Mycoplasma genitalium is a cause of nongonococcal urethritis, particularly in patients not infected withChlamydia trachomatis. A quantitative 5 nuclease assay (TaqMan PCR) was developed and validated. The assay detected a fragment of the MgPa adhesin gene by use of a TaqMan MGB (minor groove binder) probe and included an internal processing control to detect PCR inhibition. Urethral swab specimens and first-void urine samples from M. genitalium-positive men were examined, and the M. genitalium DNA load was correlated to symptoms and signs. The assay consistently detected <5 genome copies without cross-reactions with other mycoplasmas. Urine and urethral swab specimens from men with urethritis had higher M. genitalium DNA loads than specimens from men without urethritis. However, a very broad overlap of DNA loads between patients with and without urethritis was observed. Urethral swab specimens from patients with urethral discharge had a significantly higher DNA load than specimens from patients without discharge. This correlation was not found in first-void urine specimens.Mycoplasma genitalium was first isolated after prolonged incubation from 2 of 13 men with urethritis (26, 27). Despite repeated attempts by conventional culture techniques (21, 23), other urogenital isolates have been extremely rare and have been difficult to obtain (10, 17). Isolation of M. genitalium in cultures of throat and synovial fluid specimens has also been recorded, although these isolates were mixed with Mycoplasma pneumoniae (1, 25).M. genitalium and M. pneumoniae share several structural properties, such as the flask shape and the terminal tip-like structure, and a significant antigenic relationship between the two mycoplasma species has hampered diagnostic serology (15,16).Because traditional procedures for the diagnosis of M. genitalium infection, like culture and serology, have failed, more extensive studies indicating that M. genitalium is a cause of sexually transmitted infections had to await the development of the PCR (11, 18).The first two PCR-based studies of M. genitalium in patients with nongonococcal urethritis (NGU) were reported in 1993 (12). In those studies (16) M. genitalium was found significantly more often in men with NGU than in those without the condition. Furthermore, M. genitalium was found more often in men with Chlamydia trachomatis-negative NGU than in those with chlamydial NGU, indicating that the two microbes may act as separate causes of urethritis (6, 12). Several case-control studies of NGU patients and controls have confirmed this finding (for a review, see references 8 and 24). Subsequent studies have shown that M. genitalium is more closely associated with symptomatic urethritis than with asymptomatic urethritis (2; P. J. Horner and D. Taylor-Robinson, Letter, Lancet 343:790-791, 1994), particularly in those patients with an observable discharge (7). Recently, chronic NGU was also demonstrated to be associated with M. genitalium infection (5) The detection of M. genitalium in higher numbers in ur...

M. genitalium infection is unlikely to be a major risk factor for clinical pelvic inflammatory disease in this population.

Mycoplasma genitalium causes male nonchlamydial, nongonococcal urethritis and is associated with cervicitis and pelvic inflammatory disease in women. Epidemiological studies indicate that M. genitalium is sexually transmitted, and the aim of the present study was to further substantiate this by means of a DNA typing system. A typing assay based on a diagnostic mgpB gene PCR was developed, evaluated, and applied directly to urogenital specimens. The assay had a low limit of detection and hence a high typeability. Sequences of isolates from 52 unrelated patients were divided into 29 different sequence types, giving a discriminatory index of 0.95. Two to six M. genitalium-positive specimens were collected from each of 44 patients over a median interval of 56 days (range, 11 to 1,395). Forty had the same sequence type in consecutive specimens. Specimens collected from two men were repeatedly positive at intervals of 472 and 1,395 days, respectively, but the sequence types had changed. A new strain was introduced in one sexual dyad, and the sequence types changed subsequently. Seventy-nine M. genitalium-positive specimens from 19 couples were investigated, and all partners initially had concordant sequence types, but one couple had discordant types at one time point before a newly introduced strain took over. The present typing system is simple and reproducible and has an excellent discriminatory capacity which might prove useful in studies of sexual networks and for evaluation of treatment failures. In the laboratory, this system may document the uniqueness of newly isolated M. genitalium strains.

Urine appeared to be a better diagnostic specimen than the urethral swab for M. genitalium and C. trachomatis detection by PCR in this cohort of sexually transmitted disease clinic attendees but should be supplemented with a cervical specimen in women.

No abstract

In order to develop a species-specific PCR for the detection of Mycoplasma genitalium, the sequence of 1,490 bases of the 16S rRNA gene was determined for M. genitalium G37 (type strain) and four Danish isolates of M. genitalium. The sequences of the four Danish strains, mutually different with respect to their MgPa gene, were 100% homologous, although they carried a single common base substitution compared to the type strain. Among members of the Mycoplasma pneumoniae phylogenetic cluster, M. genitalium showed the most-prominent homology to the 16S rRNA sequence of M. pneumoniae (98% homology). From regions showing the least homology to the M. pneumoniae 16S rRNA gene sequence, primers were chosen to amplify DNA from M. genitalium only. Two sets of primers were selected for their ability to detect <10 to 50 M. genitalium genome copies without cross-reactions with M. pneumoniae. The performance of these primers was compared to the performance of two pairs of primers amplifying parts of the MgPa adhesin gene; 1,030 randomly selected specimens submitted for Chlamydia trachomatis culture were screened with one of the 16S rRNA gene primer sets. A total of 41 specimens were found to be positive for this gene; 40 of these could be confirmed by one of the MgPa primer sets, whereas the other MgPa primer set detected only 21 positive specimens out of 40. These results indicate that estimates of the prevalence of M. genitalium in various populations using MgPa PCR primers could be incorrectly low if the PCR primers are located in variable regions of the MgPa gene.Two Mycoplasma genitalium strains were originally isolated in 1980 from the urogenital tracts of 2 of 13 men with nongonococcal urethritis (NGU) (27). Despite repeated attempts with conventional culture techniques, no other urogenital isolates have been reported (21, 24). Using a cell-culture-based method, however, we succeeded in isolating four new strains from the urethras of male patients with NGU who were PCR positive for M. genitalium (11). M. genitalium and Mycoplasma pneumoniae share several structural properties, such as the flask shape and the terminal tip-like structure, and a significant antigenic relationship between the two mycoplasma species has hampered diagnostic serology (15; K. Lind, Letter, Lancet ii:1158-1159, 1982).Because traditional diagnostic procedures for M. genitalium such as culture and serology have failed, other methods have been investigated. The development of a DNA probe provided some evidence for the presence of M. genitalium in the male urogenital tract (8), but data indicating that M. genitalium is a potential cause of NGU have only recently been demonstrated by the use of PCR (2, 9, 10, 13, 26).M. genitalium strains recently isolated from Danish patients (11) show a significant degree of sequence diversity of the main adhesin (MgPa) gene. This variability has not yet been sufficiently characterized to determine the presence of conserved regions in the gene so that PCR primers covering all M. genitalium strains can be designed....

Mycoplasma genitalium is associated with cervicitis and pelvic inflammatory disease but little is known about its role in pregnancy. We investigated the prevalence of M. genitalium by polymerase chain reaction assay on urine specimens from 1216 pregnant women (mean age 31years) presenting before 10 weeks of gestation in 32 general practices. The prevalence of M. genitalium was 0.7% (6/915, 95% CI 0.1 -1.2). It was more common in women aged <20 years, women of Afro-Caribbean or black African ethnic origin, women in social classes 3 -5 and single women. Only one woman with M. genitalium infection miscarried, and none of those followed up to term had a preterm birth, although the numbers were small. The low prevalence of M. genitalium infection suggests it is unlikely to be an important risk factor in adverse pregnancy outcome in healthy women in the community.

A PCR assay for the detection of Bordetella pertussis and Bordetella parapertussis was compared with the conventional culture method under routine laboratory conditions. Detection of B. pertussis was based on the amplification of a section of the IS481 insertion sequence and confirmation of positive results was based on a sequence of the pertussis toxin promoter region. Detection of B. parapertussis was based on the amplification of a section of the IS1001 insertion sequence. An internal control was included. Data were available for the period 28 November 2000 to 9 July 2003. In this period, 3096 patients were examined for infection with B. pertussis and B. parapertussis by culture and PCR on the same day. B. pertussis was found in 496 (16 %) patients; 208 (42 %) were diagnosed by PCR alone whereas 17 (3 %) were diagnosed by culture alone. B. parapertussis was found in 64 (2 %) patients. The sensitivity of the PCR was 97 % and of culture 58 %. The specificity of PCR was 93 % when regarding culture as 100 % sensitive. There was a significant relationship between laboratory method and age, as the superiority of PCR was most marked in the age group 0 . 5-3 years. The PCR assay proved highly sensitive for the diagnosis of pertussis. The specificity estimate of the PCR assay suffers from the influence of a gold-standard method with a low sensitivity. The PCR assay is considered highly specific due to the amplification of two different sequences in two separate assays.