Newcastle disease (ND) is caused by virulent strains of avian paramyxovirus type 1, also known as Newcastle disease virus (NDV). Despite vaccination, the frequency of reported outbreaks in Ethiopia has increased. From January to June 2022, an active outbreak investigation was conducted in six commercial chicken farms across areas of central Ethiopia to identify the circulating NDV strains. Thirty pooled tissue specimens were collected from chickens suspected of being infected with NDV. A questionnaire survey of farm owners and veterinarians was also carried out to collect information on the farms and the outbreak status. NDV was isolated using specific-pathogen-free (SPF)-embryonated chicken eggs and detected using haemagglutination and the reverse transcriptase–polymerase chain reaction (RT–PCR). The genotype and virulence of field NDV isolates were determined using phylogenetic analysis of fusion (F) protein gene sequences and the mean death time (MDT) test in SPF-embryonated chicken eggs. The questionnaire results revealed that ND caused morbidity (23.1%), mortality (16.3%), case fatality (70.8%), and significant economic losses. Eleven of thirty tissue specimens tested positive for NDV using haemagglutination and RT–PCR. The MDT testing and sequence analysis revealed the presence of virulent NDV classified as genotype VII of class II velogenic pathotype and distinct from locally used vaccine strains (genotype II). The amino acid sequences of the current virulent NDV fusion protein cleavage site motif revealed ¹¹²RRQKR↓F¹¹⁷, unlike the locally used avirulent vaccine strains (¹¹²GRQGR↓L¹¹⁷). The epidemiological data, MDT results, cleavage site sequence, and phylogenetic analysis all indicated that the present NDV isolates were virulent. The four NDV sequences were deposited in GenBank with accession numbers F gene (PP726912-15) and M gene (PP726916-19). The genetic difference between avirulent vaccine strains and circulating virulent NDV could explain the low level of protection provided by locally used vaccines. Further studies are needed to better understand the circulating NDV genotypes in different production systems.

The current study was carried out to develop a chicken embryo fibroblast (CEF) and Vero cell-adapted Newcastle disease virus thermostable strain (NDV-I-2) vaccine and evaluate its efficacy against challenge infection. CEF and Vero cells were infected with the vaccine virus, and serial passage was continued until passage 8 and 15, respectively. A characteristic cytopathic effect (CPE) induced by the virus was noted beginning from passages 3 and 10 in the CEF and Vero cell monolayers at 72 h post-infection. The infectious titer of the adapted virus demonstrated a consistent increase along the passage level. The culture supernatant of CEF and Vero cells adapted to NDV I-2 containing a higher concentration of infectious titer was formulated to generate a trial vaccine. CEF-adapted NDV-I-2, Vero cell-adapted NDV-I-2, commercially available NDV-I-2, and negative control were the designations given to groups one, two, three, and four, respectively. The virus-induced specific antibody titer was determined using a hemagglutination inhibition assay after vaccination of chickens through the intraocular route. The antibody titers measured from CEF, Vero cells, and commercially available NDV-I-2 vaccinated chicks were 75.8 ± 6.8, 50.0 ± 4.8, and 86.5 ± 9.2, respectively. Chickens immunized with the CEF-adapted NDV-I-2 vaccine develop higher antibody levels than those immunized with the Vero cell-adapted virus, and all vaccinated groups showed a significant increase in antibody titers, and 100 % safety was achieved. Chickens vaccinated with the CEF-adapted virus and the commercially available vaccine showed complete protection against challenge infection, while the Vero cell-adapted virus showed 95 % protection against very virulent NDV. All unvaccinated groups had 100 % morbidity and mortality. To sum up, the NDV-I-2 vaccine strain is well adapted to CEF and Vero cells, immunogenic, capable of inducing antibody development, and effectively protects chicks against challenge with the circulating field NDV isolate. Therefore, it is advised that the NDV-I-2 vaccine be produced on an industrial scale using CEF or Vero cell culture in order to overcome the drawbacks of using SPF embryonated eggs.