Recent advances in the epidemiology, diagnosis, and management of Trichomonas vaginalis infection

Trichomonas vaginalis is the most common, curable non-viral sexually transmitted infection (STI) worldwide. Despite this burden of disease, it is not currently a reportable disease in the United States. Recent advances in the epidemiology, diagnosis, and management of T. vaginalis infection are described in this article. This includes updated global and U.S. prevalence data in women and men as well as recent epidemiological data in HIV-infected individuals and pregnant women. Advances in molecular diagnostics are also reviewed, as are data from recent clinical trials regarding the treatment of trichomonas in women.


Introduction
Trichomonas vaginalis is the most common, non-viral sexually transmitted infection (STI) worldwide 1 . Patients with symptomatic T. vaginalis report various symptoms, including vaginal discharge and dysuria in women and urethral discharge and dysuria in men 2-4 . Many infected patients, however, never experience symptoms 3 . Untreated or persistent T. vaginalis in women has been associated with infertility and adverse birth outcomes [5][6][7] . While less is known about T. vaginalis in men, it has been described as a cause of nongonococcal urethritis (NGU), prostatitis, and epididymitis 4 . T. vaginalis has also been linked to an increased risk of HIV, posing a major public health threat 8 . Nevertheless, T. vaginalis remains understudied given the lack of public health attention it has received 9 . It is not currently a reportable disease in the U.S., as it has previously been found to meet only three out of seven criteria 10 . This review aims to provide an update on recent advances in the epidemiology, diagnosis, and treatment of T. vaginalis.

Epidemiology
The World Health Organization (WHO) estimated 156 million cases of T. vaginalis worldwide in 2016, accounting for almost half of the global STI incidence that year 1 . Updated epidemiological data on the national prevalence of trichomonas among women and men in the U.S. was published in 2018 11 . These data were collected during 2013-2014 in the National Health and Nutrition Examination Survey (NHANES) using the Hologic Gen-Probe Aptima® T. vaginalis assay on urine specimens. T. vaginalis prevalence was 1.8% in women and 0.5% in men aged 18-59 years. Prior to this study, the national prevalence of T. vaginalis had been poorly characterized among U.S. men as a result of diagnostic challenges; NHANES did not test men for T. vaginalis until 2013-2014. Spontaneous resolution of T. vaginalis is known to occur at relatively high rates (36-69%) in men 12,13 , which may explain the lower prevalence compared to women in the recent NHANES study. While T. vaginalis is less common in men 11 , it is readily passed between sexual partners during penile-vaginal sex, even when the infected partner is asymptomatic. Thus, treatment of infected men is an important public health concern 14 . In men who have sex with men (MSM), T. vaginalis rarely causes urethral or rectal infection, and screening in asymptomatic individuals is of low utility 15 .
A marked racial disparity regarding T. vaginalis was noted among African American women and men in the recent NHANES study, with an estimated prevalence of 6.8% among the black population compared to 0.4% among other groups 11 . This is consistent with 2001-2004 NHANES data which found higher rates of T. vaginalis in African American women compared to women of other races/ethnicities 16 . This pronounced racial disparity is likely multifactorial, involving differences in sexual networks, individual-level sexual risk behaviors such as larger numbers of sexual partners, and structural disparities (i.e. inadequate access to healthcare resources) [17][18][19] .
In the recent NHANES study, T. vaginalis was found to be significantly associated with older age, lower educational level, lower socioeconomic status, and having two or more sexual partners in the past year 11 . Compared to other high-income countries such as the United Kingdom, T. vaginalis prevalence in the U.S. is higher, possibly a function of the lack of public health consideration it receives 20 .
The only population for which routine T. vaginalis screening is currently recommended in the U.S. is HIV-infected women 21 . Even in the absence of symptoms, T. vaginalis in this population has been associated with high rates of adverse events such as pelvic inflammatory disease (PID) and poor birth outcomes 5,22 . Several recent studies have found high T. vaginalis prevalence (17.4-20%) 23-26 and repeat infection rates (up to 22.7% over a median of 16 months) among HIVinfected women 23 . Similar to HIV-uninfected men, T. vaginalis is less common in HIV-infected men and infrequently seen in HIV-infected MSM 24 .
Since T. vaginalis has been associated with adverse birth outcomes, its effect on pregnant women is important to consider 6 . Currently, there are no recommendations for screening of asymptomatic pregnant women for T. vaginalis. This is largely owing to results from a prior randomized controlled trial (RCT) of asymptomatic pregnant women with T. vaginalis 27 . This study found a higher risk of preterm delivery among women treated with two doses (2 grams each) of metronidazole (MTZ) 48 hours apart between 16 and 23 and between 24 and 29 weeks of gestation, respectively, versus placebo. However, this trial had several limitations including atypical MTZ dosing. In addition, the second round of MTZ was given between 24 and 29 weeks' gestation, whereas the greatest increase of pre-term delivery was at 35-36 weeks in this study. Because of this, definitive conclusions regarding an association between MTZ treatment of asymptomatic T. vaginalis during pregnancy and pre-term birth cannot be made and further studies are needed 27,28 .
The global prevalence of T. vaginalis in pregnant women varies geographically. A 2016 systematic review of 75 studies of STI prevalence among pregnant women found that T. vaginalis prevalence ranged from 3.9-24.6% in low-to middle-income countries (i.e. Latin America and Southern Africa) 29 . Recent studies found a T. vaginalis prevalence of 20% among HIVinfected pregnant women in South Africa 26 as well as high rates of incident infection in pregnant women (9.2/100 person-years) in South Africa and Zimbabwe 30 . Unexpectedly high rates of persistent T. vaginalis (44% at 21 days or more following treatment) by nucleic acid amplification test (NAAT) in pregnant women in the Southern U.S. have been noted by Lazenby et al. 31 . This is higher than the rate of 7% in pregnant women noted in a previous U.S. study 27 . Based on their data, Lazenby et al. suggested that all pregnant women with T. vaginalis should be re-tested with NAATs approximately 3 weeks post-treatment 31 .

Diagnosis
The primary diagnostic modality for T. vaginalis has traditionally been microscopic examination of a wet mount of vaginal fluid, looking for motile trichomonads 32 . While wet mount is inexpensive and rapid, its use is limited by low sensitivity, which ranges from 44-68% compared to culture 33 . Prior to the advent of NAATs, culture was the gold standard for diagnosis of T. vaginalis, with a sensitivity of 81-94% 34-36 . Diamond's medium is the traditional culture method used for the isolation of T. vaginalis 37 . However, contamination with vaginal bacteria is common, making this technique difficult 34 . Culture systems, such as the InPouch® system (BioMed Diagnostics, White City, OR), have been developed to avoid contamination by placing the specimen in a two-chambered bag, allowing for simultaneous sampling for wet mount and culture while maintaining similar efficacy to Diamond's medium 38 .
Despite relatively high sensitivity and specificity, T. vaginalis culture remains time consuming, as it requires incubation and reading of the InPouch® over several days 39 . It is categorized by the Clinical Laboratory Improvement Amendments (CLIA) as moderately complex 40 . Over the past several years, molecular testing for T. vaginalis has become the preferred diagnostic modality 21 . There are three US Food and Drug Administration (FDA)-approved T. vaginalis NAATs currently available in the U.S., all of which are able to detect co-infection with gonorrhea and chlamydia from the same specimen 33,41-43 . The Aptima® T. vaginalis assay (Hologic, Bedford, MA) was the first NAAT to be approved by the FDA for T. vaginalis detection in asymptomatic and symptomatic women 41 . This assay detects an rRNA target via transcription-mediated amplification (TMA), with sensitivity ranging from 88-100% and specificity from 98-100% 44 . It can be performed on clinician-obtained vaginal and endocervical swab specimens, urine specimens, and ThinPrep PreservCyt specimens with results in less than 8 hours 41 . Importantly, it has not been FDA approved for use in men and must be internally validated before use 33 . The BD ProbTec T. vaginalis Q x (TVQ) amplified DNA assay (BD Diagnostics, Baltimore, MD) was the second T. vaginalis NAAT approved by the FDA for use in female urine, endocervical swab specimens, and patient-or clinicianobtained vaginal specimens 42 . Similar to the Aptima® TV assay, this test is only FDA approved in women and must be internally validated prior to use in men. The TVQ assay is able to yield results in less than 8 hours 42 .
The Xpert® TV assay (Cepheid, Sunnyvale, CA) was the first T. vaginalis NAAT FDA approved for use in female urine, endocervical swab, and patient-and clinician-collected vaginal specimens as well as male urine 43 . Diagnostic sensitivity and specificity for the Xpert® TV assay range from 99.5-100% and 99.4-99.9% for female genital specimens and 97.2-99.9% for male urine specimens 43 . Once collected and placed in the testing platform, the Xpert® TV assay yields results in 60-90 minutes, allowing for point-of-care (POC) diagnosis and management 45 .
POC STI diagnostics are powerful tools, allowing providers to accurately diagnose and provide appropriate treatment for patients in the same visit. Given the communicability and frequency of T. vaginalis, rapid diagnosis and treatment could have a substantial impact on its public health burden 46 . The Solana® Trichomonas assay (Quidel, San Diego, CA) is a new rapid test for the qualitative detection of T. vaginalis DNA and can yield results within 40 minutes of specimen collection 47 . This assay was FDA approved in 2017 for the diagnosis of T. vaginalis from female vaginal and urine specimens from asymptomatic and symptomatic women, with sensitivity compared to NAAT of over 98% for vaginal specimens and over 92% for urine 47 . Solana® requires a specific testing instrument to process samples; thus, similar to NAATs, there is upfront cost associated with its use. After a specimen is collected, it is lysed by heat, diluted, and added to a reaction tube containing helicase-dependent amplification (HDA) reagents including primers specific for the amplification of a T. vaginalis-specific target sequence (https://www.quidel.com/molecular-diagnostics/ solana-trichomonas-assay). The AmpliVue® Trichomonas Assay (Quidel, San Diego, CA) is another POC test providing qualitative detection of T. vaginalis which is FDA approved for vaginal specimens from symptomatic and asymptomatic women 48 . Similar to Solana®, AmpliVue® also uses HDA technology but testing can be performed in a small handheld cartridge that requires no additional equipment (http://www.quidel.com/molecular-diagnostics/amplivueproducts). AmpliVue® has demonstrated comparable sensitivity and specificity to the Aptima® TV assay, at 90.7% and 98.9%, respectively 48 .
The OSOM® Trichomonas Rapid Test (Sekisui, Framingham, MA) is a qualitative antigen-detection immunochromatographic assay with a processing time of 10-15 minutes 49 . It is validated for the diagnosis of T. vaginalis in women from clinician-obtained vaginal specimens with a sensitivity of 83-92% and specificity of 99% 50,51 . This test has not performed well in identifying infection in male urine when compared to the Aptima® TV assay 51 ; thus, it is currently recommended in women only 21 . Because it requires no special equipment and is low cost, the OSOM® test is appealing in the setting of STI testing campaigns in low-resource settings 52 .
With the advent of these numerous high-quality T. vaginalis diagnostics, clinicians should be aware of which of the above tests are currently available in their laboratories beyond wet mount. In choosing which of the above diagnostic tests to use, the need for a rapid diagnosis should be balanced with the need for a highly sensitive test. Furthermore, there are several additional trichomoniasis diagnostic tests in the pipeline whose performance characteristics have not yet been published.

Management
The management of trichomoniasis continues to evolve, particularly in women. Per the 2015 Centers for Disease Control and Prevention (CDC) sexually transmitted disease treatment guidelines 21 , first-line treatment for trichomoniasis in HIVnegative women and men includes a single 2 gram dose of oral MTZ or tinidazole (TIN). Oral MTZ 500 mg twice daily for 7 days is an alternative therapy. These recommendations were based on several small trials conducted over 30 years ago 53-56 , several of which had a high potential for bias, as well as a desire to reduce adherence issues with multi-dose therapy. A metaanalysis of prior trichomonas treatment trials recently showed that women receiving the 7-day regimen of oral MTZ were 50% less likely to be positive at test of cure (TOC) compared to those receiving single-dose therapy 57 . In addition, RCTs of HIV-infected women with T. vaginalis 58 and HIV-negative women with T. vaginalis 59 found that the 7-day MTZ dose was more effective than single-dose therapy at TOC. While the CDC recommends only 7-day MTZ for HIV-infected women, it is highly likely that this regimen will be recommended for all women moving forward 60 . Of note, a recent literature review found no increased risk of teratogenicity with the use of MTZ (a class B drug) during pregnancy 61 . TIN has not been evaluated in pregnancy and remains a class C drug.
There have been no head-to-head comparisons of the single 2 gram dose of oral MTZ and the 7-day regimen in men. One observational study of 325 men with trichomoniasis found that treatment with the 2 gram dose of oral MTZ was unsuccessful in 42.9% of cases 62 . A second study found that the microbiological efficacy of treatment with the 2 gram dose of oral MTZ in men was 77.1% 63 . While these cure rates seem to be suboptimal, neither of these studies had a comparison group with the 7-day MTZ regimen; thus, no conclusions can be made.
In addition to MTZ, TIN is another 5-nitroimidazole medication that is FDA approved and recommended by the CDC for the treatment of trichomoniasis. TIN has better absorption and fewer gastrointestinal side effects than MTZ 64 but is ten times more expensive (with an approximate retail price of $44.66 per 2 gram dose, compared with $3.47 per 2 gram dose of MTZ at the time of writing) 3 and less likely to be adopted by clinicians. Other nitroimidazoles, such as secnidazole 65 and ornidazole 63,66 , have been used in other countries but are not currently FDA approved for the treatment of trichomoniasis in the U.S. Persistent or recurrent infection due to antimicrobial-resistant T. vaginalis or other causes should be differentiated from reinfection from an untreated or insufficiently treated sexual partner. A detailed patient history should be taken to assess the likelihood of reinfection. Following treatment failure, and if reinfection has been excluded, persistent or recurrent trichomoniasis 3 has been treated successfully with longer courses or additional doses of the same medications used in standard therapy (i.e. high-dose oral MTZ or TIN 2 grams orally daily for 7 days) 21 . Single-dose MTZ or TIN therapy should be avoided 21 . If drug resistance is suspected, the isolate can also be sent to the CDC for drug resistance testing (https://www.cdc.gov/laboratory/specimen-submission/detail.html?CDCTestCode=CDC-10239). If the patient fails the 7-day regimen of high-dose oral MTZ or TIN, two additional treatment options are available which have had successful results. The first is high-dose oral TIN 2-3 grams daily (in divided doses) plus intravaginal TIN 500 mg twice daily for 14 days 67 . The second is high-dose oral TIN (1 gram three times daily) plus intravaginal paromomycin (4 g of 6.25% intravaginal paromomycin cream nightly) for 14 days 68,69 . Paromomycin is an aminoglycoside with a different mechanism of action (destruction of ribosomal RNA) than MTZ (inhibition of nucleic acid synthesis by DNA disruption) 70 ; successful treatment of MTZ-resistant trichomoniasis with both high-dose oral TIN and intravaginal paromomycin may suggest a synergistic effect. It should be noted, however, that intravaginal paromomycin cream may cause vaginal ulceration(s) 71 . These ulcerations may spontaneously regress when therapy is stopped. Expert consultation should be sought for additional treatment options for patients who fail the above treatment options. There are currently no data regarding optimal treatment of male sexual partners of women with MTZ-resistant trichomoniasis.
Another complicated treatment scenario for women with trichomoniasis is the setting of an IgE-mediated-type hypersensitivity reaction to 5-nitroimidazoles. These patients should be managed by MTZ desensitization according to a published regimen 72,73 and in consultation with an allergy specialist 21 . Treatment of patients with T. vaginalis who are unable to be desensitized is difficult and mainly based on anecdotal data 21,74 . One option for which we have had success is intravaginal boric acid 600 mg twice daily for 60 days 75,76 .

Partner treatment
Per current CDC sexually transmitted disease treatment guidelines, concurrent treatment of all sexual partners of patients with trichomoniasis is critical for symptomatic relief, microbiologic cure, and prevention of transmission and reinfection 21 . In the last several years, the evidence for expedited partner therapy (EPT) as a mechanism of prevention of T. vaginalis infection has grown 77 . EPT, or the treatment of sexual partners of a patient diagnosed with an STI by providing treatment prescriptions to the patient without clinical assessment of the partners 78 , has been recognized as an effective option for partner treatment of chlamydia and gonorrhea 79 . The CDC currently recommends EPT as an option for partner therapy for these STIs in women and heterosexual men 21,79 . Interestingly, two prior RCTs conducted on the use of EPT in partners of women with trichomoniasis had mixed results 80,81 . Schwebke et al. found EPT to be well accepted and safe with rates of repeat infection in the EPT arm lower than those of the public health disease intervention and partnerreferral arms 81 . In an RCT by Kissinger et al., however, randomization to T. vaginalis EPT did not lead to increased partner treatment uptake or lower follow-up rates compared to standard partner referral 80 . Nevertheless, given high reinfection rates and frequent asymptomatic infection in men, EPT is still recommended as a valid means of partner therapy of T. vaginalis-infected patients 21 . EPT has also recently been shown to be effective in decreasing repeat T. vaginalis infection rates when used in conjunction with POC testing strategies in South African women 52 . Despite some data supporting EPT for patients with T. vaginalis, implementation has been limited because of legal restrictions in many states (https://www.cdc.gov/std/ept/legal/default.htm).
Additionally, EPT acceptance has been found to vary with patient and provider characteristics 82 . It is currently unknown how often providers prescribe EPT for trichomoniasis in the U.S.

Conclusion
In recent years, many advances have been made in the epidemiology, diagnosis, and treatment of T. vaginalis. The focus of these efforts, however, has largely been on women.
More study is needed on the epidemiology of trichomoniasis in men as well as how to best diagnose and treat men who are infected, particularly given its high prevalence and communicability. transport medium. J Clin Microbiol. 1999; 37(11): 3749-50.