Newcastle disease (ND) is considered to be one of the most economically significant avian viral diseases. It has a worldwide distribution and a continuous diversity of genotypes. Despite its limited zoonotic potential, Newcastle disease virus (NDV) outbreaks in Egypt occur frequently and result in serious economic losses in the poultry industry. In this study, we investigated and characterized NDV in wild cattle egrets and house sparrows. Fifty cattle egrets and fifty house sparrows were collected from the vicinity of chicken farms in Kafrelsheikh Governorate, Egypt, which has a history of NDV infection. Lung, spleen, and brain tissue samples were pooled from each bird and screened for NDV by real-time reverse transcriptase polymerase chain reaction (RRT-PCR) and reverse transcriptase polymerase chain reaction (RT-PCR) to amplify the 370 bp NDV F gene fragment. NDV was detected by RRT-PCR in 22 of 50 (44%) cattle egrets and 13 of 50 (26%) house sparrows, while the conventional RT-PCR detected NDV in 18 of 50 (36%) cattle egrets and 10 of 50 (20%) of house sparrows. Phylogenic analysis revealed that the NDV strains identified in the present study are closely related to other Egyptian class II, sub-genotype VII.1.1 NDV strains from GenBank, having 99.7%–98.5% identity. The pathogenicity of the wild-bird-origin NDV sub-genotype VII.1.1 NDV strains were assessed by experimental inoculation of identified strains (KFS-Motobas-2, KFS-Elhamoul-1, and KFS-Elhamoul-3) in 28-day-old specific-pathogen-free (SPF) Cobb chickens. The clinical signs and post-mortem changes of velogenic NDV genotype VII (GVII) were observed in inoculated chickens 3 to 7 days post-inoculation, with 67.5%–70% mortality rates. NDV was detected in all NDV-inoculated chickens by RRT-PCR and RT-PCR at 3, 7, and 10 days post-inoculation. The histopathological findings of the experimentally infected chickens showed marked pulmonary congestion and pneumonia associated with complete bronchial stenosis. The spleen showed histocytic cell proliferation with marked lymphoid depletion, while the brain had malacia and diffuse gliosis. These findings provide interesting data about the characterization of NDV in wild birds from Egypt and add to our understanding of their possible role in the transmission dynamics of the disease in Egypt. Further research is needed to explore the role of other species of wild birds in the epidemiology of this disease and to compare the strains circulating in wild birds with those found in poultry.

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Newcastle disease virus (NDV) remains a constant threat to the poultry industry. There is scarce information concerning the pathogenicity and genetic characteristics of the circulating velogenic Newcastle disease virus (NDV) in Egypt. In the present work, NDV was screened from tracheal swabs collected from several broiler chicken farms (N = 12) in Dakahlia Governorate, Egypt. Real-time reverse transcriptase polymerase chain reaction (RRT-PCR) was used for screening of velogenic and mesogenic NDV strains through targeting F gene fragment amplification, followed by sequencing of the resulting PCR products. The identified strain, namely, NDV-CH-EGYPT-F42-DAKAHLIA-2019, was isolated and titrated in the allantoic cavity of 10 day old specific pathogen-free (SPF) embryonated chicken eggs (ECEs), and then their virulence was determined by mean death time (MDT) and intracerebral pathogenicity index (ICPI). The pathogenicity of the identified velogenic NDV strain was also assessed in 28 day old chickens using different inoculation routes as follows: intraocular, choanal slit, intranasal routes, and a combination of both intranasal and intraocular routes. In addition, sera were collected 5 and 10 days post inoculation (pi) for the detection of NDV antibodies by hemagglutination inhibition test (HI), and tissue samples from different organs were collected for histopathological and immunohistochemical examination. A series of different clinical signs and postmortem lesions were recorded with the various routes. Interestingly, histopathology and immunohistochemistry for NDV nucleoprotein displayed widespread systemic distribution. The intensity of viral nucleoprotein immunolabeling was detected within different cells including the epithelial and endothelium lining, as well as macrophages. The onset, distribution, and severity of the observed lesions were remarkably different between various inoculation routes. Collectively, a time-course comparative pathogenesis study of NDV infection demonstrated the role of different routes in the pathogenicity of NDV. The intranasal challenge was associated with a prominent increase in NDV lesions, whereas the choanal slit route was the route least accompanied by severe NDV pathological findings. Clearly, the present findings might be helpful for implementation of proper vaccination strategies against NDV.

Avian influenza virus (AIV) remains one of the enzootic zoonotic diseases that challenges the poultry industry in Egypt. In the present study, a total of 500 tissue samples were collected from 100 chicken farms (broilers and layers) suspected to be infected with AIV through the period from 2009 to 2019 from Dakahlia governorate, Egypt. These samples were pooled in 100 working samples and screened for AIV then the positive samples were subjected to histopathological examination combined with real time-polymerase chain reaction (RRT-PCR). RRT-PCR positive samples were also subjected to conventional reverse transcriptase-polymerase chain reaction (RT-PCR) for detection of H5 AIV and some of these resulting positive samples were sequenced for detection of the molecular nature of the studied virus. Interestingly, the histopathological examination revealed necrotic liver with leukocytic infiltration with degenerative changes with necrotic pancreatitis, edema, and intense lymphoid depletion of splenic tissue and hyperplastic tracheal epithelium. Likewise, edema and congested sub mucosal blood vessels and intense bronchial necrosis with hyalinized wall vascular wall and heterophils infiltration were reported. Pneumonic areas with intense leukocytic aggregation mainly and vasculitis of the pulmonary blood vessels were also detected in lung. Collectively, these significant pathological changes in examined tissues cohered with AIV infection. Regarding the molecular characterization, 66 samples were positive for AIV by RRT-PCR and 52 of them were positive for H5 AIV by RT-PCR. The phylogenetic analysis revealed that the H5 viruses identified in this study were aligned with other Egyptian H5N1 AIVs in the Egyptian sub clade 2.2.1, while some of the identified strains were aligned with other Egyptian H5N8 strains in the new Egyptian sub clade 2.3.4.4. Taken together, our present findings emphasize the wide spread of AIV in Egypt and the importance of developing an efficient surveillance and periodical screening program for controlling such disease of public health concern.

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Bovine papillomatosis is a viral disease of cattle causing cutaneous warts. A diagnosis of this viral infection is very mandatory for combating the resulting economic losses. Given the limited data available about bovine papillomavirus (BPV) in Egypt, the present study involved the molecular diagnosis of bovine papillomavirus type-1 (BPV-1), -2, -4, -5, and -10 in cattle presenting cutaneous warts on the head and neck from New Valley Province, Egypt. The phylogenetic analysis of the detected types of BPV was also performed, followed by developing a point-of-need molecular assay for the rapid identification of identified BPV types. In this regard, a total of 308 cattle from private farms in Egypt were clinically examined, of which 13 animals presented cutaneous warts due to suspected BPV infection. The symptomatic animals were treated surgically, and biopsies from skin lesions were collected for BPV-1, -2, -4, -5, and -10 molecular identification using polymerase chain reaction (PCR). The presence of BPV-1 DNA was confirmed in 11 collected samples (84.6%), while BPV-2, -4, -5, and -10 were not detected. Sequencing of the PCR products suggested the Egyptian virus is closely related to BPV found in India. An isothermal nucleic acid amplification test (NAAT) with labeled primers specific for the BPV-1 L1 gene sequence, and based on recombinase polymerase amplification (RPA), in combination with a lateral flow strip assay for the detection of RPA products, was developed and tested. The point-of-need molecular assay demonstrated a diagnostic utility comparable to PCR-based testing. Taken together, the present study provides interesting molecular data related to the occurrence of BPV-1 in Egypt and reveals the genetic relatedness of the Egyptian BPV-1 with BPV-1 found in buffalo in India. In addition, a simple, low-cost combined test was also validated for diagnosis of the infection. The present study suggests the necessity of future investigations about the circulating strains of the virus among the cattle in Egypt to assess their genetic relatedness and better understand the epidemiological pattern of the disease.

Avipoxviruses (APVs) are among the most complex viruses that infect a wide range of birds’ species. The infection by APVs is often associated with breathing and swallowing difficulties, reduced growth, decreased egg production, and high mortalities in domestic poultry. In the present study, 200 cutaneous nodular samples were collected from different avian species (chicken, pigeon, turkey, and canary) suspected to be infected with APVs from Dakahlia Governorate, Egypt. Pooled samples (n = 40) were prepared and inoculated in embryonated chicken eggs (ECEs). APVs were then identified by polymerase chain reaction (PCR) and sequence analysis of the APV P4b gene. Furthermore, the forty strains of APVs were screened for the presence of reticuloendotheliosis virus (REV)-5’LTR in their genomes. Interestingly, the phylogenic tree of the APV P4b gene was separated into 2 clades: clade 1, in which our fowlpox virus (FWPV), turkeypox virus (TKPV), and canarypox virus (CNPV) isolates were grouped, along with reference FWPVs and TKPVs retrieved from GenBank, whereas, in clade2, the pigeonpox virus (PGPV) isolate was grouped with PGPVs retrieved from GenBank. Likewise, REV-5’LTR was amplified from 30 strains isolated from chicken, turkey, and canary, while PGPV strains were free from REV-5’LTR integration. To the best of our knowledge, this study involved the detection and characterization of REV-5’LTR insertions in the APVs field isolates in Egypt for the first time. Given the above information, further future research seems recommended to understand the impact of the resulting REV-5’LTR insertions on the pathogenesis, virulence, and inadequate vaccine protection against APVs.

In the present study, a total of 90 cutaneous lesions samples were collected from chickens, pigeons, and turkeys farms in Dakahlia Governorate, Egypt during summer 2016. These farms suspected to be infected with Avipoxviruses (APVs).Thirty pooled samples were created (10 from chickens, 10 from pigeons and 10 from turkeys). Hyperimmune serum was prepared against standard fowlpox virus in adult white New Zealand rabbits. APV were identified in the collected samples using agar gel precipitation test (AGPT), indirect immunoperoxidase, and polymerase chain reaction (PCR) based on 4b gene of APVs. The results revealed that out of 30 tested samples there were 16 samples (53.3%) tested positive via AGPT including, 6 chicken samples (60%) , 5 pigeon samples (50%) and 5 turkey samples (50%). while using indirect immunoperoxidase, positive results were detected in 23 samples (76.7%) including, 8 chicken samples (80%), 8 pigeon samples (80%) and 7 turkey samples (70%).The 4b gene of APVs was detected using PCR in all tested samples (100%). In conclusion, Indirect immunoperoxidase is superior over AGPT in APVs detection in collected samples from chickens, pigeons and turkeys. PCR could be efficiently used in molecular diagnosis of the virus.