水牛巴贝斯虫病病原—牛巴贝斯虫体外培养的研究

STUDY ON CONTINUOUS CULTIVATION IN VITRO OF BABESIA BOVIS PARASITIZED IN THE BLOOD OF BUFFALO

本研究采用微气静相培养技术,对一株采自人工感染的去脾脏水牛犊的牛巴贝斯虫(Babesia bovis)进行了80天的连续体外培养·继代26次,累积增殖6.51×10^((?))倍;累积稀释2.19×10^(35)倍。培养24小时红细胞的染虫率为2.63±0.50％;48小时为7.18±1.39％;72小时为20.78±4 52％。最高红细胞染虫率达33.50％。体外培养的牛巴贝斯虫与采自病牛血液内的牛巴贝斯虫形态一致,说明已建立了水牛巴贝斯虫病病原——牛巴贝斯虫的体外培养。 对影响牛巴贝斯虫体外培养的几种因素进行的实验结果表明:培养液pH值显著地影响虫体的繁殖,最适的体外连续培养的pH为7.2;合适的血清浓度为40％;血清灭活后,支持牛巴贝斯虫生长繁殖的能力明显降低;脱纤血分离的血清和自然凝固血析出的血清用于培养的效果相同;RPMI-1640和TCM-199培养液支持牛巴贝斯虫生长繁殖的能力没有差异。

Adapting the technology of microaerophilous stationary phase(MASP) culture developed by Levy and ristic(1980),Babesia bovis, collected from an artificially infected splenectomized buffalo calf,has been continuously cultivated in vitro for 80 days, and subcultured for 26 times.The mean percentages of infected erythrocyte of subculture at 24, 48 and 72 hours were 2.63±0.50,7.18±1.39 and 20.78±4.52 respectively.The maximum infectious rate of erythrocyte was 33.5%. The commulative increase amounted to 6.51×1038 folds and commulative dilution reached to 2.19×1035 times. After several subcultures, culture-derived B. bovis were simillar to form seen in the blood of affected bufallo. It is suggested that the continuous in vitro cultivation of B. bovis in the blood of bufallo has been established, The pH of cultures markedly influenced the growth of the parasites and the optimum pH value of the culture was 7.2; optimum serum ratio in culture was 40% ; serum reduced their ability to support the growth of B. bovis when inactivated , and no difference was found between the serum from defibrinated blood and the serum from coagulated blood. A comparison growth in the culture contained mediums RPMI-1640 or TCM-I99 showed no significant difference.