Literature review > Issue 9 > Review on Stellrecht  et al. 

Stellrecht and her colleagues report on a sensitive and specific multiplex PCR test they have developed for the detection of Ureaplasma spp., Mycoplasma genitalium and Mycoplasma hominis. The PCR test is somewhat cumbersome, with a lot of hands-on time. It includes phenol-chloroform extraction and alcohol precipitation of the DNA, followed by a hot-start PCR in a thermocycler. Detection of the PCR product was via an ELOSA method. Identification of M. hominis from M. genitalium required a further digestion and identification of separate bands. They claim that all 3 of these mollicutes may be detected by a single amplification reaction in less than 8 hours, compared to 2 to 56 days for culture.

Culture of genital mycoplasmas is extremely difficult and time-consuming. However, while their paper's title suggests that the study design would utilize an optimized method for culture comparison, they did not include a broth-based medium in their methods, one that they claim would have increased the sensitivity of their culture for Mycoplasma spp. by up to 76%. In addition, the culture incubation time of only 5 days was entirely too short to recover M. genitalium. As they report, that organism may require 8 weeks of incubation.

In spite of this faulty study design, the new PCR test was demonstrated to be very sensitive and specific. The analytical limit of detection was excellent for U. urealyticum, M. hominis and M. genitalium (less than 12.5 CFU for each, whether alone or in a mixture). The results for the analytical specificity testing with 500,000 CFU of closely related organisms showed no cross-reactions.

The clinical specimens (cervical, vaginal, semen, and urine) were collected from 85 patients seen at a fertility clinic in 1998 and 1999. They were apparently stored frozen for an unknown number of years before the PCR testing was done. Agreement between the culture and PCR methods was lower than might be expected for a very sensitive PCR test - only 17 of the 21 culture positive specimens were also positive by PCR. Seventeen samples grew Ureaplasma spp. and 4 grew Mycoplasma spp. by culture. An additional 11 specimens were PCR positive but culture negative. Overall, the PCR method identified 23 samples as containing Ureaplasma spp. and 3 containing M. hominis, with 2 samples containing both. Interestingly, no samples were identified to contain M. genitalium by PCR or culture.

The authors are to be congratulated for their efforts to resolve discrepant samples. All samples were re-tested by a second PCR method. In this manner, the definition of a "true infection" may be resolved. Remarkably, all of the PCR-positive/culture-negative discordant samples were also positive by the second PCR method. Therefore, the overall prevalence was 36%.

Prospective trials, using fresh specimens and a final, optimized approach for specimen processing, would help to further define the use of this multiplex PCR test on clinical specimens. At this point, their conclusions appear valid for only Ureaplasma spp. The number of positives for the Mycoplasma spp., in particular for the important M. genitalium, is insufficient to draw any valid conclusions at this point.

The role of these mycoplasmas in genital and reproductive disease remains controversial. Overall, this new multiplex PCR method has been well-characterized, and should be a useful tool to further elucidate the epidemiology, pathogenicity and treatment aspects of infection with these mycoplasmas.

about SDI | newsletters | grants | publications | literature reviews

WHO Home - WHO Search - TDR Home - SDI Home - SDI Contact us
(c) WHO/OMS 2001