USE OF A NONRADIOACTIVE DNA-PROBE FOR DETECTION OF ANAPLASMA-MARGINALE INFECTION IN-FIELD CATTLE - COMPARISON WITH COMPLEMENT-FIXATION SEROLOGY AND MICROSCOPIC EXAMINATION

A sensitive Anaplasma marginale-specific 409-base pair DNA probe was d eveloped in a previous study for detection of A. marginale infection i n experimentally infected cattle with a test that employed slot-blot a nd in situ hybridization. To test the suitability of the probe to dete ct A. marginale in the blood of naturally infected carrier cattle, slo t-blot hybridization was used to determine the infection rate of A. ma rginale in cattle from 3 geographic areas in Oklahoma. For comparison, blood samples from the same cattle were also examined by light micros copy and were tested by the complement fixation test. For the DNA hybr idization assay, the probe was labeled with digoxigenin 11-dUTP by pol ymerase chain reaction (PCR). DNA was extracted from blood using the Q IAamp blood kit and then applied to a nylon membrane and hybridized wi th the probe. The study herds consisted of 31 beef cows in Harper Coun ty, OK, and 42 and 70 dairy cows from Payne and Pittsburg counties, OK , respectively, In the 3 herds, 80.6%, 92.8%, and 57.1% of the cows we re positive for A. marginale as assessed with the DNA hybridization as say. In contrast, only 25.8% and 2.86% were complement fixation positi ve in 2 herds, and no complement fixation positives were found in 1 he rd. Uncountable parasitemia that was too low to accurately determine ( <0.01%) from 29.0%, 4.8%, and 11.4% of the samples, respectively, was demonstrated by microscopic examination. All samples positive by compl ement fixation and microscopic examination had positive probe reaction s in the DNA hybridization assay. Therefore, the PCR-mediated nonradio active DNA probe described here may be useful in epidemiologic investi gations and in identification of carrier cattle. This assay could be a dapted for use in diagnostic laboratories because it is sensitive, spe cific, nontoxic, quickly executed, and inexpensive.