Prevalence of Atopobium vaginae and Porphyromonas somerae in the Genital Tract of Women Undergoing Hysterectomy for Benign and Malignant Disease at a South Indian Tertiary Hospital.

Endometrial cancer is the third most common cause of death secondary to cancers affecting women. Endometrial cancer contributes to 20%–30% of malignant tumors of the female genital tract and the incidence is increasing by 1%–2% annually.[ 1 ] This may be attributed to the growing population of elderly women and the rising prevalence of obesity and other medical conditions, largely due to sedentary lifestyles. As a result, even premenopausal women are increasingly predisposed to endometrial cancer.[ 2 ] Around 67% of endometrial cancers are diagnosed at an early stage, as they usually present with postmenopausal bleeding and have good survival benefits, whereas advanced stages of disease carry a grave prognosis.[ 3 ] The causative or triggering factors for endometrial malignancy remain elusive. Host factors explain only 20% of endometrial cancer in the form of microsatellite instability or abnormalities in aerobic glycolysis. Obesity, medical disorders, and unopposed estrogen stimulation might also contribute.[ 4 ] Hence, exploring the possibility of a highly specific agent or pathogen playing a definitive role in the etiology of endometrial cancer is of paramount importance. A recent study showed a significant association between endometrial cancer and the presence of Porphyromonas species and Atopobium vaginae in the female reproductive tract, along with an alkaline vaginal pH. It is postulated that A. vaginae creates a chronic inflammatory profile that eventually leads to disruption of local immunity and enhances intracellular infection by Porphyromonas somerae , which is capable of dysregulating normal cell functions, eventually acting as a trigger for malignancy. This might be further facilitated by the anoxic microenvironment created by these microorganisms.[ 5 ] Although the role of microbial triggers has been well established in carcinoma of the cervix and gastric cancer, there is a paucity of data on the role of microbes in the etiopathogenesis of endometrial cancer.[ 6 ] This study aimed to assess the frequency and occurrence of A. vaginae and P. somerae in the lower genital tract of women with endometrial cancer, hyperplasia, and benign gynecological diseases and to determine the predominant aerobic and facultative anaerobic flora related to the same.

Out of 64 women, 32 had endometrial cancer, while the remaining 32 had benign gynecological diseases. Among the 32 benign lesions, more than half (21; 65.6%) were leiomyomas, 5 (15.6%) had uterovaginal prolapse, 3 (9.4%) had adenomyosis, 2 (6.3%) had simple hyperplasia, and 1 (3.1%) had serous cystadenoma. The mean (standard deviation) age was 56.6 ± 7.9 and 47.1 ± 5.3 in both groups, respectively. More than half of the women (17; 53.1%) in the malignant group had DM, and half of the women had (16; 50%) hypertension. In the benign group, nearly three-fourths of the women did not have DM or hypertension, and this association was statistically significant with P = 0.03. About 26 (81.2%) women were in their postmenopausal state in the malignant group, and 25 (78%) were in their premenopausal stage in the benign group. Nearly 30% of them had a vaginal pH of ≤4.5 in the benign group [ Tables 1 and 2 ]. Baseline characteristics between the groups BMI: Body mass index, SD: Standard deviation Comparison of baseline characteristics and risk factors between both the groups In the current study, we found that 25 (78.1%, 95% confidence interval [CI]: 60.0–90.7) and 14 (43.5%, 95% CI: 26.4–62.3) women in endometrial cancer and benign gynecological disease groups had A. vaginae , P . somerae , or both organisms. The presence of organisms is depicted in Table 3 . Distribution of organisms in both the groups Vaginal colonization of A . vaginae was observed in 38.5% of women between the ages of 40–49 and 50–59 years. The presence of P . somerae was slightly higher in the age group of 40–49 years. Colonization of both organisms had been seen in higher proportions in the endometrial cancer group compared to benign diseases (69.2 vs. 30.8 for A . vaginae and 65.4 vs. 34.6 for P . somerae ). There was a significantly higher occurrence of endometrial cancer in women harboring both organisms (83.3%), in comparison to those having either only one or none (42.3%) [ Tables 4 and 5 ]. Association between coexistence of Atopobium vaginae and Porphyromonas somerae and endometrial cancer Comparison of risk factors between the presence and absence of organisms Colonization of the lower genital tract with organisms either isolated (56% for A . vaginae and 53% for P . somerae ) or in coexistence (31%) had poor sensitivity, but good specificity ranging between 75% and 93% in the prediction of endometrial cancer [ Figure 1 ]. The diagnostic sensitivity (31%) and specificity (67%) of the presence of A. vaginae and P. somerae as predictors of endometrial cancer indicate that while these organisms are more frequently associated with the disease, they were not consistently present in all cases. Sensitivity is limited by heterogeneity in disease mechanisms and variable colonization rates of these bacteria in the endometrial cancer group. The low sensitivity observed could be attributed to the multifactorial etiology of endometrial cancer, the overlap in microbial presence between groups, and the small sample size. This highlights the need for larger studies and a broader examination of microbial profiles to validate these findings. Atopobium vaginae and Porphyromonas somerae positivity between the two study groups Among the 32 women with endometrial cancer, 29 were diagnosed with the endometrioid subtype, while one case each of clear cell, serous, and undifferentiated carcinoma were observed. Hence, the correlation of organisms with each subtype was not feasible. Future studies with larger and more diverse cohorts are required to explore histotype-specific microbial associations [ Table 6 ]. Diagnostic validity of the presence of Atopobium vaginae and Porphyromonas somerae in the prediction of endometrial cancer

The occurrence of A . vaginae and P . somerae in the lower genital tract of women with endometrial cancer was significantly higher in comparison to the benign gynecological disease group. Although the lower genital tract of women with endometrial cancer showed higher colonization with A. vaginae and P . somerae , the sample size was small due to logistical constraints and further large-scale studies in diverse populations are required in the future to substantiate the association between A. vaginae and P . somerae with endometrial cancer and also in implicating these organisms as a causative agent for triggering carcinogenesis in endometrium. There are no conflicts of interest.

Endometrial cancer is one of the most common cancers of the female genital tract in the western population. However, only 10% of women with postmenopausal bleeding harbor malignancy, with endometrial atrophy being the most common cause of postmenopausal bleeding.[ 7 ] Our study demonstrated that colonization by A. vaginae , P. somerae , or both was observed in 25 women with endometrial cancer (78.1%, 95% CI: 60.0–90.7) and in 14 women with benign gynecological disease (43.5%, 95% CI: 26.4–62.3). Vaginal pH was >4.5 in 87.5% of women as compared to 88.2% in the study done by Walther-António et al .[ 4 ] The prevalence of diabetes and hypertension was higher in women (25% and 21.9%) with benign gynecological diseases in the present study in comparison to the rest, probably due to ethnicity.[ 8 ] Although they play a role in creating anoxic microenvironment and carcinogenesis, this study did not collect data on multiple sexual behaviors, human papillomavirus (HPV) vaccination status, smoking history due to cultural sensitivities, low prevalence of HPV vaccination in India, and the scope of the study design. In India, HPV vaccination uptake remains low, particularly in older women. This limits the ability to analyze its impact on the vaginal microbiome in this population. As vaccination programs expand, future research may reveal new insights into its role in modulating microbial profiles. Moreover, A . vaginae and P . somerae are not sexually transmitted infections. These bacteria are part of the vaginal microbiota and are associated with dysbiosis or microbial imbalances, rather than direct sexual transmission. Postmenopausal bleeding was the most common presentation of women with endometrial cancer in the present study, as compared to Pakish et al .[ 9 ] However, the results were in contrast with those of Zhao et al . and Walker et al .[ 10 11 ] The mean vaginal pH of postmenopausal women in the present study was slightly higher than the other studies, probably due to the higher mean age in the present study, as compared to Panda et al .[ 12 ] The occurrence of A . vaginae and P . somerae in the lower genital tract of women with endometrial cancer was higher than that of Walther-António et al .,[ 4 ] where they recruited 10 and 17 women with benign gynecological diseases and endometrial cancer, respectively. The proportion of coexistence of both organisms in women with endometrial cancer was also lower than that of this study. In the same study, the sensitivity and specificity ranged between 73%–93% and 67%–90%, respectively, in addressing the prediction of endometrial cancer by the presence of A . vaginae and P . somerae in the lower genital tract. On the contrary, though we observed a statistically significant association between A . vaginae and P. somerae occurrence and endometrial cancer ( P = 0.02 and 0.03), the specificity was comparable. We have included an equal number of women with endometrial cancer and benign gynecological diseases. We have used a highly sensitive diagnostic tool for the identification of A . vaginae and P. somerae from the lower genital tract, which was done through a highly sensitive real-time PCR. In the current study, 1/3 rd of the women did not harbor any organisms on vaginal swabs, which may be attributed to the delay in transportation, drying up of swab specimens, alkaline vaginal pH, and irregular vaginal bleeding.[ 12 ] Moreover, we could not study the organisms separately in the cervix, vagina, and endometrium; hence, PCR was done from the pooled specimens of the cervix and vagina of the lower genital tract. Because of smaller numbers, the association of A. vaginae and P . somerae and endometrial cancer with vaginal pH was not studied.

This was a hospital-based cross-sectional study, conducted among 40–80-year-old women with endometrial cancer or women undergoing hysterectomy for any other gynecological disease. Women with active vaginal bleeding and antibiotic use 2 weeks before the enrollment were excluded from the study to minimize contamination of samples with blood, which could interfere with the appropriate detection of bacterial DNA of A. vaginae and P . somerae through the molecular methods. Assuming the proportion of women with endometrial cancer and benign gynecological diseases had A. vaginae and P. somerae at 40% and 10%, a power of 80%, and an alpha error of 5%, the calculated sample size was 64.[ 4 ] Convenient sampling was used to select the study participants, of which 32 were women with endometrial cancer and 32 were women with benign gynecological diseases. After obtaining informed consent and ethical clearance, a data collection pro forma was used to collect the age, weight, height, marital status, parity, family history of cancer, prior hormonal treatment, and presence of diabetes mellitus (DM) and hypertension. History of the last antibiotic use and duration of use were extracted from the women’s clinical records. A gentle pelvic examination was done on the day of admission to confirm the findings and plan management. The following day, four sterile swabs—one from the cervix and three from the lateral wall of the mid-vagina—were aseptically collected using a speculum without lubricant and transported under a proper cold chain to the Department of Microbiology at JIPMER. Vaginal pH was recorded simultaneously using a pH strip. Gram staining, culture with blood and MacConkey agar, and antibiotic susceptibility test were done using the smears. The third vaginal swab and cervical swab were stored in phosphate-buffered saline at −80°C and used for molecular identification of A . vaginae and P . somerae by real-time polymerase chain reaction (PCR). For all the symptomatic patients, appropriate treatment was given based on the microbial flora. The collected data were entered into Microsoft Excel and analyzed using SPSS Version 24.0 developed by IBM Corporation, headquartered in Armonk, New York, United States. Categorical variables were expressed as frequencies and percentages. The association of categorical variables was carried out using the Chi-square test or Fisher’s exact test. The comparison of continuous variables was done using independent Student’s t -test. P <0.05 was considered statistically significant.