Detection of Brucella melitensis in semen using the polymerase chain reaction assay

An evaluation of the polymerase chain reaction (PCR) for detection of Bruce lla melitensis DNA in bovine and ovine semen was performed. Since semen con tains different components that inhibit PCR amplification, a protocol was u sed to purify Brucella-DNA from bovine and ovine semen samples prior to con ducting amplification of the targeted DNA. When separated fractions of natu rally Brucella contaminated semen were analyzed by the PCR, most of B. meli tensis DNA were present in the seminal fluid and non-sperm fractions. The PCR examination results for detection of B. melitensis DNA in different semen fractions were compared with the results for traditional cultural me thods of Brucella from semen. The PCR was more sensitive than the tradition al cultural methods since it detected Brucella-DNA in 12 (10%) out of 120 s emen samples while direct culture detected only 7 (5.8%) in the same semen samples. The limit of detection by PCR was 100 CFU/ml of semen. In addition , the results of PCR were available in one day, whereas isolation and ident ification of Brucella organisms required days or even weeks. The PCR may be used as a supplementary test for detection of B. melitensis in semen. (C) 2001 Elsevier Science B.V. All rights reserved.