Detection of Theileria annulata in cattle and vector ticks by PCR using the Tams1 gene sequences

A Polymerase Chain Reaction (PCR) and Southern blot hybridization for the d etection of Theileria annulata are described. The PCR used primers amplifyi ng a 785 base-pair fragment of the T. annulata gene which encodes the 30 kD a major merozoite surface antigen, Tams1. The sensitivity of the PCR in bov ine blood was 1 piroplasm in 1 mu l of blood. T. buffeli, T. parva, Babesia bigemina, B. bovis and B. divergens were not detected. The PCR detected do wn to 1 infected acinus/tick in resting and partially fed adult Hyalomma an atolicum anatolicum ticks and was negative for T. lestoquardi and T. equi, which are transmitted by this tick but are not infective to cattle. The spe cificity of the PCR was checked using 30 stocks of T. annulata, all of whic h were detected. Three stocks of T. lestoquardi, 4 of T. equi and 1 each of T. buffeli, T. parva, B. bigemina, B. bovis and B. divergens were used to ascertain there were no cross-reactions. A nested PCR using separate primer s for the first reaction and the same primers for the second reaction detec ted T. annulata to the same sensitivity and specificity in saponin-extracte d DNA samples stored for long periods at - 20 degrees C.