CONTINUOUS IN-VITRO CULTIVATION OF ERYTHROCYTIC STAGES OF BABESIA-EQUI

The protozoan parasite Babesia egui, a causative agent of equine pirop lasmosis, was continuously cultivated in horse erythrocytes. The paras ites were isolated from a carrier horse at a time when no parasite was detected in a thin blood smear. The culture medium consisted of modif ied medium 199 supplemented with 40% non-heat-inactivated horse serum in a humidified atmosphere containing 5% CO2, 2% O-2, and 93% N-2 at 3 7 degrees C. parasites were detected after 2 days in culture, When the percentage of parasitized erythrocytes (PPE) reached 1%, the cultures were transferred into a humidified atmosphere of 5% CO2 in air. After 7 days the cultures were split at a ratio of 1:2, and after another 5 days they were split at a ratio of 1:4. From them on, cultures were s plit at a ratio of 1:4 routinely at 2-day intervals. The PPE ranged be tween 10% and 25%. Supplementation with hypoxanthine was essential for the initiation and propagation of cultures. In established cultures, hypo xanthine could be replaced by equimolar concentrations of adenosi ne or guanosine. Parasites from cultures could be cryopreserved and re suscitated. Cultures were maintained for more than 300 days.