Received: 10-Jan-2022, Manuscript No. gpmp-22-51437; Editor assigned: 11-Jan-2022, Pre QC No. P-51437; Reviewed: 15-Jan-2022, QC No. Q-51437; Revised: 27-Jan-2022, Manuscript No. R-51437; Published: 29-Mar-2022
Recent data from the human microbiome project show that endometrium, previously considers to be sterile, has its own unique microbiome. Although the uterine cervix serves as barrier for ascending microorganisms from the vagina new wide spread non-culturable techniques show presence of Lactobacillus-dominant environment. Rapidly accumulating data show that this microbiome is qualitatively and quantitatively different from the microbiome in lower genital tract. It plays role in embryo adhesion and development. In some conditions changes in uterine microbiome could cause disturbances in normal physiological processes and subsequent gynecological diseases.
microbiome; endometrium; lactobacilli; gynecologic conditions
Data from the human microbiome project show that body areas traditionally considered to be sterile, such as the placenta and endometrium, are inhabited by a unique microbiome [1, 2]. Bacteria inhabiting the urogenital tract make up about 9% of the human microbiota [3, 4] and most of them are not easy to identify and cultivate. Until recently, the uterus was considered to be sterile and most microbiological tests were based on vaginal samples. The dominant flora in reproductive age in a healthy woman consists of lactobacilli, with some variations depending on age and hormonal environment [5, 6]. This is an interesting characteristic of humans, as the genital tract of other mammals has a vaginal microbiome not dominated by lactobacilli .
During childhood, the vaginal flora is a mixture of aerobic and anaerobic bacterial populations including Prevotella, Enterobacteria, Streptococcus, and Staphylococcus species . With the development of an estrogenic environment during puberty, an increase in the glycogen levels and a pH decrease, the dominance of the lactobacilli types begins. Vaginal pH varies from 3.5 to 4.5 and is the second most acidic area in the human body after the stomach. This low pH created by the lactobacilli involvement significantly inhibits the development of pathogens.
What happens to the upper genital tract – does it have its own microbiome and then, eventually, what is it? For more than a century, a consensus has been established, based on Henry Tissier's 1900 work, that the uterus is a sterile environment . This sterility is thought to be maintained by the cervical obstruction, compared to the Colossus of Rhodes, providing n insurmountable barrier to ascending bacteria from the vagina . Isolation of pathogens from endometrial samples was initially considered to be contamination from vaginal contents or was attributed to various gynecological diseases [11, 12]. The colonization of the uterine cavity is mainly due to the ascension of normal vaginal flora. Other routes of colonization by microorganisms of the upper genital tract are possible: hematogenous dissemination of microorganisms from distant body sites, retrograde infection of the uterus by peritoneal microorganisms through the fallopian tubes, direct iatrogenic inoculation, and microorganisms attached to sperm. Notably, although the endometrium can be accessed in many different ways, it has not been classified as a non-sterile medium before .
Data on the upper genital tract microbiome have been accumulating rapidly in recent years, with the conclusion that the upper genital tract microbiome is qualitatively and quantitatively different from that of the lower genital tract .
In a study by Mitchell et al.  using next-generation sequencing of the 16s rRNA gene, bacterial cultures from endometrial samples in healthy women were detected in 100%, with lactobacilli being dominant, followed by Gardnerella, Prevotella, Atopobium, and Sneathia, which were also found in vaginal samples. Moreno et al. performed a simultaneous comparison of the vaginal and endometrial microbiome . They found that the endometrial microbiome was not equal to the vaginal one. Vaginal lactobacilli produce lactic acid and short-chain fatty acids causing lower pH levels. However, this is not the case in the uterine cavity, the pH in the endometrium is 6.6 – 8.51. These data suggest other biochemical processes in the endometrium, where the embryo adheres and develops. Non-Lactobacillus dominant microbiome triggers an inflammatory process in the endometrium affecting embryo implantation because inflammatory mediators are tightly regulated in the blastocyst adhesion to the endometrial wall .
The female reproductive system is an open system. During ovulation, the mature egg released from the ovary passes into the peritoneal cavity, where it is seized by the fimbriae of the fallopian tubes. In the fallopian tube, the egg is fertilized developing into an embryo, which is moved by the peristaltic waves and the oscillations of the ciliated epithelium of the fallopian tube into the uterine cavity where it implants. The vagina is home to billions of bacteria and the cervix should be a perfect barrier to their ascension to the uterine cavity and tubes. The physical barrier provided by the cervical mucus, the high levels of antimicrobial peptides, inflammatory cytokines, immunoglobulins, and matrix-degrading enzymes is considered to be protection from ascending bacteria [17 - 23]. During the menstrual cycle, the composition and pH of cervical mucus change, which under certain conditions, can lead to its overcoming as a barrier. The uterine peristaltic pump aids in the transport of sperm to the fallopian tubes and may be involved in the spread of bacteria in the uterus . During the follicular phase of the menstrual cycle, uterine contractions are at their highest frequency. Additionally, different uterine conditions can lead to hyper- and dysregulation of uterine contractions .
In current review 57 articles identified from Pubmed using the following search criteria: “microbiome”, “endometrium”, “lactobacilli”, “gynecologic conditions” are included. Gynecologic conditions as chronic endometritis, endometriosis, endometrial polyps, dysfunctional menstrual bleeding and others are discussed in relation to endometrial microbiome.
The most telling example of pathology as a result of damaged endometrial microbiota is chronic endometritis. It is characterized by prolonged inflammation of the endometrial mucosa produced by the colonization of the uterus by common bacteria: Enterococcus faecalis, Escherichia coli, Gardnerella vaginalis, Klebsiella pneumoniae, Proteus spp., Pseudomonas aeruginosa, Staphylococcus heptacuscopa, Streptococcus spp. Mycoplasma and Ureaplasma spp., and fungi such as Saccharomyces cerevisiae and Candida spp. [26, 27]. The incidence of chronic endometritis in the general population is estimated at 19%  and 45% in the infertile population . It should be noted that this high incidence is more highly associated with recurrent implantation failure (RIF) and recurrent pregnancy loss (RPL) than with other causes of infertility [30 - 33]. As chronic endometritis is asymptomatic and undetectable by vaginal ultrasound, it is rarely suspected and diagnosed. Current diagnosis of chronic endometritis is based on histopathological identification of plasma cells by conventional staining or immunohistochemical examination of CD 138 in endometrial stroma obtained by in-office endometrial biopsy. In addition to this gold standard for the diagnosis of chronic endometritis, some authors suggest hysteroscopy as a reliable method of diagnosis [34, 35]. Hysteroscopic diagnosis is based on the detection of micropolyps, stromal edema, focal or diffuse hyperemia [36, 37]. Endometrial tissue culture testing obtained by endometrial biopsy is not a routine procedure as it takes a long time and because not all microorganisms causing chronic endometritis can be cultivated.
Endometriosis is a condition characterized by the growth of endometrial epithelial and stromal tissue outside the uterine cavity. It affects about 10% of women of reproductive age and is clinically manifested by dysmenorrhea, pelvic pain, dyspareunia, infertility, impaired quality of life of patients . Despite extensive research on endometriosis, its origin remains unclear. Recent results from some studies indicate bacterial contamination of the endometrium as a potential new factor in the development of endometriosis. This theory comes from a study showing that the menstrual blood of women with endometriosis is highly contaminated with E. coli  with correspondingly higher levels of endotoxins in menstrual blood and peritoneal fluid as a result of reflux to the small pelvis. In addition to the traditional transplantation and coelomic metaplasia theory, the authors propose a novel hypothesis of bacterial contamination causing the development and maintenance of endometriosis. The same authors found that pathogenic genera such as Gardnerella, Enterococcus, Streptococcus, and Staphylococcus were the main ones identified in endometrial samples from women diagnosed with endometriosis, followed by other taxonomic species such as Actinomyces, Corynebacterium, Fusobacterium, and Prevotellapion. For comparison, mainly lactobacilli were found in the control group .
Bacteria from the Streptococcaceae and Staphylococcaceae families are significantly elevated in the cystic fluid obtained from women with ovarian endometrioma compared to control groups . A similar finding with significant changes in the composition of the microbiome has been found in patients with adenomyosis . Changes in the microbiome can trigger endometriosis by changing the microenvironment. Significantly lower levels of lactobacilli in the endometrial microbiome were found in women with endometriosis after the administration of GnRH agonist treatment compared to women who did not receive such a treatment . Endometriosis is associated with higher levels of the proinflammatory cytokine IL-6 in follicular fluid  and IL-1 beta, which causes inflammation in the peritoneum . The link between endometriosis and inflammation is also evidenced by a Danish study showing that women with endometriosis had a significantly higher risk of developing inflammatory bowel disease, ulcerative colitis, and Crohn's disease compared to healthy controls . Even 20 years after initial hospitalization for endometriosis, these patients remain at increased risk of developing ulcerative colitis and Crohn's disease.
Cicinelli et al. found that patients with endometriosis treated with antibiotics before implantation had a better pregnancy outcome than patients who were not treated, suggesting that partially the negative impact of endometriosis on fertility may be associated with bacteria in the uterus . These results support the association of chronic endometritis with endometriosis. It can be explained by the examination of cultures of ectopic endometrial cells in response to inflammatory-induced dysperistalsis and impairment of uterine contractility in patients with chronic endometritis [46, 47].
Endometrial polyps are a common gynecological disease characterized by local overgrowth of the endometrial mucosa, associated with chronic endometritis. Long-term stimulation by biological inflammatory factors is thought to contribute to the disease [48, 49]. Elevated Lactobacillus, Bifidobacterium, Gardnerella, Streptococcus, Alteromonas, and Euryarchaeota (Archaea) and decreased Pseudomonas and Enterobacteriaceae were found in endometrial samples of women with endometrial polyps compared to healthy women .
Dysfunctional menstrual bleeding
Certain microbial species are thought to play a role in gynecological pathologies such as menorrhagia and dysmenorrhea. In nulipari and virgo intacta undergoing surgical treatment for menorrhagia, Pelzer et al. found obligate anaerobic microorganisms – Fusobacterium spp., Jonquetella spp., which can be considered members of the spectrum of bacteria associated with bacterial vaginosis, instead of the expected lactobacilli . The authors found no evidence of inflammation and suggested that the microorganisms lead to dysbiosis presented with excessive menstrual bleeding. It is the result of damaged angiogenic mediators known to be upregulated in conditions of infection. On this occasion, the authors suggest the use of probiotics or antibiotics in cases of menstrual abnormalities when no organic cause is identified.
Bacterial vaginosis has been shown to affect the onset of pelvic inflammatory disease  and cervical intraepithelial neoplasia . Walther-António et al.  investigated the importance of the microbiome in genital cancer by performing sequencing of bacterial 16S rRNA gene. For this, they examined vaginal, cervical, tubal, ovarian, peritoneal samples, and urine in 31 women who underwent hysteroscopy for endometrial carcinoma, endometrial hyperplasia, or other benign conditions. An abundance of taxa, such as Anaerostipes, Dialister, Peptoniphilus, Ruminococcus, Anaerotruncus, Atopobium, Bacteroides, and Porphyromonas, has been found in the reproductive tract of patients with endometrial hyperplasia and carcinoma compared to patients with benign conditions. This suggests an infectious/inflammatory role of bacteria in the occurrence of endometrial cancer . The microbiota can activate a malignant process through various mechanisms: suppression of apoptosis, stimulation of proliferation, or by causing genomic instability . The effect of microorganisms on triggering a pathological process may not only be expressed in inflammation and secretion of cytokines by the host cells, but may also be influenced by the hormonal status of the host. Sex hormones, in particular estrogens, are important factors in certain cancers.
Concerning the reproductive tract, the question is whether they can affect the uterine microbiome like the vaginal microbiome. The administration of a GnRH agonist has been associated with a change in the composition of the uterine microbiome, indicating that it can be hormonally regulated . The intestinal microbiota facilitates estrogen reuptake, thus being related to estrogen-dependent cancer . In support of this, both intestinal microbiota composition and systemic estrogen levels have been found to be significantly different in breast cancer patients compared to healthy controls [56, 57].
A lot of data is available nowadays showing the uterine cavity as non-sterile compartment. Current studies on the microbiome provide basic point for future research to understand its role in uterine physiology in health and disease. Determination and qualification of its microbiome plays a great role in development of some gynecologic diseases and also in maintaining of women’s health. On the other hand, it is unclear whether described gynecological conditions may provoke changes in otherwise normal endometrial microbiome and consequent uterine dysbiosis. More data is needed to clarify the relationship between uterine normo- and dysbiosis and gynecological diseases as this research area is new and lot of questions is steadily arising.
Conceptualization T.B.; Methodology E.K., S.P.; Formal Analysis S.P.; Investigation T.B.; Resources T.B., E.K.; Writing—Original &Draft Preparation T.B..; Writing—Review & Editing A.Y., E.K..; Visualization T.B.; Supervision A.Y.
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