The Virulence Determination And Plaque Cloning Of Different NDV Isolates In Guizhou Province

Newcastle disease (ND) is an acute, highly contiguous infectious disease caused by Newcastle disease virus(NDV). Newcastle disease virus is a member of Rubulaviru, Paramyxovirinae, Paramyxoviridae, and a negative RNA virus. It exists in all tissues, apparatus, body fluid, secretion and excrement.In this paper, the detection of Newcastle disease virus by RT-PCR and virulence determination of different NDV isolates in Guizhou province were carried out. All these improved the detection technology for detecting Newcastle disease virus quickly and accurately.1. The detection by RT-PCR and virulent determination on NDVOn the basis of difference in compositions and sequence of aminoacidat the cleavagesit of F protein of virulent and non-virulent strains of Newcastle disease virus (NDV) three primer pairs A+B,A+C and A+D were designed according to relevant references. The virulence of NDV identified by RT-PCR accorded with those by conventionally biological methods. Furthermore the method of RT-PCR to diagnose ND have sensitive, simple and rapid.RT-PCR was used in detection of clinical case; the results confirmed the results of serology, such as hemagglutination test and hemagglutination inhibition test. The results showed that RT-PCR can be used to determine cases which caused by Newcastle disease virus and determine the virulent in 6 hours; the detection rate was higher than isolation rate with 53.84%. compared with RT-PCR, the progress of traditional methods such as MDT, ICPI and IVPI were trouble and tedious, and we need about 7-10 days to wait the results, at the same time, the possibility cross-contamination still exist in operative procedure. In this experience, RT-PCR was used to avoid the shortcomings effectively.2. Purified of NDV strains by plaque clone technology. Purified of 2 NDV Guizhou isolated strains (GN, ND₉₉) and 2 vaccine strains by plaque clone technology. 12 purified strains were received. The results of RT-PCR detected showed: 7 are the mesogenic virus, and the size of the plaque which they formed was 0.3-1.0 mm; 5 are the Velogenic virus, the plaque size was 1.0-2.0 mm, in which I/1 was 0.4 mm red plaque. In the non-pancreas enzyme or in the magnesium ion situation, low virulent NDV becomes on CEF not to be able to form the plaque, the full strength and medium can form plaques.3. The constructed of virulent spectrum.According to the determination results of conventional virulent index, and in view of the HN protein monoclonal antibody, and RT-PCR to appraisal the NDV strains, the quality synthetic evaluation different scene popular virulent, to construct the Newcastle disease virus isolation virulent spectrum. The quality synthetic evaluation three methods to the different NDV separation virulent determination result as follow:F₄₈E₈ >FW>BY> H₂>ND₉₈> P₂> P₁> P₃>L₂>GN> II > IV > ND₉₉ > ND₈₉...