Effect of host selective pressure on Newcastle disease virus virulence

Newcastle disease virus (NDV) causes an economically important disease that can vary from clinically inapparent to highly virulent forms. Generally, NDV strains isolated from wild birds are non-pathogenic for chicken. However, there are evidences supporting the fact that avirulent viruses maintained in feral birds could have caused outbreaks of virulent NDV in poultry. The strain-specific difference in virulence is determined by structural variations on the fusion glycoprotein (F). More basic amino acids are present in the F cleavage site of virulent strains. Nevertheless, other regions have been involved in virulence determination. When we subjected an avirulent NDV isolated from a wild bird to a host change we found that the virus arose was virulent for chicken. Nucleotide changes in the F protein cleavage site amino acid sequence and in the hemagglutinin-neuraminidase protein sequence are reported.     

The avian response to Newcastle disease virus

Newcastle disease virus (NDV) is classified as a member of the superfamily Mononegavirales in the family Paramyxoviridae. This virus family is divided into two subfamilies, the Paramyxovirinae and the Pneumovirinae. In 1993 the International Committee on the Taxonomy of Viruses rearranged the order of the Paramyxovirus genus and placed NDV within the Rubulavirus genus among the Paramyxovirinae. The enveloped virus has a negative sense single-stranded RNA genome of 15,186 kb which codes for an RNA directed RNA polymerase, hemagglutinin-neuraminidase protein, fusion protein, matrix protein, phosphoprotein and nucleoprotein in the 5' to 3' direction. The virus has a wide host range with most orders of birds reported to have been infected by NDV. Isolates are characterized by virulence in chickens and are categorized into three main pathotypes depending on severity of disease. Lentogenic isolates are of low virulence while viruses of intermediate virulence are termed mesogenic. Highly virulent viruses that cause high mortality in birds are termed neurotropic or viscerotropic velogenic. Velogenic NDV are List A pathogens that require reporting to the Office of International Epizootics and outbreaks result in strict trade embargoes. The primary molecular determinant for NDV pathogenicity is the fusion protein cleavage site amino acid sequence. Vaccination for NDV is primarily by mass application of live-virus vaccines among commercial poultry. Although protection is measured by presence of antibodies to NDV, vaccinated B-cell depleted chickens are resistant to disease. Consequently, immune protection involves responses that are presently incompletely defined.     

Phylogenetic Relationships among Highly Virulent Newcastle Disease Virus Isolates Obtained from Exotic Birds and Poultry from 1989 to 1996

Newcastle disease virus {NDV (avian paramyxovirus type 1 [APMV1])} isolates were recovered from imported exotic birds confiscated following importation into the United States, from waterbirds in the United States, and from poultry. The exotic birds probably originated from Central and South America, Asia, and Africa. The NDV isolates were initially characterized as highly virulent because of a short mean death time in embryonated chicken eggs. The isolates were typed as neurotropic or viscerotropic velogenic by intracloacal inoculation of adult chickens. Intracerebral pathogenicity index values for the virulent NDV isolates ranged from 1.54 to 1.90, compared to a possible maximum value of 2.0. These isolates had a dibasic amino acid motif in the fusion protein cleavage site sequence required for host systemic replication. Sequence differences were detected surrounding the fusion protein cleavage site and the matrix protein nuclear localization signal, indicating evolution of highly virulent NDV. Phylogenetically, these isolates were categorized with other highly virulent NDV strains that caused outbreaks in southern California poultry during 1972 and in cormorants in the north central United States and southern Canada during 1990 and 1992. These isolates are related to NDV that may have the APMV1 strain chicken/Australia/AV/32 or a related virus as a possible progenitor. Recent virulent NDV isolates and those recovered during disease outbreaks since the 1970s are phylogenetically distinct from current vaccine viruses and standard challenge strains.     

The epidemiology of Newcastle disease in rural poultry: A review

Rural poultry are the dominant form of poultry kept in the developing world. They are a natural resource whose potential is not fully exploited for the welfare of rural populations. The most devastating disease of rural poultry is Newcastle disease (ND). ND is an epidemic disease in intensive poultry and is responsible for high economic losses. Its epidemiology in intensive poultry is well understood, but little has been published on its behaviour in village poultry populations. Some research has been carried out during the past few years and it now appears that rural poultry are receiving increasing support for research and development from many government and international funding agencies throughout the world. All strains of Newcastle disease virus (NDV) occur in rural poultry, but velogenic strains are reported to be more common. Serological surveys in conjunction with isolation studies have shown that velogenic NDV strains are endemic in rural poultry populations even in isolated villages and possibly in isolated flocks. Although NDV is endemic in village poultry, the clinical disease usually follows an epidemic pattern. ND outbreaks often occur once or twice a year at regular intervals affirming the endemicity of the virus, however, 'mini' outbreaks in individual flocks and sporadic cases in individual birds may occur. Epidemics usually occur at times of climatic stress, leading to seasonal occurrence. The spread of NDV within and between village poultry populations is relatively slow due to a low contact rate. The major mode of transmission appears to be by the faecal-oral route. The respiratory route may also play a role in flocks where close bird-to-bird associations exist. Other poultry species, wild and feral birds, wild animals, communal water reservoirs and domestic animals may play a role in transmission; however, their role has not been properly investigated. In non-immune, intensively reared poultry, introduction of a sufficient quantity of virulent NDV is sufficient to cause severe outbreaks, but in rural poultry epidemic disease depends upon a number of factors. These include NDV pathotype, age structure and immunity of the host population, breed susceptibility, concurrent disease and seasonal influences. Although ND is endemic in rural poultry, many aspects of the epidemiology and economics of ND have not been fully understood. Well-designed, longitudinal studies made within a socio-economic framework are required to improve our understanding of the main problems of rural poultry systems.     

The long view: a selective review of 40 years of Newcastle disease research

This review is written for the series celebrating the 40th year since the first issue of Avian Pathology. The aim of the authors was to cover the developments in Newcastle disease (ND) research over the last 40 years that they considered significant. During those 40 years there have been several panzootics of this serious disease in poultry and for the last 30 years there has been a continuing panzootic in domestic pigeons, which has spread to wild birds and poultry. The 40-year period began with worldwide outbreaks of severe ND, which served as an important impetus for ND research work. Although early work was concerned with controlling the disease, specifically by improving and developing new vaccines and vaccine regimens, even prior to the 1970s ND virus was seen as a useful laboratory virus for replication and virulence studies. This review covers the historical developments in the following areas: understanding the molecular basis of virulence; epidemiology and relatedness of different ND strains, both antigenically and genetically; the emergence of virulent strains and their relationship with viruses of low virulence; sequencing and understanding the viral genome and genes; the development of rapid molecular-based diagnostic tests; and the phylogeny and molecular taxonomy of ND virus. The authors suggest areas in which future research could or should be undertaken.     

Effect of phylogenetic diversity of velogenic Newcastle disease virus challenge on virus shedding post homologous and heterologous DNA vaccination in chickens

Newcastle disease (ND) is a highly devastating disease for the poultry industry as it causes high economic losses. In this present study, a DNA vaccine containing the F and HN surface antigens of a highly virulent Newcastle disease virus (NDV), NDV/1/Chicken/2005 (FJ939313), was successfully generated. Cell transfection test indicated that the vaccine expressed the F and HN genes in Hep-2 cells. The main objective of this study was to compare the extent of protection induced by DNA vaccination after homologous and heterologous NDV-challenge as determined by the amount of NDV shedding after challenge. NDV-antibody-negative chickens were vaccinated either once, twice or thrice intramuscularly at 7, 14 and 21 days old and were challenged 14 days post vaccination with either homologous virus (vaccine-matched velogenic viscerotropic Newcastle disease virus (vvNDV) strain, FJ939313), phylogenetically related to group VII, or a phylogenetically divergent heterologous virus (unmatched vvNDV strain, AY968809), which belongs to genogroup VI and shows 84.1% nucleotide similarity to the NDV-sequences of the DNA vaccine. Our data indicate that birds, which received a single dose of the DNA vaccine were poorly protected, and only 30–40% of these birds survived after challenge with high virus shedding titre. Multiple administration of the DNA vaccine induced high protection rates of 70–90% with reduced virus shedding compared to the non-vaccinated and challenged birds. Generally, homologous challenge led to reduced tracheal and cloacal shedding compared to the heterologous vvNDV strain. This study provides a promising approach for the control of ND in chickens using DNA vaccines, which are phylogenetically closely related to the circulating field strains.     

Generation of a velogenic Newcastle disease virus from cDNA and expression of the green fluorescent protein

Summary Newcastle disease virus (NDV) is a pathogen that is important in the poultry industry worldwide. Specifically, the virulent (velogenic) NDV is aparticular threat because it has now occurred frequently worldwide. The outbreaks caused by highly virulent NDV in waterfowl and especially in goose flocks, have led to greater concern in recent years as aquatic birds were previously resistant to most virulent NDV strains from chickens. The molecular determinants of host tropism, virulence and emergence of NDV isolated from diseased goose flocks are poorly understood. In the present study, we rescued a highly virulent NDV isolated from a goose using the reverse genetics approach. Infectious virus was successfully generated by cotransfection of a full-length cDNA clone of the NDV strain ZJ1 with helper plasmids. The recombinant NDV was indistinguishable from the parental wild-type virus with respect to its growth kinetics in cell culture as well as its biological properties. A recombinant NDV expressing green fluorescent protein (GFP) was generated, and GFP was subsequently detected in cells and various organs from the infected chickens.     

Enhanced phylogenetic resolution of Newcastle disease outbreaks using complete viral genome sequences from formalin-fixed paraffin-embedded tissue samples

Virus Genes - Highly virulent Newcastle disease virus (NDV) causes Newcastle disease (ND), which is a threat to poultry production worldwide. Effective disease management requires approaches to...     

Complete genome and clinicopathological characterization of a virulent Newcastle disease virus isolate from South America

Newcastle disease (ND) is one of the most important diseases of poultry, negatively affecting poultry production worldwide. The disease is caused by Newcastle disease virus (NDV) or avian paramyxovirus type 1 (APMV-1), a negative-sense single-stranded RNA virus of the genus Avulavirus, family Paramyxoviridae. Although all NDV isolates characterized to date belong to a single serotype of APMV-1, significant genetic diversity has been described between different NDV isolates. Here we present the complete genome sequence and the clinicopathological characterization of a virulent Newcastle disease virus isolate (NDV-Peru/08) obtained from poultry during an outbreak of ND in Peru in 2008. Phylogenetic reconstruction and analysis of the evolutionary distances between NDV-Peru/08 and other isolates representing established NDV genotypes revealed the existence of large genomic and amino differences that clearly distinguish this isolate from viruses of typical NDV genotypes. Although NDV-Peru/08 is a genetically distinct virus, pathogenesis studies conducted with chickens revealed that NDV-Peru/08 infection results in clinical signs characteristic of velogenic viscerotropic NDV strains. Additionally, vaccination studies have shown that an inactivated NDV-LaSota/46 vaccine conferred full protection from NDV-Peru/08-induced clinical disease and mortality. This represents the first complete characterization of a virulent NDV isolate from South America.     

Newcastle disease virus: An evolving pathogen?

Australia experienced outbreaks of virulent Newcastle disease (ND) in chickens in the state of New South Wales in the years 1998, 1999 and 2000. The disease had occurred previously in Australia in 1930 and 1932 but the country was free of it until the recent outbreaks. Avirulent strains of Newcastle disease virus (NDV) were detected in 1966 and, during the next two to three decades, strains (so-called lentogenic strains) able to induce mild respiratory disease equivalent to that induced by vaccine strains such as LaSota were also detected. Nucleotide sequence analysis of the genes encoding the haemagglutinin and fusion proteins of Australian isolates of the virus during this time demonstrated that Australian chicken strains of NDV could be differentiated from NDV isolated elsewhere. Analysis in this way demonstrated that NDV isolates causing the recent outbreaks of virulent disease were Australian viruses that were so closely related to a recognized Australian lentogenic strain, termed the Peat's Ridge strain, that it was considered to be the precursor of the virulent virus. The outbreaks of virulent disease in 1998 and 1999 were controlled by an official "stamping out" eradication campaign. This was subsequently replaced by strategic use of ND vaccines when virulent virus was again detected on some farms that had been restocked following depopulation. The national situation with regard to ND is now being assessed through a structured national survey of ND viruses, particularly to determine the distribution of the precursor strain. No new outbreaks of virulent ND have been recognized since February 2000, although immunization of flocks in areas where the disease was recognized has occurred.     

Six-year surveillance of Newcastle disease virus in wild birds in north-eastern Spain (Catalonia)

Given that Newcastle disease (ND) is one of the major threats for the poultry industry, testing of Newcastle disease virus (NDV) has been carried out since 2010 in cases of mortality in wild birds (passive surveillance) in Catalonia. The objective is to provide an early warning system to prevent the infection of poultry. Since 2010, 35 episodes of mortality in wild birds were attributed to NDV infection. Throughout this period there was a progressive expansion of NDV to new areas, with an increase in the episodes of mortality, although it is not clear whether they were the result of the spread of the virus, or of the improvement of the surveillance. Phylogenetic analyses indicate that two distinct sublineages of NDV, 4a and 4b, were circulating in Catalonia. Both sublineages seem to be endemic in the wild bird population, affecting mainly Eurasian-collared doves, with a clear pattern in relation to its spatial distribution (coincident with the distribution of this species), and its temporal distribution (with the majority of cases between September and February). So far, endemicity in wild birds has not resulted in ND outbreaks in poultry. However, there are still many uncertainties about, for example, whether NDV may expand to new areas of Catalonia (with higher poultry density), or about the threat that the apparently more novel sublineage 4a may represent. Hence, efforts should be made so that measures to prevent infection of poultry farms (particularly in high-risk areas and periods) are encouraged, and surveillance is maintained.     

Identification of a natural multi-recombinant of Newcastle disease virus

Newcastle disease (ND), caused by ND virus (NDV), is one of the most serious illnesses of birds, particularly chickens, and has been one of the major causes of economic losses in the poultry industry. Live vaccines are widely used to prevent chicken from NDV all over the world. Given the implications that recombination has for RNA virus evolution, it is clearly important to determine the extent to which recombination plays a role in NDV evolution. In this study, we performed the phylogenetic and recombination analysis on complete NDV genomes. A natural multi-recombinant cockatoo/Indonesia/14698/90 (AY562985) was identified. Its two minor parental-like strains might be from the NDV vaccine lineage and anhinga/U.S.(Fl)/44083/93 lineage, respectively. Our study suggests that recombination plays a role in NDV evolution. Especially, the study also suggests that live vaccines have capacity to play roles in shaping NDV evolution by homologous recombination with circulating virus.     

Molecular Epidemiology of Outbreak-Associated and Wild-Waterfowl-Derived Newcastle Disease Virus Strains in Finland,Including a Novel Class I Genotype

Newcastle disease (ND) is a highly contagious, severe disease of poultry caused by pathogenic strains of Newcastle disease virus (NDV; or avian paramyxovirus-1). NDV is endemic in wild birds worldwide and one of the economically most important poultry pathogens. Most of the published strains are outbreak-associated strains, while the apathogenic NDV strains that occur in wild birds, posing a constant threat to poultry with their capability to convert into more virulent forms, have remained less studied. We screened for NDV RNA in cloacal and oropharyngeal samples from wild waterfowl in Finland during the years 2006 to 2010: 39 of 715 birds were positive (prevalence, 5.5%). The partial or full-length F genes of 37 strains were sequenced for phylogenetic purposes. We also characterized viruses derived from three NDV outbreaks in Finland and discuss the relationships between these outbreak-associated and the wild-bird-associated strains. We found that all waterfowl NDV isolates were lentogenic strains of class I or class II genotype I. We also isolated a genetically distinct class I strain (teal/Finland/13111/2008) grouping phylogenetically together with only strain HIECK87191, isolated in Northern Ireland in 1987. Together they seem to form a novel class I genotype genetically differing from other known NDVs by at least 12%.     

Separate Evolution of Virulent Newcastle Disease Viruses from Mexico and Central America

An outbreak of Newcastle disease (ND) in poultry was reported in Belize in 2008. The characteristics of three virulent Newcastle disease virus (NDV) isolates from this outbreak (NDV-Belize-3/08, NDV-Belize-4/08, and NDV-Belize-12/08) were assessed by genomic analysis and by clinicopathological characterization in specific-pathogen-free (SPF) chickens. The results showed that all three strains belong to NDV genotype V and are virulent, as assessed by the intracerebral pathogenicity index and the polybasic amino acid sequence at the fusion protein cleavage site. In 4-week-old SPF chickens, NDV-Belize-3/08 behaved as a typical velogenic viscerotropic NDV strain, causing severe necrohemorrhagic lesions in the lymphoid organs, with systemic virus distribution. Phylogenetic analysis of multiple NDV genotype V representatives revealed that genotype V can be divided into three subgenotypes, namely, Va, Vb, and Vc, and that all tested Belizean isolates belong to subgenotype Vb. Furthermore, these isolates are nearly identical to a 2007 isolate from Honduras and appear to have evolved separately from other contemporary viruses circulating in Mexico, clustering into a new clade within NDV subgenotype Vb.     

Newcastle disease in the European Union 2000 to 2009

Newcastle disease (ND) is a devastating disease of poultry that has to some extent been neglected by those working in the field in the past 10 to 15 years while attention has been focused on the emergence and spread of highly pathogenic avian influenza caused by a H5N1 subtype virus. During 2000 to 2009 in the European Union (EU) member states, ND viruses virulent for chickens have been detected in wild birds, domesticated pigeons and poultry. Based on these isolations it appears that the epizootic in racing pigeons caused by the variant viruses termed pigeon avian paramyxovirus type 1, which form the genetic group 4b(VIb) first seen in Europe in 1981, continued during 2000 to 2009, and the virus is probably enzootic in racing pigeons in some EU countries. This virus appears to have spread regularly to wild birds, especially those of the Columbidae family, and has been the cause of significant outbreaks in poultry. Other avian paramyxovirus type 1 viruses responsible for ND outbreaks in the EU during 2000 to 2009 have been those from genetic groups 5b(VIIb) and 5d(VIId). There is evidence that the former may well represent spread from a wild bird source and these viruses have also been isolated from wild birds, while the latter represents continuing spread from the East. Future legislation or recommendations aimed at the control and eradication of ND will need to encompass these three sources of virulent ND viruses.     

Use of a heteroduplex mobility assay to detect differences in the fusion protein cleavage site coding sequence among Newcastle disease virus isolates

Newcastle disease virus (NDV) is an economically important pathogen of poultry that may cause clinical disease that ranges from a mild respiratory syndrome to a virulent form with high mortality, depending on an isolate's pathotype. Infections with virulent NDV strains are required to be reported by member nations to the Office of International Epizootes (OIE). The primary determinant for virulence among NDV isolates is the presence or absence of dibasic amino acids in the fusion (F) protein cleavage activation site. Along with biological virulence determinations as the definitive tests, OIE accepts reporting of the F protein cleavage site sequence of NDV isolates as a virulence criterion. Nucleotide sequence data for many NDV isolates recently isolated from infected chickens and other avian species worldwide have been deposited in GenBank. Consequently, viral genomic information surrounding the F protein cleavage site coding sequence was used to develop a heteroduplex mobility assay (HMA) to aid in further identification of molecular markers as predictors of NDV virulence. Using common vaccine strains as a reference, we were able to distinguish virulent viruses among NDV isolates that correlated with phylogenetic analysis of the nucleotide sequence. This technique was also used to examine NDV isolates not previously characterized. We were able to distinguish vaccine-like viruses from other isolates potentially virulent for chickens. This technique will help improve international harmonization of veterinary biologics as set forth by the OIE and the Veterinary International Cooperation on Harmonization of Technical Requirements of Veterinary Medicinal Products. Ultimately, the HMA could be used for initial screening among a large number of isolates and rapid identification of potentially virulent NDV that continue to threaten commercial poultry worldwide.     

Emergence of a virulent genotype VIIi of Newcastle disease virus in Iran

Newcastle disease (ND) is a contagious viral disease affecting numerous avian species, particularly domestic poultry, and causes devastating outbreaks. In spite of its endemicity and importance in Iran, data on the genetic characterization of ND virus (NDV) are scarce. An alarming issue that has just been raised is the occurrence of ND outbreaks with unexpected high mortality and severe clinical signs. The present study was conducted to characterize the emerging NDV genetically. An NDV strain, isolated in 2017 from commercial broilers showing severe nervous and enteric signs, was completely sequenced and found to be 15,192 nucleotides in length. The phylogenetic analysis demonstrated that the virus belonged to subgenotype VIIi, a subgenotype with potential panzootic features which has recently emerged in the Middle East and Asia. The supporting genetic pattern obtained from the complete genome, fusion and haemagglutinin gene analysis showed close relationship of the isolate with Pakistani VIIi NDVs. The analysis of the F protein showed a polybasic amino acid motif and a phenylalanine at position 117 at the cleavage site, which is a characteristic of virulent strains. The isolate showed significant differences from the previously characterized NDV strains from commercial and rural chickens in Iran. This may describe the importance of the illegal trade of pet birds from neighbouring countries leading to the emergence of new genotypes. This study introduces a newly emerging NDV VIIi subgenotype in Iran. This investigation emphasizes the necessity of effective control strategies.     

Sequencing and analysis of the complete genome of Newcastle disease virus isolated from a commercial poultry farm in 2010

Newcastle disease virus (NDV) infects wild and domestic birds but causes contagious and lethal disease in domestic poultry. ND is currently endemic in Pakistan, but no complete genome sequence of a Pakistani NDV isolate has been reported. An NDV strain isolated from a commercial poultry farm was completely sequenced. Phylogenetic analysis revealed that the isolate is closely related to genotype VII and, more specifically, to subgenotype VIIb, yet with substantial enough differences to be regarded as new subgenotype (VIIf). These findings provide insight into the genetic nature of NDV circulating in Pakistan and are useful for both laboratory diagnosis and vaccine development for NDV.