The Study Of Optimal Culture Conditions And The Methods Of Inactivating Infectious Spleen And Kidney Necrosis Virus In Siniperca Chuatsi

The mandarin fish(Siniperca chuatsi), has "freshwater grouper" name in the international market of aquatic products, and is one of the important high-quality varieties of freshwater fish consumption and export in Guangdong Province. The artificial breeding and intensive farming scale of Mandarin fish has developed rapidly in Guangdong Province in recent years.With the rapid development of the artificial breeding and culture, the stocking density of mandarin fish increases. There are many mandarin fish disease, especially since 1994 caused by infectious spleen and kidney necrosis virus(Infectious spleen and kidney necrosis virus, ISKNV). ISKNV infection resulted in an acute and high mortality disease and have caused huge economic losses in Guangdong Province every year, and the mortality rate can reach as high as 60% in some cases. The mandarin fish vaccine is the only effective method to prevent virus disease, despite the economical importance of the fish and significant losses caused by diseases. This study cultivated the brain cells of Siniperca chuatsi with diverse media culture medium like M199, L15, DMEM and MEM at multiple serum concentrations of 10%, 8%, 6%, 4% and 2%. Subsequently, trypsinization is utilized to collect the resulted cells and the number of cells was confirmed while their curves of growth were drawn. Through the curves of growth as well as the cell viability, this study measured the impacts of the diverse conditions on the cellular proliferation for the brain cells of Siniperca chuatsi.The results showed that L-15 was the most suitable medium, 10%fetal bovine serum(FBS) was the best concentration. And the optimum p H was 7.0, the optimum temperature was 28 degrees. The surface of rolling bottle is covered at the third day. Until one week the cells will not fall off. The CPB cells can be applied to spinner flasks production. As the experimental results revealed, ISKNV would be fully inactivated under the condition of 0.1% concentration of BPL for 12 hours. There were no cytopathic effect(CPE) observed in cell culture and the artificial infection test in healthy mandarin fish did not cause any disease.The effect of inactivated vaccine indicates, although the time of the inactivation of BEI might take longer, but the BEI inactivated ISKNV vaccine was highly efficious in vaccinated mandarin fish challenged by injected by ISKNV 3, 21 or 48 days after immunization. The BEI inactivated whole virus vaccine was efficacious 3, 21 days or 48 days after vaccination of mandarin fish, which is expected since BEI does not interfere with the virus structure if a low concentration is used.