Comparative immunocompetence and interspecies transmission of avian orthoavulavirus-1 in feral birds originating from rural and urban settings.

Feral birds residing close to urban settings exhibit higher immunocompetence against environmental pathogens than their counterparts in rural areas. In this study, we comprehensively evaluated the immunocompetence of five specific feral bird species and investigated the potential for interspecies transmission and pathogenicity of Avian orthoavulavirus-1 (AOAV-1) originating from the Anseriformes order. The immunocompetence assessment involved administering the phytohemagglutinin (PHA) test to individual groups of birds from rural and urban settings, measuring patagium thickness at specific time intervals (12, 24, 36, 48, and 60 h) following the administration of 0.1 mL (1 mg/mL) of PHA. Urban birds displayed significantly enhanced mean swelling responses, particularly urban pigeons, which exhibited a significant difference in patagium thickness at all-time intervals except for 24 h (p = 0.000, p = 0.12). Similarly, rural and urban quails and crows showed substantial differences in patagium thickness at all-time intervals except for 12 h (p = 0.542, p = 0.29). For the assessment of interspecies transmission potential and pathogenicity, each feral bird group was separately housed with naive broiler birds (n = 10 each) and challenged with a velogenic strain of AOAV-1 isolate (Mallard-II/UVAS/Pak/2016) at a dose of 1 mL (108 EID50/mL). Urban birds demonstrated higher resistance to the virus compared to their rural counterparts. These findings highlight the specific immunocompetence of feral bird species and their potential contributions to AOAV-1 transmission and pathogenicity. Continuous monitoring, surveillance, and strict implementation of biosafety and biosecurity measures are crucial for effectively controlling AOAV-1 spillover to the environment and wild bird populations in resource-limited settings, particularly Pakistan.

A comparative evaluation of serum biochemistry profile and antigenic relatedness among velogenic and mesogenic Avian avulavirus 1 infection in chickens and pigeons.

Newcastle disease (ND), caused by virulent Avian avulavirus 1 (AAvV 1), affects variety of avian species around the globe. Several AAvV 1 viruses of different genotypes have recently emerged with varying clinical impacts on their susceptible hosts. Although experimental infection with velogenic and mesogenic strains in chickens and pigeons is well-studied, nevertheless, there exists a paucity of data for comparative variations in serum biochemistry profile of susceptible hosts upon challenge with isolates of varying pathogenicities. With this background, a comparative assessment of a range of serum biochemical parameters was made following challenge with duck-originated velogenic strain (sub-genotype VIIi; MF437287) and pigeon-originated mesogenic strain (sub-genotype VIm; KU885949) in chickens and pigeons. For each of the isolate, commercial broiler chickens and wild pigeons were challenged (10-6.51 EID50/0.1 mL for sub-genotype VIIi and 10-6.87 EID50/0.1 mL sub-genotype Vim) separately via intranasal and intraocular route. Sera were collected on 0, 3rd, 5th, 7th, and 9th day post-infection (dpi), and processed for quantitative analysis of different biochemical parameters. By day 3 post-infection (pi), a substantial decrease (p < 0.0001) in serum alkaline phosphatase (ALP) was observed in chickens and pigeons challenged with velogenic isolate. On the other hand, from day 5 pi and onward, a significant increase (p < 0.001) in serum ALP and total protein concentration was observed exclusively in pigeons challenged with mesogenic isolate. For serum aspartate aminotransferase (AST), a significant increase (p < 0.05) in concentration was observed on day 3 pi which decreased from day 5 pi and onward in pigeons and chickens challenged with mesogenic isolate. Also, to reveal antigenic differences among homologous and heterologous vaccine and field-prevalent strains, cross-hemagglutination inhibition assay demonstrated antigenically diverse nature (R-value < 0.5) of both strains from vaccine strain (LaSota, genotype II). The study concludes antigenic differences among prevalent genotypes than vaccine strain and, although requires further studies to ascertain study outcomes, the serum biochemical profile may facilitate presumptive diagnosis of disease in their susceptible hosts.

Genomic and biological characterization of Newcastle disease viruses isolated from migratory mallards (Anas platyrhynchos).

Given the global evolutionary dynamics of Newcastle disease viruses (NDVs), it is imperative to continue extensive surveillance, routine monitoring and characterization of isolates originating from natural reservoirs (waterfowls). In this report, we isolated and characterized two virulent NDV strains from clinically healthy mallard (Anas platyrhynchos). Both isolates had a genome of 15,192 nucleotides encoding six genes in an order of 3´-NP-P-M-F-HN-L-5´. The biological characteristics (mean death time: 49.5-50 hr, EID50108.5 ml-1) and presence of a typical cleavage site in the fusion (F) protein (112R-R-Q-K-R↓F117) confirmed the velogenic nature of these isolates. Phylogenetic analysis classified both isolates as members of genotype VII within class-II. Furthermore, based upon the hypervariable region of the F gene (375 nt), isolates showed clustering within sub-genotype VIIi. Similarity index and parallel comparison revealed a higher nucleotide divergence from commonly used vaccine strains; LaSota (21%) and Mukteswar (17%). A comparative residues analysis with representative strains of different genotypes, including vaccine strains, revealed a number of substitutions at important structural and functional domains within the F and hemagglutinin-neuraminidase (HN) proteins. Together, the results highlight consistent evolution among circulating NDVs supporting extensive surveillance of the virus in waterfowl to better elucidate epidemiology, evolutionary relationships and their impacts on commercial and backyard poultry.

Infectivity of wild bird-origin avian paramyxovirus serotype 1 and vaccine effectiveness in chickens.

Newcastle disease virus, a prototype avian paramyxovirus serotype 1 (APMV-1), causes economically devastating disease in avian species around the world. Newcastle disease is enzootic in Pakistan and recurrent outbreaks are frequent in multiple avian species even after continuous and extensive use of vaccines. A number of APMV-1 and pigeon paramyxovirus serotype 1 (PPMV-1) strains have been isolated and genetically characterized in recent years. However, the impact of recently characterized wild bird-origin APMVs in domestic poultry, and the potency of routinely used vaccines against these novel and genetically diverse viruses remain unknown. Here, we applied next-generation sequencing for unbiased complete genome characterization of APMV-1 and PPMV-1 strains isolated from clinically diseased peacocks (Pavocristatus) and pigeons (Columbalivia), respectively. Global phylodynamics and evolutionary analysis demonstrates Pigeon/MZS-UVAS-Pak/2014 is clustered into lineage 4 (or genotype VI) and Peacock/MZS-UVAS-Pak/2014 into lineage 5 (or genotype VII). The genomes of both isolates encoded for polybasic residues (112RRQKR↓F117) at the fusion protein cleavage motif along with a number of important substitutions in the surface glycoproteins compared with the vaccine strains. Clinicopathological and immunological investigations in domesticated chickens indicate that these isolates can potentially transmit between tested avian species, can cause systemic infections, and can induce antibodies that are unable to prevent virus shedding. Collectively, the data from these genomic and biological assessments highlight the potential of wild birds in transmitting APMVs to domesticated chickens. The study also demonstrates that the current vaccine regimens are incapable of providing complete protection against wild bird-origin APMVs and PPMVs.

Avian rotavirus enteritis - an updated review.

Rotaviruses (RVs) are among the leading causes of enteritis and diarrhea in a number of mammalian and avian species, and impose colossal loss to livestock and poultry industry globally. Subsequent to detection of rotavirus in mammalian hosts in 1973, avian rotavirus (AvRV) was first reported in turkey poults in USA during 1977 and since then RVs of group A (RVA), D (RVD), F (RVF) and G (RVG) have been identified around the globe. Besides RVA, other AvRV groups (RVD, RVF and RVG) may also contribute to disease. However, their significance has yet to be unraveled. Under field conditions, co-infection of AvRVs occurs with other infectious agents such as astroviruses, enteroviruses, reoviruses, paramyxovirus, adenovirus, Salmonella, Escherichia coli, cryptosporidium and Eimeria species prospering severity of disease outcome. Birds surviving to RV disease predominantly succumb to secondary bacterial infections, mostly E. coli and Salmonella spp. Recent developments in molecular tools including state-of-the-art diagnostics and vaccine development have led to advances in our understanding towards AvRVs. Development of new generation vaccines using immunogenic antigens of AvRV has to be explored and given due importance. Till now, no effective vaccines are available. Although specific as well as sensitive approaches are available to identify and characterize AvRVs, there is still need to have point-of-care detection assays to review disease burden, contemplate new directions for adopting vaccination and follow improvements in public health measures. This review discusses AvRVs, their epidemiology, pathology and pathogenesis, immunity, recent trends in diagnostics, vaccines, therapeutics as well as appropriate prevention and control strategies.

Genetic diversity of Newcastle disease virus in Pakistan: a countrywide perspective.

BACKGROUND: Newcastle disease (ND) is one of the most deadly diseases of poultry around the globe. The disease is endemic in Pakistan and recurrent outbreaks are being reported regularly in wild captive, rural and commercial poultry flocks. Though, efforts have been made to characterize the causative agent in some of parts of the country, the genetic nature of strains circulating throughout Pakistan is currently lacking. MATERIAL AND METHODS: To ascertain the genetics of NDV, 452 blood samples were collected from 113 flocks, originating from all the provinces of Pakistan, showing high mortality (30-80%). The samples represented domesticated poultry (broiler, layer and rural) as well as wild captive birds (pigeons, turkeys, pheasants and peacock). Samples were screened with real-time PCR for both matrix and fusion genes (1792 bp), positive samples were subjected to amplification of full fusion gene and subsequent sequencing and phylogenetic analysis. RESULTS: The deduced amino acid sequence of the fusion protein cleavage site indicated the presence of motif (112RK/RQRR↓F117) typical for velogenic strains of NDV. Phylogenetic analysis of hypervariable region of the fusion gene indicated that all the isolates belong to lineage 5 of NDV except isolates collected from Khyber Pakhtunkhwa (KPK) province. A higher resolution of the phylogenetic analysis of lineage 5 showed the distribution of Pakistani NDV strains to 5b. However, the isolates from KPK belonged to lineage 4c; the first report of such lineage from this province. CONCLUSIONS: Taken together, data indicated the prevalence of multiple lineages of NDV in different poultry population including wild captive birds. Such understanding is crucial to underpin the nature of circulating strains of NDV, their potential for interspecies transmission and disease diagnosis and control strategies.

Genetic analysis of Newcastle disease virus from Punjab, Pakistan.

The strains of Newcastle disease virus (NDV) were isolated from five suspected outbreaks of ND in broiler (n = 3) and layers (n = 2) poultry farms. The egg-isolated viruses were subjected to biological and genetic characterization. Based on the biological characterization, isolates showed haemagglutination titer ≥log 2(7), mean death time <55 h, intracerebral pathogenecity index ≤1.8, and egg lethal dose 50 from 10(-7.15) to 10(-9.31)/1 ml. Genetic characterization of the fusion (F) gene revealed that the isolates clustered with NDV strains of genotype VII (VIIf) within class II, which remained predominant genotype in the domestic poultry of Asia. The deduced amino acid sequence of the isolates confirmed virulent motif (112)RRQKRF(117) at the F protein cleavage site. A bioinformatics and pairwise comparison approach was applied to estimate the synonymous and non-synonymous substitution rates (dN/dS) and selective evolutionary pressure for the F protein. The dN/dS calculated for genotype VII indicate purifying selection, which resulted in a low evolution rate in F gene. The F protein shows a strong negative pressure throughout the length of the protein and no recombination event was determined. Collectively, the results suggest that very similar virulent strains of NDV are involved during current wave of disease outbreak throughout the country. From these results, in conjunction with our recent reports of NDV from Pakistan, it is possible to conclude that emergence of novel group may require revisiting the diagnostics and vaccine control strategies.

Biological characterization and phylogenetic analysis of a novel genetic group of Newcastle disease virus isolated from outbreaks in commercial poultry and from backyard poultry flocks in Pakistan.

Newcastle disease (ND) is a contagious viral disease of many avian species particularly domestic poultry, and is responsible for devastating outbreaks in the poultry industries around the globe. In spite of its importance and endemicity in Southern Asia, data on the genetic nature of the viruses and epizootiological information of the disease is scarce. In this study, six isolates from an emerging wave of ND outbreaks in the north of Pakistan and two isolates from healthy poultry flocks were biologically and genetically characterized. Based on pathogenicity indices such as intracerebral pathogenicity index (ICPI), mean death time (MDT) and cleavage motifs in the fusion protein, all these isolates were classified as virulent. Phylogenetic analysis of the fusion (F), hemagglutinin-neuraminidase (HN) and matrix (M) genes indicated the emergence of a novel genetic group within lineage 5, distinct from isolates previously reported in the region. Several mutations in the neutralizing epitopes and functionally important motifs of the F and HN genes pose a need for re-evaluation of the currently used vaccine and vaccination practices. The characteristics of Newcastle disease virus (NDV) as virulent (F protein cleavage site, ICPI and MDT) in apparently healthy backyard poultry (BYP) explain that BYP can play crucial role in the epizootiology and spread of the disease. The present investigation provides essential information on the genetic nature of NDV circulating in Pakistan and its implication on disease diagnosis and control. Furthermore, these investigations emphasize the importance of continuous surveillance of ND in developing countries.