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Molecular characterization of full genome sequences of Newcastle disease viruses circulating among vaccinated chickens in Egypt during 2011-2013.

ABSTRACT: Although intensive vaccination programs have been implemented, Newcastle disease (ND) outbreaks, accompanied by severe economic losses, are still reported in Egypt. The genetic characterization of ND virus (NDV) strains isolated from ND-vaccinated chicken flocks provides essential information for improving ND control strategies. Therefore, here, 38 NDV strains were isolated and identified from outbreaks among vaccinated flocks of broiler chickens located in the provinces of Qena, Luxor, and Aswan of Upper Egypt during 2011-2013. The investigated broiler chicken flocks (aged 28 to 40 days) had high mortality rates of up to 80%. All NDV isolates were genetically analyzed using next-generation DNA sequencing. From these isolates, 10 representative NDV strains were selected for further genetic analyses. Phylogenetic analysis of full-length coding genes revealed that the Egyptian NDV isolates belonged to a single sub-genotype, VII.1.1. These isolates were phylogenetically distant from the vaccine strains, including La Sota or Clone 30 (genotype II), which have been commonly used to vaccinate chicken flocks. Amino acid substitution K78R was observed in the neutralizing epitopes of the F proteins; whereas several mutations were found in the neutralizing epitopes of the hemagglutinin-neuraminidase proteins, notably, E347K. Overall, our results suggested that the occurrence of neutralizing epitope variants may be one of potential reasons for ND outbreaks. Further studies are needed to determine the protective effect of current vaccines against circulating virulent NDV strains.

SUBMITTER: Tran GTH

PROVIDER: S-EPMC7324829 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The aim of this study was to optimise and evaluate an intracellular cytokine staining (ICS) assay for assessment of T cell IFN-γ responses in chickens vaccinated against Newcastle disease (ND). We aimed to validate currently available antibodies to chicken IFN-γ using transfected CHO cells. Moreover, this ICS assay was evaluated for use to detect mitogen and antigen induced IFN-γ production in chicken peripheral blood leucocytes. Chickens from an inbred white leghorn line containing two MHC haplotypes, B19 and B21, were divided into three experimental groups; one group was kept as naive controls, one group was vaccinated intramuscularly twice with a commercial inactivated ND virus (NDV) vaccine, and the last group was vaccinated orally twice with a commercial live attenuated NDV vaccine. PBMC were ex vivo stimulated with ConA or with NDV antigen. The ICS assay was used to determine the phenotype and frequency of IFN-γ positive cells. ConA stimulation induced extensive IFN-γ production in both CD3+TCRγδ+ (γδ T cells) cells and CD3+TCRγδ- cells (αβ T cells), but no significant differences were observed between the experimental groups. Furthermore, a large proportion of the IFN-γ producing cells were CD3- indicating that other cells than classic T cells, secreted this cytokine. NDV antigen stimulation induced IFN-γ production but to a lower extent than ConA and with a large variation between individuals. The CD3+TCR1γδ-CD8α+ (CTL) population produced the highest NDV specific IFN-γ responses, with significantly elevated levels of IFN-γ producing cells in the B19 chickens vaccinated orally with live attenuated NDV vaccine. This was not the case in the B21 animals, indicating a haplotype restricted variation. In contrast, the CD3+TCR1γδ-CD4+ (Th) population did not show a significant increase in IFN-γ production in NDV stimulated samples which was in part due to a high number of IFN-γ producing cells after incubation with medium alone. In conclusion, an ICS assay for phenotyping of IFN-γ producing chicken leukocytes was set up that proved useful in identifying cytokine producing cells upon either mitogen or antigen-specific stimulation.

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Molecular characterization of full genome sequences of Newcastle disease viruses circulating among vaccinated chickens in Egypt during 2011-2013.

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