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FOCUS: MOLECULAR DIAGNOSIS OF SEXUALLY TRANSMITTED INFECTIONS

Trichomonas vaginalis: Common, curable and in the diagnostic spotlight

CHRIS L. MCGOWIN, RODNEY E. ROHDE, GERALD REDWINE

LEARNING OBJECTIVES tology, 1901 Perdido St.; MEB 6214, New Orleans, LA 1. Discuss the historical and epidemiological spectra 70112-2822, 504 568-7281, [email protected] of vaginalis. 2. Define the clinical characteristics of Trichomonas vaginalis is the most common curable Downloaded from in men and women. sexually transmitted infection (STI) worldwide.1 The 3. Explain the methods for clinical diagnosis of T. 2001-2004 National Health and Nutrition vaginalis infection. Examination Surveys (NHANES) report that American 4. Compare and contrast the treatment strategies for women between the ages of 14 and 49 are infected with trichomoniasis in women. T. vaginalis with a prevalence of approximately 3% - 2,3 greater than that of C. trachomatis and N. gonorrhoeae. http://hwmaint.clsjournal.ascls.org/ ABBREVIATIONS: CDC - Centers for Disease Additional studies have highlighted that, unlike C. Control and Prevention, FDA - Food and Drug trachomatis and N. gonorrhoeae, T. vaginalis affects all Administration, MDx - molecular diagnostics, NAAT - age groups and the prevalence is highest in women over nucleic acid amplification test, NGU - non-gonococcal 40 years of age.4-7 Annually, it is estimated that more , NHANES - National Health and Nutrition than half of the approximately 248 million new T. Examination Surveys, PID - pelvic inflammatory vaginalis infections occur in males worldwide.1 disease: POC - point of care, PPV - positive predictive However, the majority of prevalent T. vaginalis value, STD - sexually transmitted disease, STI - sexually infections are detected in women, which is consistent transmitted infection with the chronic nature of these infections in the female urogenital tract. Most infections in women are on September 24 2021 INDEX TERMS: Molecular diagnostics, sexually asymptomatic, however T. vaginalis has been linked to transmitted disease, sexually transmitted infections, lower and upper reproductive tract disease syndromes Trichomonas vaginalis, trichomoniasis including , and pelvic inflammatory disease (PID). Pregnancy-related complications include Clin Lab Sci 2014;27(1):53 pre-term birth and infertility. In men, T. vaginalis is most commonly a cause of non-gonococcal urethritis Chris L. McGowin, PhD, Louisiana State University (NGU) but infections tend to be transient and Health Sciences Center, Department of Microbiology, frequently asymptomatic. In this article, we concisely Immunology and Parasitology, New Orleans, LA discuss the current knowledge on T. vaginalis epidemiology and the developing trends in diagnosis of Rodney E. Rohde, PhD, MS, SV, SM(ASCP)CM, MBCM, this incredibly prevalent STI. Clinical Laboratory Science Program, College of Health Professions, Texas State University, San Marcos, TX Disease spectrum of T. vaginalis infections Currently, trichomoniasis is not a notifiable disease in Gerald Redwine MEd, MT(ASCP), Clinical Labora- the USA and so therefore, compared to other STIs, tory Science Program, College of Health Professions, Texas relatively little is known about the epidemiology and State University, San Marcos, TX pathogenesis of T. vaginalis infections. The spectrum of disease in women is characterized by symptoms of Address for Correspondence: Chris L. McGowin, PhD, vaginitis including discharge, and localized vulvar Louisiana State University Health Sciences Center, pruritus. Classical disease signs include purulent vaginal Department of Microbiology, Immunology and Parasi- discharge, and vaginal and vulvar erythema. The

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FOCUS: MOLECULAR DIAGNOSIS OF SEXUALLY TRANSMITTED INFECTIONS

is commonly infected with accompanying symptoms of Although both techniques rely upon a trained dysuria in women with vaginal infection. In addition, microscopist, wet mount microscopy and culture colpitis macularis, or “strawberry cervix” is a highly methods are highly specific for T. vaginalis detection specific sign of infection but is only accurately (>99%)11,13 primarily due to the characteristic tumbling diagnosed during colposcopy. The dogmatic notion that motility of viable organisms. T. vaginalis infections are overtly symptomatic in women has been unequivocally overturned. In fact, up Microscopy-based techniques were followed by the first to 80% of T. vaginalis infections are asymptomatic in generation of nucleic acid amplification tests (NAATs). women depending on the clinical setting.3 Despite the Collectively, NAATs have dramatically enhanced our asymptomatic nature of infection, T. vaginalis has been understanding of T. vaginalis epidemiology, linked to several inflammatory syndromes in women pathogenesis, and helped to identify a previously thus highlighting the importance for screening underappreciated population of asymptomatic initiatives in subjects at risk for STIs. T. vaginalis infections in men and women. Contemporary NAATs Downloaded from infection in women is also associated with increased are both highly specific and sensitive, and several have susceptibility to HIV acquisition and HIV shedding in been adapted to high-throughput instrumentation. co-infected individuals. In males, T. vaginalis is most Unlike microscopy, NAATs require no subjective commonly a cause of NGU but infection is frequently interpretation of the test results and have faster turn- asymptomatic and the majority of cases are reported to around times compared to culture. Enhanced sensitivity http://hwmaint.clsjournal.ascls.org/ clear spontaneously within 10 days.8-11 Together, the is the clear advantage of NAATs compared to culture importance of asymptomatic males as transmitters to and wet mount microscopy. The most widely used women is enormous since T. vaginalis infections in culture methods utilize the InPouch TV (BioMed women, with or without symptoms, can persist for at Diagnostics, White City, OR) device or inoculation of least several months.11 Like the bacterial STIs, specimen material into Diamond’s medium. These tests complicated T. vaginalis infections in men can lead to are moderately sensitive ranging from 44-75%13 and more severe inflammatory outcomes including require three to seven days before results are available. prostatitis, urethral stricture, and infertility.12 It was not until the late 1990’s when the first NAAT was FDA-approved for diagnostic testing in the USA.

Diagnosis of Trichomoniasis Currently, development and implementation of T. on September 24 2021 The history of T. vaginalis diagnostics dates back to the vaginalis testing platforms into clinical laboratories are early part of the 19th century when motile organisms top industry priorities in the realm of STI diagnostics. were visible on wet mount preparations of vaginal swab As of this 2014 publication, only two NAATs are FDA- specimens. Strikingly, direct microscopic observation approved and marketed for T. vaginalis detection in the remained the standard of diagnostic care until in vitro USA: 1) the APTIMA TV test (GenProbe, Inc., San culture methods were introduced in 1949.11 Several Diego, CA) for use on the TIGRIS and PANTHER studies have reported low sensitivities of T. vaginalis platforms; and 2) the ProbeTec Trichomonas vaginalis detection using wet mount microscopy that range from Qx Amplified DNA Assay for use on the BD Viper 44-68%.13 In addition, sensitivity can be negatively platform (Beckton Dickinson, Franklin Lakes, NJ). impacted by even short time intervals (10-30 minutes) Targeting a conserved region of 16s rRNA molecules, between swab collection and microscopic examination.14 the APTIMA TV test utilizes the combined Despite low sensitivity and the need for rapid technologies of transcription medicated amplification processing, microscopy remains the most commonly (TMA) and a hybridization protection assay (HPA). By used technique for diagnosis of trichomoniasis due to targeting a high-copy transcript in the T. vaginalis cell, ease and cost-effectiveness. Culture of trichomonads the FDA-approved APTIMA TV test is characterized by from female urogenital specimens using Diamond’s very high sensitivity values ranging from 95-100% broth medium is substantially more sensitive compared depending on the specimen type and gold standard to wet mount microscopy (96% vs. 36-82%, comparator.13 According to the manufacturer’s package respectively)11 but requires additional supplies and insert, sensitivity values range from 92-100% for the equipment including an incubator for growing cultures. ProbeTec Trichomonas vaginalis Qx Amplified DNA

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Assay among both symptomatic and asymptomatic paradigms for men are lacking considering the women when compared to a composite comparator of substantial number of incident infections. wet mount and culture. The ProbeTec assay utilizes strand displacement assay (SDA) technology as Treatment of T. vaginalis infections described previously.15 Taken together, modern NAATs Effective therapies for T. vaginalis infections are substantially improve sensitivity of T. vaginalis available for men and women diagnosed with detection from several specimen types, and serve both as trichomoniasis. The current CDC-recommended first- diagnostic and screening tools in symptomatic and line therapy for T. vaginalis infection is a single 2-gram asymptomatic women. Unfortunately, no NAAT has dose of or – both of which are been approved for T. vaginalis detection in men. FDA-approved for trichomoniasis therapy in men and However, as clinical research clarifies the value of women. Current recommendations for patients who fail screening in asymptomatic men and women, combined stat metronidazole are to treat with a single dose of with the high frequency of vaginitis, it is predicted that tinidazole or an extended regimen of metronidazole Downloaded from the volume of NAATs for T. vaginalis will exceed those (500 mg twice daily for seven days). Fortunately, for C. trachomatis and N. gonorrhoeae in the coming treatment failures are infrequent following single dose years. metronidazole therapy in patients for whom re-infection can be excluded. In short, non-susceptibility rates to the In contrast to high-throughput NAATs typically run at currently available therapies are low,9 and parasitological http://hwmaint.clsjournal.ascls.org/ hospital or reference laboratories, point of care (POC) cure rates are high (86-100%) in women.16 Failure of tests for STIs facilitate the initiation of treatment during the recommended second line regimens should be the initial clinic visit. This approach ensures compliance followed by treatment with oral metronidazole or with the treatment regimen since most first line tinidazole at 2 grams daily for five days. In patients with therapies are single STAT doses of antibiotics. Ideally, persistent T. vaginalis infection after extended POC tests allow differentiation of etiologic agents to metronidazole/tinidazole regimens, a combination of substantially reduce the number of patients that are oral and topically applied agents have been anecdotally managed syndromically. In the accompanying effective and may be indicated in cases of chronic Mycoplasma genitalium article in this series, we describe treatment failure.16 how POC testing could substantially reduce syndromic management of NGU in men and non-gonococcal Subjects with T. vaginalis infection are up to twice as on September 24 2021 cervicitis in women where treatment guidelines fall likely to acquire HIV infection17 thus underscoring the short to effectively treat all causative agents. Over the global importance for continued investigation of this last decade, POC tests for T. vaginalis have been prevalent STI. Interestingly, a recent randomized available that utilize either immunochromatographic clinical trial showed that women with HIV responded test strips (OSOM Trichomonas Rapid Test; Sekisui better to extended metronidazole (500 mg twice daily Diagnostics, California, USA) or non-amplification for seven days) compared to a single 2-gram dose and nucleic acid detection technology (BD Affirm VPIII; had lower rates of re-infection upon follow up.18 It is Beckton Dickinson, Maryland, USA). The sensitivities possible that special considerations should be applied to of the rapid diagnostic tests parallel those of culture- subjects with HIV, AIDS or who are based tests and are consistently higher than wet mount immunocompromised. However, data are lacking microscopy (40-95%).13 Despite lower sensitivity, the regarding T. vaginalis disease and therapies in HIV- positive predictive value (PPV; the ability to detect truly positive individuals despite high rates of co-infection. positive subjects) for the available POC tests can be increased to levels of contemporary NAATs by using Conclusions them in populations where prevalence is high (e.g. As the most common parasitic in women with signs of disease). As for NAATs, no FDA- developed countries, T. vaginalis infections are approved POC tests are available for T. vaginalis widespread and disproportionally affect women in both diagnosis in men. Unfortunately, wet mount prevalence and disease sequelae. Considering the microscopy is insensitive for detection of trichomonads substantial proportion of asymptomatic infections, the from male urogenital specimens and, in general, testing true impact of T. vaginalis testing will be realized fully

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when these assays are used as screening tests in both correlates of Trichomonas vaginalis among incarcerated persons men and women. In lieu of relatively low sensitivities, assessed using a highly sensitive molecular assay. Sex Transm Dis 2010;37(3);165-8. POC tests for T. vaginalis including wet mount 6. Miller WC, Swygard H, Hobbs MM, et al. The prevalence of microscopy remain valuable tools to initiate stat therapy trichomoniasis in young adults in the United States. Sex in clinical settings where follow up is unlikely. Transm Dis 2005;32(10);593-8. Interestingly, because T. vaginalis infections are more 7. Schwebke JR, Burgess D. Trichomoniasis. Clin Micro Rev common in women over 40 years of age, the apparent 2004;17(4);794-803. 8. Paisarntantiwong R, Brockmann S, Clarke L, et al. The risk for pathological sequelae without symptoms is high relationship of vaginal trichomoniasis and pelvic inflammatory and efforts to screen should be guided to all age groups disease among women colonized with trachomatis. rather than specifically to young adults. We still have Sex Transm Dis 1995;22(6);344-7. much to learn regarding the epidemiology and 9. Cudmore SL, Delgaty KL, Hayward-McClelland SF, et al. Treatment of infections caused by metronidazole-resistant pathogenesis of T. vaginalis infections. With FDA- Trichomonas vaginalis. Clin Micro Rev 2004;17(4);783-93. approved NAATs entering clinical laboratories in the 10. Schwebke JR, Rompalo A, Taylor S, et al. Re-evaluating the Downloaded from USA and abroad, we are on the cusp of a deeper treatment of nongonococcal urethritis: emphasizing emerging understanding of clinical disease and optimized --a randomized clinical trial. Clin Infect Dis therapeutic approaches in both men and women. 2011;52(2);163-70. 11. Poole DN, McClelland RS. Global epidemiology of Trichomonas vaginalis. Sex Transm Dis 2013;89(6);418-22. REFERENCES 12. Holmes KK. 1999. Sexually Transmitted Diseases, Third http://hwmaint.clsjournal.ascls.org/ 1. World Health Organization. Prevalence and incidence of Edition. New York: McGraw-Hill Press. selected sexually transmitted infections, , 13. Hobbs MM, Sena AC. Modern diagnosis of Trichomonas , , and Trichomonas vaginalis: vaginalis infection. Sex Transm Dis 2013;89(6);434-8. methods and results used by the WHO to generate 2005 14. Kingston MA, Bansal D, Carlin EM. 'Shelf life' of Trichomonas estimates. Geneva, Switzerland: World Health Organization, vaginalis. Int J STD AIDS 2003;14(1);28-9. 2011. 15. Little MC, Andrews J, Moore R, et al. Strand displacement 2. Allsworth JE, Ratner JA, Peipert JF. Trichomoniasis and other amplification and homogeneous real-time detection sexually transmitted infections: results from the 2001-2004 incorporated in a second-generation DNA probe system, National Health and Nutrition Examination Surveys. Sex BDProbeTecET. Clin Chem 1999;45(6);777-84. Trans Dis 2009;36(12);738-44. 16. Bachmann LH, Hobbs MM, Sena AC, et al. Trichomonas 3. Sutton M, Sternberg M, Koumans EH, et al. The prevalence of vaginalis genital infections;progress and challenges. Clin Infect Trichomonas vaginalis infection among reproductive-age Dis 2011;53:Suppl 3;S160-72.

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