中国水产科学2018，Vol.25Issue(1)：211-219,9.DOI:10.3724/SP.J.1118.2018.17058
大鲵细胞与大鲵虹彩病毒的微载体规模化培养工艺及优化
Technologies for large-scale cultivation of giant salamander cells and iridovirus by the Cytodex 3 microcarrier
摘要
Abstract
The Chinese giant salamander Andrias davidianus, a member of the family Cryptobranchidae, is the largest extant amphibian species in the world, which is valued for food, medicine, and research on animal evolu-tion and biodiversity because of its unique phylogenetic position and physiological features. Within the last decade, Chinese giant salamander farming has increased rapidly in China. However, with the rapid development of Chi-nese giant salamander aquaculture, a severe epizootic recently occurred in cultured Chinese giant salamanders in Hubei, Hunan, Sichuan, Shannxi, and Zhejiang Provinces of China, causing tremendous economic losses. The causative pathogen has been identified as the giant salamander iridovirus (GSIV). At present, no effective methods are available for the control of the disease. Immunoprophylaxis is considered the best measure in controlling viral diseases in aquatic animals, and the large-scale cultivation technology of giant salamander cells and GSIV are of significance for the immunoprophylaxis of the disease. In this study, by using Cytodex 3 microcarriers to culture giant salamander muscle cells (GSM) and GSIV at a large scale, the GSM cell morphology, proliferative charac-terization, and GSIV growth dynamics were investigated. The results showed that the attachment efficacy reached 95% after 10 h of cultivation with intermittent agitation of 30 r/min for 2 min followed by 40 min still culture during the cell attachment period in the Cytodex 3 microcarrier culture system. The optimal culture conditions are as follows: serum concentration 5%, microcarrier concentration 2 g/L, and initial inoculation cell density of 1.2 × 105cells/mL. During the cell growth period, the continuous stirring speed was 25 r/min. Under inverted micros-copy and electron microscopy, the GSM cells on the Cytodex 3 microcarriers were long, spindle-shaped, and well adhered. After infection with GSIV at a multiplicity of infection of 0.5, the GSM cells on Cytodex 3 microcarriers showed the typical cytopathic effect at 48 h post infection, and the highest virus titer (TCID50/mL) of 10–8.50±0.20/mL was reached at 72 h post infection. This study established a solid foundation for further investiga-tion on the large-scale technologies of GSIV vaccine production in the future.关键词
大鲵肌肉细胞(GSM)/大鲵虹彩病毒(GSIV)/Cytodex3/规模化培养/工艺Key words
giant salamander muscle cell line (GSM)/giant salamander iridovirus (GSIV)/Cytodex 3 microcarrier/large-scale cultivation/technologies分类
农业科技引用本文复制引用
贾路路,周勇,马杰,范玉顶,刘文枝,刘学芹,曾令兵..大鲵细胞与大鲵虹彩病毒的微载体规模化培养工艺及优化[J].中国水产科学,2018,25(1):211-219,9.基金项目
公益性行业(农业)科研专项(201203086-05). （农业）
