A New Real-Time PCR Assay for Improved Detection of the Parasite Babesia microti

Babesiosis is an emerging zoonosis with important public health implications, as the incidence of the disease has risen dramatically over the past decade. Because the current gold standard for detection of Babesia is microscopic examination of blood smears, accurate identification requires trained personnel. Species in the genus cannot be distinguished microscopically, and Babesia can also be confused with the early trophozoite stage (ring forms) of Plasmodium parasites. To allow more accurate diagnosis in a format that is accessible to a wider variety of laboratories, we developed a real-time PCR assay targeting the 18S rRNA gene of Babesia microti, the dominant babesiosis pathogen in the United States. The real-time PCR is performed on DNA extracted from whole-blood specimens and detects Babesia microti with a limit of detection of similar to 100 gene copies in 5 mu l of blood. The real-time PCR assay was shown to be 100% specific when tested against a panel of 24 organisms consisting of Babesia microti, other Babesia species, Plasmodium species, tick-borne and other pathogenic bacteria, and other blood-borne parasites. The results using clinical specimens show that the assay can detect infections of lower parasitemia than can be detected by microscopic examination. This method is therefore a rapid, sensitive, and accurate method for detection of Babesia microti in patient specimens.