| Porcine Reproductive and Respiratory Syndrome (PRRS) broke out first in America in 1987, and then went through worldwide quickly with clinical signs of abortion, such as death, mummy and respiratory diseases in pigs with various age. The pathogen of PRRS, porcine reproductive and respiratory syndrome virus (PRRSV), was first detected by Wensvoort in 1991 in Holland, and named Lalystad virus. The virus is one of the members of the artivirus family. The genome of PRRSV is 15kb in length, encoding 9 ORFs. ORFla and ORFlb, the longest ORF, encode the polymerase of PRRSV, ORF2-ORF7 encode the structural gene of PRRSV.The first PRRSV in China was isolated in swine with suspected PRRS in 1995 by Baoqing Guo, confirming that PRRSV had spreaded in China, and then followed by other PRRSV found in most of China. This paper acquired the target gene fragments of the different structural genes by PCR after designing different pairs of primers which were based on the published sequences from Genebank and optimizing the amplifying conditions after acquiring the whole sequence of CH-la strain in order to clarify the source and genetic background of PRRSV from different parts of China, thus providing molecular basis for vaccine research. The sequences of the structural genes of PRRSV from Shandong province and Daqing area were analyzed and compared with those of CH-la strain, BJ-4 strain, VR-2332 and LV strain. Result: The base composition was basically the same among different PRRSV strains in China Among those nucleosides that changed, more than 80% were transformed between A-G and T-C. The results show that all PRRSV strains in China are closely related to the strain VR-2332. Among different isolates in China, ORF6 displayed the most significant changes, while the ORF7 was the most conservative. The obvious CPE could not be seen until the fourth passage when Mark-145 cells were vaccinated with PRRSV field strain, and 70% - 80% cells showed CPE in the sixth passage in only 2~3 days, indicating that PRRSV litres could increase.Localization of PRRSV in vivo and the pathology of PRRSV-infected porcine have been studied extensively, however, no exact criterion has been achieved. Therefore, this study used VR-2332 to inoculate the pregnant swine and piglets. The result indicates that the patient had the symptoms of pulmonary cirrhosis and wide microcirculation malfunction, which may be used as an index for the diagnosis of PRRS. With the indirect immune fluenscence we found that VR-2332 located mainly in immune tissue, such as spleen, and stayed in macrophages and vascular endothelial cells determined by IFA.The dynamic changes of CD4+ and CD8+ T lymphocytes after vaccinated with rivulent and weak VR-2332 strains were studied using flow cytometry. The results was that the virulent strain could not cause significant changes of CD4+ and CD8+ lymphocytes, indicating that rivulent virus did not have obvious effects on the cellular inmmunity. But after the weak virus strain infection the CD4+ and CD8+ T lymphocytes significant changes. It indicated that weak virus had very strong effects on the cellular immunity. The results indicated that this weakIIIVR-2332 strain had distinct immune-regulation to piglets, so the weak virus was the best vaccine against PRRS.In summary, mis experiment finished the analysis of structural genes of PRRSV from different areas of China The determination of the site of PRRSV infection in vivo gives a new way to sample the patient and the immunological changes of the patient were snowed through the detection of cell immunity and humoral immunity after infected by PRRSV, which all belong to American strain. Through monitoring the changes of cell immunity and humoral immunity after infected with PRRSV, we have obtained the immune regular patterns. The results can set biological foundation for vaccine researching, and provide excellent requirement for establishing immediately reasonable immune program and high efficiency, accurately diagnosis methods.
|
PDF Full Text Request