolates within genotype XII. Our data provide essential information on the genetic diversity of AMPV-1 isolates circulating in Colombia.

Avaliação sazonal do perfil sanitário de pombos-domésticos (Columba livia) em áreas de armazenamento de grãos e sementes no Estado de São Paulo / Seasonal health status survey of feral pigeons (Columba livia) in areas used for the storage of grains and seeds in São Paulo State

Columbiformes sinantrópicos podem ter um importante papel na epidemiologia de patógenos com potencial zoonótico ou de impacto econômico para a indústria avícola. Dentre eles destacam-se: Mycoplasma spp., Salmonella spp., paramixovírus aviário tipo 1 (APMV-1), inseridos no Programa Nacional de Sanidade Avícola (PNSA) e a Chlamydophila psittaci, agente de uma das principais zoonoses relacionada com aves silvestres. Dentro desse contexto, este trabalho objetivou pesquisar, sazonalmente, a ocorrência destes patógenos em pombos-domésticos (Columba livia) em dois entrepostos no Estado de São Paulo. Ao longo de um ano, mensalmente 10 pombos foram capturados em cada entreposto para a colheita de amostras de suabe cloacal e sangue. A técnica de soroaglutinação rápida em placa (SAR) foi utilizada para a detecção de anticorpos anti-M. synoviae, anti-M. gallisepticum e anti-S.Pullorum/Gallinarum; para a confirmação dos sororeagentes foram utilizadas a prova de inibição da hemaglutinação e soroaglutinação lenta, respectivamente. Para a detecção do DNA de C. psittaci e RNA de AMPV-1 foram utilizados métodos moleculares, PCR e RT-PCR. Para investigação de anticorpos anti-APMV-1 foi empregada a técnica de HI. Na SAR, 3,3% dos soros foram reagentes para M. synoviae; 2,5% para M. gallisepticum e 0,4% para S. Pullorum/Gallinarum. No entanto, essas amostras foram negativas nas técnicas confirmatórias. A ocorrência do APMV-1 não foi detectada. O DNA de C. psittaci foi detectado em 13,3% das amostras sendo 10,8% provenientes de aves capturadas na estação seca e 15,8% na estação chuvosa. Tais resultados são relevantes, pois demonstram que a C. psittaci ocorre em pombos presentes em áreas públicas frequentadas por um grande número de pessoas. Frente à escassez de pesquisas realizadas em Columbiformes no país, novos estudos são necessários para a determinação do real risco que pombos-domésticos podem representar quanto à transmissão de patógenos para aves comerciais e a influência da sazonalidade na disseminação desses microrganismos

Columbiformes may play an important role in the epidemiology of pathogens with zoonotic potential or economic impact in the poultry industry. Among these pathogens there are Mycoplasma spp., Salmonella spp., Avian Paramyxovirus type 1 (APMV-1), included in the National Poultry Health Program (PNSA) and Chlamydophila psittaci, etiologic agent of an important zoonosis associated with wild birds. Thus, the aim of this study was to investigate, seasonally, the occurrence of the pathogens listed above in feral pigeons (Columba livia) in two warehouses in São Paulo State. During one year, 10 birds were captured monthly in each locality and cloacal swabs and blood samples were collected from each pigeon. The rapid seroagglutination test was performed for the detection of antibodies against M. synoviae, M. gallisepticum and Salmonella Pullorum/Gallinarum. Positive results were submitted to the hemagglutination inhibition and slow seroagglutination test, respectively. For the C. psittacis DNA and APMV-1s RNA diagnosis, molecular techniques PCR and RT-PCR were performed. Hemagglutination inhibition test was also performed in order to detect antibodies against APMV-1. From the serum samples analyzed by rapid seroagglutination test, 3.3% were positive for M. synoviae, 2.5% for M. gallisepticum and 0.4% for S. Pullorum/Gallinarum. However, none of these samples was positive on the confirmatory tests. APMV-1 was not detected in any of the laboratory tests used. C. psittacis DNA was detected in 13.3% of the samples being, 10.8% from pigeons captured during the dry season and 15.8% in the rainy season. These results are relevant since they indicate that C. psittaci occurs in birds living in public areas frequented by a large number of people. The occurrence of the other pathogens was not detected. Nevertheless, due to lack of information about the pigeons sanitary status in the country, additional researches are necessary to determine the risk that feral pigeons can pose in the transmission of pathogens for poultry and the influence of each season in the spread of these microorganisms

Molecular surveillance of the Newcastle disease virus in domestic and wild birds on the North Eastern Coast and Amazon biome of Brazil

Brazil is one of the world's largest countries with a rich diversity of wildlife, including resident and migratory wild birds, which may be natural reservoirs of the Newcastle disease virus (NDV). Because Brazil is a major global exporter of chicken meat, the emergence of such a disease may have a huge negative impact not only on the economy due to trade restrictions and embargoes, but also on the quality of life of the population. Samples were collected from 1,022 asymptomatic domestic and wild birds from the Brazilian coast and the Amazon region using tracheal/cloacal swabs and tested by RT-qPCR. The results showed 7 (0.7%) birds were positive for NDV. The positive samples were then isolated in embryonated chicken eggs and their matrix protein genes were partially sequenced, revealing a low-pathogenicity NDV. This study confirms the maintenance of the velogenic-NDV free status of Brazil.(AU)

Molecular surveillance of the Newcastle disease virus in domestic and wild birds on the North Eastern Coast and Amazon biome of Brazil

Brazil is one of the world's largest countries with a rich diversity of wildlife, including resident and migratory wild birds, which may be natural reservoirs of the Newcastle disease virus (NDV). Because Brazil is a major global exporter of chicken meat, the emergence of such a disease may have a huge negative impact not only on the economy due to trade restrictions and embargoes, but also on the quality of life of the population. Samples were collected from 1,022 asymptomatic domestic and wild birds from the Brazilian coast and the Amazon region using tracheal/cloacal swabs and tested by RT-qPCR. The results showed 7 (0.7%) birds were positive for NDV. The positive samples were then isolated in embryonated chicken eggs and their matrix protein genes were partially sequenced, revealing a low-pathogenicity NDV. This study confirms the maintenance of the velogenic-NDV free status of Brazil.