Genomic and comparative clinico-pathological assessment of two Pakistani pigeon-derived newcastle disease virus sub-genotypes XXI.1.1 and XXI.1.2 isolated in 2017.

Pigeon paramyxovirus type-1 (PPMV-1) is an antigenic-variant of Newcastle disease virus (NDV) which is associated with infection in Columbidae family. In this study, we isolated two pigeon-derived strains pi/Pak/Lhr/SA_1/17 (designed as SA_1) and pi/Pak/Lhr/SA_2/17 (designed as SA_2) from diseased pigeons collected in Punjab province in 2017. We performed the whole genome, phylogenetic analysis and comparative clinico-pathological evaluation of two viruses in pigeons. Phylogenetic analysis based on fusion (F) gene and complete genome sequences showed that SA_1 belonged to sub-genotype XXI.1.1 and SA_2 clustered in sub-genotype XXI.1.2. SA_1 and SA_2 viruses contributed to morbidity and mortality in pigeons. Remarkably, although the two viruses resulted in comparatively similar pattern of pathogenesis and replication ability in various tissues of infected pigeons, SA_2 could cause more severe histopathological lesions and had comparatively high replication ability in pigeons than SA_1. Moreover, pigeons infected with SA_2 had higher shedding efficiency than that of pigeons infected with SA_1. Moreover, several aa substitutions in the major functional domains of the F and HN proteins might be contributed to the pathogenic differences between the two isolates in pigeons. Overall, these findings provide us with important insight into the epidemiology and evolution of PPMV-1 in Pakistan and laid the foundation for the further elucidation of the mechanism underlying the pathogenic difference of PPMV-1 in pigeons.

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.

Isolation and Characterization of Avian Influenza H9N2 Viruses from Different Avian Species in Pakistan 2016-17.

Avian influenza (AI) virus (AIV) subtype H9N2 continues to cause significant outbreaks among commercial and backyard poultry in Pakistan. Despite this, the characterization of H9N2 viruses in avian hosts other than chickens in Pakistan has not been thoroughly investigated. In this study, 12 low pathogenicity avian influenza viruses subtype H9N2 were isolated from peacocks (n = 4), ducks (n = 4), pheasants (n = 2), geese (n = 1), and black swans (n = 1) in Pakistan during 2016 and were characterized on the basis of the hemagglutinin (HA) and neuraminidase genes. All of the viruses possessed an amino acid substitution Q226L in the receptor-binding site of the HA protein, which is known to contribute to increased viral replication and virulence in mammals. In addition, phylogenetic studies showed that these H9N2 AIVs belonged to the Middle East B genetic group of sublineage G1 and were very similar to viruses isolated from an outbreak in chickens in Pakistan in 2017. This demonstrates an epidemiologic link between poultry and other avian species, which is a fact to consider in future H9N2 disease management programs.

Nota de investigación ­ Aplicación in ovo y al primer día de edad de una vacuna viva contra el virus de la enfermedad infecciosa de la bolsa en pollos de engorde comerciales. La enfermedad infecciosa de la bolsa (IBD) es una enfermedad económicamente importante de pollos jóvenes causada por un Avibirnavirus, el virus de la enfermedad infecciosa de la bolsa (IBDV). El virus causal es altamente resistente a los ambientes avícolas y la vacunación es la medida más efectiva para el control de esta enfermedad. Sin embargo, la presunta neutralización de las cepas altamente atenuadas por los anticuerpos maternos y la supuesta virulencia de las cepas parcialmente atenuadas ha limitado la implementación de cepas de vacunas vivas convencionales contra la enfermedad de Gumboro en pollitos antes o después de la eclosión. Sin embargo, los datos preliminares han generado dudas sobre la validez de este dogma prevaleciente. Para analizar la posible aplicación de una cepa de vacuna viva intermedia plus del virus de Gumboro, cepa MB-1, en embriones de pollo con inmunidad materna y a pollos de un día de edad, se realizaron cuatro ensayos de campo a gran escala en distintos lugares del mundo. Los cuatro ensayos midieron la seguridad relativa, los parámetros de inmunización para Gumboro y los rendimientos de producción de MB-1 frente a las vacunas de Gumboro con complejos inmunes establecidas en una variedad de sistemas comerciales de engorde. El desempeño general de la salud y la producción en los cuatro ensayos ha sido similar o mejor en los grupos con la cepa MB-1. Los resultados desafían la noción prevaleciente de que las cepas vivas de del virus de la enfermedad infecciosa de la bolsa pueden neutralizarse o sobrepasar la inmunidad materna e inducir daño permanente a la respuesta inmune de los pollos de engorde jóvenes. Se ha observado un fenómeno de replicación retardada después de la administración parenteral de la cepa de vacuna viva contra el virus de la enfermedad de Gumboro, sin embargo, este mecanismo de replicación retardada aún no se ha dilucidado. Los resultados de este estudio justifican una mayor investigación de las cepas de vacunas vivas convencionales contra la enfermedad de Gumboro como una alternativa para la inmunización activa de pollos de engorde antes y después de la eclosión.

Immunoprotective role of LaSota vaccine under immunosuppressive conditions in chicken challenged with velogenic avian avulavirus-1.

The first objective of the present study was to evaluate if the antibodies induced by the live LaSota and killed Newcastle disease (sub-genotype VIIi) vaccines protect the chickens against exposure with pathogenic avian avulavirus-1 (AAvV-1) of chicken and/or pigeon origins. The second objective was to study the effect of vaccines on stressed birds (dexamethasone, aflatoxin, and heat stressed) with respect to antibody production and protection against pathogenic AAvV-1 challenge. Sixty-one-day-old Hubbard chickens were divided into six groups (gA-gF) with ten animals each. All the groups received LaSota (105 EID50, 0.1 ml per chick) on days 7 and 27 via eye drop and one intramuscular injection of a killed vaccine (sub-genotype VIIi) (107.5 EID50, 1 ml) on day 18, except the control birds received the PBS only. Moreover, group gC-DEX received dexamethasone intramuscularly at a dose rate of 1-mg/kg body weight daily; gD-AFLA had received aflatoxin as oral gavage at a dose rate of 30 ppb daily, and gE-HEAT was kept under heat stressed (38 °C) till challenged. All the groups were challenged with AAvV-1 strain of chicken origin of sub-genotype VIIi, except the group gA-pigeon was challenged with pigeon-origin strain (sub-genotype VIm). The result showed that the gA-pigeon and gB-chicken vaccinate showed 100% and 80% protection. The immunosuppressive birds produced low pre-challenge HI titer, and protection was observed at 40%, 50%, and 70% in gC-DEX, gD-AFLA, and gE-HEAT, respectively. Our findings suggest the stress factors such as aflatoxin in the feed and dexamethasone are immunosuppressive in nature and suppress the immune response and its associated protective role during infection.