Quantitative real-time PCR for detection of members of the Ehrlichia phagocytophila genogroup in host animals and Ixodes ricinus ticks

A TaqMan PCR was established for identification and quantitation of members of the Ehrlichia phagocytophila group in experimentally infected cows and in Ixodes ricinus ticks. The TaqMan PCR identified a 106-bp section of the 16S rRNA gene by use of a specific fluorogenic probe and two primers. This technique was specific for members of the E. phagocytophila group, which in clude E. phagocgtophila, Ehrlichia equi, and the agent of human granulocyti c ehrlichiosis. The TaqMan system identified 10 copies of a cloned section of the 16S rRNA gene of E. phagocytophila. The sensitivity and specificity of the TaqMan PCR were similar to those of conventional nested PCR The numb ers of ehrlichiae in leukocytes of the two cows experimentally infected wit h E. phagocytophila were measured daily by TaqMan PCR and had a course simi lar to that of the percentages of infected leukocytes determined daily by L ight microscopy. The prevalence of infected free-living ticks, which were c ollected from areas where bovine ehrlichiosis is endemic and from regions w ith sporadic occurrences of granulocytic ehrlichiosis in dogs and horses, w as identical as determined by nested PCR and TaqMan PCR.