Pathology and Host Immune Response in Pigeons Affected with Newcastle Disease Virus

Abstract

With the intent to study the prevalence and pathology of Newcastle disease in pigeons and backyard fowl flocks, localities around Ranbir Singh Pura and Jammu city were investigated for outbreaks of the disease from March 2021 to September 2022. The disease was suspected in sixteen pigeon (Columba livia domestica) flocks and five flocks of backyard fowl (Gallus gallus domesticus), with 89.66% and 100% morbidity and approximately 81.8% and 91.5% mortality rates, respectively. Chiefly neurological signs in pigeons, and predominantly respiratory and/or enteric clinical signs in fowls were recorded. Gross lesions in pigeon were subdural haemorrhages, meningeal congestion or haemorrhages in the brain stem and lungs but rarely in the proventriculus. In fowls, gross lesions were mainly manifested by vascular derangement, causing haemorrhages on proventriculus and enteric mucosa and corresponding necrotizing enteritis. Haemorrhages were evident in most of the organs of the respiratory tract, while mild congestion of meninges was noticeable in few birds. Microscopic lesions in both pigeons and fowl validated the gross lesions typical of Newcastle Disease, pertaining to the neurotropic form in pigeons and viscerotropic form in fowls. Preliminary diagnosis was also based on Haemagglutination assay showing 78.7% positivity with clinical samples. Confirmation of the disease in all clinical samples and infected allantoic fluids was done using Reverse Transcriptase PCR (RT-PCR) targeting a partial Fusion protein gene. Six isolates, three from each pigeon and fowl ND outbreaks from different geospatial locations of the region were sequenced and allotted accession numbers from GenBank. Sequence comparison of isolates showed five of the six isolates with close homology (99.6-100%) to each other. Deducted amino acid sequence at the Fusion protein cleavage site showed a 112R-R-Q-K-R*F117 velogenic motif for all pigeon and fowl isolates. Biological characterization also showed velogenic pathotypic traits with indicators like Mean Death Time (MDT lt60 hrs), and pronounced cytopathic effect (CPE) in chicken embryo fibroblasts (CEF). Phylogenetic analysis showed that the three pigeon isolates and two fowl isolates clustered within genotype II, and one of the fowl isolate clustered with sub genotype VII.1.1. Experimental trial was conducted in 45 healthy pigeons divided into three groups consisting of pigeons inoculated with genotype II NDV pigeon isolate (Group-I) and genotype VII NDV fowl isolate (Group-II), respectively via intranasal and intraocular route. Control group was mock infected with PBS. Signs, gross and histopathological lesions were recorded on 1st, 3rd and 7th day post infection along with mRNA expression of selected innate immune proteins (Pattern Recognition Receptors( PRR s): TLR-7, TLR-3 and RIG-1), anti-viral inflammatory cytokines/ chemokines (CCL-5, IFN-and#947;, IL-10, IL-6 and IL-1and#946;) and apoptotic factors (BCL-2). Progressive signs and lesions were apparent in both the groups after 3 dpi. Lesions were predominantly associated with nervous system in Group-I birds and respiratory and enteric system in-Group-II birds. In general, increased mRNA expression of PRRs was evident in both groups; and expression of pro-inflammatory cytokines (IL-6, IL-1and#946;, and IFN-and#947;) and chemokines (CCL-5) in the spleen, lung and brain was fairly up-regulated as compared to the mock-infected control birds in both the groups, signifying a robust innate and anti-viral inflammatory response. Delayed expression of BCL-2, an anti-apoptotic factor was evident in lungs while early expression in spleen in both the groups. Strain homology, unique mutations and establishment of experimental infection in heterologous host could suggest the possible cross-transmission potential between avian species and emerging threats of circulating ND viruses in the region. newline