Detection and Identification ofEhrlichia,Borrelia burgdorferiSensu Lato, andBartonellaSpecies in DutchIxodes ricinusTicks

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Abstract
A sensitive and specific PCR hybridization assay was developed for the simultaneous detection and identification ofEhrlichiaandBorrelia burgdorferisensu lato. In separate assays the 16S rRNA gene ofEhrlichiaspecies and the 23S-5S rRNA spacer region ofB. burgdorferisensu lato were amplified and labeled by PCR. These PCR products were used in a reverse line blot hybridization assay in which oligonucleotide probes are covalently linked to a membrane in parallel lines. Hybridization of the samples with the oligonucleotide probes on this membrane enabled the simultaneous detection and identification ofEhrlichia,B. burgdorferi, andBartonellaspecies in 40 different samples. The application of the assay to DNA extracts from 121Ixodes ricinusticks collected from roe deer demonstrated that 45% of these ticks carriedEhrlichiaDNA. More than half of these positive ticks carried species with 16S rRNA gene sequences closely related to those ofE. phagocytophilaand the human granulocytic ehrlichiosis agent. The majority of the other positive ticks were infected with a newly identifiedEhrlichia-like species. In addition, 13% of the ticks were infected with one or moreB. burgdorferigenospecies. In more than 70% of the ticks 16S rRNA gene sequences forBartonellaspecies or other species closely related toBartonellawere found. In five of the ticks bothEhrlichiaandB. burgdorferispecies were detected.