Invasive Species Compendium

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Abstract

Detection of Theileria annulata in blood samples of carrier cattle by PCR.

Abstract

The detection of Theileria annulata in blood samples obtained from carrier cattle by the polymerase chain reaction (PCR) is reported. The assay used primers specific for the gene encoding the 30 000 MW major merozoite surface antigen of T. annulata. A 721-bp fragment was amplified from blood samples taken monthly from calves experimentally infected with one of 4 different stocks of T. annulata originating in either Mauritania, Portugal, Spain or Turkey. At the end of the experiment, 5 animals carried the infection for 12 months and 2 animals remained infected for 15 months. DNAs from 6 other Theileria species, T. parva, T. mutans, T. sergenti, T. buffeli, T. velifera, and T. taurotragi, were not amplified. Moreover, DNAs from 4 other haemoparasites (Anaplasma centrale, A. marginale, Babesia bovis and B. bigemina) were also not amplified. As a control, primers derived from the small subunit rRNA gene of Theileria spp. amplified a 1.1-kb DNA fragment from all Theileria species examined but not from the other 4 haemoparasites. As few as 2 to 3 parasites/µl of infected blood in a 50-µl sample volume were detected by Southern or microplate hybridization with a T. annulata-specific cDNA probe. In addition, 92 field samples obtained from cattle in Spain were tested; 22% were positive in blood smears, 40% were positive by immunofluorescent antibody test, and 75% were positive for T. annulata by PCR. It is concluded that this method provides a useful diagnostic tool for detecting T. annulata carrier cattle.