Detection and characterisation of Ureaplasma spp in men with and without urogenital symptoms

Abstract

Ureaplasma spp colonize the lower genital tract in many healthy men, yet can also cause urethritis and infertility. Since Ureaplasma spp are frequently isolated from the normal urogenital tract, it has been suggested that only certain serotypes are associated with diseases. The genus Ureaplasma consists of 14 serotypes that can be divided into two biotypes, biotype 1 (U. parvum) and biotype 2 (U. urealyticum). Biotype 1 includes four serotypes (1, 3, 6 and 14) and biotype 2 includes ten serotypes (2, 4, 5 and 7 to 13). Identification and characterization of Ureaplasma spp are mainly performed by traditional culture methods that are difficult to perform and take two to seven days. Commercial kits have been an alternative method for the quick identification of Ureaplasma spp (24 to 48 hrs) but these kits cannot speciate the bacteria. However, polymerase chain reaction (PCR) assays have been developed for characterization and speciation of Ureaplasma spp. In this study, 200 first-void urine specimens were collected, 100 from symptomatic men (discharge/dysuria) and 100 from asymptomatic (without discharge/dysuria) men. The specimens were all cultured on U9 broth then subcultured on A2 agar for confirmation of growth. Antibiotic susceptibility determination was performed on the positive isolates using the Mycofast evolution 3 kit (ELITech Microbiology, France). Molecular detection of U. urealyticum was performed using a commercial kit, the Ureaplasma urealyticum real-time PCR kit. Detection and characterization of U. parvum was performed using a multiplex Taqman real-time PCR assay targeting the multiple banded antigen genes (MBA). Culture results of all specimens showed 40% (79/200) urease production on Shepard’s U9 broth and of these specimens 35% (28/79) were positive when subcultured on A2 medium, 25% (20/79) were contaminated with either yeast or Mycoplasma hominis or both and 39% (31/79) of the U9 positive specimens did not grow when subcultured on Shepard’s A2 agar medium. When determining the susceptibility profile of Ureaplasma spp, 32% (9/28) of the isolates did not grow with the kit and four were contaminated. None of the isolates was resistant to all of the antibiotics and all isolates were sensitive to doxycycline, pristinamycin, roxycycline and azithromycin. One isolate was resistant to ciprofloxacin and josamycin but intermediately resistant to ofloxacin and another isolate was resistant to ofloxacin, while a second isolate was only resistant to ciprofloxacin. Ureaplasma urealyticum was detected in 16% (31/100) of the symptomatic men and 15% (15/100) of the asymptomatic men whereas Ureaplasma parvum was detected in 11% (11/100) of the symptomatic men and 18% (18/100) of the asymptomatic men. There was no significant difference between the two groups (p= 01598). The distribution of serotypes did not differ significantly between the two groups (p= 0.309). The predominant serotype was serotype 6 followed by 1, 3 and 14. In conclusion, the real-time PCR assay was rapid, not prone to contamination and implementation of the assay in diagnostic laboratories will help in the rapid detection and administering of treatment to patients (especially neonates) infected with Ureaplasma species. The determination of the susceptibility profiles of Ureaplasma spp will assist in the monitoring of the antibiotic resistant profile trends in Pretoria, so that the correct antibiotic regimens can be administered to people with Ureaplasma infections.