THEILERIA-ANNULATA - IN-VITRO CULTIVATION OF SCHIZONT-INFECTED BOVINELYMPHOCYTES

Experimental parameters for optimization of the in vitro growth condit ions for Theileria annulata schizont-infected bovine lymphoid cells ar e presented, Of the nine different media tested in the course of 14 su ccessive passages, Leibovitz L-15 (L-15) and a combination of McCoy an d Leibovitz L-15 (ML) were preferable to McCoy, Dulbecco (DMEM), RPMI 1640 or Eagle's minimum essential medium (MEM) based on either Hank's or Earle's salts. The lowest multiplication rate was obtained with M-1 99 medium based on Hank's or Earle's salts. The highest yield of cells was obtained when L-15 was supplemented with 20% newborn bovine serum , while lowering the serum concentration by half resulted in a 25% dec rease in cell yield, There was no effect on multiplication rate observ ed during ten passages when 2-mercaptoethanol and a mixture of oxaloac etate, sodium pyruvate and insulin were added to the growth medium. On substitution of conditioned medium for 20 or 50% of the growth medium , the yield of cells decreased by 12 or 20%, respectively, a factor wh ich might be considered in calculations of the cost of anti-T. annulat a vaccine production. When cells were grown in stationary cultures in Roux bottles for 7 days without change of medium, the highest yield re sulted from seeding at 10(7) cells per vessel (100 ml) in L-15 supplem ented with 20% serum, In Roller bottles, with the same type and volume of medium and cultivation for 7 days without medium change, best yiel d resulted from seeding with the highest inoculum size of 4 x 10(7) ce lls per vessel.