Differentiation between Vaccinal and Iranian Virulent Isolates of Newcastle Disease Virus based on F Region Genotyping by HRM Analysis

The purpose of this study is investigate Differentiation Between Vaccinal and Iranian Virulent Isolates of Newcastle Disease Virus based on F Region Genotyping by HRM Analysis. Discrimination of circulating virulent strains of Newcastle Disease Virus (NDV) from low pathogenic and vaccine stains is the basis for implementation of strategies to control and eradication of that aims at the eradication of NDV in poultry. At the present study the applicability of Real time RT-PCR followed High-Resolution Melting-Curve Analysis (HRM) was evaluated for differentiation of pathogenic ND viruses from the live vaccinal strains. Five virulent NDV isolates and 6 live vaccines samples of NDV were used to develop and evaluate the tests. Based on the nucleotide sequence of Fusion (F) gene, 3 primer pairs (B, E and H) were designed and utilized for HRM analysis. The melting temperature of vaccinal strains higher than the virulent isolates were obtained using the B and H primer pairs, but it was not the same as E. Based on the analysis of HRM curves in this study, the B and H primer pairs could better differentiate the vaccinal strains from each other and from virulent isolate than E. As it is found in this study, HRM analysis can differentiate virulent NDVs from live vaccines. In comparison to the existing methods for discrimination of different pathotypes of the ND viruses, the developed test takes a shorter time, needs less infrastructure and facilities and costs significantly less than the other traditional methods like intra cerebral pathogenicity index (ICPI).