|Year : 2013 | Volume
| Issue : 1 | Page : 25-31
Trichomoniasis: How do we diagnose in a resource poor setting?
R Sivaranjini1, TJ Jaisankar1, Devinder Mohan Thappa1, Rashmi Kumari1, Laxmisha Chandrasekhar1, M Malathi1, Subhash Chandra Parija2, Syed Habeebullah3
1 Department of Dermatology, Sexually Transmitted Diseases and Leprosy, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry, India
2 Department of Microbiology, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry, India
3 Department of Obstetrics and Gynecology, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducherry, India
|Date of Web Publication||4-Jun-2013|
Devinder Mohan Thappa
Department of Dermatology, Sexually Transmitted Diseases, Venereology and Leprosy, Jawaharlal Institute of Postgraduate Medical Education and Research, Puducheery - 605 006
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Diagnosis of Trichomonas vaginalis vaginalis infection based solely on clinical symptoms and signs is unreliable because the spectrum of infection is broad and other sexually transmitted pathogens cause similar signs and symptoms. Aims: Our study was undertaken to study the frequency of T. vaginalis infection in women presenting with vaginal discharge, to characterize the clinical features, and to study the sensitivity and specificity of microbiological investigations in the diagnosis of the same. Materials and Methods: This was a hospital-based descriptive study done on 400 female patients with vaginal discharge attending the Gynecology out-patient department (OPD) of JIPMER, Puducherry, from May 2010 to July 2011. Women of age between 20 years and 50 years presenting with vaginal discharge irrespective of marital status, were included, and detailed history was elicited and thorough examination was performed. Results: In 400 women presenting with vaginal discharge from Gynecology out-patient department (OPD) included in the study, T. vaginalis infection was found in 27 (6.75%) women. The risk factors for trichomoniasis included history of pre- or extramarital sexual contact in the woman or her partner, symptomatic partner, and alcohol consumption. A positive association with pelvic inflammatory disease was also observed. The most frequent symptoms included lower abdominal pain, dysuria, and dyspareunia. Combining of Whiff test, pH > 4.5, and pus cells in Gram-stained smear, the specificity in diagnosing the infection (97.3%) approached that of the reference standard, i.e., culture. On combining wet mount with Papanicolaou smear, the sensitivity increased to 92.6%, which was higher than that individually done. Conclusion: To conclude, diagnosis of T. vaginalis infection based solely on clinical symptoms and signs is unreliable, and combination of simple laboratory tests increases the diagnostic performance close to the reference standard (culture), especially in resource poor settings.
Keywords: Culture, Papanicolaou smear, Trichomonas vaginalis, vaginal discharge, wet mount
|How to cite this article:|
Sivaranjini R, Jaisankar T J, Thappa DM, Kumari R, Chandrasekhar L, Malathi M, Parija SC, Habeebullah S. Trichomoniasis: How do we diagnose in a resource poor setting?. Indian J Sex Transm Dis 2013;34:25-31
|How to cite this URL:|
Sivaranjini R, Jaisankar T J, Thappa DM, Kumari R, Chandrasekhar L, Malathi M, Parija SC, Habeebullah S. Trichomoniasis: How do we diagnose in a resource poor setting?. Indian J Sex Transm Dis [serial online] 2013 [cited 2019 Dec 8];34:25-31. Available from: http://www.ijstd.org/text.asp?2013/34/1/25/112866
| Introduction|| |
Trichomoniasis, caused by parasitic protozoan Trichomonas vaginalis, is the most common non-viral sexually transmitted infection (STI). Diagnosis of T. vaginalis infection based solely on clinical symptoms and signs is unreliable because the spectrum of infection is broad and other sexually transmitted pathogens cause similar signs and symptoms.  Therefore, the laboratory plays a key role in the diagnosis of this infection; and more so, it is compelling to identify T. vaginalis infection reliably in resource poor settings with the readily available tests. The present study was proposed to address these issues. Our study was undertaken to study the frequency of T. vaginalis infection in women presenting with vaginal discharge, to characterize the clinical features, and to study the sensitivity and specificity of microbiological investigations in the diagnosis of the same.
| Materials and Methods|| |
This was a hospital-based descriptive study done on 400 female patients with vaginal discharge attending the Gynecology out-patient department (OPD) of JIPMER, Puducherry, from May 2010 to July 2011. The Institute Ethics Committee clearance was obtained and informed consent was taken from the recruited women. Women of age between 20 years and 50 years presenting with vaginal discharge, irrespective of marital status, were included, and detailed history was elicited and thorough examination was performed. Women who had used antibiotics or vaginal medication in the previous 14 days and pregnant women were excluded from the study.
The vaginal pH was measured and a sterile cotton swab was used to collect five samples of vaginal discharge from the posterior fornix under direct vision. In virgins, the specimen was obtained from the introitus. The Papanicolaou smears ( Pap smear More Detailss) were taken and a bimanual examination was done in all, except in virgins, to look for adnexal tenderness. The first swab was subjected immediately to wet mount microscopy to observe for motile trichomonads under 100× and 400× magnifications. With the second sample, a 10% potassium hydroxide (KOH) mount was prepared and Whiff test was done and the same was examined for the presence of budding yeast cells under 100× and 400× magnifications. The third sample was immediately inoculated directly and swirled into Diamond medium (Himedia labs, Mumbai, India) for T. vaginalis culture. The culture tubes with 5 ml of the broth were incubated in an anaerobic atmosphere at 35°C. The fourth sample was inoculated on to Sabouraud's dextrose agar medium for candidia culture. The fifth sample was streaked onto a microscopic slide for Gram staining to check for the presence of clue cells, yeast cells, and pus cells under 1000× magnification. Women with high-risk sexual behavior were counseled and tested for HIV using enzyme-linked immunosorbent assay (ELISA).
A diagnosis of trichomoniasis was made based if the wet mount showed pear-shaped organisms with characteristic jerky movements or if the Pap smear showed blue or grey pear-shaped organisms with bright-red granules.  From the culture, diagnosis was made by performing wet mounts for evidence of motile trichomonads by examining cultures after 24 h of incubation and then daily up to 7 days. Bacterial vaginosis was diagnosed based on Amsel's criteria,  and a diagnosis of candidiasis was made based on positive microscopy and/or culture. A diagnosis of pelvic inflammatory disease was made if, in addition to the presenting symptoms of abnormal vaginal discharge and lower abdominal pain, adnexal tenderness was elicited on examination. 
The data collected was tabulated in Microsoft Excel worksheet and computer-based analysis was performed using the SPSS 13.0 software (SPSS, Chicago, IL, USA). For comparison of means, unpaired t-test and one-way analysis of variance (ANOVA) were used for two and more groups, respectively. For comparison of proportions, Chi-square test was used. In cases where any one of cell value was <5; Fisher's exact test was used.
| Results|| |
Among the 400 women enrolled in our study, 27 (6.75%) had infection with T. vaginalis, identified by culture technique, the "reference standard," the mean age being 35.37 ± 7.66 years. Eighteen (66.6%) were positive for T. vaginalis on screening with the wet mount of vaginal discharge; 19 of 26 (73.1%) available Papanicolaou smear had evidence of trophozoites of Trichomonas. [Table 1] shows the sensitivity, specificity, and positive and negative predictive values of the abovementioned tests and the ancillary tests in the diagnosis of T. vaginalis infection. Of the 27 women with trichomoniasis, 12 (44.4%) had trichomoniasis alone and the other 15 (55.5%) had trichomoniasis with concurrent infections. Bacterial vaginosis was the most common concurrent infection with trichomoniasis with an odds ratio of 1.864 (95% confidence interval: 0.8453-4.112). The demographic and risk behavior characteristics of patients with trichomoniasis are summarized in [Table 2]. The sensitivity, specificity, and positive predictive and negative predictive values of the various symptoms and signs are presented in [Table 3].
|Table 1: Performance of diagnostics tests for trichomoniasis (individual tests and combinations)|
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|Table 2: Demographic and risk behavior characteristics of patients with Trichomonas vaginalis infection as compared to those without infection|
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|Table 3: Diagnostic value of clinical features associated with Trichomonas vaginalis infection (N=27)|
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| Discussion|| |
T. vaginalis , a pathogenic protozoan of urogenital tract causes vaginitis in women, although a considerable proportion of infections are asymptomatic. Prevalence estimates of T. vaginalis infection varies considerably in female population around the world, depending on the population studied and techniques employed in diagnosis, ranging from 5% to 74%. , Our study, which recruited women from the Gynecology OPD revealed that 27 out of the 400 women (6.75%) with vaginal discharge had infection with T. vaginalis by culture. Amongst the South-Asian studies, the rate observed in our study was comparable with the population-based study on community prevalence of STIs and HIV infections in rural south India done by Thomas et al.,  which reported a prevalence of 5.25% for trichomoniasis by direct wet mount of vaginal swabs. Among mixed infections occurring with T. vaginalis, concurrent bacterial vaginosis was the most common as observed by Bhalla et al.,  probably because T. vaginalis alters the vaginal ecology and facilitates the development of bacterial vaginosis or women with bacterial vaginosis have lost the natural protection against genital tract infection, leading to the acquisition of STIs like T. vaginalis infection. 
Unlike chlamydia and gonorrhea, the prevalence of T. vaginalis has been shown to increase with age.  The increased prevalence of trichomonal infection in older women suggests longer duration of infectiousness and the predominant asymptomatic nature of infection.  This was evident from our study as the mean age at presentation for women with T. vaginalis infection was 35.37 years. This was comparable with the prospective study from Srilanka by Fernando et al.,  who reported a mean age of 33 years.
Infection with T. vaginalis can be a marker for high-risk behavior, and the predominant asymptomatic nature of the infection underscores the need for identification of risk factors associated with the infection. Significant risk factors associated with T. vaginalis infection in our study were a history of pre/extramarital sexual contact in the women (OR = 8.250, 95% CI 2.594-26.234), husband having extramarital contact (OR = 5.916, 95% CI 2.559-13.679), a history of symptomatic partner in the form of dysuria and urethral discharge suggestive of trichomoniasis in men (OR = 6.899, 95% CI 1.974-24.110), and alcohol consumption by women (OR = 106.71, 95% CI 5.356-2126.3). Kaur et al.,  in their study in north India to assess the prevalence of T. vaginalis infection in symptomatic women as well as in women with carcinoma cervix and HIV infection, observed that being a housewife, belonging to the middle socioeconomic status, and non-use of contraception were significantly associated with trichomoniasis.
The signs and symptoms associated with the infection have a relatively low-positive predictive values for trichomoniasis because of the frequent occurrence of these signs and symptoms among women with other infections. , Also, Fouts et al.,  in their study on women attending a STI clinic in Georgia demonstrated that, if the clinical features alone were used to diagnose trichomoniasis, 88% of the infected women would be missed and 29% would be falsely indicated as having infection. In our study on women presenting with vaginal discharge, the reported symptoms had low-positive predictive values for trichomoniasis. Among the clinical signs in women with vaginal discharge, having a profuse vaginal discharge (OR = 4.426, 95% CI 1.994-9.823), malodorous discharge (OR = 3.554, 95% CI 1.507-8.381), frothy discharge (OR = 273.29, 95% CI 15.071-4955.7), and mucopurulent or purulent discharge (OR = 27.013, 95% CI 6.292-115.98) were significantly associated with trichomoniasis. Frothy discharge was observed in only 25.9% women in our study, but this was the most specific sign with a positive predictive value of 100%. Colpitis macularis was not observed in any of the women by naked eye examination in our study as colposcopy was not done on a routine basis. Women with T. vaginalis infection have a significantly higher risk of pelvic inflammatory disease (PID) than women without trichomoniasis.  Adnexal tenderness, as an evidence of PID was present in 40.7% patients, which was significantly higher than that seen in women without trichomoniasis.
The sensitivity and specificity of microscopic and cytologic examination were assessed considering culture of T. vaginalis as the "reference standard." Whiff test and pH testing are simple tests recommended in the evaluation of vaginal discharge and can be used as screening tools. , pH testing was reported to be a highly sensitive, but less specific screening tool for diagnosis of trichomoniasis.  A similar finding was observed in our study, where pH of >4.5 had 100% sensitivity, but a low specificity of 41.3% in the diagnosis of trichomoniasis. Whiff test was observed to have low sensitivity (51.8%) and low specificity (78.8%) in predicting T. vaginalis infection in our study. Women with trichomoniasis have a significantly higher concentration of white blood cells in the vaginal wet mount than those without trichomoniasis.  In our study, 55.6% cases with trichomoniasis had pus cells in Gram-stained smear of vaginal discharge.
In our study, we attempted to increase the specificity of these simple screening tests in the diagnosis of trichomoniasis by studying the performances of the combinations of these tests [Table 1]. pH > 4.5 had a sensitivity of 100% and a specificity of 41.3%. On combining positive Whiff test with pH > 4.5 and including cases that were positive for both tests, the specificity was increased by two folds (78.8%). Similar finding was observed on combining pus cells in Gram-stained smear with pH > 4.5. When all three (pH > 4.5, Whiff test, pus cells in Gram-stained smear) tests were combined, the specificity (97.3%) approached to that of our reference standard diagnostic test. This offers an advantage in settings where culture is not available and where the patient may be lost to follow-up due to it being time-consuming.
The diagnosis of trichomoniasis in women is most commonly made by wet mount examination; the Papanicolaou smear represents a common, readily available cytological technique for primary screening of trichomoniasis.  In our study, the sensitivity of wet mount and Papanicolaou smear was observed to be 66.7% and 76%, respectively; both being 100% specific with 100% positive predictive value in the diagnosis of T. vaginalis infection. Thus, culture of direct wet mount positive vaginal specimens may be unnecessary owing to the 100% specificity of the test. Since our study was restricted to women with symptomatic T. vaginalis infection, it is probable that these women harbored greater numbers of organisms and thus had a higher rate of positivity for wet mount. On combining wet mount with Papanicolaou smear, the sensitivity increased to 92.6%; this has an implication in resource-limited setting where culture may not be available, combining Papanicolaou smear with wet mount would help identify few more cases of trichomoniasis that would have been missed by wet mount alone. [Figure 1] shows an algorithm that can be followed for diagnosis of trichomoniasis.
|Figure 1: Algorithm for diagnosis of trichomoniasis derived from our study|
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To conclude, in our study, a moderate prevalence of T. vaginalis infection was observed, though the prevalence in the community might be higher as we had included only symptomatic women when majority of the infection is asymptomatic. The majority of the women with trichomoniasis had co-infections with other organisms. We observed a significant association of trichomoniasis with PID, resulting in reproductive morbidity. The poor clinical value of symptoms and signs in the diagnosis of T. vaginalis infection could be corrected by appropriate combination of simple tests that increases the performance of these screening tests comparable to that of the diagnostic tests, especially, in the setting of non-availability of culture for T. vaginalis.
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[Table 1], [Table 2], [Table 3]