Prevalence and risk factors associated with Chlamydia trachomatis,Neisseria gonorrhoeae,and Mycoplasma genitalium among women in Pelotas,Southern Brazil

Abstract

The frequently asymptomatic sexually transmitted infections (STIs) caused by Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Mycoplasma genitalium (MG) are poorly diagnosed in Brazil and can lead to severe complications/sequelae without timely detection and treatment. We investigated prevalence of CT, NG, and MG infections and associated demographic, behavioral, and clinical factors in consecutive women attending a gynecology and obstetrics outpatient clinic in Pelotas, Southern Brazil. Vaginal swab samples were prospectively obtained from asymptomatic and symptomatic women (n = 498) from August 2015 to December 2016 and tested with Aptima Combo2 and Aptima M. genitalium assays (Hologic). The prevalence of CT, NG, and MG was 6.8% (34/498), 1.0% (5/498), and 4.2% (21/498), respectively. Three (0.6%) cases of CT and NG co-infection and one (0.2%) case of CT and MG co-infection were identified. The risk factors associated with these bacterial STIs were youth (<30 years), no steady sexual partner, infection with additional STI, and lack of income. Bacterial STIs, particularly CT and MG, were prevalent among women, including pregnant women (60% of positive cases), in Pelotas, Brazil. Sensitive and specific diagnostic testing and early treatment are essential to control STIs, limit transmission chains, avoid future complications/sequelae, and reduce health and cost burdens on the population.

Keywords

Sexually transmitted infections diagnosis screening women Brazil

Introduction

Sexually transmitted infections (STIs) are global public health problems. In Brazil, STIs are considered a priority in the Brazilian public health system due to magnitude, vulnerability, and control issues. Major concerns include the potential for rapid dissemination, the severe complications and sequelae of untreated infections, as well as the lack of an efficient screening and testing system that can detect and treat symptomatic and particularly asymptomatic patients in a timely manner.^1,2

In 2012, the World Health Organization (WHO) estimated 357 million new cases worldwide of four curable non-viral STIs. Among the estimated bacterial STIs, Chlamydia trachomatis (CT) caused the most frequent infections (131 million cases), followed by Neisseria gonorrhoeae (NG) (78 million cases).^1,3 In Brazil, national data on STI incidence and prevalence remain scarce. Notification is mandatory only for some STIs (syphilis and HIV) and sensitive and specific laboratory diagnosis of non-viral STIs is very limited.^2,4,5 However, 12 million new cases of curable STIs are estimated to occur in Brazil each year.⁴ The prevalence of Mycoplasma genitalium (MG) infections is not estimated globally but these are likely also very common worldwide,⁶ including in Brazil.^7,8 Undetected and/or untreated infections with CT, NG, and/or MG could lead to severe complications and sequelae, disproportionally affecting women, such as pelvic inflammatory disease, abortion, ectopic pregnancy, infertility, neonatal conjunctivitis leading to blindness, and increase HIV acquisition and transmission.¹ Asymptomatic CT, NG, and MG infections are very common, particularly in women and at extragenital anatomical sites, which makes sensitive and specific testing of these STIs essential.

In Brazil, an old study performed in 2005 and examining urogenital specimens from women being treated at an STI clinic in five Brazilian state capitals using nucleic acid amplification tests (NAATs) reported prevalence rates of 7.3% for CT and 3.3% for NG.⁴ The prevalence of MG in Brazil remains mainly unknown, but according to some sporadic NAAT studies the prevalence appears to be high.^9,10 The lack of appropriate STI prevalence data in Brazil remains a major problem, particularly because the few prevalence studies are mainly from larger STI centers and cities, and the prevalence in the general population and in smaller cities is mainly unknown.⁵ Furthermore, sensitive and specific NAATs are the recommended methods for detection of CT, NG, and MG in all international guidelines.^11–14 However, appropriate NAATs are rarely used for diagnosis of non-viral STIs in Brazil and have not previously been used in the smaller city of Pelotas in Southern Brazil, where accurate prevalence data for CT, NG, or MG are lacking. In settings lacking adequate laboratory diagnostics of STIs, mainly syndromic management is used, which misses the STIs in all asymptomatic individuals that might progress into serious reproductive tract complications and sequelae and also increase the transmission of HIV. Furthermore, syndromic management results in overtreatment and incorrect treatment. Accordingly, appropriate laboratory diagnosis of non-viral STIs is crucial for individual case management as well as epidemiological surveillance.^{1,2,4,11–14} Nevertheless, in settings lacking etiologically-based diagnostics for STIs, asymptomatic individuals at higher risk for STIs can be identified by evaluation of risk factors.

The aims of the present study were to determine the prevalence of and risk factors associated with CT, NG, and MG infections in consecutive symptomatic and asymptomatic women in Pelotas, Southern Brazil diagnosed with the US FDA-approved Aptima Combo2 Assay (for CT and NG) and the new US FDA-approved Aptima Mycoplasma genitalium Assay (both assays Hologic Inc., San Diego, CA, USA).

Material and methods

Study population

A cross-sectional study was conducted from August 2015 to December 2016 examining consecutive sexually-active women, 15 years of age or older, who attended a gynecological and obstetrical care unit of the Federal University of Pelotas (Rio Grande do Sul, Brazil). Women were attending the clinic mainly because of having urogenital symptoms, having had unprotected sex, or for antenatal care. Briefly, these visits were recorded as routine checkup (57.2%), prenatal care (22.5%), cervical and breast cancer prevention (11.3%), infection (7.2%), and hormonal problems (1.8%). All women attending the clinic were invited to participate and participating women were enrolled after information about the study had been provided and they had signed informed consent (probability sampling). The exclusion criteria were being younger than 18 years old and/or had not had their sexual début. A face-to-face interview using a structured questionnaire was conducted to collect socio-demographic, behavioral, and clinical data from each patient. The questionnaire was only slightly adapted from a questionnaire previously used and validated by the Brazilian Ministry of Health (‘Brasil. Ministério da Saúde. Secretaria de Vigilância em Saúde. Programa Nacional de DST e Aids. Prevalências e frequências relativas de Doenças Sexualmente Transmissíveis [DST] em populações selecionadas de seis capitais brasileiras, 2005/Ministério da Saúde, Secretaria de Vigilância em Saúde, Programa Nacional de DST e Aids. – Brasília: Ministério da Saúde, 2008.’).

Specimen collection, laboratory diagnostic tests, and treatment

All participants underwent a speculum examination and vaginal samples were collected (one swab per woman) using the Aptima Vaginal Swab Specimen Collection Kit (Hologic), according to manufacturer’s instructions. The specimens were stored at −20°C prior to shipment to the WHO Collaborating Centre for Gonorrhoea and other STIs in Örebro, Sweden for NAAT analysis.

The US FDA-approved Aptima Combo2 Assay and the new US FDA-approved Aptima Mycoplasma genitalium Assay (Hologic) were performed, according to the instructions of the manufacturer, on the Panther system (Hologic).

All women diagnosed with any of the investigated STIs, along with their sexual partners, received medical treatment according to the Brazilian Ministry of Health guidelines.²

Independent variables

The socioeconomic, demographic, behavioral, and clinical characteristics of the women collected were as follows: age, in full years and grouped as 15–19, 20–30, 31–40, and >40 years; self-declared ethnicity, grouped as white and other (brown/black); education level in years, grouped as 0–4, 5–8, 9–11, and >11 years (in accordance with the levels of the Brazilian educational system)^2,4; marital status (married or consensual/steady union, single, widowed, separated/divorced); own income (yes, no); HIV status (yes, no); symptoms (yes, no); vaginal discharge (yes, no); smoking currently (yes, no); any alcohol use (yes, no); drug use (yes, no).

Statistical analysis

All data were analyzed using the Statistical Package for Social Sciences version 17.0. A preliminary analysis was performed using exploratory techniques to check for distribution patterns and trends among the variables. Bivariate analysis was then performed to determine associations between the variables. Chi square tests were used to compare proportions and Student’s t-tests and variance analysis were used for testing differences between mean values. Bivariate and multivariate odds ratios (ORs) (adjusted for potential confounders) and 95% confidence intervals (CIs) were reported. Variables significant at p < 0.20 in bivariate analysis and known confounders (e.g. age and education) were included in the multivariate analysis, using a stepwise multiple logistic regression model. In the multivariate analysis, p < 0.05 was considered statistically significant.

Ethical approval

This study was approved by the Universidade Federal de Pelotas School of Medicine’s Ethics Committee (873.180) and the National Commission on Ethics in Research (1.838.829).

Results

In total, 498 women, of which 123 (24.7%) attended for antenatal care, were enrolled in the study. The socioeconomic, demographic, behavioral, and clinical characteristics of the women have been summarized in Table 1. Their mean (range) age was 36.5 (15–77) years and their mean education was 5.5 years. The majority of the women were symptomatic (59.8%), white (75%), had low education levels, and were either married or in a consensual union (Table 1). The main symptoms were vaginal discharge (78.9%, 235 of the 298 symptomatic women), itching (39.9%, 119/298), malodor (35.2%, 105/298), dyspareunia (24.8%, 74/298), and dysuria (19.8%, 59/298).

Table 1.

Distribution of demographic, socioeconomic, and behavioral characteristics of women aged 15–77 years (N = 498), Pelotas, Southern Brazil, 2015–2016.

Variable	N ^a	%
Age (years; N = 498)^b
15–19	45	9.0
20–30	124	24.9
31–40	127	25.5
>40	202	40.6
Ethnicity (N = 493)^c
White	367	74.4
Other	126	25.6
Education level (years; N = 487)^d
0–4	75	15.4
5–8	181	37.2
9–11	169	34.7
>11	62	12.7
Marital status (N = 494)
Married or consensual/steady union	342	69.2
Single	105	21.3
Widowed	26	5.3
Separated/divorced	21	4.2
Own income (minimum wage) (N = 486)
Yes	268	61.3
No	188	38.7
HIV status (N = 498)
Yes	129	25.9
No	369	74.1
Symptoms (N = 474)^e
Yes	298	62.9
No	176	37.1
Vaginal discharge (N = 298)
Yes	235	78.9
No	63	21.1
Smoking currently (N = 489)
Yes	99	20.2
No	390	79.8
Any alcohol use (N = 475)
Yes	26	5.5
No	449	94.5
Drug use (N = 475)^f
Yes	13	2.7
No	462	97.3

^aTotals do not match due to lack of information for some variables.

^bAge categorization used at the recruitment clinic and in a recent Trichomonas vaginalis study examining the same sample of women.¹⁵

^cBlack and Brown were grouped to allow bivariate and multivariable analysis.

^dIn accordance with the levels of the Brazilian educational system.^2,⁴

^eVaginal discharge (78.9%), itching (39.9%), malodor (35.2%), dyspareunia (24.8%), and dysuria (19.8%).

^fIllicit drug use, e.g. marijuana, cocaine, crack, etc.

The prevalence of urogenital CT, NG, and MG is shown in Table 2. Briefly, the prevalence of CT, NG, and MG was 6.8% (34/498), 1.0% (5/498), and 4.2% (21/498), respectively. Among these cases, three (0.6%) cases of CT and NG co-infection and one (0.2%) case of CT and MG co-infection were identified. Of the pregnant women (n = 123), 29.3% (n = 36) were positive for any of the tested STIs, that is CT (n = 18), NG (n = 1), or MG (n = 17). Overall, 434 (87.1%) of the women were negative for all three tested STIs (Table 2).

Table 2.

Prevalence of urogenital Chlamydia trachomatis, Neisseria gonorrhoeae, and Mycoplasma genitalium in women (n = 498), Pelotas, Southern Brazil, 2015–2016.

	N	%	95% CI
C. trachomatis only	34	6.83	4.59–9.01
N. gonorrhoeae only	5	1.00	0.13–1.87
M. genitalium only	21	4.22	2.44–5.96
Mixed infection^a	4	0.80	0.02–1.58

CI: confidence interval.

^aAmong the infected cases, three (0.6%) cases of CT and NG co-infection and one (0.2%) case of CT and MG co-infection were identified.

According to the bivariate analysis, young women (15–19 years of age) were 20.6 times more likely to be infected by CT, and women aged 20–30 years were 9.9 times more likely to be infected with MG. Women with NG infection were 22.4 times more likely to be infected with CT (Table 3). In bivariate analysis for NG, some variables were grouped due to the low number of positive samples (n = 5). Being adolescent (15–19 years) (OR = 10.1 [95% CI 1.4–75.6]), being single (OR = 11.9 [95% CI 1.2–116]), and having co-infection with CT (OR = 22.4 [95% CI 3.6–138.8]) were risk factors for NG (data not shown).

Table 3.

Crude analysis of the risk factors for Chlamydia trachomatis and Mycoplasma genitalium infection in women (n = 498), Pelotas, Southern Brazil, 2015–2016.

Variable	C. trachomatis(%)	Bivariate analysis		M. genitalium(%)	Bivariate analysis
Variable	C. trachomatis(%)	OR 95% CI	P-value	M. genitalium(%)	OR 95% CI	P-value
Age (years)			<0.001			0.026
15–19	11 (24.4%)	20.6 (5.5–77.7)		2 (4.4%)	4.5 (0.6–32.6)
20–30	10 (8.4%)	5.9 (1.6–21.9)		11 (9.3%)	9.9 (2.1–45.4)
31–40	8 (6.5%)	4.4 (1.1–16.9)		6 (4.8%)	4.9 (1.0–24.6)
>40	3 (1.5%)	1		2 (1.0%)	1
Marital status			0.001			0.751
Married/consensual union	14 (4.2%)	1		12 (3.6%)	1
Single	17 (17.3%)	4.8 (2.3–10.1		6 (6.1%)	1.7 (0.6–4.8)
Widowed	2 (7.4%)	1.8 (0.4–1.4)		1 (3.7%)	1.0 (0.1–8.2)
Separated/divorced	1 (5%)	1.2 (0.1–9.6)		1 (5%)	1.4 (0.2–11.4)
Own income (minimum wage)			0.06			0.002
Yes	13 (4.5%)	1		5 (1.7%)	1
No	21 (11.5%)	2.7 (1.3–5.6)		15 (8.2%)	5.1 (1.8–14.2)
NG infection			0.001			0.703
Yes	3 (8.3%)	22.4 (3.6–138.8)		0 (0%)	1
No	2 (0.4%)	1		21 (4.3%)	1.5 (0.2–11.4)
Ethnicity			0.512			0.13
White	27 (7.6%)	1.3 (0.6–3.1)		12 (3.4%)	0.5 (0.2–1.2)
Other	7 (5.8%)	1		8 (6.7%)	1
Education level			0.437			0.259
0–4	2 (2.8%)	1		2 (2.8%)	1.5 (0.1–17.5)
5–8	16 (8.8%)	3.4 (0.8–15.1)		12 (6.6%)	3.8 (0.5–30.0)
9–11	11 (6.6%	2.5 (0.5–11.4)		5 (3.0%)	1.7 (0.2–14.6)
>11	4 (7.3%)	2.7 (0.5–15.6)		1 (1.8%)	1
HIV status			0.38			<0.001
Yes	11 (8.5%)	0.7 (0.3–1.5)		13 (10.1%)	5.0 (2.0–12.5)
No	23 (6.2%)	1		8 (2.2%)	1
Symptoms			0.99			0.265
Yes	26 (8.7%)	0.5 (0.2–1.1)		15 (5.0%)	1.8 (0.6–5.0)
No	8 (4.6%)	1		5 (2.9%)	1
Smoking currently			0.878			0.265
Yes	7 (7.5%)	0.9 (0.4–2.2)		6 (6.5%)	1.8 (0.6–5.0)
No	27 (7.1%)	1		14 (3.7%)	1
Any alcohol use			0.921			0.924
Yes	2 (7.7%)	0.9 (0.2–4.1)		1 (3.8%)	0.9 (0.1–7.0)
No	32 (7.1%)	1		19 (4.2%)	1
Drug use			0.99		0.99
Yes	0 (0%)			1 (7.7%)
No	34 (7.4%)			19 (4.1%)

CI: confidence interval; NG: Neisseria gonorrhoeae; OR: odds ratio.

Table 4 shows the variables that remained statistically significant (p < 0.05) in the multivariate analysis for CT, NG, and MG. Variables significantly associated with urogenital CT infection were as follows: age under 19 years (p = 0.025), being single (p = 0.004), having no income (p = 0.021; e.g. being housewife or unemployed), and having co-infection with NG (p = 0.014). Only one significant association was found for urogenital NG, co-infection with CT (p = 0.008), while the significant associations with urogenital MG infection were having no income (p = 0.004) and HIV co-infection (p = 0.001).

Table 4.

Multivariate analysis of factors associated with Chlamydia trachomatis, Mycoplasma genitalium, and Neisseria gonorrhoeae in women (n = 498), Pelotas, Southern Brazil, 2015–2016.

Factors	Adjusted odds ratio	95% CI	P-value
C. trachomatis
Age
15–19 years	9.8	2.2–43.2	0.025
Marital status
Single	4.6	1.9–11.0	0.004
Own income (minimum wage)
No	2.8	1.2–6.9	0.021
N. gonorrhoeae co-infection
Yes	28.8	2.0–417.9	0.014
M. genitalium
Has own income (at least minimum wage)
No	5.2	1.7–15.5	0.004
HIV infection
Yes	5.4	2.0–14.4	0.001
N. gonorrhoeae
C. trachomatis co-infection
Yes	26.3	2.4–294.0	0.008

CI: confidence interval.

Discussion

In the present study, the prevalence of CT and MG was high (6.8 and 4.2%, respectively) among women, including pregnant women, in Pelotas, Brazil. NG prevalence was lower but still significant (1%). Youth (particularly 15–19 years), being single or infected with additional STI, and lacking own income were risk factors associated with CT infection. MG infection was associated with lacking own income and living with HIV.

In Brazil, STI studies should be the main source of infection prevalence data; however, these are mostly rare multicenter studies that offer time-definitive testing for specific groups, such as HIV-positive patients and pregnant women.^5,16 Accordingly, extremely few studies have simultaneously investigated the occurrence of several non-viral STIs¹⁷ in mixed, high-risk, non-specific clinical populations, as was done in the present study.

In the present study, the prevalence of CT, NG, and MG infections determined with the highly sensitive and specific Aptima Combo2 Assay and Aptima Mycoplasma genitalium Assay (Hologic) was 6.8, 1.0, and 4.2%, respectively. Of these women positive for any tested STI (n = 60), 36 (60.0%) were pregnant, which means that 29.3% (36/123) of all pregnant women were infected with CT (n = 18), NG (n = 1), or MG (n = 17). If these STIs are not effectively treated, they can cause several severe adverse pregnancy outcomes.^1,2 Previous Brazilian prevalence studies using NAATs have found CT infection rates ranging from 1.1 to 13.9% and NG infection rates from 0.5 to 21.5%. Some studies have been conducted on populations similar to this one (i.e. public health system patients in hospitals or university clinics), while the majority have been conducted targeting specific groups of patients, such as only HIV patients, pregnant women, and young people.^5,8,17–25 The CT and NG infection prevalence identified in the present study is in concordance with those reported by the Brazilian Ministry of Health.⁴ Our sample reflected the most varied population types, since it was drawn from a reference center for women seeking various types of care: routine examinations, STI treatment, antenatal care, etc.

Regarding MG, studies in Brazil remain very scarce,¹⁷ the WHO has no global infection estimates, and MG infection is not mandatorily reported internationally.^1,13 Nevertheless, a number of prevalence studies worldwide have identified significant rates in both men and women.^6,13,26–28 In the present study, the MG rate among women was 4.2%. Other Brazilian studies have reported rates from 0.9 to 28.1%.^{8,10,17,20,28} Accordingly, MG infections are prevalent worldwide. In addition, resistance in MG to the first- and second-line treatments (azithromycin and moxifloxacin, respectively) has significantly increased internationally during the last decade.^1,29

The Brazilian Ministry of Health has developed a clinical protocol of therapeutic guidelines for health professionals, which proposes combined strategies of testing and treatment for asymptomatic STIs as well as the management of symptomatic cases through syndromic management flowcharts.² Some infections, such as syphilis, receive priority treatment, but the protocol also strongly supports strategies to reduce other STIs. In the absence of laboratory diagnosis, the guidelines recommended that the sexual partners of men with urethritis should be treated.² However, in the present study, the presence of symptoms was not significantly associated with infection, which makes the recommended flowcharts of limited value.

A total of 129/498 (25.9%) of the participants were HIV-positive and had routine outpatient care. We found an association between MG infection and HIV infection (adjusted OR = 5.4, 95% CI 2.0–14.4). Other studies have shown that infection with this pathogen, as well as other inflammatory STIs such as gonorrhoea and CT infection (not observed in the present study), can be a co-factor in acquiring HIV, more than doubling the risk.^30–32 Regarding risk factors, we found that CT and NG infections were more prevalent among youth than MG infection, which is more frequently reported in patients of higher ages. CT infection was also associated with lacking own income and being single. The associations between CT infection and younger ages and being single are well-known associations, likely mainly explained by the increased risk for STIs in these populations due to factors such as a higher number of unprotected contacts with new and/or multiple partners and a lower condom usage. The association between CT infection and lacking own income might be at least partly explained by socioeconomic factors such as lower access to health care in general, STI diagnosis and treatment, prevention, and awareness. This is all mainly in concordance with previous studies.^{1,13,19,33–35} Finally, among CT and NG infections, infection with one was a risk factor for the other.

The limitations of the present study included that no sample size estimation or representativeness analysis for the study or follow-up after treatment of the women, which can be inherent limitations of this type of cross-sectional study, was performed. Furthermore, response and/or memory biases when responding to the structured questionnaire during the interview can never be excluded. Finally, no systematic testing of all women for syphilis and HIV was conducted.

In conclusion, bacterial STIs, particularly CT and MG, were prevalent among women, including pregnant women (60% of positive cases), in Pelotas, Southern Brazil, which is very worrying since laboratory diagnosis of these STIs is not currently offered in the Brazilian public health system. It is essential to implement sensitive and specific diagnostic testing, using validated and quality-assured NAATs, and early treatment of STIs in Brazil to control the frequently asymptomatic STIs, limit the transmission chains, avoid future complications/sequelae, and reduce the health and cost burden on the population. In resource poor settings, NAAT diagnostics can be cost-effectively centralized at larger, high-throughput reference or general diagnostic laboratories.

Footnotes

Declaration of conflicting interests

The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

The authors disclosed receipt of the following financial support for the research, authorship and/or publication of this article: This work was supported by grants from the Programa de Apoio a Pós-Graduação and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, Brasília, DF, Brazil; Conselho Nacional de Desenvolvimento Cientifico e Tecnologico, Brazil; the Örebro County Council Research Committee and the Foundation for Medical Research at Örebro University Hospital, Örebro, Sweden.

ORCID iD

M Unemo

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