Metritis and the uterine disease microbiome are associated with long-term changes in the endometrium of dairy cows†.

Cows with metritis (uterine disease) during the first 1 to 2 weeks postpartum have lower pregnancy rates when inseminated later postpartum (typically >10 weeks). We hypothesized that metritis and the disease-associated uterine microbiome have a long-term effect on endometrial gene expression. Changes in gene expression may inform a mechanism through which disease lowers pregnancy rates. A total of 20 cows were enrolled at 1 to 2 weeks postpartum to either metritis (clinical disease; n = 10) or healthy (control; n = 10) groups and randomly assigned to be slaughtered at approximately 80 and 165 dpp (mid-lactation). The microbiome of the reproductive tract was sampled to confirm the presence of pathogens that are typical of metritis. In addition to the original clinical diagnosis, study cows were retrospectively assigned to uterine-disease and control groups based on the composition of their microbiome. There was no effect of early postpartum uterine disease on the uterine microbiome at mid-lactation (time of slaughter). Nonetheless, early postpartum metritis and the disease microbiome were associated with a large number of differentially-expressed genes at mid-lactation primarily in the caruncular compared with the inter-caruncular endometrium. Gene enrichment analysis identified oxidative phosphorylation as the primary pathway increased in caruncular endometrium of diseased cows whereas growth factor signaling pathways were reduced. The current study demonstrated that metritis and a uterine disease microbiome leave a sustained imprint on gene expression in the caruncular endometrium that may explain lower fertility in cows with postpartum uterine disease.

The role and challenges of regulating endometrial microbiome in uterine health and diseases.

The uterine environment provides necessary conditions for the existence of endometrial microbiota, which in turn plays an important role in maintaining the homeostasis of the uterine environment. The endometrial microbiome is highly susceptible to external factors such as age, hormones, menstrual, pregnancy, etc. When the microbiota is imbalanced, it will further promote the occurrence of uterine diseases such as endometritis and endometrial cancer. Regulating the microbiome of the endometrium is of positive significance for promoting uterine health. Among them, antibiotics, probiotics, prebiotics, and microbial transplantation may be important pathways for regulating endometrial microbiota in the future. However, there is currently no unified plan for evaluating the endometrial microbiota. In addition, due to the small sample size, it is easy to be contaminated by exogenous bacterial DNA, which poses great challenges for studying the mechanism of microbial community regulating uterine health. Therefore, there are still many areas worth exploring for the future of endometrial microbiome.

Vaginal and endometrial microbiome dysbiosis associated with adverse embryo transfer outcomes.

BACKGROUND: Assisted reproductive technology (ART) is the most effective method to treat infertility and the pathogenesis of implantation failure after in vitro fertilization-embryo transfer (IVF-ET) is a challenging filed in infertility. Microbes in the female reproductive tract are considered to be associated with gynecological and obstetric diseases. However, its effects on embryo implantation failure are unsured. PURPOSE: This study aimed to investigate reproductive tract dysbiosis, identify different bacteria in reproductive tract as potential biomarkers of embryo implantation failure and demonstrate the pathogenesis through metabolites analysis. METHODS: We compared the data from 16S rRNA gene and metagenome in reproductive tracts through QIIME2 and HUMAnN2 by the times of embryo implantation failure on 239 infertile patients and 17 healthy women. RESULTS: Our study revealed a strong positive correlation between Lactobacillus abundance and embryo implantation success (IS) after IVF-ET. The microbial community composition and structure in reproductive tract showed substantially difference between the embryo implantation failure (IF) and healthy control. Moreover, we established a diagnostic model through receiver operating characteristic (ROC) with 0.913 area under curve (AUC) in IS and multiple implantation failures (MIF), verified its effectiveness with an AUC = 0.784 demonstrating microbial community alterations could efficiently discriminate MIF patients. Metagenome functional analyses of vaginal samples from another independent infertile patients after IVF-ET revealed the L-lysine synthesis pathway enriched in IF patients, along with ascended vaginal pH and decreased Lactobacillus abundance. CONCLUSIONS: This study clarifies several independent relationships of bacteria in vagina and endometrial fluid on embryo implantation failure and undoubtedly broadens the understanding about female reproductive health.

Evaluation of molecular and bacteriological detection methods performed on the formalin-fixed paraffin-embedded biopsy samples collected from endometrial and lymph node tuberculosis suspected patients.

BACKGROUND: Endometrial Tuberculosis is one of the most common gynecological problems known to have serious implications for the quality of life like infertility. The commonly practiced histopathology solely relies on the suggestive feature of Tuberculosis (TB) with low specificity. Regarding the alternative bacteriological and molecular detection tools, little evidence was generated on their utility in the diagnosis of endometrial tuberculosis in Ethiopia. Therefore, we aim to investigate the detection rate of molecular and bacteriological detection methods on formalin-fixed paraffin-embedded biopsy samples for the diagnosis of endometrial and lymph node TB. METHODS: A retrospective cross-sectional study was conducted on 90 formalin fixed paraffin embedded biopsy samples from patients with gynecologic and lymph problems collected between 2018 and 2022 at St. Paul's Hospital Millennium Medical College, Addis Ababa, Ethiopia. SPSS version 26 was used for statistical analysis. The diagnostic performance was calculated using the histopathology method as the reference standard. Cohen's Kappa value was used to measure the level of agreement. A test with a P-value of < 0.05 was considered statistically significant. RESULTS: A total of 90 samples were analyzed in the current study. Auramine O, GeneXpert MTB/RIF assay, and Real-Time PCR tests have shown a detection rate of 32/90 (36%), 43/90 (47.8%), and 54/90 (60%) respectively (P ≤ 0.01). The sensitivity and specificity of AO were 38.1% and 95% respectively. RT PCR showed superior sensitivity followed by GeneXpert MTB/RIF assay, 70% and 58.6%. AO and molecular methods have shown a similarly low level of agreement with histopathology (Kappa value = 0.2). CONCLUSIONS: In a resource-limited setting, the selection of diagnostic tools needs careful attention. Putting the patients on anti-TB treatments based solely on histopathological findings may lead to undesired and adverse complications. Therefore, applying molecular and bacteriological detection methods along with histopathology, could help minimize inappropriate antimicrobial use.

Chronic endometritis and the endometrial microbiota: implications for reproductive success in patients with recurrent implantation failure.

BACKGROUND: Chronic endometritis (CE) is associated with poor reproductive outcomes, yet the role of endometrial microbiota in patients with recurrent implantation failure (RIF) and CE remains unclear. This study aims to characterize endometrial microbiota in RIF patients with CE and assess its implications for reproductive outcomes. METHODS: In this prospective study, we enrolled RIF patients both with and without CE. Endometrial and cervical samples were collected for 16 S rRNA gene sequencing. Microbiota composition was compared between groups using diversity indices, phylum, and genus-level analysis. Canonical correlation analysis (CCA) and Spearman's correlation coefficients were used to assess relationships between CE, reproductive outcomes, and microbiota. Predictive functional profiling was performed to evaluate metabolic pathways associated with CE. RESULTS: Endometrial microbiota in CE patients exhibited greater diversity and evenness compared to non-CE patients. Principal coordinates analysis (PCoA) revealed distinct clustering between CE and non-CE groups. Linear discriminant analysis (LDA) identified Proteobacteria, Aminicenantales, and Chloroflexaceae as characteristic of CE, while Lactobacillus, Acinetobacter, Herbaspirillum, Ralstonia, Shewanela, and Micrococcaceae were associated with non-CE. CCA demonstrated associations between CE, adverse reproductive outcomes, and specific bacterial taxa. Microbial metabolic pathways significantly differed between CE and non-CE groups, with enrichment in pathways related to cofactors, vitamins, secondary metabolites, and the immune system in CE patients. CONCLUSION: RIF patients with CE exhibit distinct endometrial microbiota compositions associated with adverse reproductive outcomes. The increased microbial diversity and altered metabolic pathways in CE suggest a potential correlation with reproductive outcomes, although further studies are necessary to elucidate the causal relationship between microbiota alterations and fertility. Modulating the endometrial microbiome may represent a novel therapeutic strategy to improve IVF outcomes in patients with CE.

Comparative analysis of endometrial, vaginal, and gut microbiota in patients with and without adenomyosis.

INTRODUCTION: Alterations in microbiota composition have been implicated in a variety of human diseases. Patients with adenomyosis present immune dysregulation leading to a persistent chronic inflammatory response. In this context, the hypothesis that alterations in the microbiota may be involved in the pathogenesis of adenomyosis, by affecting the epigenetic, immunologic, and biochemical functions of the host, has recently been postulated. The aim of the present study was to compare the microbiota composition in the vagina, endometrium, and gut of individuals with and without adenomyosis. MATERIAL AND METHODS: Cross-sectional study including 38 adenomyosis patients and 46 controls, performed between September 2021 and October 2022 in a university hospital-based research center. The diagnosis of adenomyosis was based on sonographic criteria. Fecal, vaginal, and endometrial samples were collected. Study of the microbiota using 16S rRNA gene sequencing. RESULTS: Patients with adenomyosis exhibited a significant reduction in the gut microbial alpha diversity compared with healthy controls (Chao1 p = 0.012, Fisher p = 0.005, Observed species p = 0.005). Beta-diversity analysis showed significant differences in the compositions of both gut and vaginal microbiota between adenomyosis patients and the control group (Adonis p-value = 0.001; R2 = 0.03 and Adonis p-value = 0.034; R2 = 0.04 respectively). Specific bacterial taxa were found to be either overrepresented (Rhodospirillales, Ruminococcus gauvreauii group, Ruminococcaceae, and Actinomyces) or underrepresented in the gut and endometrial microbiota of adenomyosis patients compared with controls. Distinct microbiota profiles were identified among patients with internal and external adenomyosis phenotypes. CONCLUSIONS: The study revealed reduced gut microbiota diversity in adenomyosis patients, accompanied by distinct compositions in gut and vaginal microbiota compared with controls. Overrepresented or underrepresented bacterial taxa were noted in the gut and endometrial microbiota of adenomyosis patients, with variations in microbiota profiles among those with internal and external adenomyosis phenotypes. These findings suggest a potential association between microbiota and adenomyosis, indicating the need for further research to comprehensively understand the implications of these differences.

Significant increased isolation of Escherichia coli in Iranian women with endometriosis: a case control - study.

BACKGROUND: The role of bacterial contamination in the development and progression of endometriosis lesions is currently a hot topic for gynecologists. In this study, we decided to compare the endometrial cultures of women affected by endometriosis with those of non-endometriotic women, focusing on specific microbial pathogens. MATERIAL AND METHOD: In this cross-sectional case-control study, 30 women with endometriosis in stages 4 of the disease whose endometriosis was confirmed based on clinical, ultrasound, and histopathological findings, and 30 women without endometriosis who were candidates for surgery due to benign uterine diseases with regular menstrual cycle, underwent endometrial biopsy with Novak Kort in sterile conditions before starting their operation, and the results of their endometrial culture were analyzed and compared. RESULTS: Results of the study indicate that there were no significant differences in terms of age, BMI, smoking, education level, place of residency, use of the intrauterine device, or vaginal douche, and age of menarche between the case and control groups. The only demographic difference observed was in parity, where the control group had a significantly higher parity than the case group (P = 0.001). Out of the 60 cultures, only 15 samples were positive in the endometriosis group, and E. coli was the most prevalent species, with 10 (33.3%) samples testing positive for it. Klebsiella spp. and Enterobacteria spp. were also detected in 3 (10.0%) and 2 (6.7%) samples, respectively. The comparison between the two groups showed that only E. coli had a significant association with the presence of endometriosis (P = 0.001). There was no significant relationship between the location of endometriosis in the pelvic cavity and culture results. It was observed that parity among the E. coli negative group was significantly higher compared to the E. coli positive group (P < 0.001). CONCLUSION: Based on The high occurrence of E. coli in women with endometriosis, along with its potential involvement in the progression and/or recurrence of this condition, the researchers propose that treating women with endometriosis and recurrent IVF failure, as well as those with endometriosis recurrence after surgical treatment, with suitable antibiotics and repeated culture until the culture becomes negative, could be beneficial.

Histopathologic patterns of female genital tuberculosis with clinical correlation: a 10-year (2013-2022) retrospective cross-sectional study.

OBJECTIVE: Underdiagnosis of female genital tuberculosis (FGTB) often leads to infertility. In this study, we aimed to determine the site and histopathologic patterns of FGTB and its correlation with clinical presentation and acid-fast bacilli (AFB) status. METHODS: A retrospective cross-sectional study was conducted on 122 patients with a histopathological diagnosis of FGTB at the Department of Pathology, College of Health Sciences (CHS), Tikur Anbessa Specialized Hospital (TASH), Addis Ababa University (AAU), from January 1, 2013, to August 30, 2022. RESULTS: Female genital tuberculosis was found in 0.94% of the gynecology specimens examined. The most common presentations were menstrual disturbance, abdominopelvic pain, and infertility. Among patients with FGTB, 4.6% exhibited misleading clinical and radiologic findings, leading to suspicion of malignancy and subsequent aggressive surgical management. The endometrium was the most frequently affected organ, followed by the fallopian tube, ovary, cervix, and vulva. In the majority of tuberculous endometritis cases (53.3%), histopathology revealed early-stage granulomas. Acid-fast bacilli were found in a significant proportion (42.6%) of FGTB tissues with TB histopathology. The ovary had the highest rate of AFB detection, followed by the fallopian tube, endometrium, and cervix. CONCLUSION: Female genital tuberculosis should be considered in reproductive-age women presenting with menstrual irregularities, abdominopelvic pain, infertility, or an abdominopelvic mass. The endometrium is commonly affected, displaying early granulomas with low AFB positivity.

Impact of vaginal microecological differences on pregnancy outcomes and endometrial microbiota in frozen embryo transfer cycles.

PURPOSE: This prospective study investigates the correlation between vaginal microecology and pregnancy outcomes and explores their impact on endometrial microbiota composition during frozen embryo transfer (FET) cycles. Additionally, the impact of transvaginal Lactobacillus supplementation on reproductive outcomes in patients with previous failed cycles was assessed. METHODS: A total of 379 patients undergoing FET at a reproductive medicine center were categorized into clinical pregnancy (CP), miscarriage (MISC), and non-pregnant (NP) groups. Vaginal specimens were collected for microecological evaluation prior to embryo transfer. Endometrial microbiota samples were obtained during embryo transfer for 16S rRNA gene sequencing analysis to assess endometrial microbiota composition. Vaginal microecological indicators, including pH, Lactobacillus dominance, and leukocyte esterase activity, were measured. Transvaginal Lactobacillus supplementation was investigated in 60 patients with previous failed cycles. RESULTS: Vaginal microecology significantly correlated with pregnancy outcomes, with normal microecology associated with a higher clinical pregnancy rate. Vaginal pH and leukocyte esterase activity were significantly associated with clinical pregnancy. Furthermore, vaginal microecological differences significantly impacted endometrial microbiota composition. However, no significant differences were observed in endometrial microbiota composition among the CP, MISC, and NP groups. Notably, transvaginal Lactobacillus supplementation increased the clinical pregnancy rate without affecting the miscarriage rate. CONCLUSION: This study highlights that normal vaginal microecology, characterized by lower pH and leukocyte esterase negativity, is associated with a higher likelihood of clinical pregnancy following FET. Importantly, vaginal microecological differences influence endometrial microbiota composition. Moreover, transvaginal Lactobacillus supplementation appears promising in improving clinical pregnancy rates in patients with previous failed cycles. These findings contribute to a better understanding of the interplay between vaginal and endometrial microbiota and offer potential interventions to enhance reproductive success in assisted reproductive technologies.

The alteration of intrauterine microbiota in chronic endometritis patients based on 16S rRNA sequencing analysis.

BACKGROUND: Chronic endometritis (CE) is a disease of continuous and subtle inflammation occurring in the endometrial stromal area, which is often asymptomatic or present with non-specific clinical symptoms. METHODS: This study investigated the composition and distribution of the intrauterine microbiota of 71 patients who underwent hysteroscopy during the routine clinical inspection of infertility. Among them, patients who were diagnosed with chronic endometritis (CE) were allocated into CE group (n = 29) and others into non-CE group (n = 42). There was no significant difference in average age between the two groups (P = 0.19). Uterine flushing fluid was collected by the self-developed cervical trocar uterine cavity sampler and 16S rRNA sequencing was performed. RESULTS: The alpha diversity in the CE group was significantly higher than that in the non-CE group (P < 0.05). Firmicutes (newly named Bacillota) were the dominant phylum in the non-CE group (72.23%), while their abundance was much lower in the CE group (49.92%), but there was no statistically significant difference between the two groups. The abundances of Actinobacteriota and Cyanobacteria in the CE group were significantly higher than those in the non-CE group (P < 0.05). At the genus level, the abundance of Lactobacillus dominated in all samples, which presented a significantly lower abundance in the CE group (40.88%) than that in the non-CE group (64.22%) (P < 0.05). Correspondingly, the abundance of non-Lactobacillus was higher in the CE group, among which Pseudomonas and Cutibacterium increased significantly (P < 0.01). Moreover, compared with the non-CE group, the pathways involved in arginine and proline metabolism and retinol metabolism were significantly enriched in the CE group (P < 0.05), while the metabolism of lipid and prenyltransferases were significantly decreased in the CE group (P < 0.05). CONCLUSIONS: A certain microbial community was colonized in the uterine cavity, which was dominated by Lactobacillus. The structure and distribution of intrauterine microbiota in the CE group were different from those in the non-CE group by showing a lower abundance of Lactobacillus, and a significantly higher abundance of Pseudomonas and Cutibacterium. Additionally, the microbial metabolism was altered in the CE group. This study elaborated the alteration of intrauterine microbiota in CE patients, which may contribute to the diagnosis of CE and provide a reference for antibiotic treatment of CE.

Therapeutic intervention based on gene sequencing analysis of microbial 16S ribosomal RNA of the intrauterine microbiome improves pregnancy outcomes in IVF patients: a prospective cohort study.

PURPOSE: A Lactobacillus-dominated microbiota in the endometrium was reported to be associated with favorable reproductive outcomes. We investigated in this study whether 16S ribosomal RNA (rRNA) gene sequencing analysis of the uterine microbiome improves pregnancy outcomes. METHODS: This prospective cohort study recruited a total of 195 women with recurrent implantation failure (RIF) between March 2019 and April 2021 in our fertility center. Analysis of the endometrial microbiota by 16S rRNA gene sequencing was suggested for all patients who had three or more failed embryo transfers (ETs). One hundred and thirty-one patients underwent microbial 16S rRNA gene sequencing (study group) before additional transfers, while 64 patients proceeded to ET without that analysis (control group). The primary outcome was to compare the cumulative clinical pregnancy rate of two additional ETs. MAIN RESULTS: An endometrial microbiota considered abnormal was detected in 30 patients (22.9%). All but one of these 30 patients received antibiotics according to the bacterial genus detected in their sample, followed by treatment with probiotics. As a result, the cumulative clinical pregnancy rate (study group: 64.5% vs. control group: 33.3%, p = 0.005) and the ongoing pregnancy rate (study group: 48.9% vs. control group: 32.8%, p = 0.028) were significantly increased in the study group compared to the control group. CONCLUSION: Personalized treatment recommendations based on the microbial 16S rRNA gene sequencing of the uterine microbiota can improve IVF outcomes of patients with RIF. TRIAL REGISTRATION: The University Hospital Medical Information Network (UMIN) Clinical Trial Registry: UMIN000036050 (date of registration: March 1, 2019).

Comparing Vaginal and Endometrial Microbiota Using Culturomics: Proof of Concept.

It is generally accepted that microorganisms can colonize a non-pathological endometrium. However, in a clinical setting, endometrial samples are always collected by passing through the vaginal-cervical route. As such, the vaginal and cervical microbiomes can easily cross-contaminate endometrial samples, resulting in a biased representation of the endometrial microbiome. This makes it difficult to demonstrate that the endometrial microbiome is not merely a reflection of contamination originating from sampling. Therefore, we investigated to what extent the endometrial microbiome corresponds to that of the vagina, applying culturomics on paired vaginal and endometrial samples. Culturomics could give novel insights into the microbiome of the female genital tract, as it overcomes sequencing-related bias. Ten subfertile women undergoing diagnostic hysteroscopy and endometrial biopsy were included. An additional vaginal swab was taken from each participant right before hysteroscopy. Both endometrial biopsies and vaginal swabs were analyzed using our previously described WASPLab-assisted culturomics protocol. In total, 101 bacterial and two fungal species were identified among these 10 patients. Fifty-six species were found in endometrial biopsies and 90 were found in vaginal swabs. On average, 28 % of species were found in both the endometrial biopsy and vaginal swab of a given patient. Of the 56 species found in the endometrial biopsies, 13 were not found in the vaginal swabs. Of the 90 species found in vaginal swabs, 47 were not found in the endometrium. Our culturomics-based approach sheds a different light on the current understanding of the endometrial microbiome. The data suggest the potential existence of a unique endometrial microbiome that is not merely a presentation of cross-contamination derived from sampling. However, we cannot exclude cross-contamination completely. In addition, we observe that the microbiome of the vagina is richer in species than that of the endometrium, which contradicts the current sequence-based literature.

Current findings in endometrial microbiome: impact on uterine diseases.

Microbiome or microbiota is essential to regulate many mammalian physiological processes, including reproduction. Like other organs or tissues, the upper female reproductive tract used to be considered as devoid of microorganisms; however, a non-infection-related bacterial community was discovered in the uterus from humans and other mammals, and its composition is related to reproductive success. The dysbiosis of endometrial microbiota is associated with benign and malign uterine diseases. Hence, this review addressed the current knowledge about uterine microbiota alterations and their association with common endometrial diseases, including endometrial polyposis, endometriosis, uterine myomatosis, endometrial hyperplasia, and endometrial cancer. There is a specific bacterial community in the endometrium in the most-analyzed uterine diseases. However, the constant finding consists in a reduced abundance of Firmicutes and Lactobacillus, while there is an increased abundance of Proteobacteria (such as Escherichia coli and Enterococcus), Bacteroidetes (Prevotella, for example), and Actinobacteria (as Gardnerella), in contrast to healthy endometrium. Besides, we discussed the future usefulness of the endometrial microbiota components as biomarkers to diagnose uterine diseases and their probable clinical outcomes. In addition, we analyzed their potential use as probiotics since they could provide an alternative or complement to existing therapies.

Microbiome as a predictor of implantation.

PURPOSE OF REVIEW: Review the latest research on the female urogenital microbiome as a predictor of successful implantation. RECENT FINDINGS: Lactobacillus crispatus seems to be beneficial species in a healthy female genital tract, although the presence of anaerobic bacteria and their impact has yet to be determined. The vaginal microbiome is associated with assisted reproductive technology (ART) outcome in terms of successful implantation and pregnancy. Approaches restoring a dysbiotic vaginal microbiome seem promising. It is questionable if a unique endometrial microbiome exists, given the low bacterial biomass, the invasiveness of endometrial sampling, and its associated high contamination risk. Future studies should focus on the whole microbiome using proteomics and metabolomics, as well as the virome to get a more holistic understanding of its role in reproduction. SUMMARY: The vaginal and endometrial compartments are being studied to determine a healthy and unhealthy microbiome composition. Defining a healthy composition could provide insight into physiological processes related to the success of embryo implantation. The vaginal microbiome is easily accessible and its composition can be reliably assessed and can be associated with ART outcome. The existence of an endometrial or uterine microbiome is still debated, due to the combination of low biomass and unavoidable high risk of contamination during sampling.

Differences in microbial profile of endometrial fluid and tissue samples in women with in vitro fertilization failure are driven by Lactobacillus abundance.

INTRODUCTION: The endometrial microbiota has been linked to several gynecological disorders, including infertility. It has been shown that the microbial profile of endometrium could have a role in fertilization and pregnancy outcomes. In this study we aim to assess the microbial community of endometrial tissue (ET) and endometrial fluid (EF) samples in women receiving in vitro fertilization (IVF) treatment. We also search for possible associations between chronic endometritis (CE) and endometrial microbiota. MATERIAL AND METHODS: This was a cohort study involving 25 women aged between 28 and 42 years with both primary and secondary infertility and with at least one IVF failure. The ET and EF sample collection was carried out between September 2016 and November 2018. Each of the participants provided two types of samples-tissue and fluid samples (50 samples in total). A 16S rRNA sequencing was performed on both of the sample types for microbial profile evaluation. CE was diagnosed based on a CD138 immunohistochemistry where CE diagnosis was confirmed in the presence of one or more plasma cells. Microbial profiles of women with and without CE were compared in both sample types separately. RESULTS: We report no differences in the microbial composition and alpha diversity (pObserved  = 0.07, pShannon  = 0.65, pInverse Simpson  = 0.59) between the EF and ET samples of IVF patients. We show that the abundance of the genus Lactobacillus influences the variation in microbial beta diversity between and fluid samples (r2  = 0.34; false discovery rate [FDR] <9.9 × 10-5 ). We report that 32% (8/25) of the participants had differences in Lactobacillus dominance in the paired samples and these samples also present a different microbial diversity (pShannon  = 0.06, FDRweighted UniFrac  = 0.01). These results suggest that the microbial differences between ET and fluid samples are driven by the abundance of genus Lactobacillus. The microbiome of CE and without CE (ie non-CE) women in our sample set of IVF patients was similar. CONCLUSIONS: Our findings show that genus Lactobacillus dominance is an important factor influencing the microbial composition of ET and fluid samples.

The Endometrial Microbiome and Its Impact on Human Conception.

Changes in the female genital tract microbiome are consistently correlated to gynecological and obstetrical pathologies, and tract dysbiosis can impact reproductive outcomes during fertility treatment. Nonetheless, a consensus regarding the physiological microbiome core inside the uterine cavity has not been reached due to a myriad of study limitations, such as sample size and experimental design variations, and the influence of endometrial bacterial communities on human reproduction remains debated. Understanding the healthy endometrial microbiota and how changes in its composition affect fertility would potentially allow personalized treatment through microbiome management during assisted reproductive therapies, ultimately leading to improvement of clinical outcomes. Here, we review current knowledge regarding the uterine microbiota and how it relates to human conception.

Gonococcal Pelvic Inflammatory Disease: Placing Mechanistic Insights Into the Context of Clinical and Epidemiological Observations.

While infection by Neisseria gonorrhoeae is often asymptomatic in women, undetected infections can ascend into the upper genital tract to elicit an inflammatory response that manifests as pelvic inflammatory disease, with the outcomes depending on the intensity and duration of inflammation and whether it is localized to the endometrial, fallopian tube, ovarian, and/or other tissues. This review examines the contribution of N. gonorrhoeae versus other potential causes of pelvic inflammatory disease by considering new insights gained through molecular, immunological, and microbiome-based analyses, and the current epidemiological burden of infection, with an aim to highlighting key areas for future study.

16S rRNA long-read nanopore sequencing is feasible and reliable for endometrial microbiome analysis.

RESEARCH QUESTION: Full-length 16S rRNA gene sequencing using nanopore technology is a fast alternative to conventional short-read 16S rRNA gene sequencing with low initial investment costs that has been used for various microbiome studies but has not yet been investigated as an alternative approach for endometrial microbiome analysis. Is in-situ 16S rRNA gene long-read sequencing using portable nanopore sequencing technology feasible and reliable for endometrial microbiome analysis? DESIGN: A prospective experimental study based on 33 patients seeking infertility treatment between January and October 2019. A 16S rRNA gene long-read nanopore sequencing protocol for analysing endometrial microbiome samples was established, including negative controls for contamination evaluation and positive controls for bias evaluation. Contamination caused by kit and exterior sources was identified and excluded from the analysis. Endometrial samples from 33 infertile patients were sequenced using the optimized long-read nanopore sequencing protocol and compared with conventional short-read sequencing carried out by external laboratories. RESULTS: Of the 33 endometrial patient samples, 23 successfully amplified (69.7%) and their microbiome was assessed using nanopore sequencing. Of those 23 samples, 14 (60.9%) were Lactobacillus-dominated (>80% of reads mapping to Lactobacillus), with 10 samples resulting in more than 90% Lactobacillus reads. Our long-read nanopore sequencing revealed results similar to two conventional short-read sequencing approaches and to long-read sequencing validation carried out in external laboratories. CONCLUSION: In this pilot study, 16S rRNA gene long-read nanopore sequencing was established to analyse the endometrial microbiome in situ that could be widely applied owing to its cost efficiency and portable character.

The Role of Genital Tract Microbiome in Fertility: A Systematic Review.

The human microbiome plays a crucial role in determining the health status of every human being, and the microbiome of the genital tract can affect the fertility potential before and during assisted reproductive treatments (ARTs). This review aims to identify and appraise studies investigating the correlation of genital microbiome to infertility. Publications up to February 2021 were identified by searching the electronic databases PubMed/MEDLINE, Scopus and Embase and bibliographies. Only full-text original research articles written in English were considered eligible for analysis, whereas reviews, editorials, opinions or letters, case studies, conference papers, and abstracts were excluded. Twenty-six articles were identified. The oldest studies adopted the exclusive culture-based technique, while in recent years PCR and RNA sequencing based on 16S rRNA were the most used technique. Regardless of the anatomical site under investigation, the Lactobacillus-dominated flora seems to play a pivotal role in determining fertility, and in particular Lactobacillus crispatus showed a central role. Nonetheless, the presence of pathogens in the genital tract, such as Chlamydia trachomatis, Gardnerella vaginalis, Ureaplasma species, and Gram-negative stains microorganism, affected fertility also in case of asymptomatic bacterial vaginosis (BV). We failed to identify descriptive or comparative studies regarding tubal microbiome. The microbiome of the genital tract plays a pivotal role in fertility, also in case of ARTs. The standardization of the sampling methods and investigations approaches is warranted to stratify the fertility potential and its subsequent treatment. Prospective tubal microbiome studies are warranted.

Peptidoglycan disrupts early embryo-maternal crosstalk via suppression of ISGs expression induced by interferon-tau in the bovine endometrium.

Uterine infection with bacteria and the release of peptidoglycan (PGN), antigenic cell wall components of both Gram-negative and Gram-positive bacteria, can cause early pregnancy losses in ruminants, but the associated mechanisms remain unsolved. Day 7 blastocyst starts to secrete a minute amount of interferon-tau (IFNT) in the uterine horn which is required for early stage of maternal recognition of pregnancy (MRP) in ruminants, and it induces interferon stimulated genes (ISGs) for driving uterine receptivity in cows. This study investigated if PGN disrupts IFNT response through modulation of endometrial ISGs expressions. Cultured bovine endometrial epithelial cells (BEECs) were treated with embryo culture medium (ECM) or IFNT (1 ng/ml) in the presence or absence of a low level of PGN (10 pg/ml) for 24 h. A real-time PCR analyses revealed that the presence of PGN suppressed IFNT-induced ISGs (OAS1 and ISG15) and STAT1 expressions in BEECs. To visualize the impact of PGN in an ex-vivo model that resembles the in vivo status, endometrial explants were treated by IFNT (1 ng/ml) with or without PGN (10 pg/ml) for 12 h. PGN suppressed IFNT-induced gene expressions of the above factors, but not for IFNA receptor type1 (IFNAR1) or type2 (IFNAR2) in explants. Immunofluorescence analysis illustrated that PGN completely suppressed the IFNT-triggered OAS1 protein expression in the luminal epithelium of explants. Of note, PGN did not stimulate pro-inflammatory cytokines (TNFA and IL1B) or TLR2 mRNA expression in both models. These findings indicate that the presence of low levels of PGN suppresses ISGs expression induced by IFNT secreted from early embryo, at the luminal epithelium of the bovine endometrium. This could severely interfere with early stage of MRP processes in cows, leading to pregnancy failure.