Background
Chlamydia trachomatis one of the most common sexually transmitted bacterial infections worldwide [
1]. According to the World Health Organization,
C. trachomatis affects approximately 400 million people worldwide. However, the actual number of infected individuals may be higher as
C. trachomatis infection is often asymptomatic in 50% of men and 80% of women [
2].
C. trachomatis infection is more prevalent in women than in men, and its prevalence varies with age and country of residence. With a unique biphasic developmental cycle,
C. trachomatis can cause non-gonococcal urethritis and pelvic inflammatory disease, leading to ectopic pregnancy. As a result, it is gradually becoming an important public health issue for women [
2]. In some developed countries,
C. trachomatis screening programmes have been implemented to reduce its transmission and reproductive tract complications. In Sweden, extensive Chlamydia screening of asymptomatic young women in a variety of health care settings were recommended by Swedish Institute for Infectious Disease Control, and all testing and treatment are free of charge. Moreover, the Communicable Diseases Act has made it mandatory to report
C. trachomatis genital infections including contact tracing, mandatory partner notification, and compulsory testing of suspicious partners [
3]. In 1979, the US Centers for Disease Control established the first training center and model clinics for sexually transmitted disease prevention and recommended annual screening of all sexually active women aged ≤ 25 years [
4]. Following large-scale screening of asymptomatic women, the prevalence of
C. trachomatis has reduced in developed countries to some extent, but not in low-resource settings, such as Africa and Asia, including Burundi, India, and poor regions of China.
C. trachomatis genotyping is useful for monitoring re-infections and treatment efficacy. Furthermore, it provides useful information for the clinical treatment and vaccine development for
C. trachomatis infection [
3].
C. trachomatis is classified into 19 serovars on the basis of antibody specificity toward the major outer membrane protein (MOMP). These serovars exhibit distinct tissues tropism [
5], e.g., serovars A–C cause trachoma, serovars D–K cause oculogenital infections, and serovars L1–3 cause lymphogranuloma venereum. The major outer membrane protein of
C. trachomatis is encoded by a single copy gene of
omp1, which differs across serovars [
6]. This protein is an immunodominant antigen and contains four variable segments (VS1–4) [
2,
7], which are flanked and interspaced with five constant domains [
8]. Although several studies have described the
omp1 mutation sites [
6,
9], their relationship with the clinical manifestations remains unclear.
The use of antibodies to identify
C. trachomatis infections often underestimates their prevalence due to the inability to differentiate between recently acquired and previous infections. Furthermore, the likelihood of seropositivity increases with the cumulative number of infections [
10]. Pgp3 is highly conserved across isolates and rarely found in
C. pneumonia, which was recognized as a sensitive and unique serum antibody biomarker of
C. trachomatis infection due to the capability to avoid cross-reactivity with other
Chlamydia spp [
11]. Besides, 98.7% sequence identity of Pgp3 among different
C. trachomatis serovars maintains the cross-reactivity within
C. trachomatis species [
12]. Pgp3-based serological testing was then widely used to monitor
C. trachomatis infections among younger children and predict the development of infertility due to tubal dysfunction [
13]. Thus, knowledge regarding the seroepidemiology of
C. trachomatis is important for molecular epidemiological investigation and determining the prevalence and incidence of infections, for differentiating between recent and past infections, and for identifying subclinical infections.
Our previous studies have shown that
C. trachomatis infection is associated with low-grade intraepithelial neoplasia [
14]. The clinical manifestations of
C. trachomatis infection vary with the genotype, i.e., almost half of the asymptomatic patients have been infected with serovar E. Infections with serovars F and G are closely associated with a young age and lower abdominal pain, respectively [
15]. However, the effects of
C. trachomatis pgp3 antibody,
omp1 genotype, and
omp1 gene mutation (sense and nonsense mutations) on cervical intraepithelial neoplasia (CIN) and vaginal inflammation have not been well-elucidated.
Our previous studies revealed that women with infertility have a similar
C. trachomatis prevalence compared to healthy women attending physical examination center (PEC).
C. trachomatis infection is a significant risk factor for female infertility. Although
C. trachomatis infections of the female genital tract may recover spontaneously over several days, reinfection occurs in 10–20% of cases, typically within 12 months [
16]. Although it is difficult to detect
C. trachomatis DNA among women with infertility and passive infection, the antibody can persist for a long period. Accordingly, we speculated that women with infertility may have a high
C. trachomatis seroprevalence, representing both recently acquired and previous infections. To test this, we enrolled a large number of women from the infertility and gynecology clinics and PEC to assess the prevalence of
C. trachomatis infection, its subtype distribution,
omp1 mutations, seroprevalence, and associated cervical lesions.
Discussion
The prevalence rates of
C. trachomatis DNA and pgp3 antibody in a representative sample of Southern Chinese women aged 13–87 years were 3.76% and 47.46%, with the highest DNA prevalence among women aged 14–20 years and the highest seroprevalence rate among women aged 21–30 years, which was in line with the results reported from developing countries [
7]. A relatively low pgp3 seropositive rate of 28.1% was previously reported among individuals aged 18–65 years from northern China [
19], which may be due to an unbalanced age distribution and regional differences.
In our analysis, age was closely associated with
C. trachomatis infection. We found a high prevalence of
C. trachomatis infection in young women aged < 30 years, with the prevalence decreasing at older ages. The highest prevalence was reported in adult women aged 18–24 years in the general population in Germany [
19]. In southwest China,
C. trachomatis infection was more common in women aged 25–34 years [
23], suggesting that young women were more susceptible to
C. trachomatis infection, which may be due to their sexual activities, low educational status, absence of appropriate sexual education, and poor vaginal hygiene [
25]. Tailored counseling coupled with annual screening may decrease the burden of
C. trachomatis infection among the young population.
In this study, we found that
C. trachomatis serovar E was the most prevalent
C. trachomatis genotype in the female lower genital tract, which was congruent with results reported from Sweden, Stockholm County, and Finland [
3,
18], while serovar D was the most common in Liuzhou and serovar F in Thailand [
1]. Furthermore, we found that F was the most prevalent (24%) in women with positive
C. trachomatis pgp3 antibody. Genotype F was associated with a higher urogenital infections loads and greater disease progression [
26]. A high titer of antibody against the powerful
C. trachomatis pgp3 antigen may be secreted during infection, which can lead to an autoimmune response.
C. trachomatis omp1 is evolutionarily highly conserved and is thought to play a vital role in protective immunity, which consists of 5 regions of conserved sequence that alternate with 4 variable regions (VS1–VS4). The
omp1 VS1-VS2 are surface exposed and allow genotype classifications, which may alter its function, antigenicity and clinical manifestations [
27,
28]. We found 83 genetic variants of the 309
omp1 gene sequences, in accordance with previously reported genetic variability of 10–81% [
29]. Genotype J was one of the most mutable genotypes with a mutation rate of 98.3%. The identification of mutations at positions 150 and 506 in genotype J is a new finding. Although the mutations occurred at the position 369 diverging from the commonly used reference strain J/UW36, they were confirmed to be identical to the C. trachomatis J/isolate 6858 and Taiwanese genotype J strains [
2]. For the most prevalent genotype E, seven samples diverged (7/85, 8%) from the reference sequence E/Bour ( the most constant genotype), which is in line with a prior study reporting the genetic variants of E strains [
3].
With the exception of genotype B, mutations also occurred among other
C. trachomatis genotypes, with 33% (28/83) of them being sense mutations, leading to an elongated protein. Surprisingly, no statistical differences in clinical characteristics were observed between mutation and non-mutation groups, while sense mutations were more likely to result in greater severity of CIN on colposcopy. Several studies have reported the incidence of
C. trachomatis infection over the last two decades. In our phylogenetic analysis,
omp1 polymorphism was relatively stable, and the rate of genetic change was slow [
5].
Our study is unique because we assessed
C. trachomatis DNA and antibody concurrently from clinical samples, which is vital for epidemiological and vaccination studies. Pgp3-based LISA used in this study was validated to be a suitable assay for the detection of anti-
C. trachomatis antibody as our previously reported [
28]. As anticipated, women with positive
C. trachomatis DNA had a markedly higher prevalence of pgp3 antibody compared to those with negative
C. trachomatis DNA. We observed an association between cervical abnormalities and
C. trachomatis DNA, but not with the pgp3 antibody. Therefore, noninvasive nucleic acid amplification tests are recommended to detect
C. trachomatis infection of the genital tract [
30]. Antibody responses can indicate the presence of chlamydial antigens in the host, indicating past infections. Thus, seropositive women often have no obvious clinical manifestations, which may explain the lack of significant associations between the pgp3 antibody and cervical abnormalities. Intriguingly, the pgp3 antibody persists in the human body for as long as 12 years [
31], which can be used to estimate the cumulative risk of
C. trachomatis infection; considering this, our results suggest that almost half of the women in our cohort had been infected with
C. trachomatis.
C. trachomatis infection often causes few or mild symptoms, making them undetectable. Worsening and persisting infections may result in scarring and obstruction of the fallopian tubes, ultimately leading to female infertility. We found that women with infertility had a similar prevalence of
C. trachomatis DNA with those of childbearing age attending the PEC. However, a markedly higher seroprevalence was observed in in women with infertility, which was consistent with previous studies that women with infertility have 2–3 times higher levels of
C. trachomatis antibodies compared to the general female population [
32]. These findings suggest that the
C. trachomatis antibody test is a useful predictor of
C. trachomatis-mediated infertility, as most women with infertility had previous
C. trachomatis infection. Recurrent infections may also increase the risk of
C. trachomatis spreading from the genital tract to the gastrointestinal tract, leading to long-lasting colonization, and intensify the chlamydial pathogenicity in the reproductive system when it is transmitted back to the genital tract, similar to a second hit [
33]. These results highlight the usefulness and significance of
C. trachomatis screening, including genital and intestinal
C. trachomatis nucleic acid and antibody toward
C. trachomatis, for infertility.
There were several limitations to our study. First, a large female population was randomly recruited from a single hospital, which could limit the generalizability of our findings. Second, there was a lack of detailed information on demographic characteristics, such as such as ethnicity, educational level, country of birth, and sexual activities, which may have introduced bias in the prevalence estimates. Third, data were collected for only 2.5 years, whereas the distribution of C. trachomatis genotypes may have been different if additional data were collected from a larger sample and over a longer time period. Fourth, we did not collect information related to the gynecological examination and pgp3 antibodies for all participants because most refused to undergo the test, which may have impacted our results related to the association between C. trachomatis infection and cervical abnormalities.
In conclusion, we conducted the first investigation into the point DNA prevalence and seroprevalence of C. trachomatis among women in China, providing estimates for both current infection and cumulative exposure. Overall, the C. trachomatis prevalence was high. The difference between seroprevalence (47.46%) and current DNA prevalence (3.76%) in our study reflects the high clearance of C. trachomatis infections. The seroprevalence among women with infertility with negative C. trachomatis DNA was more than two-fold higher compared to that in the PEC group. These findings are helpful in elucidating the association between C. trachomatis and infertility. We also demonstrated a strong correlation between positive C. trachomatis DNA and leucorrhea cleanliness, CIN on cervical cytology, and CIN on colposcopy. These data bear important implications for the continuous improvement of primary and secondary prevention strategies as well as the development of subunit vaccines to decrease C. trachomatis infections in China.
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