title: Evaluation of PCR methods for detection of Brucella strains from culture and tissues.
abstract: The genus Brucella causes significant economic losses due to infertility, abortion, stillbirth or weak calves, and neonatal mortality in livestock. Brucellosis is still a zoonosis of public health importance worldwide. The study was aimed to optimize and evaluate PCR assays used for the diagnosis of Brucella infections. For this aim, several primers and PCR protocols were performed and compared with Brucella cultures and biological material inoculated with Brucella. In PCR assays, genus- or species-specific oligonucleotide primers derived from 16S rRNA sequences (F4/R2, Ba148/928, IS711, BruP6-P7) and OMPs (JPF/JPR, 31ter/sd) of Brucella were used. All primers except for BruP6-P7 detected the DNA from reference Brucella strains and field isolates. In spiked blood, milk, and semen samples, F4-R2 primer-oriented PCR assays detected minimal numbers of Brucella. In spiked serum and fetal stomach content, Ba148/928 primer-oriented PCR assays detected minimal numbers of Brucella. Field samples collected from sheep and cattle were examined by bacteriological methods and optimized PCR assays. Overall, sensitivity of PCR assays was found superior to conventional bacteriological isolation. Brucella DNA was detected in 35.1, 1.1, 24.8, 5.0, and 8.0% of aborted fetus, blood, milk, semen, and serum samples by PCR assays, respectively. In conclusion, PCR assay in optimized conditions was found to be valuable in sensitive and specific detection of Brucella infections of animals.
authors.affiliation: Department of Microbiology, Faculty of Veterinary Medicine, University of Ondokuz Mayıs, Samsun, Turkey. aciftci@omu.edu.tr.
authors.forename: Alper
authors.initials: A
authors.lastname: Çiftci
authors.affiliation: Department of Art and Sciences, Faculty of Biology, University of Dumlupınar, Kütahya, Turkey.
authors.forename: Tuba
authors.initials: T
authors.lastname: İça
authors.affiliation: Department of Microbiology, Faculty of Veterinary Medicine, University of Adnan Menderes, Aydın, Turkey.
authors.forename: Serap
authors.initials: S
authors.lastname: Savaşan
authors.affiliation: Department of Microbiology, Faculty of Veterinary Medicine, University of Ankara, Ankara, Turkey.
authors.forename: Barış
authors.initials: B
authors.lastname: Sareyyüpoğlu
authors.affiliation: Department of Microbiology, Faculty of Veterinary Medicine, University of Ankara, Ankara, Turkey.
authors.forename: Mehmet
authors.initials: M
authors.lastname: Akan
authors.affiliation: Department of Microbiology, Faculty of Veterinary Medicine, University of Ankara, Ankara, Turkey.
authors.forename: Kadir Serdar
authors.initials: KS
authors.lastname: Diker
mesh_terms: D037881:Aborted Fetus / Q000737:chemistry; D000034:Abortion, Veterinary; D000818:Animals; D001431:Bacteriological Techniques; D002002:Brucella / Q000235:genetics / Q000302:isolation & purification*; D002006:Brucellosis / Q000175:diagnosis / Q000662:veterinary*; D002417:Cattle; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis*; D005260:Female; D008892:Milk / Q000737:chemistry; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D011247:Pregnancy; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D012680:Sensitivity and Specificity; D012756:Sheep; D012757:Sheep Diseases / Q000175:diagnosis; D034561:Sheep, Domestic; D013045:Species Specificity; D015047:Zoonoses / Q000175:diagnosis
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
keywords: Brucella; Cattle; Detection; PCR; Sheep