الفهرس 
IntroductionOnly 14 pages are availabe for public view
 |  | from | 180 |  |  |
| | | from | 180 | | |
Abstract
NDV is an acute disease of chicken causing respiratory, digestive and nervous signs. The Newcastle disease caused by Avian orthovula virus1 that related to orderMononegavirales,family Paramyxovidae, subfamily Avulavirinae and genus Orthoavulavirus.it is an endemic disease and one of the limiting factors for the poultry production in Egypt as it characterized by high mortality and high morbidity rates.This study conducted on a total of 120 clinical samples (24 pooled samples) including (larynx, trachea ,lung, proventriculus , spleen and brain) collected from different farms in Dakahlia Governorate from clinically diseased birds suspected to be infected with Newcastle disease virus. Diseased birds were showing respiratory, digestive and nervous signs with rapid DROP of egg productions in layers.A trial for NDV isolation from collected samples into the allontoic fluid of 10 days old ECEs was conducted and three serial passages were carried out for each sample.The isolated virus was identified by HA, HIT and AGPT.Molecular confirmation of NDV was done by RT-PCR amplification of 362bp NDV F gene fragment from both velogenic and lentogenic strains of NDV. While differentiation between both velogenic and lentogenic strains of NDV was done by RT-PCR amplification 254bp NDV F gene fragment which was amplified from velogenic NDV strains only together with sequencing and phylogenic analysis of 362bp fragment of NDV F gene.The results revealed that:Twenty samples causing dwarfism, congestion and death of some inoculated embryos. Twenty sample gave positive result with HA and the positive result appear as lattice shape due to agglutination of chicken RBCs. Eighteen samples gave positive result with HIT using hyperimmune serum prepared against standard NDV. The positive result appear as button shape due to binding of antibodies with NDV which inhibit the agglutination of RBCs while the other two samples were negative as they may have other HA virus not NDV. Sixteensamples gave positive result with AGPT which appear as clear line of precipitin.Agarose gel electrophoresis of the RT-PCR products obtained from amplification reaction of RNA extracted from third egg passage of the allantoic fluid revealed the positive amplification of 365 bp and 254 bp product fragment of F gene from 18samples, lasota strain gave only 365 bp band but failed to amplify 254 bp fragment of F gene of virulent NDV. Sequencing and phylogenetic analysis of 362bp F gene fragment can confirm the results obtained by RT-PCR by the detection of the NDV genotype. NDV isolated in this study are more closely related to Egyptian velogenic NDV isolates in class II, genotype VII of NDV with 97.00 % to 99.3% identity. Our isolates are also closely related to velogenic NDV isolates in class II, genotype VII NDVs from China, South Korea, Indonesia, Peru and Togo with 95.4 to 83.2% identities. While standard LaSota NDV was more closely related to lentogenic NDV strains in class II, genotype II NDV obtained from GenBank with 100% identity 8- The deduced amino acid sequence of F gene cleavage site of the selected isolates was 112R-R-Q-K-R116at the C terminus of F2 protein and phenyle alanine at residue 117(N-terminus of F1) with exception of K11SE in mans 2 strain. This amino acid sequence is characteristic of virulent NDVs. The deduced amino acid sequence of lasota strain cleavage site was 112G-R-Q-G-R↓L117).at F2-C terminus and leucine at N terminus of F1 protein which is characteristic of lentogenic NDV. 10- Comparing used methods revealed that RT-PCR , HIT and AGPT respectively could be used for laboratory diagnosis of ND in Egypt.