title: Modified real-time PCR for detecting, differentiating, and quantifying Ureaplasma urealyticum and Ureaplasma parvum.
abstract: We evaluated a previously described quantitative real-time PCR (qPCR) for quantifying and differentiating Ureaplasma parvum and U. urealyticum. Because of nonspecific reactions with Staphylococcus aureus DNA in the U. parvum PCR, we developed a modified qPCR and designed new primers. These oligonucleotides eradicated cross-reactions, indicating higher specificity. The detection limits of the qPCR were determined at 1 and 3 colony-forming units/ml for U. parvum and U. urealyticum, respectively. The quantification limits of the assay for both Ureaplasma species ranged from 2.10(6) to 2.10(1) copy numbers per PCR. A total of 300 patient samples obtained from the lower genital tract were tested with this newly designed qPCR assay and compared with culture results. Of the samples, 132 (44.0%) were culture positive, whereas 151 (50.3%) tested positive using qPCR. The U. parvum and U. urealyticum species were present in 79.5% and 12.6% of the qPCR-positive samples, respectively. Both species were found in 7.9% of those samples. Quantification of U. parvum and U. urealyticum in the samples ranged from less than 2.5 × 10(3) to 7.4 × 10(7) copies per specimen. In conclusion, the modified qPCR is a suitable method for rapid detection, differentiation, and quantification of U. parvum and U. urealyticum.
authors.affiliation: Department of Microbiology and Infection Control, Universitair Ziekenhuis Brussel, Brussels, Belgium. ellen.vancutsem@uzbrussel.be
authors.forename: Ellen
authors.initials: E
authors.lastname: Vancutsem
authors.forename: Oriane
authors.initials: O
authors.lastname: Soetens
authors.forename: Maria
authors.initials: M
authors.lastname: Breugelmans
authors.forename: Walter
authors.initials: W
authors.lastname: Foulon
authors.forename: Anne
authors.initials: A
authors.lastname: Naessens
mesh_terms: D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014509:Ureaplasma / Q000235:genetics*; D016869:Ureaplasma Infections / Q000175:diagnosis* / Q000382:microbiology; D016990:Ureaplasma urealyticum / Q000235:genetics*
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial