الفهرس 
Introduction and aim of the workOnly 14 pages are availabe for public view
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Abstract
In the present study efforts were conducted in order to identify the correlation between some commercially used vaccinal strains and local isolates of IBDV at the molecular level of both antigenic and genomic structure. Briefly both vaccinal strains (D78 and Bursa Vac.) and local isolates, (Local 1 and Local 2) were propagated in vero cell culture for three passages and titrated. Titers were 4.3, 3.6, 3.6, and 6.1 log10
TCIDso/ml for 078, B. V., local1 and Local2 respectively and confirmed by using Dot Immuno-Blot Assay (DIA).
The genomic dsRNA of IBDVs were extracted and analyzed by agarose gel electrophoresis to detect variations in size of the dsRNA genome of IBDV.The sizes determined by agarose gel electrophoresis were similar for the tested strains and were about 3.1 kilo base for segment A and 2.8 kilo base for segment B.
Reverse Transcription/ polymerase Chain Reaction (RT/PCR) was conducted to amplify a 643 base pair fragment covering the hyper variable region of VP2 gene, by analysis of the amplified product using agarose gel electrophoresis. We estimated the amplified fragment to be of the expected size without significant differences between the strains indicating no nucleotides deletion or insertion at this region of the VP2 gene.
Restriction Fragment Length Polymorphism (RFLP) using the restriction enzymes Mbol and BstXI for the PCR product were done to compare the polymorphism of the fragments obtained by digestion using these enzymes.
RFLPs obtained by digestion with BstXI are similar for the different strains giving rise to three fragments of about 330 bp, 200 bp and 113 bp sizes for the vaccinal and local isolates, which indicates the presence of the BstXI restriction site in the tested viruses. The BstXI restriction site correlates with the antigenicity of the IBDV, as BstXI cuts in the position occupied by the amino acid residue 222 which is proline and any mutation in this position lead to change from proline to threonine which means mutation of the classial strains to anti genically variant strains. As the results of RFLPs obtained by BstXI are the same so we can predict that the locally isolated strains are anti genically related to the classical vacc inal strains.
Digestion using Mbol enzyme were identical for the viruses D78, Bursa vac., and Local:! giving rise to three fragments of 456, 124, and 64 base pairs sizes. Oifferent R FLPs using Mhol were ohserved, however, for the Local! isolate giving rise to only two fragments of about 500 bp and 143 bp sizes indicating that the MboI restriction site is present only in one position in the VP2 gene hypervariable region and in a different position giving rise to restriction fragments of lengths different from those obtained for vaccinal strains. This genetic difference can help us in identification of the locall isolate from the vaccinal strain inspite of their antigenic relatedness.
The immuonogcnic relationship between these strains were detected by applying westem blot assay on the fractionated proteins of IBDV s using hyper immune serum against lBDV and the results of western blot showed high correlation with that of the RT/PCR-RFLPs, as all of the proteins of IBDV reacted with IBDV antiserum, indicating that there is no antigenic differences between the locally isolated strains and the tested vaccinal strains.
In conclusion nucleotide sequences differences are the basis for the antigenic differences of IBDV strains and these were confirmed in this study from the correlation between Western Blot results and RFLPs patterns, confirming that the RT/PCR-RFLP assay has the potentiality to identify IBDV strains and serves as a new typing method. If an RFLPs data base on IBDV strains including virulent and vaccinal strains is developed, this information might be useful in the future to quickly identify lBDV strains circulating in commercial chicken flocks but this can be achieved by development of a sequencing data base of the hypervariable region of the VP2 gene of the locally isolated strains and the commercially used vaccinal strains to provide a valuable information on the evolution of these isolates and the molecular mechanisms for antigenic and pathogenic variations. Then one can develop the restriction sites data base which will help us in detection and differentiation of IBDV isolates using RTIPCR-RFLP assay.