伊氏锥虫HGPRT缺陷株驯化及生物学特性观察

Induced Cultivation and Biological Characters of HGPRT deficient Trypansoma evans i

为了获得伊氏锥虫次黄嘌呤鸟嘌呤磷酸核糖转化酶（ＨＧＰＲＴ）缺陷株，用不同浓度的８－氮鸟嘌呤（８－ＡＧ）和８－ＡＧ加紫外线照射的方法，对伊氏锥虫体外培养株进行了为期６０～１０１ｄ诱导驯化。结果，以每毫升含８－ＡＧ４０μｇ的培养液培养驯化，再配合紫外线照射，以及以每毫升含８－ＡＧ６０μｇ的培养液培养驯化，均获得了伊氏锥虫ＨＧＰＲＴ缺陷株。试验表明，培养虫体经过２～３个死亡期后逐步进入适应期，虫数升高到（１．５～２．５）×１０６／ｍＬ，饲养细胞通常于２～５ｄ变黑、崩解，应适时更换。培养驯化的１３株克隆虫株，经６－巯基鸟嘌呤（６－ＴＧ）抗性测定，１０株存活。克隆株在３倍量氨基喋呤的ＨＡＴ选择性培养液中死亡。生物学试验表明，ＨＧＰＲＴ缺陷株在无ＨＴ的培养液中生长良好，对小鼠致病力有下隆的趋势，虫血症和小鼠死亡时间比对照组延长１倍多。抗体凝集试验表明，ＨＧＰＲＴ缺陷株未发生抗原变异现象。超微结构显示，ＨＧＰＲＴ缺陷株的动基体、核和核仁比对照组明显增大，核膜电子密度区也明显增宽。

A HGPRT deficient Trypanosoma evansi strain (T. evansi/HGPRT -) was obtained after T. evansi in vitro culture strain had been induced with different doses of 8 AG and ultraviolet rays for 60 101 days. The results showed that the cultivated parasites had gradually adapted to the induced environment after 2 to 3 time dead periods. When the parasites reached up to (1.5 2.5)×10 6/mL the feeder layer cells got dark and disolved in 2 5 days. The parasites of 40 mg/L and 60 mg/L 8 AG groups survived by determination of anti 6 TG and then ten cloning strains were obtained respectively. The cloning strains would be died in HAT selective medium of triple aminopurine. The biological test proved that T. evansi/HGPRT - could proliferate in the medium without HT and its pathogenicity to mice declined with the parasitemia and death of the inoculated mice to be delayed double than the control group. The antibody agglutination reaction showed that the antigenity of T. evansi/HGPRT - had not been changed. The ultramicroscopic structure proved that the size of kinetoplast, nucleus, nucleolus and electron dense of nuclear membrane of the parasite were large obviousely than that of the control.