Stability of Trichomonas vaginalis DNA in Urine Specimens

Abstract

Trichomonas vaginalis is an important pathogen in both men and women. Culture is considered the diagnostic gold standard, although studies have shown that PCR is more sensitive than either culture or wet mount for the diagnosis of T. vaginalis infections. We sought to identify a simple method for stabilizing T. vaginalis DNA in urine samples that could be easily applied to molecular testing. The stability of T. vaginalis DNA in 40 urine samples was assessed by storage for various times at either 4°C or room temperature with or without the Becton Dickinson urine preservative transport (UPT) kit. Overall, there was better stability of T. vaginalis DNA when specimens were stored at 4°C than when they were stored at 20 to 22°C and when the UPT system was used. T. vaginalis DNA was stable in specimens stored without using the UPT at 4°C for about 3 days and at room temperature for only 1 day. For specimens placed in the UPT within 24 h (times of 1, 6, and 24 h) of collection, the DNA was stable for up to 30 days when stored at 4°C. For specimens stored at room temperature, the urine should be added to the UPT ideally within 1 hour of collection, and in this case the DNA remained stable for up to 30 days. When storing specimens at room temperature, a delay of 24 h prior to adding to UPT led to an unacceptably high loss of assay sensitivity.Trichomonas vaginalis is a common sexually transmitted disease, with an estimated 7.4 million new cases annually in the United States and as many as 180 million cases worldwide (13). T. vaginalis can cause symptoms in both men and women and can cause severe complications in pregnant women. Symptoms in women include vaginal discharge and vulval irritation; if untreated, more-severe complications may occur, including endometritis, cervical erosion, and infertility (3, 9). T. vaginalis infections in pregnant women can lead to premature rupture of membranes, low-birth-weight infants, and preterm deliveries (2). In men, infection with T. vaginalis may cause nongonococcal urethritis, with untreated infections causing chronic prostatitis, epididymitis, infertility, or urethral strictures (3, 9). Moreover, T. vaginalis infection can lead to an increased risk of acquiring human immunodeficiency virus type 1 infection due to the local inflammation caused by T. vaginalis infections in the genital tract (5,7,8,11).There are numerous reports supporting the role of PCR in the diagnosis of T. vaginalis infections from genital specimens; these reports have shown that the detection of T. vaginalis DNA by PCR is significantly more sensitive than both culture and wet mount examination (1,4,6,12,14,15). We have previously reported that vaginal swabs collected with the BDProbeTec dry swab system were a reliable specimen for PCR testing and allowed the detection of Chlamydia trachomatis, Neisseria gonorrhoeae, and T. vaginalis from a single specimen (1). For clinical studies using urine specimens, our procedure for stabilizing T. vaginalis was cumbersome and involved adding an aliquot of urine to Fuji me...

Genotyping of urinary samples stored with EDTA for forensic applications

“…Cannas et al (2009) discovered that only study location and the addition of EDTA correlated with urinary DNA stability. EDTA preservatives for urine are commonly prepared to a final concentration of 10 mM using commercially available urine transport tubes sold specifically for downstream molecular analysis (Ingersoll et al, 2008). Milde et al (1999) found that storing urine at room temperature with sodium azide for 30 days or at -20°C with EDTA for 72 days provided the best protocol for the analysis of human DNA.…”

Current Issues and Considerations Regarding Trichomoniasis and Human Immunodeficiency Virus in African-Americans

“…Ingersoll et al recently suggested that the stability of T. vaginalis DNA in urine could be improved with the use of a urine preservative transport kit. This method, while useful only for molecular diagnosis, could serve to substantially improve the sensitivity of molecular testing (36). PCR for the diagnosis of T. vaginalis is also available, but unlike for infections such as gonorrhea and chlamydia, in which PCR appears to have greater sensitivity than culture methods, PCR for trichomoniasis in women does not appear to offer a diagnostic advantage.…”

Laboratory evaluation of a specimen transport medium for downstream molecular processing of sputum samples to detect Mycobacterium tuberculosis

“…However, studies on other pathogens in different clinical specimens have shown a negative effect on molecular testing when kept at ambient temperature for extended periods of time (Hasan et al, 2012, Ingersoll et al, 2008. In that context, the manufacturer of the Xpert assay recommends that sputum samples be kept for no longer than 3 days at ambient temperatures up to 35° C until processing (Banada, et al, 2010).…”