Duplex Reverse-Transcription Real-Time Polymerase Chain Reaction Assay Targeting 23S rRNA Single Nucleotide Polymorphisms for the Detection of Flea-Borne Rickettsioses

Flea-borne spotted fever and flea-borne (murine) typhus are rickettsioses caused by Rickettsia felis and Rickettsia typhi , respectively, and typically present as undifferentiated febrile illnesses. The relative contribution of these agents to flea-borne rickettsioses in California is unclear. We have developed a duplex reverse transcription real-time polymerase chain reaction (RT-rtPCR) assay targeting R. felis – and R. typhi –specific 23S ribosomal RNA single nucleotide polymorphisms to better understand the respective roles of these agents in causing flea-borne rickettsioses in California. This assay was compared with an established duplex R. felis – and R. typhi–ompB rt-PCR assay and was shown to have 1,000-fold and 10-fold greater analytical sensitivity for the detection of R. felis and R. typhi , respectively. Retrospective testing of clinical specimens with both assays established R. typhi as the major etiologic agent of flea-borne rickettsioses in California.