A cross-sectional study was conducted among 439 HPV-positive women who underwent colposcopy in Hunan province, China, to investigate the prevalence and age distribution of C. trachomatis and M. genitalium infections, as well as the impact of co-infection with HPV on cervical lesions. The aim of the study was to contribute to the existing research in this field. C. trachomatis was detected in 17 participants (3.87%), while M. genitalium was detected in 16 participants (3.64%). Co-infection of C. trachomatis and M. genitalium was not observed. The prevalence of M. genitalium was highest in women aged 19–30 years (10.20%; 95% CI, 1.41-18.99%) and decreased steadily with age (Ptrend = 0.014). It was observed that HPV16, HPV31, and HPV33 may exhibit higher cervical pathogenicity in Hunan province, China. Additionally, a surprising finding was the association between elevated levels of leukocytes in vaginal secretions and cervical lesions. However, neither C. trachomatis nor M. genitalium infection, alone or in co-infection with HPV16, were found to be associated with the severity of cervical lesions. This study provided insights into the pathogenicity of different HPV subtypes on cervical lesions and explored the age distribution of M. genitalium and its relationship with cervical lesions in the HPV-positive population in Hunan province, China. These findings are expected to contribute to the understanding of the epidemic pathology of sexually transmitted pathogens and aid in the prevention, screening, and control of such pathogens.

The population included in our study had certain unique characteristics. Samples were collected from the colposcopy laboratory, and patients who underwent colposcopy were excluded if they had bacterial vaginitis, fungal vaginitis, trichomonas vaginitis, HIV, or syphilis. This exclusion was done to prevent worsening of the patients’ condition and to avoid the risk of infection for healthcare workers. Therefore, bacterial, fungal, trichomonas, HIV, and syphilis infections were not considered in our study population to eliminate any potential synergistic pathogenic effects they might have with C. trachomatis or M. genitalium.

The prevalence of both C. trachomatis and M. genitalium was relatively low, with C. trachomatis often having a higher prevalence compared to M. genitalium. For instance, among the general female population visiting gynecology departments in Sichuan province, China, the infection rates of C. trachomatis and M. genitalium were reported as 6.5% and 2.6%, respectively [10]. In a study on women undergoing cervical cancer screening in Beijing, China, the infection rates of C. trachomatis and M. genitalium were 11.3% and 1.0%, respectively [9]. In our study, the infection rate of C. trachomatis (3.87%) was slightly lower than that reported in other studies, while the infection rate of M. genitalium (3.64%) was slightly higher. Co-infection of M. genitalium with other sexually transmitted pathogens has been reported. In young high-risk women with asymptomatic bacterial vaginosis in the United States, the prevalence of M. genitalium co-infection with C. trachomatis was 29.9%, and co-infection with Neisseria gonorrhoeae was 23.6% [14]. However, in a study conducted in Belgium, Germany, Spain, and the United Kingdom, the co-infection rate of M. genitalium with C. trachomatis was only 0.6%, and with N. gonorrhoeae was 0.1% [15]. In our study, co-infection of C. trachomatis and M. genitalium was not observed, likely due to differences in the study population and the low positive rates of C. trachomatis and M. genitalium.

Studies have indicated that sexually transmitted pathogen infections are more prevalent among young women [6]. Our study revealed that the highest prevalence of M. genitalium was observed in women aged 19 to 30 years (10.20%; 95% CI, 1.41–18.99%), and this prevalence declined steadily with age, consistent with previous research [16, 17]. However, the prevalence of C. trachomatis was higher among women over 50 years old (5.16%; 95% CI, 1.64–8.68%), but not in young women under 30 years of age. This finding contradicts Chen’s research, as Chen et al. discovered that the prevalence of C. trachomatis infection was highest among the group aged ≤ 25 years and gradually decreased with age [8]. However, a study conducted in Shenzhen, China, found that the prevalence of C. trachomatis infection was higher in the group aged > 35 years compared to the group aged ≤ 35 years [18]. This finding aligns with our study and suggests that C. trachomatis is not necessarily exclusive to young women. The inconsistent results may be attributed to differences in study areas and populations. In conclusion, our findings suggest that the growing openness towards sexual concepts and the younger age at first sexual intercourse among Chinese women may increase the likelihood of sexually transmitted pathogen infections in young women, but the health of elderly women should not be disregarded.

Few studies have explored the relationship between leukocyte levels in vaginal secretions and the severity of cervical lesions. A study conducted in southwest China found that patients infected with C. trachomatis or M. genitalium were more likely to have elevated leukocyte levels in vaginal secretions [10]. In our study, we observed that increased leukocyte levels in vaginal secretions may be associated with cervical lesions. Leukocyte levels are often indicative of inflammation, but our study excluded patients with bacterial vaginitis, fungal vaginitis, and trichomonal vaginitis. The elevated leukocyte levels in the vaginal secretions of the subjects may have resulted from an imbalance in the vaginal microecology. In recent years, numerous studies have focused on the relationship between vaginal microbiota and female reproductive tract diseases [19, 20]. Some studies have revealed that an imbalance in vaginal microecology may be associated with persistent HPV infection and cervical lesions [21, 22]. In the future, metagenomics technology can be employed to further investigate the vaginal microbiota and its relationship with cervical lesions.

Persistent infection with high-risk HPV can result in cervical lesions and potentially cervical cancer. The prevalence and distribution of HPV subtypes vary across countries, races, and populations. In our study, the most prevalent HPV genotype was HPV52 (30.79%), followed by HPV16 (18.62%), HPV58 (16.95%), and HPV53 (10.02%). These results align with data from other Chinese populations. A study conducted in southern China identified HPV52, HPV16, and HPV58 as the three most common HPV subtypes [8]. In the Inner Mongolia region of China, HPV16 was the most prevalent genotype, followed by HPV58 and HPV52 [23]. This indicates that HPV16, HPV52, and HPV58 are predominant among the general Chinese population. However, a high prevalence of HPV subtypes does not necessarily imply their strong ability to cause cervical lesions. In our study, HPV16 (OR = 3.43, 95% CI, 2.13–5.53), HPV31 (OR = 3.70, 95% CI, 1.44–9.50), and HPV33 (OR = 3.71, 95% CI, 1.43–9.67) infections were associated with an increased severity of cervical lesions, whereas HPV53 infection was not likely to progress to advanced cervical lesions (OR = 0.45, 95% CI, 0.23–0.89). Therefore, it is recommended to pay more attention to patients infected with more pathogenic HPV subtypes based on local HPV epidemiological data when managing HPV-positive patients.

Many previous studies have primarily focused on the relationship between cervical cytology results and sexually transmitted pathogens [23, 24]. However, cervical cytology results do not fully represent the cervical status. Cervical biopsy serves as the gold standard for assessing cervical status, highlighting the importance of exploring the relationship between sexually transmitted pathogens, HPV infection, and cervical biopsy. In our study, we found no association between C. trachomatis and M. genitalium infection, either alone or in co-infection with HPV16, and the severity of cervical lesions. Regarding C. trachomatis infection, its association with HPV infection and cervical lesions is currently controversial. Several studies have indicated an association between C. trachomatis infection and HPV infection as well as cervical lesions [8, 23]. However, other studies have suggested that C. trachomatis infection does not increase the risk of HPV infection and cervical lesions [9, 10, 25], which aligns with our research findings. The conflicting results may stem from differences in cohorts or limitations in sample size. As for M. genitalium infection, there is limited research on its association with HPV infection and cervical lesions. A study by A et al. found no association between M. genitalium infection and HPV infection or cervical lesions [9], which is consistent with our findings. Currently, most studies are cross-sectional or retrospective. Large-scale, multicenter prospective cohort studies are warranted to explore the relationship between co-infection of sexually transmitted pathogens with HPV and cervical lesions.

This study had several limitations. Firstly, it was a cross-sectional study, which cannot accurately capture the dynamic changes in HPV infection and cervical lesions among the subjects. Secondly, the sample size was small, and due to the low infection rates of C. trachomatis and M. genitalium, the number of positive cases was insufficient to conduct comprehensive stratified analysis. Additionally, the samples collected were primarily from Changsha city, Hunan Province, which may affect the generalizability of the findings regarding the prevalence of sexually transmitted pathogens. Future research should be conducted using large longitudinal cohorts, including women from different locations, to identify and validate risk factors for HPV infection and the progression of cervical lesions, ultimately aiming to prevent cervical cancer.

In conclusion, our study suggests that greater attention should be given to M. genitalium infection among young women, while C. trachomatis infection in older women should not be overlooked. Increased levels of leukocytes in vaginal secretions may be associated with cervical lesions. In Hunan province of China, HPV16, HPV31, and HPV33 appear to have higher pathogenicity in relation to cervical lesions. The findings of this study highlight the ongoing controversy regarding whether C. trachomatis and M. genitalium infection, either alone or in co-infection with HPV, contribute to an increased severity of cervical lesions. Further clarification is needed through large longitudinal cohort studies in the future.