Showing papers on "Newcastle disease published in 2012"

Preparation and efficacy of a live newcastle disease virus vaccine encapsulated in chitosan nanoparticles

Abstract: Background Newcastle disease (ND) is a highly contagious viral disease of poultry caused by pathogenic strains of the Newcastle disease virus (NDV). Live NDV vaccines are administered by drinking water, eyedrops or coarse aerosol spray. To further enhance mucosal immune responses, chitosan nanoparticles were developed for the mucosal delivery of a live NDV vaccine. Methodology/Principal Findings A lentogenic live-virus vaccine (strain LaSota) against NDV encapsulated in chitosan nanoparticles were developed using an ionic crosslinking method. Chitosan nanoparticles containing the lentogenic live-virus vaccine against NDV (NDV-CS-NPs) were produced with good morphology, high stability, a mean diameter of 371.1 nm, an encapsulation rate of 77% and a zeta potential of +2.84 mV. The Western blotting analysis showed that NDV structural proteins were detected in NDV-CS-NPs. The virus release assay results of NDV-CS-NPs indicated that NDV was released from NDV-CS-NPs. Chickens immunized orally or intranasally with NDV-CS-NPs were fully protected whereas one out of five chickens immunized with the LaSota live NDV vaccine and three out of five chickens immunized with the inactivated NDV vaccine were dead after challenge with the highly virulent NDV strain F48E9. Conclusions/Significance NDV-CS-NPs induced better protection of immunized specific pathogen free chickens compared to the live NDV vaccine strain LaSota and the inactivated NDV vaccine. This study lays a foundation for the further development of mucosal vaccines and drugs encapsulated in chitosan nanoparticles.

In vitro characterization of the antiviral activity of fucoidan from Cladosiphon okamuranus against Newcastle Disease Virus

Abstract: Background Newcastle Disease Virus (NDV) causes a serious infectious disease in birds that results in severe losses in the worldwide poultry industry. Despite vaccination, NDV outbreaks have increased the necessity of alternative prevention and control measures. Several recent studies focused on antiviral compounds obtained from natural resources. Many extracts from marine organisms have been isolated and tested for pharmacological purposes, and their antiviral activity has been demonstrated in vitro and in vivo. Fucoidan is a sulfated polysaccharide present in the cell wall matrix of brown algae that has been demonstrated to inhibit certain enveloped viruses with low toxicity. This study evaluated the potential antiviral activity and the mechanism of action of fucoidan from Cladosiphon okamuranus against NDV in the Vero cell line.

Co-infection of broilers with Ornithobacterium rhinotracheale and H9N2 avian influenza virus

Abstract: Since 2008, a progressive pneumonia has become prevalent in broilers and laying hens. This disease occurrs the first day after hatching and lasts more than 30 days, resulting in approximately 70% morbidity and 30% mortality in broilers. The objective of this study was to isolate and identify the pathogens that are responsible for the progressive pneumonia and establish an animal model for drug screening. 193 serum samples were collected from 8 intensive farms from 5 provinces in China and analysed in the current research. Our clinical survey showed that 65.2% to 100% of breeding broilers, breeding layers, broilers and laying hens were seropositive for ORT antibodies. From 8 intensive farms, six ORT isolates were identified by PCR and biochemical assays, and two H9N2 viruses were isolated. Newcastle Disease Virus (NDV) and Infectious BronchitisVirus (IBV) were excluded. Typical pneumonia and airsacculitis were observed both in broilers inoculated intraperitoneally with an ORT isolate alone and in those co-infected with ORT and H9N2 virus isolates. Specifically, the survival rate was 30%, 20%, 70%, 50% and 90% in birds inoculated with ORT+H9N2 virus, ORT followed by H9N2 virus, H9N2 virus followed by ORT, and ORT or H9N2 virus alone, respectively. The results of this study suggest that ORT infections of domestic poultry have been occurring frequently in China. ORT infection can induce higher economic losses and mortality if H9N2 AIV is also present. Although the isolation of ORT and H9N2 virus has been reported previously, there have been no reported co-infections of poultry with these two pathogens. This is the first report of co-infection of broilers with ORT and H9N2 virus, and this co-infection is probably associated with the outbreak of broiler airsacculitis in China, which has caused extensive economic losses.

Complete Genome and Clinicopathological Characterization of a Virulent Newcastle Disease Virus Isolate from South America

Abstract: 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.

Generation by Reverse Genetics of an Effective, Stable, Live-Attenuated Newcastle Disease Virus Vaccine Based on a Currently Circulating, Highly Virulent Indonesian Strain

Abstract: Newcastle disease virus (NDV) can cause severe disease in chickens. Although NDV vaccines exist, there are frequent reports of outbreaks in vaccinated chickens. During 2009–2010, despite intense vaccination, NDV caused major outbreaks among commercial poultry farms in Indonesia. These outbreaks raised concern regarding the protective immunity of current vaccines against circulating virulent strains in Indonesia. In this study, we investigated whether a recombinant attenuated Indonesian NDV strain could provide better protection against prevalent Indonesian viruses. A reverse genetics system for the highly virulent NDV strain Banjarmasin/010/10 (Ban/010) isolated in Indonesia in 2010 was constructed. The Ban/010 virus is classified in genotype VII of class II NDV, which is genetically distinct from the commercial vaccine strains B1 and LaSota, which belong to genotype II, and shares only 89 and 87% amino acid identity for the protective antigens F and HN, respectively. A mutant virus, named Ban/AF, was developed in which the virulent F protein cleavage site motif “RRQKR↓F” was modified to an avirulent motif “GRQGR↓L” by three amino acid substitutions (underlined). The Ban/AF vaccine virus did not produce syncytia or plaques in cell culture, even in the presence of added protease. Pathogenicity tests showed that Ban/AF was completely avirulent. Ban/AF replicated efficiently during 10 consecutive passages in chickens and remained genetically stable. Serological analysis showed that Ban/AF induced higher neutralization and hemagglutination inhibition antibody titers against the prevalent viruses than the commercial vaccines B1 or LaSota. Both Ban/AF and commercial vaccines provided protection against clinical disease and mortality after challenge with virulent NDV strain Ban/010 (genotype VII) or GB Texas (genotype II). However, Ban/AF significantly reduced challenge virus shedding from the vaccinated birds compared to B1 vaccine. These results suggest that Ban/AF can provide better protection than commercial vaccines and is a promising vaccine candidate against NDV strains circulating in Indonesia.

Biological and Phylogenetic Characterization of a Genotype VII Newcastle Disease Virus from Venezuela: Efficacy of Field Vaccination

Abstract: Here we report the biological and molecular characterization of a virulent genotype VII Newcastle disease virus (NDV) circulating in Venezuela and the assessment of the vaccination efficacy under field conditions compared to controlled rearing conditions. Biological pathotyping showed a mean embryo dead time of 50 h and an intracerebral pathogenicity index of 1.86. Sequence-based phylogenetic analysis demonstrated that the virus belongs to genotype VII in class II (a genotype often found in Asia and Africa), representing the first report of the presence of this genotype in the continent of South America. A vaccine-challenge trial in commercial broilers reared in fields or in a experimental setting included dual (live/killed) priming of 1-day-old chicks plus two live NDV and infectious bursal disease virus (IBDV) field vaccinations at days 7 and 17, followed by a very stringent genotype VII NDV challenge at day 28. Serology for NDV and IBDV, bursal integrity, and protection against NDV lethal challenge were assessed. At 28 days, field vaccinates showed significantly lower NDV (1,356 versus 2,384) and higher IBD (7,295 versus 1,489) enzyme-linked immunosorbent assay (ELISA) antibody titers than the experimentally reared birds. A lower bursal size and bursa-body weight ratio (P < 0.05) and higher bursa lesion score were also detected in the field set. Only 57.1% of field vaccinates survived the lethal challenge, differing (P < 0.05) from 90.5% survival in the experimental farm. Overall, results confirmed the presence of the genotype VII viruses in South America and suggest that field-associated factors such as immunosuppression compromise the efficacy of the vaccination protocols implemented.

Advancement in Vaccination Against Newcastle Disease: Recombinant HVT NDV Provides High Clinical Protection and Reduces Challenge Virus Shedding with the Absence of Vaccine Reactions

Abstract: SUMMARY. Newcastle disease (ND) is a highly contagious disease of chickens causing significant economic losses worldwide. Due to the limitation in their efficacy, current vaccination strategies against ND need improvements. This study aimed to evaluate a new-generation ND vaccine for its efficacy in providing clinical protection and reducing virus shedding after challenge. Broiler chickens were vaccinated in ovo or subcutaneously at hatch with a turkey herpesvirus-based recombinant vaccine (rHVT) expressing a key protective antigen (F glycoprotein) of Newcastle disease virus (NDV). Groups of birds were challenged at 20, 27, and 40 days of age with a genotype V viscerotropic velogenic NDV strain. Protection was 57% and 81%, 100% and 95%, and 100% and 100% after the subsequent challenges in the in ovo and subcutaneously vaccinated chickens, respectively. Humoral immune response to vaccination could be detected from 3–4 wk of age. Challenge virus shedding was lower and gradually decreased over time in the va...

Complete genome sequences of Newcastle disease virus strains circulating in chicken populations of Indonesia.

Abstract: Eight highly virulent Newcastle disease virus (NDV) strains were isolated from vaccinated commercial chickens in Indonesia during outbreaks in 2009 and 2010. The complete genome sequences of two NDV strains and the sequences of the surface protein genes (F and HN) of six other strains were determined. Phylogenetic analysis classified them into two new subgroups of genotype VII in the class II cluster that were genetically distinct from vaccine strains. This is the first report of complete genome sequences of NDV strains isolated from chickens in Indonesia.

Genomic and biological characterization of a velogenic Newcastle disease virus isolated from a healthy backyard poultry flock in 2010

Abstract: Newcastle disease virus (NDV) causes severe and economically important disease in poultry around the globe. None of NDV strains in Pakistan have been completely characterized and the role of rural poultry in harbouring NDV is unclear. Since they have a very important role for long-term circulation of the virus, samples were collected from apparently healthy backyard poultry (BYP) flocks. These samples were biologically analyzed using mean death time (MDT) and intracerebral pathogenicity index (ICPI), whereas genotypically characterized by the real-time PCRs coupled with sequencing of the complete genome. Despite of being non-pathogenic for BYP, the isolate exhibited MDT of 49.6 h in embryonated chicken eggs and an ICPI value of 1.5. The F gene based real-time PCR was positive, whereas M-gene based was negative due to substantial changes in the probe-binding site. The entire genome of the isolate was found to be 15192 nucleotides long and encodes for six genes with an order of 3'-NP-P-M-F-HN-L-5'. The F protein cleavage site, an indicative of pathogenicity, was 112RRQKRF117. Complete genome comparison indicated that the RNA dependent RNA polymerase gene was the most and the phosphoprotein was least conserved gene, among all the genes. The isolate showed an Y526Q substitution in the HN protein, which determines neuraminidase receptor binding and fusion activity of NDV. Phylogenetic analysis, based on F and HN genes, classified this isolate into genotype VII, a predominant genotype responsible for ND outbreaks in Asian countries. However, it clustered well apart from other isolates in this genotype to be considered a new subgenotype (VII-f). These results revealed that this isolate was similar to virulent strains of NDV and was avirulent in BYP either due to resistance of local breeds or due to other factors such as substantial mutations in the HN protein. Furthermore, we have characterized the first isolate of NDV, which could act as domestic reference strain and could help in development and selection of appropriate strain of NDV for vaccine in the country.

Characterization of Live LaSota Vaccine Strain–Induced Protection in Chickens upon Early Challenge with a Virulent Newcastle Disease Virus of Heterologous Genotype

Abstract: SUMMARY. Newcastle disease (ND) is a major threat to the international poultry industry, causing bird mortality, reduction in growth and egg production, and trade restrictions. The primary strategy available to the poultry industry to control virulent Newcastle disease virus (NDV), the causative agent of ND, is vaccination. LaSota and other commonly used live-virus NDV vaccine strains were developed in the 1950s and 1960s and show a great degree of genetic divergence from currently circulating NDV strains. In order to characterize protective immunity induced by LaSota against a heterologous NDV strain, we vaccinated groups of specific-pathogen-free (SPF) chickens with LaSota (virus titers ranging from 10 2 to 10 8 egg infective dose 50 [EID50 ]i n 10-fold increments) and challenged the birds 14 days later with ZJ1 strain, an NDV strain that was isolated in the year 2000 from geese in China. We monitored multiple parameters of immunity, including serum antibody titers, antigen-specific lymphocyte proliferation, and splenic cytokine expression and determined that SPF birds vaccinated with an adequate titer of LaSota strain live vaccine are fully protected from morbidity and mortality due to challenge with ZJ1 strain NDV, and we concluded that in the absence of interfering maternal antibody, protection due to vaccination increases with vaccine titer until a threshold titer is reached, beyond which, little or no further benefit can be elucidated.

Characterization of Newcastle Disease Viruses Isolated from Cormorant and Gull Species in the United States in 2010

Abstract: SUMMARY. Newcastle disease virus (NDV), a member of the genusAvulavirus of the family Paramyxoviridae ,i s the causative agent of Newcastle disease (ND), a highly contagious disease that affects many species of birds and which frequently causes significant economic losses to the poultry industry worldwide. Virulent NDV (vNDV) is exotic in poultry in the United States; however, the virus has been frequently associated with outbreaks of ND in cormorants, which poses a significant threat to poultry species. Here, we present the characterization of 13 NDV isolates obtained from outbreaks of ND affecting cormorants and gulls in the states of Minnesota, Massachusetts, Maine, New Hampshire, and Maryland in 2010. All 2010 isolates are closely related to the viruses that caused the ND outbreaks in Minnesota in 2008, following the new evolutionary trend observed in cormorant NDV isolates since 2005. Similar to the results obtained with the 2008 isolates, the standard United States Department of Agriculture F-gene real-time reverse-transcription PCR (RRT-PCR) assay failed to detect the 2010 cormorant viruses, whereas all viruses were detected by a cormorant-specific F-gene RRTPCR assay. Notably, NDV-positive gulls were captured on the eastern shore of Maryland, which represents a significant geographic expansion of the virus since its emergence in North America. This is the first report of vNDV originating from cormorants isolated from wild birds in Maryland and, notably, the first time that genotype Vv NDV has been isolated from multiple wild bird species in the United States. These findings highlight the need for constant epidemiologic surveillance for NDV in wild bird populations and for consistent biosecurity measures to prevent the introduction of the agent into domestic poultry flocks.

Isolation and detection of newcastle disease virus from field outbreaks in broiler and layer chickens by reverse transcription-polymerase chain reaction.

Abstract: The present research work was carried out to isolate and identify Newcastle disease virus (NDV) by using haemagglutination inhibition (HI) test and reverse transcription-polymerase chain reaction (RT-PCR) assay. A total of 160 clinical (blood, tracheal and cloacal swabs) and post-mortem (brain, lung, colon and spleen) samples were collected from chickens of two field outbreaks of Newcastle disease (ND) in 2006, one in a broiler (Cobb-500) farm of Mymensingh district and other one in a layer (Sonali) farm of Gazipur district. All the samples were inoculated onto 10-day-old embryonated chicken eggs through allantoic sac route and in the chicken embryo fibroblasts (CEFs) cell culture. The allantoic fluid (AF) of the dead embryos and the infected culture fluid (ICF) of the CEF were harvested at 48 and 96 hours of post-infection, respectively. The HI and RT-PCR were employed to detect NDV in tissue homogenates of all the clinical and post-mortem samples as well as laboratory samples (AF and ICF). Among the clinical samples, virus isolation rate was found higher from tracheal swab (90%) compared to those of cloacal swab (85%) and serum (65%). On the other hand, among the four different types of post-mortem samples, virus isolation rate was found higher in spleen (100%) compared to those of lungs (80%), colon (60%), and brain (80%) samples. In CEF cell culture system, the rate of virus isolation from all the aforesaid samples was found 100% with the exception of serum samples. The isolation rate of NDV was higher in CEF culture system (93.8%) compared to that of avian embryos (80%). Among the clinical and post-mortem samples, inoculum of only cloacal swab and colon showed HA and HI activities. The anti-NDV hyperimmune serum revealed complete inhibition of the 4 haemagglutination unit of each isolate of viruses isolated from broiler and layer chickens present in the laboratory samples (AF and ICF). The NDV specific primers used in the direct RT-PCR for genome detection of NDV showed equal sensitivity and specificity with the RNA extracted from the clinical, post-mortem and laboratory samples (AF and ICF) as with the genomic RNA of reference NDV. Higher rate of detection of NDV was recorded with RT-PCR assay than HI test. Therefore, the molecular method (RT-PCR) can be introduced for rapid and confirmatory detection of NDV from any form of outbreak of ND in the field level of Bangladesh. DOI = http://dx.doi.org/10.3329/bjvm.v8i2.9618 Bangl. J. Vet. Med. (2010). 8 (2) : 87–92

Virulent Newcastle disease virus in Nigeria: identification of a new clade of sub-lineage 5f from livebird markets.

Abstract: Newcastle disease (ND), caused by Avian Paramyxovirus Type 1, is a highly contagious and devastating viral disease of poultry of worldwide distribution with an enormous economic impact. Although ND is reported to be endemic in Nigeria, little information exists on the molecular epidemiology and the lineage distribution of the Newcastle disease viruses (NDVs) in the country, especially in the live bird markets (LBMs). Recent studies reported the identification of three unique sub-lineages. namely; 5f, 5g and 5h in West Africa, and sub-lineages 5f and 5g in particular in non-commercial farms in Nigeria. In this study, 33 NDV isolates, which included NDVs recovered from LBMs in Nigeria, during active surveillance from 2007 to 2008 and viruses recovered from outbreaks in backyard and commercial chicken farms within the same period were analysed. Based on determination of the F0 cleavage site amino acid sequence and phylogenetic analysis, the isolates were classified as virulent; 16 strains were identified as sub-lineage 5g and 17 as sub-lineage 5f. Interestingly, 13 strains from the 5f group formed a distinct cluster that was not identified by other groups in similar studies. The close genetic similarities identified, provided evidence for the first time of the epidemiological link between the viruses circulating in the LBMs and those recovered from outbreaks in backyard and commercial chicken farms in Nigeria between 2007 and 2008. The emergence and identification of new sub-lineages provide an insight into the high rate of genetic drift occurring in NDV strains in Nigeria, and raises a lot of concerns about the efficacy of current ND control measures in the country.

Review on major viral diseases of chickens reported in Ethiopia

Abstract: In Ethiopia, the major poultry products come from backyards chickens. But in recent times, more commercialized poultry farms are flourishing having considerable contribution to the supply of poultry products, especially to urban areas. There are also attempts to upgrade the productivity of local chickens through distribution of exotic and cross breeds to the rural areas. These endeavors, however, are hampered from providing the expected benefits due to various constraints, among which viral diseases are of greater concern. Some of the viral diseases are thought to be introduced concurrent with intensification of poultry industry. In addition, the growing numbers of exotic flocks in the backyard system increases the number of birds which are at risk of getting infected with pathogens in the environment. The present review article deals with major viral diseases of chickens, their current status and future challenges to the poultry industry in Ethiopia. Among these, Newcastle disease, infectious bursal disease and Marek’s disease become serious threats to poultry production. Due to limited research activities, the epidemiology and the total economic damage caused by this disease are not fully known. Frequent outbreaks and occurrence of new strains for these viral diseases became a challenge to the juvenile poultry industry in Ethiopia. Key words: Chickens, Ethiopia, infectious bursal disease, Marek’s disease, Newcastle disease, viral diseases.

Evaluating viral interference between Influenza virus and Newcastle disease virus using real-time reverse transcription–polymerase chain reaction in chicken eggs

Abstract: Background Simultaneous and sequential allantoic cavity inoculations of Specific-pathogen-free (SPF) chicken eggs with Influenza virus (AIV) and Newcastle disease virus (NDV) demonstrated that the interaction of AIV and NDV during co-infection was variable. Our research revisited the replication interference potential of AIV and NDV using real-time reverse transcription–polymerase chain reaction (real-time RT-PCR) for AIV and NDV to specifically detect the viral genomes in mixed infections.

Investigation of aqueous extract of Moringa oleifera lam seed for antiviral activity against newcastle disease virus in ovo

Abstract: Investigation was made into the effect of aqueous seed extract of Moringa oleifera against Newcastle Disease Virus (NDV) using an in ovo assay. Phytochemical analysis revealed the presence of pharmacologically active and nutritionally relevant compounds in the seed. Nine-day-old embryonated chicken eggs were divided into ten groups of fives and received various treatments. Groups 1 to 6 received 100 EID50/0.1 ml NDV pre-treated with M. oleifera seed extracts at final concentrations of 250, 200, 100, 50, 25 and 10 mg/ml in that order, controls were included. Embryo survival was observed daily. Allantoic fluid from treated eggs and serum from hatched chicks were collected for spot hemagglutination (HA) and hemagglutination inhibition (HI) tests to detect NDV in the eggs and antibodies against NDV in the hatched chicks respectively. Results showed that embryo survival was directly proportional to increasing extract concentration. Just as increase in extract concentration was directly proportional to virus death and inversely proportional to production of antibody against NDV. The current findings have clearly demonstrated that M. oleifera seed extract has nutritional value as well as strong antiviral activity against NDV in ovo. In vivo trials are needed to validate the use of resin from the tree in controlling Newcastle disease in chickens. Key words: Moringa oleifera, newcastle disease virus, in ovo, phytochemical analysis.

Velogenic Newcastle Disease Virus as an Oncolytic Virotherapeutics: In Vitro Characterization

Abstract: Cancer is one of the killer diseases in humans and needs alternate curative measures despite recent improvement in modern treatment modalities. Oncolytic virotherapy seems to be a promising nonconventional way to treat cancers. Newcastle disease virus (NDV), a poultry virus, is nonpathogenic to human and domestic animals and has a long history of being used in oncotherapy research in several preclinical studies. The ability of NDV to successfully infect and destroy cancer cells is dependent on the strain and the pathotype of the virus. Adaptation of viruses to heterologous hosts without losing its replicative and oncolytic potential is prerequisite for use as cancer virotherapeutics. In the present study, velogenic NDV was adapted for replication in HeLa cells, and its cytotoxic potential was evaluated by observing morphological, biochemical, and nuclear landmarks of apoptosis. Our results indicated that the NDV-induced apoptosis in HeLa cells was dependent on upregulation of TNF-related apoptosis-inducing ligand (TRAIL) and caspases activation. Different determinants of apoptosis evaluated in the present study indicated that this strain could be a promising candidate for cancer therapy in future.

Adjuvant activity of Sargassum pallidum polysaccharides against combined Newcastle disease, infectious bronchitis and avian influenza inactivated vaccines.

Abstract: This study evaluates the effects of Sargassum pallidum polysaccharides (SPP) on the immune responses in a chicken model. The adjuvanticity of Sargassum pallidum polysaccharides in Newcastle disease (ND), infectious bronchitis (IB) and avian influenza (AI) was investigated by examining the antibody titers and lymphocyte proliferation following immunization in chickens. The chickens were administrated combined ND, IB and AI inactivated vaccines containing SPP at 10, 30 and 50 mg/mL, using an oil adjuvant vaccine as a control. The ND, IB and AI antibody titers and the lymphocyte proliferation were enhanced at 30 mg/mL SPP. In conclusion, an appropriate dose of SPP may be a safe and efficacious immune stimulator candidate that is suitable for vaccines to produce early and persistent prophylaxis.

Strong innate immune response and cell death in chicken splenocytes infected with genotype VIId Newcastle disease virus.

Abstract: Background Genotype VIId Newcastle disease virus (NDV) isolates induce more severe damage to lymphoid tissues, especially to the spleen, when compared to virulent viruses of other genotypes. However, the biological basis of the unusual pathological changes remains largely unknown.

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

Abstract: 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%.

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

Abstract: 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.

A Multiplex RT-PCR for the Detection of Astrovirus, Rotavirus, and Reovirus in Turkeys

Abstract: This study was undertaken to develop and validate a multiplex reverse transcription-polymerase chain reaction (mRT-PCR) for simultaneous detection of avian rotavirus, turkey astrovirus-2 (TAstV-2), and avian reovirus. Primers targeting the conserved regions of NSP4 gene of avian rotavirus, polymerase gene of TAstV-2, and S4 gene of avian reovirus were used. The position of bands at 630, 802, and 1120 base pairs on agarose gel confirmed the presence of rotavirus, TAstV-2, and reovirus, respectively. This mRT-PCR was found to be specific as no amplification was observed with avian influenza virus, Newcastle disease virus, turkey coronavirus, avian metapneumovirus, and intestinal contents of uninfected turkey poults. Intestinal contents of poults from flocks suspected of exhibiting "poult enteritis syndrome" were pooled and tested. Of the 120 pooled samples tested, 70% were positive for TAstV-2, 45% for avian rotavirus, and 18% for avian reovirus. These three viruses were detected alone or in different combinations. Of the samples tested, 20% were negative for these three viruses, 38% were positive for a single virus (TAstV or rotavirus or reovirus), and 42% were positive for two or three viruses. This single-tube mRT-PCR assay has the potential to serve as a rapid diagnostic method for the simultaneous detection of the three enteric viruses in turkeys.