Evaluation of a TaqMan real-time PCR for the detection of Theileria parva in buffalo and cattle
Onderstepoort Veterinary Instititute, Agricultural Research Council, Private Bag X05, Onderstepoort 0110, South Africa; TIB MOLBIOL Syntheselabor GmbH, Eresburgstraße 22-23, D-12103 Berlin, Germany
A real-time PCR assay based on TaqMan probe chemistry was developed for the detection of Theileria parva DNA in blood samples. It uses a Theileria genus-specific PCR primer set and a T. parva-specific probe to amplify and hybridize with a species-specific part of the 18S rRNA gene of the parasite. The test was evaluated using positive and negative reference blood samples and shown to be specific for T. parva. Analytical sensitivity was determined by testing a dilution series of T. parva positive blood. It was shown to be able to detect parasitaemia as low as 2×10 -6%. The Taqman assay results were also compared with that obtained with the real-time hybridization probe PCR assay, which is currently employed as the official test for the diagnosis of T. parva infections in buffalo and cattle and was shown to be equally sensitive. A panel of 1164 field samples was screened using both assays and 164 samples tested positive in both tests, indicating a good correlation. © 2010 Elsevier B.V.
RNA 18S; article; blood sampling; buffalo; cattle; controlled study; diagnostic accuracy; diagnostic test accuracy study; DNA determination; East Coast fever; hybridization polymerase chain reaction; intermethod comparison; nonhuman; nucleotide sequence; parasite identification; polymerase chain reaction; real time polymerase chain reaction; screening test; sensitivity and specificity; Theileria parva; Animals; Buffaloes; Cattle; Cattle Diseases; DNA, Protozoan; Nucleic Acid Hybridization; Parasitemia; Real-Time Polymerase Chain Reaction; RNA, Ribosomal, 18S; Sensitivity and Specificity; Species Specificity; Theileria parva; Theileriasis; Bos; Theileria; Theileria parva