title: Development and performance of a microwell-plate-based polymerase chain reaction assay for Mycoplasma genitalium.
abstract: BACKGROUND
  Mycoplasma genitalium is associated with, and could be the cause of, idiopathic cases of urethritis, endometritis, and cervicitis. Further epidemiologic studies on this organism are needed, but currently used polymerase chain reaction (PCR) assays are labor-intensive and culture is insensitive.
  
  
  OBJECTIVE
  The goal was to develop and evaluate a microwell-plate-based PCR assay for M. genitalium.
  
  
  METHODS
  We adapted an M. genitalium PCR assay targeting the MgPa gene to a 96-microwell plate format with colorimetric detection of PCR products and incorporation of an internal inhibition control to determine the limit of detection of this assay (termed MgPa-IMW) for M. genitalium DNA and evaluate its performance on cervical and male urine specimens.
  
  
  RESULTS
  The MgPa-IMW PCR assay detected 1 and 17 genome copies of M. genitalium (with 27% and 95% confidence) and was able to detect specimens inhibited for amplification. This assay was 100% concordant (50 positive and 50 negative) with the Southern-blot-based PCR assay with cervical specimens. Similarly, this test was 89% concordant with the Southern-blot-based assay for 64 male urine specimens (25 positive, 32 negative, 7 discordant), 97% concordant after correcting for specimens no longer positive by the Southern blot-based assay after freezer storage.
  
  
  CONCLUSIONS
  The MgPa-IMW assay is sensitive and specific for the detection of M. genitalium in patient specimens and should facilitate large-scale screening for this organism.
authors.affiliation: Department of Medicine, University of Washington, Seattle, USA.
authors.forename: Susan M
authors.initials: SM
authors.lastname: Dutro
authors.forename: Jennifer K
authors.initials: JK
authors.lastname: Hebb
authors.forename: Cresley A
authors.initials: CA
authors.lastname: Garin
authors.forename: James P
authors.initials: JP
authors.lastname: Hughes
authors.forename: George E
authors.initials: GE
authors.lastname: Kenny
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Totten
mesh_terms: D015139:Blotting, Southern; D002584:Cervix Uteri / Q000382:microbiology; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis*; D005260:Female; D006801:Humans; D008297:Male; D009175:Mycoplasma Infections / Q000175:diagnosis* / Q000382:microbiology; D045704:Mycoplasma genitalium / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods / Q000592:standards*; D011237:Predictive Value of Tests; D012680:Sensitivity and Specificity; D014556:Urine / Q000382:microbiology
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial