Effect of select medium supplements on in vitro development of Cryptosporidium andersoni in HCT-8 cells.
We evaluated the effect of fetal calf serum (FCS), glucose, ascorbic acid, calcium pantothenate, folic acid, and insulin on the growth of Cryptosporidium andersoni in human colon tumor (HCT-8) cells. After being incubated for 48 h, the proliferation of parasites was determined by real-time polymerase chain reaction (PCR) assay, and the development of C. andersoni was observed by transmission electron microscopy (TEM). Ten percent FCS was the best concentration for C. andersoni culture. Glucose, ascorbic acid, and insulin had a significant effect on the growth of C. andersoni when added into 10% FCS RPMI 1640. Calcium pantothenate had no significant effect and folic acid had the inhibited effect. We also observed the stages of trophozoite, macrogamont, microgamont, type I meront, type II meront, and sporozoite of C. andersoni in HCT-8 cells by TEM. Our results indicated that the best medium for C. andersoni was 10% FCS RPMI 1640 medium containing 50 mM glucose, 50 microg/ml ascorbic acid, and 0.3 U/ml insulin. Real-time PCR could provide a quick and precise technique to determine the proliferation of parasites. Cultivation of C. andersoni in HCT-8 cells will facilitate the study of interactions between parasites and host cells as well as provide a reliable system for evaluating anticryptosporidial compound efficacy.
Wu
Liang
L
School of Medical Science and Laboratory Medicine, Jiangsu University, 212013 Zhenjiang, People's Republic of China.
Chen
Sheng-xia
SX
Jiang
Xu-gan
XG
Shen
Yu-juan
YJ
Lu
Zhao-xi
ZX
Tu
Guo-hua
GH
Fu
Xing-li
XL
Cao
Jian-ping
JP
D000818:Animals; D045744:Cell Line, Tumor; D003458:Cryptosporidium / Q000235:genetics / Q000254:growth & development* / Q000648:ultrastructure; D003470:Culture Media / Q000737:chemistry; D006801:Humans; D046529:Microscopy, Electron, Transmission; D010274:Parasitology / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods
D003470:Culture Media