Rapid Identification and Differentiation ofBartonellaSpecies Using a Single-Step PCR Assay

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Abstract
Five species ofBartonellahave been reported to infect humans and cause a variety of diseases that can be difficult to diagnose. Four species ofBartonellahave been reported to infect cats and dogs, and two of these species are considered to be zoonotic pathogens. Diagnosis ofBartonellainfections is hampered by the slow, fastidious growth characteristics ofBartonellaspecies. We report on the development of a single-step PCR-based assay for the detection and differentiation of medically relevantBartonellaspecies. PCR-mediated amplification of the 16S-23S rRNA intergenic region resulted in a product of a unique size for eachBartonellaspecies, thereby allowing differentiation without the necessity of restriction fragment length polymorphism analysis or sequencing of the amplified product. The ability of the single-step PCR assay to differentiate betweenBartonellaspecies was determined with characterized isolates and blood samples from animals known to be infected with eitherBartonella henselae,B. clarridgeiae, orB. vinsoniisubsp.berkhoffii. The sensitivity of the single-step PCR assay relative to that of in vitro culture was determined with blood samples fromB. henselae-infected cats.B. henselaetarget DNA was amplified from 100% of samples with greater than 50 CFU/ml and 80% of samples with 10 to 30 CFU/ml. The single-step assay described in the report expedites PCR-based detection and differentiation of medically relevantBartonellaspecies.