PCR DETECTION OF ACUTE EHRLICHIA-CANIS INFECTION IN DOGS

A polymerase chain reaction (PCR)-based detection assay that specifica lly detected Ehrlichia canis in dogs with acute infections was develop ed. A region of the 16S ribosomal RNA gene of E. canis was targeted fo r PCR amplification and chemiluminescent hybridization (CH) with a com plementary internal 287-base pair (bp) oligonucleotide probe. The CH i mproved the PCR assay sensitivity 1,000-fold as compared with visualiz ation on ethidium bromide-stained agarose gels. The PCR assay with CH (PCR/CH) detected as little as 30 fg of E. canis genomic DNA, the equi valent of approximately 150 E. canis organisms. The 495-bp product def ined by the specific primers was not detected when genomic DNA from E. platys, E. chaffeensis, E. risticii, and E. equi were used in the PCR /CH assay. The PCR/CH assay was tested with unfractionated blood sampl es collected from 9 dogs experimentally infected with E. canis. The PC R/CH assay had greater detection sensitivity than did cell culture iso lation (CCI) from infected blood. PCR/CH detected E. canis 7 days prio r to CCI in 4 of 6 experimentally infected dogs. The results obtained with the PCR/CH assay otherwise consistently matched the results obtai ned by CCI. This PCR/CH assay is a rapid, sensitive, and specific meth od for E. canis detection with sensitivity comparable to or exceeding that of CCI. A diagnosis of E. canis using this PCR/CH assay can be ma de in 2 days as compared with 1-4 weeks for CCI. The PCR/CH assay appe ars to be an acceptable alternative or complement to current diagnosti c techniques.