An observational study in Repeated IVF Failure and Microbiota, sponsored by Assisting Nature. Enrolling by invitation at 1 site in Greece. Open to female participants aged 18 Years to 54 Years. Per ClinicalTrials.gov, last updated 2026-05-26.
Sponsored by Assisting Nature · Observational
The goal of this observational study is to investigate the influence and contribution of the normal composition and balance of the endometrial microbiota on the achievement of embryo implantation and the maintenance of pregnancy in women undergoing IVF, with comparable clinical profiles, including patients with a history of RIF or RPL, evaluated under identical treatment protocols and hormonal conditions.
The main questions it aims to answer are:
Participants will not undergo any additional interventions beyond standard clinical care.They will provide clinical data already collected as part of routine IVF procedures.
Endometrial biopsies were collected from patients using a Pipelle catheter during the designated phase of the menstrual cycle. Each sample was immediately placed in a specialized preservative solution and transported to two external laboratories for analysis.
In the first laboratory, bacterial DNA was extracted using a CE-IVD kit, followed by targeted amplification and barcoded sequencing of seven hypervariable regions (V2, V3, V4, V6, V7, V8, V9) of the 16S rRNA gene using the 16STM metagenomic kit and Ion Torrent sequencing system (Ion GeneStudio S5 with Ion Chef Instrument). Data were analyzed with Ion Reporter™ software and MicroSEC ID, using MicroSEC ID and Greengenes databases for taxonomic classification.
In the second laboratory, total DNA was extracted, and targeted amplification of the 16S rRNA gene was performed using the 16S Barcoding Kit 24 V14 (Nanopore), followed by NGS on the MinION Mk1C analyzer. Bioinformatic analysis was conducted using the EPI2ME Labs 16S workflow. Bacterial composition was classified at the genus level, with results expressed as relative abundance percentages. OTU-level data were not disclosed.
Interpretation of microbiota profiles was performed by the treating physician or clinical geneticist. Patients whose samples showed decreased Lactobacillus abundance or the presence of potentially pathogenic microorganisms received individualized interventions. These interventions included a two-week course of antibiotics tailored to the patient's clinical history, followed by probiotics administered orally or via combined oral and vaginal routes for two weeks to two months. Antifungal agents were administered when clinically indicated.
Embryo transfer schedules were adapted based on microbial profiles and clinical assessment. Patients with normal microbial profiles proceeded with IVF in the next cycle. Patients with abnormal profiles, signs of inflammation, or potential pathogenic colonization had embryo transfer postponed for at least one month following completion of the therapeutic regimen.
The study provides a detailed methodological framework for endometrial microbiota analysis, including sample collection, DNA extraction, sequencing strategies, bioinformatic pipelines, and clinical decision-making based on microbial composition, aiming to support personalized management in assisted reproduction.
Assisting Nature is the lead sponsor of 10 studies on the registry; 1 is open to participants now.
Counted across the registry records on this site, refreshed daily.
Participants were required to be between 18 and 54 years of age, in accordance with the legally permitted age limits for embryo transfer in Greece
Exclusion Criteria:
Participants whose endometrial microbiota is dominated by Lactobacillus species. Data were collected from previously obtained endometrial biopsies, analyzed via 16S rRNA gene sequencing. No interventions were performed beyond standard IVF clinical care.
Participants whose endometrial microbiota shows depletion of Lactobacillus species or presence of potentially pathogenic microorganisms. Data were collected from previously obtained endometrial biopsies, analyzed via 16S rRNA gene sequencing. In some cases, participants received individualized clinical interventions (e.g., antibiotics or probiotics) based on their microbial profile as part of routine care.
Implantation Rate according to microbiome profile
The study formally assesses differences in embryo implantation rates between women with Lactobacillus-dominant and non-Lactobacillus-dominant endometrial microbiota profiles.
Time frame: From the day of the embryo transfer to the day of confirmation of implantation (2-4 weeks)
Ongoing Pregnancy/Live Birth Rate according to microbiome profile
Association between endometrial microbiota composition and the likelihood of achieving an ongoing pregnancy or live birth
Time frame: From the day of embryo transfer up to the end of the first trimester (up to 12 weeks of gestation) and live birth until delivery (up to 40 weeks of gestation)
Biochemical pregnancy rate and microbiome status
Rate of biochemical pregnancies in relation to endometrial microbiota profile
Time frame: From the day of embryo transfer to the day of confirmation of implantation (2-4 weeks)
Endometrial microbiota composition
Relative abundance of Lactobacillus versus non-Lactobacillus genera in endometrial biopsies analyzed by 16S rRNA sequencing.
Time frame: At time of biopsy collection (during procedure)
Plan to share: Undecided
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