Introduction and Objectives: The purpose of this study was to evaluate and compare the analytical sensitivity of three primer pairs broadly used (B4/B5, F4/R2 and JPF/JPR) for detection of Brucella genus by PCR, in human and animal serum samples with DNA isolation using boiling method. Material and Methods: Brucella abortus strain S19 was cultured on blood agar with CO2 and was incubated for 3 days. Using sterile distilled water, a cell suspension from bacteria with a concentration equivalent to 3 McFarland (9×108 bacteria per ml) was prepared and then 10 serial dilutions 10-fold (dilutions 100 to 10-9) was prepared. 200 µl of each dilution was poured into separate tubes. Also, 150 µl from suspension with 150 µl of Brucella negative serum was mixed, vortexed, and 5 serial dilutions 10-fold (from 100 to 10-4) was prepared. The serum dilutions were incubated overnight in order to inhibitors in serum completely effect on DNA and then were processed for DNA isolation. Results: Brucella was confirmed by microbiological tests including catalase, oxidase and gram staining. B4/B5 was able to identify dilutions 10-6 of bacterial dilutions, 100 of serum dilutions and 10-4 of dilutions 1/200 resulting DNA serum dilutions. F4/R2 was able to identify dilutions 10-8 of bacterial dilutions, 100 of serum dilutions and 10-3 of dilution 1/200. Finally, JPF/JPR was able to identify dilution 10-3 of bacterial dilutions, 100 of serum dilutions and dilution 10-1 of dilution 1/200. Conclusion: In our study, the number 9 bacteria was detectable in pure culture which according to studies is lesser than 20 fg. Also, in our study, B4/B5 was able to detect 9×108 bacteria in 200 µL of serum.

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