DETECTION OF TRITRICHOMONAS-FETUS BY PCR AND DNA ENZYME-IMMUNOASSAY BASED ON RIBOSOMAL-RNA GENE UNIT SEQUENCES

Tritrichomonas foetus is the causative agent of bovine tritrichomonosi s, a sexually transmitted disease leading to infertility and abortion. Diagnosis is hampered by putative contamination of samples with intes tinal or coprophilic trichomonadid protozoa which might be mistaken fo r T. foetus. Therefore, we developed a PCR test optimized for applicab ility in routine diagnosis. Amplification is based upon primers TFR3 a nd TFR4 directed to the rRNA gene units of T. foetus. In order to avoi d potential carryover contamination by products of previous amplificat ion reactions, conditions were adapted to the use of the uracil DNA gl ycosylase system. Furthermore, documentation and interpretation of res ults were facilitated by including a DNA enzyme immunoassay for the de tection of amplification products. Specificity was confirmed with geno mic material from different related trichomonadid protozoa. The high s ensitivity of the test allowed the detection of a single T. foetus org anism in diagnostic culture medium or about 50 parasites per mt of pre putial washing fluid. The present methods are thus proposed as (i) con firmatory tests for microscopic diagnosis following diagnostic in vitr o cultivation and (ii). a direct T. foetus screening test with diagnos tic samples.