The Prevalence of Mycoplasma genitalium and Association With Human Immunodeficiency Virus Infection in Symptomatic Patients, Johannesburg, South Africa, 2007-2014.

BACKGROUND: Mycoplasma genitalium is associated with genital discharge syndrome, but limited prevalence data are available in South Africa. The prevalence rates of M. genitalium infection and human immunodeficiency virus (HIV) coinfection were determined in urogenital specimens collected from male and female patients presenting with genital discharge syndrome to a primary health care center in Johannesburg, South Africa from 2007 through 2014.
METHODS: Genital specimens from 4731 patients were tested by a validated in-house multiplex real-time polymerase chain reaction assay for the detection of Neisseria gonorrhoeae, Chlamydia trachomatis, Trichomonas vaginalis, and M. genitalium. Sera were tested for HIV infection using the Determine HIV 1/2 and Unigold assays.
RESULTS: The relative prevalence of M. genitalium in males and females was 8.9% and 10.6%, respectively. The prevalence of HIV infection in those infected with M. genitalium, without other sexually transmitted infections (STIs), was significantly higher than in those without M. genitalium infection (48.9% vs. 40.5%, P = 0.014). This significant difference in HIV seroprevalence was particularly observed among females in the study cohort.
CONCLUSIONS: The relative prevalence of M. genitalium and its association with prevalent HIV among females with vaginal discharge syndrome (VDS) calls for further research on the potential role of M. genitalium in the transmission and acquisition of HIV.

Mycoplasma genitalium is an emerging cause of genital discharge and has been implicated in male and female urogenital infections globally.[1] More than 25 years after its initial isolation from men with non-gonococcal urethritis, data supporting the role of M. genitalium as an etiological agent of acute and persistent male non-gonococcal urethritis, has increased over the years.[2] Mycoplasma genitalium has also been significantly associated with both cervicitis and pelvic inflammatory disease (PID) in women.[1]

The prevalence of M. genitalium infection varies widely in different countries. Studies conducted in Western Europe, North America and Australia, have estimated the prevalence of M. genitalium in men to range between 1% and 3.3%[3,4] and among women to range between 1% and 6.4%.[5] In a systematic review conducted by Baumann et al., the prevalence estimates of M. genitalium in the general population were reported to be low, ranging from 1.3% in countries with a high human development index to 3.9% in those with lower human development index.[6] Prevalence estimates were highest in female commercial sex workers ranging from 13.2% in community-based samples to 26.3% in clinic-based samples.

In comparison with other sexually transmitted infections (STIs), there is a paucity of information on the prevalence of M. genitalium in South Africa.[7-11] In a recent study conducted in South Africa, among adolescents and young adults with asymptomatic genital tract infections, the prevalence among females and males was 9.6% and 3.3%, respectively.[10] In another study performed among asymptomatic people living with human immunodeficiency virus (HIV)/acquired immune deficiency syndrome (AIDS), the overall M. genitalium prevalence was 6.1%.[11] Mycoplasma genitalium has been associated with HIV acquisition and coinfection in a number of studies.[11-14] Investigators have documented that M. genitalium may be an independent risk factor for the acquisition of HIV in women in Zimbabwe and Uganda, respectively.[12,13]

Mycoplasma genitalium is a fastidious and slow growing STI pathogen, which makes it difficult to culture; therefore, molecular methods have been the predominant means of detecting M. genitalium in clinical specimens.[1,15] The use of nucleic acid amplification tests, such as polymerase chain reaction (PCR)[16] and transcription-mediated amplification (TMA)[17] for the detection of M. genitalium DNA and 16S rRNA targets, respectively, has increased an understanding of the epidemiology of M. genitalium infection. The center for HIV and STIs at the National Institute for Communicable Diseases, in South Africa, has been conducting STI microbiological surveillance in Johannesburg since 2007. We describe trends in the prevalence of M. genitalium infection, and association with HIV infection, in patients presenting with genital discharge syndrome to a primary healthcare facility in Johannesburg, South Africa, spanning a period of 8 years (2007–2014).

MATERIALS AND METHODS

Study Participants

Consecutive patients presenting with male urethral syndrome (MUS) or VDS were enrolled, with written informed consent, as part of annual STI surveys at Alexandra Health Center in Johannesburg between 2007 and 2014. Alexandra Health Center is a community-orientated primary health care facility that provide services, such as HIV, AIDS, and tuberculosis treatment; STI syndromic treatment; and HIV counseling, testing, and prevention services. Demographic data were collected by a nurse-administered questionnaire. STI syndromes were treated according to national syndromic management guidelines.[18] All patients were offered on-site HIV voluntary counseling and testing.

Clinical Specimens

Each year between 2007 and 2014, urine and endocervical swab specimens were collected from males and females, respectively. Additionally, a 10-mL whole blood sample was collected from each participant.

Laboratory Methodology

DNA extraction was undertaken with an automated DNA extraction method (X-tractor Gene; QIAGEN, Hilden, Germany). A validated in-house real-time multiplex PCR assay was performed to detect Neisseria gonorrhoeae, Chlamydia trachomatis, Trichomonas vaginalis, and M. genitalium.[19] Those DNA extracts with inconclusive M. genitalium results on the in-house PCR assay, were re-tested using a commercial M. genitalium real-time PCR assay (Sacace Biotechnologies, Como, Italy) targeting the DNA subunit B of M. genitalium.[13] Human immunodeficiency virus screening was performed on sera (prepared from whole blood) by the use of two rapid tests, namely the DetermineHIV 1/2 assay (Abbott Laboratories, Japan) and Unigold™ assay (Trinity Biotech PLC, Ireland).

Statistical Analysis

Data were captured onto a study specific Microsoft Access database and exported into Stata 14.2 (Stata Corporation, College Station, TX), for analysis. Descriptive statistics were used to describe the prevalence of STI pathogens in MUS and VDS. The χ2test was used to test for the association of STI pathogens and HIV serostatus. A P value less than 0.05 was considered statistically significant. Univariable and multivariable piecewise logistic regressions with splines at each of the calendar years were used to determine demographic and clinical factors independently associated with M. genitalium infection over the defined period.

RESULTS

A total of 4731 specimens were obtained from patients presenting with genital discharge syndrome in the surveillance period 2007 to 2014. Of these, 2509 (53%) were from males and 2222 (47%) from females. The median age of both males and females at enrolment was 28 years (interquartile range, 24–32 and 23–34, respectively). At enrolment, a significantly higher percentage of females had a history of STI syndrome in the preceding 12 months compared to males (63.7% vs. 29.7%, P < 0.001). None of the demographic and clinical variables studied (ie, age, sex, year of specimen collection, history of previous STI in the preceding 1-year period, history of multiple sexual partners) was significantly associated with M. genitalium infection.

During the 8-year surveillance period, the overall relative prevalence of M. genitalium infections among males and females was 8.9% and 10.6%, respectively (Fig. 1). The highest prevalence in males was observed in the year 2007 (15.1%), and in females in 2013 (13.6%). Trend analysis was performed across all 8 years, which showed a statistically significant decline in the relative prevalence of M. genitalium infection in males (15.1% to 5.3%, P = 0.010). However, no such trend was observed among females in the 8-year period (P = 0.389) (Fig. 1). Our in-house multiplex real-time PCR also tested for other STI pathogens in the genital discharge specimens. In males, N. gonorrhoeae, C. trachomatis, and T. vaginalis were detected in 76.7%, 24.4%, and 5.4% of participants, respectively. The 8-year trend analyses for other STIs revealed a statistically significant upward trend for N. gonorrhoeae (P = 0.024) and C. trachomatis (P = 0.039) and a significant downward trend for T. vaginalis (P = 0.0007) in males. Among females, the prevalence of N. gonorrhoeae, C. trachomatis and T. vaginalis was 13.5%, 16.1%, and 22.7%, respectively (data not shown), with a statistically significant upward and downward trend for C. trachomatis (P = 0.021) and T. vaginalis (P = 0.031), respectively. However, trend analysis for N. gonorrhoeae, revealed an upward trend approaching statistical significance (P = 0.059).

Figure 1

Prevalence of M. genitalium infection detected in symptomatic males (n = 222) and females (n = 231). MG, Mycoplasma genitalium; MUS, Male Urethral Syndrome.

The association of HIV seropositivity and M. genitalium infection status among males and females is depicted in Table 1. Over the 8-year period, in the absence of other STIs, the overall prevalence of HIV in those infected with M. genitalium, was significantly higher than in those without M. genitalium infection (48.9% vs. 40.5%, P = 0.014) (Table 1). A similar trend was observed in the presence of other STIs, whereby, the prevalence of HIV coinfection in those with M. genitalium, was significantly higher than in those without M. genitalium infection (49.6% vs. 40.5%, P = 0.006) (Table 1).

TABLE 1

Association of HIV Coinfection With M. genitalium Among Males and Females (2007–2014)

When stratified by gender, there was no significant difference in the prevalence of HIV-coinfection observed in males with or without M. genitalium, either in the presence (38.6% vs. 33.2%, P = 0.207) or absence (27.4% vs. 33.7%, P = 0.201) of other STIs. In females, the prevalence of HIV infection in those coinfected with M. genitalium was significantly higher than in those without M. genitalium infection, regardless of the presence (62.9% vs. 48.8%, P = 0.005) or absence (65.1% vs. 48.5%, P < 0.001) of other STIs (Table 1).

DISCUSSION

This study sought to assess trends in the relative prevalence of M. genitalium infection in a population of adult males and females seeking care at Alexandra primary health care facility in Johannesburg. A population with symptoms of genital discharge was chosen as a preferential STI target population to address the association of HIV infection in those infected with M. genitalium, with or without other STI pathogens. These findings were reviewed in the context of previously published data.

The relative prevalence of M. genitalium among females in our study, was higher than that reported in a study performed in Zimbabwe, where 7.0% of women with symptomatic vaginal discharge were infected with M. genitalium.[20] This prevalence is comparable to that found in studies of high-risk women in Uganda (14%),[21] Kenya (12.9%–16%),[22,23] and the United States (11.3%).[24] However, these rates were higher than those found in the general female population of other countries such as Denmark and Australia (3%–4%).[4,5] The variation in M. genitalium prevalence across studies may be due to differences in study methods, study populations, specimen sampling methods, and diagnostic assays used.

In our study, the 8.9% M. genitalium prevalence in males was lower than the prevalence of 13.7% in males with visible symptoms of urethritis presenting to a family practitioner in Pretoria, South Africa,[9] but similar to the prevalence of 9.0% reported among symptomatic males in Belgium.[25] Over the 8-year period, there was a decline in the relative prevalence of M. genitalium from 15.1% to 5.3% in males. This decline may, in part, have been due to a statistical significant increase in the relative prevalence of other STI pathogens such as N. gonorrhoeae (69.3% to 80.1%, P = 0.024) and C. trachomatis (23.4% to 29.6%, P = 0.039), over the 8-year period (data not shown).

In South Africa, the total number of persons living with HIV increased from an estimated 5.09 million in 2007 to 7.52 million by 2018.[26] Approximately one-fifth of South African women in the reproductive age group (15–49 years) is HIV positive.[26] Our study showed that the prevalence of HIV infection among M. genitalium infected females was higher than the prevalence of HIV in M. genitalium uninfected females regardless of the absence and presence of other STIs. The high prevalence of HIV infection in M. genitalium infected females supports the idea of further research into this association. Infection with M. genitalium has been reported to lead to cervical inflammation, and it is possible that the infection also increases the risk of HIV-acquisition, as is the case in other STIs.[27] Other African studies have reported a high prevalence of HIV coinfection in M. genitalium in both high and low risk women.[28,29] In our M. genitalium infected cohort, there was no significant difference in the prevalence of HIV among those who were coinfected with other STIs versus those who had no STI coinfection (49.6% and 48.9%, respectively).

Studies of HIV-STI interactions have been conducted mostly on individuals not receiving antiretroviral therapy (ART). Less is known about the impact of STI coinfections on HIV shedding from individuals on ART. The extended life expectancy associated with ART and the potentially higher exposure of the HIV-infected individuals to STIs may increase the prevalence of STI coinfections, particularly with public awareness of the decreased infectiousness of HIV while on ART, leading to risk compensation with a consequential decrease in condom usage.[30] A prospective study would be required to better understand the effects of coinfection with M. genitalium on HIV transmission.

Ciprofloxacin and doxycycline were the recommended syndromic treatment options for patients presenting with MUS and VDS, before 2008, but due to the rapid emergence of fluoroquinolone-resistant Neisseria gonorrhoeae within South Africa the treatment regimen was changed in 2008 to cefixime and doxycycline. Currently, the STI Syndromic Management Guidelines (2015) for South Africa incorporate the use of 1 g azithromycin, orally, as a single dose in the treatment of MUS and VDS. Therefore, before 2015, a 7-day doxycycline course was used to provide syndromic cover for M. genitalium infection; and since 2015, single-dose azithromycin therapy has been used. Using the same samples from this article, we have recently conducted a study on the prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium among symptomatic patients (article in press).

Strengths of the study include the large number of participants enrolled and tracking the prevalence of M. genitalium over an 8-year surveillance period. The major limitation of this study is that patients were only recruited in one public health care center in Johannesburg; hence, the results may not be generalizable to other provinces or populations.

In conclusion, M. genitalium was found to represent an important microbial pathogen among patient presenting with genital discharge syndromes in Johannesburg. The significant association of M. genitalium with HIV among females with VDS calls for further research on the potential role of M. genitalium in the transmission and acquisition of HIV.