Vaginal dysbiosis - the association with reproductive outcomes in IVF patients: a systematic review and meta-analysis.

PURPOSE OF REVIEW: To examine impact of vaginal dysbiosis (VD), including bacterial vaginosis (BV) and aerobic vaginitis (AV) on reproductive outcomes of in vitro fertilization (IVF) patients. RECENT FINDINGS: BV-bacteria (e.g. Gardnerella ) and AV-bacteria (e.g. Streptococci and Enterococci ) have been identified in the endometrium. However, there is inconclusive evidence whether IVF patients with VD have lower success rates. SUMMARY: The present systematic review and meta-analysis of PubMed/Medline, until December 2023 included 25 studies, involving 6835 IVF patients. Overall VD was defined as an approximation of community state type IV, including BV and AV-type dysbiosis based on either molecular or microscopy methods. Outcomes were live birth rate (LBR), early pregnancy loss (EPL), clinical pregnancy rate (CPR), and biochemical pregnancy rate (BPR).Vaginal dysbiosis prevalence was 19% [1271/6835, 95% confidence interval (CI) 18-20%]. Six studies examined AV-type dysbiosis with a prevalence of 4% (26/628, 95% CI 3-6%). Vaginal dysbiosis correlates with a higher EPL [relative risk (RR) = 1.49, 95% CI 1.15-1.94] and lower CPR (RR = 0.82, 95% CI 0.70-0.95). No statistically significant impact of VD, BV, or AV was found on LBR and BPR.Thus, the association between VD and reproductive outcome remains puzzling as it is difficult to explain how VD impacts CPR and EPL but not LBR and BPR.

GnRHa trigger-the story of the ugly duckling.

The objective of this invited review is to describe the introduction and development of the GnRH agonist (GnRHa) trigger protocol in modern in vitro fertilization, focusing on ovarian hyperstimulation syndrome (OHSS) prevention and, equally important on the role of GnRHa trigger as an opener of the luteal phase "black box." The GnRHa trigger and freezing of all embryos is the ultimate weapon against OHSS in the OHSS-risk patient. In the non OHSS-risk patient, GnRHa trigger followed by a modified luteal phase support with lutein hormone activity and subsequent fresh embryo transfer results in excellent reproductive outcomes. Thus, the GnRHa trigger has paved the way for a virtually OHSS-free clinic, and equally important is the fact that the early lessons learned from studying the GnRHa trigger opened the "black box" of the luteal phase enabling improved reproductive outcomes in both fresh and frozen embryo transfer cycles.

Lactobacilli and Their Probiotic Effects in the Vagina of Reproductive Age Women.

In the present narrative review, the probiotic effects of vaginal Lactobacillus spp. are described in detail, covering the importance of the differential production of lactic acid, the lactic acid D/L isoforms, the questionable in vivo effect of hydrogen peroxide, as well as bacteriocins and other core proteins produced by vaginal Lactobacillus spp. Moreover, the microbe-host interaction is explained with emphasis on the vaginal mucosa. To understand the crucial role of Lactobacillus spp. dominance in the vaginal microbiota, different dysbiotic states of the vagina are explained including bacterial vaginosis and aerobic vaginitis. Finally, this review takes on the therapeutic aspect of live lactobacilli in the context of bacterial vaginosis. Until recently, there was very low-quality evidence to suggest that any probiotic might aid in reducing vaginal infections or dysbiosis. Therefore, clinical usage or over the counter usage of probiotics was not recommended. However, recent progress has been made, moving from probiotics that are typically regulated as food supplements to so-called live biotherapeutic products that are regulated as medical drugs. Thus, recently, a phase 2b trial using a Lactobacillus crispatus strain as a therapeutic add-on to standard metronidazole showed significant reduction in the recurrence of bacterial vaginosis by 12 weeks compared to placebo. This may constitute evidence for a brighter future where the therapeutic use of lactobacilli can be harnessed to improve women's health.

The relationship of total progressive motile sperm count with the outcome of IUI? An analysis of 5171 cycles.

Background: The role of motile sperm count in intrauterine insemination (IUI) success rate is controversial. This retrospective cohort study performed among unselected infertile couples undergoing IUI was to explore the association between the total progressive motile sperm count (TPMSC) and the live birth rate (LBR) following IUI.Methods: The total cohort of 5363 cycles, 2666 infertile couples between January 2015 and December 2018 and finally 5171 cycles, 2647 couples were included for analysis in Sun Yat-sen memorial hospital of Sun Yat-sen University. The primary outcome was LBR per cycle. And the secondary outcome measure was clinical pregnancy rate (CPR) per cycle.Results: From the receiver operating characteristic (ROC) analysis of female age predicting live birth, female age cutoff was defined as 28 years. With a female age of ≤28 years, the CPRs were 11.5%, 14.9%, 16.1%, and 15.8% in quartile groups of pre-wash TPMSC, respectively. For the LBRs the values were 9.4%, 12.9%, 14.4%, and 11.3%, and there were also no significant differences in quartile groups of pre-wash TPMSC with ≤24 million (M), [24M-50M], [50M-97M], >97M. No statistically significant differences in the CPRs (p = .051) and LBRs (p = .088) were also observed in the quartiles groups of post-wash TPMSC. With a female age of >28 years, the CPR in couples with post-wash TPMSC ≤22.32 M was significantly lower than with post-wash TPMSC >81.0 M (p = .007). There was an obvious trend in which CPRs and LBRs increased with the post-wash TPMSC during the <81 M interval in women >28 years.Conclusions: The optimal female age cutoff for live birth was 28 years in IUI cycles. Pre-wash and post-wash TPMSC were not significantly associated with CPR and LBR per cycle. When female age >28 years, there was a better outcome with post-wash TPMSC >22.32 million.

The combined effect of lifestyle intervention and antioxidant therapy on sperm DNA fragmentation and seminal oxidative stress in IVF patients: a pilot study

ABSTRACT Purpose: Sperm DNA fragmentation (SDF) and seminal oxidative stress are emerging measurable factors in male factor infertility, which interventions could potentially reduce. We evaluated (i) the impact of lifestyle changes combined with oral antioxidant intake on sperm DNA fragmentation index (DFI) and static oxidation-reduction potential (sORP), and (ii) the correlation between DFI and sORP. Materials and Methods: We conducted a prospective study involving 93 infertile males with a history of failed IVF/ICSI. Ten healthy male volunteers served as controls. Semen analysis was carried out according to 2010 WHO manual, whereas seminal sORP was measured using the MiOXSYS platform. SDF was assessed by sperm chromatin structure assay. Participants with DFI >15% underwent a three-month lifestyle intervention program, primarily based on diet and exercise, combined with oral antioxidant therapy using multivitamins, coenzyme Q10, omega-3, and oligo-elements. We assessed changes in semen parameters, DFI, and sORP, and compared DFI results to those of volunteers obtained two weeks apart. Spearman rank correlation tests were computed for sORP and DFI results. Results: Thirty-eight (40.8%) patients had DFI >15%, of whom 31 participated in the intervention program. A significant decrease in median DFI from 25.8% to 18.0% was seen after the intervention (P <0.0001). The mean DFI decrease was 7.2% (95% CI: 4.8-9.5%; P <0.0001), whereas it was 0.42% (95%CI; -4.8 to 5.6%) in volunteers (P <0.00001). No differences were observed in sperm parameters and sORP. Based on paired sORP and DFI data from 86 patients, no correlation was observed between sORP and DFI values (rho=0.03). Conclusion: A 3-month lifestyle intervention program combined with antioxidant therapy reduced DFI in infertile men with elevated SDF and a history of failed IVF/ICSI. A personalized lifestyle and antioxidant intervention could improve fertility of subfertile couples through a reduction in DFI, albeit controlled trials evaluating reproductive outcomes are needed before firm conclusions can be made. Trial registration number and date: clinicaltrials.gov NCT03898752, April 2, 2019.

Vaginal dysbiosis in pregnancy associates with risk of emergency caesarean section: a prospective cohort study.

OBJECTIVES: To investigate changes in vaginal microbiota during pregnancy, and the association between vaginal dysbiosis and reproductive outcomes. METHODS: A total of 730 (week 24) and 666 (week 36) vaginal samples from 738 unselected pregnant women were studied by microscopy (Nugent score) and characterized by 16S rRNA gene sequencing. A novel continuous vaginal dysbiosis score was developed based on these methods using a supervised partial least squares model. RESULTS: Among women with bacterial vaginosis in week 24 (n = 53), 47% (n = 25) also had bacterial vaginosis in week 36. In contrast, among women without bacterial vaginosis in week 24, only 3% (n = 18) developed bacterial vaginosis in week 36. Vaginal samples dominated by Lactobacillus crispatus (OR 0.35, 95% CI 0.20-0.60) and Lactobacillus iners (OR 0.40, 95% CI 0.23-0.68) in week 24 were significantly more stable by week 36 when compared with other vaginal community state types. Vaginal dysbiosis score at week 24 was associated with a significant increased risk of emergency, but not elective, caesarean section (OR 1.37, 955 CI 1.15-1.64, p < 0.001), suggesting a 37% increased risk per standard deviation increase in vaginal dysbiosis score. CONCLUSIONS: Changes in vaginal microbiota from week 24 to week 36 of pregnancy correlated with bacterial vaginosis status and vaginal community state type. A novel vaginal dysbiosis score was associated with a significantly increased risk of emergency, but not elective, caesarean section. This was not found for bacterial vaginosis or any vaginal community state type and could point to the importance of investigating vaginal dysbiosis as a nuanced continuum instead of crude clusters.

The combined effect of lifestyle intervention and antioxidant therapy on sperm DNA fragmentation and seminal oxidative stress in IVF patients: a pilot study.

PURPOSE: Sperm DNA fragmentation (SDF) and seminal oxidative stress are emerging measurable factors in male factor infertility, which interventions could potentially reduce. We evaluated (i) the impact of lifestyle changes combined with oral antioxidant intake on sperm DNA fragmentation index (DFI) and static oxidation-reduction potential (sORP), and (ii) the correlation between DFI and sORP. MATERIALS AND METHODS: We conducted a prospective study involving 93 infertile males with a history of failed IVF/ICSI. Ten healthy male volunteers served as controls. Semen analysis was carried out according to 2010 WHO manual, whereas seminal sORP was measured using the MiOXSYS platform. SDF was assessed by sperm chromatin structure assay. Participants with DFI >15% underwent a three-month lifestyle intervention program, primarily based on diet and exercise, combined with oral antioxidant therapy using multivitamins, coenzyme Q10, omega-3, and oligo-elements. We assessed changes in semen parameters, DFI, and sORP, and compared DFI results to those of volunteers obtained two weeks apart. Spearman rank correlation tests were computed for sORP and DFI results. RESULTS: Thirty-eight (40.8%) patients had DFI >15%, of whom 31 participated in the intervention program. A significant decrease in median DFI from 25.8% to 18.0% was seen after the intervention (P <0.0001). The mean DFI decrease was 7.2% (95% CI: 4.8-9.5%; P <0.0001), whereas it was 0.42% (95%CI; -4.8 to 5.6%) in volunteers (P <0.00001). No differences were observed in sperm parameters and sORP. Based on paired sORP and DFI data from 86 patients, no correlation was observed between sORP and DFI values (rho=0.03). CONCLUSION: A 3-month lifestyle intervention program combined with antioxidant therapy reduced DFI in infertile men with elevated SDF and a history of failed IVF/ICSI. A personalized lifestyle and antioxidant intervention could improve fertility of subfertile couples through a reduction in DFI, albeit controlled trials evaluating reproductive outcomes are needed before firm conclusions can be made. Trial registration number and date: clinicaltrials.gov NCT03898752, April 2, 2019.

The exogenous progesterone-free luteal phase: two pilot randomized controlled trials in IVF patients.

RESEARCH QUESTION: Is the reproductive outcome similar after gonadotrophin-releasing hormone agonist (GnRHa) trigger followed by luteal human chorionic gonadotrophin (HCG) boluses compared with HCG trigger and a standard luteal phase support (LPS)? DESIGN: Two open-label pilot randomized controlled trials (RCT) with 250 patients from 2014 to 2019, with a primary outcome of ongoing pregnancy per embryo transfer. Patients with ≤13 follicles on the trigger day were randomized (RCT 1) to: Group A (n = 65): GnRHa trigger followed by a bolus of 1500 IU HCG s.c. on the oocyte retrieval day (ORD) and 1000 IU HCG s.c. 4 days later, and no vaginal LPS; or Group B (n = 65): 6500 IU HCG trigger, followed by a standard vaginal progesterone LPS. Patients with 14-25 follicles on the trigger day were randomized (RCT 2) to Group C (n = 60): GnRHa trigger followed by 1000 IU HCG s.c. on ORD and 500 IU HCG s.c. 4 days later, and no vaginal LPS; or Group D (n = 60): 6500 IU HCG trigger and a standard vaginal LPS. RESULTS: In RCT 1, the ongoing pregnancy rate was 44% (22/50) in the GnRHa group versus 46% (25/54) in the HCG trigger group (RR 0.95, 95% CI 0.62-1.45). No ovarian hyperstimulation syndrome (OHSS) was seen in Groups A or B. In RCT 2, the ongoing pregnancy rate was 51% (25/49) in the GnRHa group versus 60% (31/52) in the HCG trigger group (RR 0.86, 95% CI 0.60-1.22). The OHSS rates were 3.3% and 6.7%, respectively. CONCLUSIONS: Although a larger-scale study is needed before standard clinical implementation, the present study supports that the exogenous progesterone-free LPS is efficacious, simple and patient-friendly.

The Association between Vaginal Dysbiosis and Reproductive Outcomes in Sub-Fertile Women Undergoing IVF-Treatment: A Systematic PRISMA Review and Meta-Analysis.

Recent advances in molecular microbiology have enabled refined studies of the genital tract microbiota. This constitutes the basis of the present updated systematic review and meta-analysis which investigate vaginal dysbiosis (VD) as defined by either microscopy (e.g., Nugent score for bacterial vaginosis) or molecular methods (qPCR and Next Generation Sequencing) to evaluate the impact of VD on the reproductive outcomes in women undergoing IVF-treatment. A total of 17 studies were included, comprising 3543 patients and with a VD prevalence of 18% (95%CI 17-19). Across all methods, VD is a significant risk factor for early pregnancy loss in IVF (Relative risk (RR) = 1.71 95%CI 1.29-2.27). Moreover, a predefined sub-analysis of studies using molecular methods for VD diagnosis showed a significant reduction in the clinical pregnancy rate when compared to normal vaginal microbiota patients (RR = 0.55 95%CI 0.32-0.93). However, regardless of diagnostic methodology, VD did not significantly influence live birth rate (LBR). In conclusion, molecular tools have provided a more detailed insight into the vaginal microbiota, which may be the reason for the increased adverse effect estimates in IVF patients with molecularly defined VD. However, the quality of evidence was very low across all outcomes according to GRADE and thus, more studies are warranted to understand the impact of VD in IVF.

Analysing endometrial microbiome: methodological considerations and recommendations for good practice.

There is growing evidence that the upper female genital tract is not sterile, harbouring its own microbial communities. However, the significance and the potential effect of endometrial microorganisms on reproductive functions remain to be fully elucidated. Analysing the endometrial microbiome, the microbes and their genetic material present in the endometrium, is an emerging area of study. The initial studies suggest it is associated with poor reproductive outcomes and with different gynaecological pathologies. Nevertheless, studying a low-biomass microbial niche as is endometrium, the challenge is to conduct well-designed and well-controlled experiments in order to avoid and adjust for the risk of contamination, especially from the lower genital tract. Herein, we aim to highlight methodological considerations and propose good practice recommendations for future endometrial microbiome studies.

The POSEIDON stratification - moving from poor ovarian response to low prognosis.

Poor ovarian response remains one of the most challenging tasks for an IVF clinician. In this review, we aim to highlight the ongoing research for optimizing the prognosis in poor ovarian response patients. The newly introduced POSEIDON criteria argue that the first step is to move from a poor response to a poor prognosis concept, while improving identification and stratification of the different sub-types of poor prognosis patients prior to ovarian stimulation. The immediate marker of success is the ability of the ovarian stimulation to retrieve the number of oocytes needed to obtain at least one euploid blastocyst for transfer in each patient. This surrogate marker of success should not replace live birth as the most important outcome, but it should be approached as a useful tool for clinicians to evaluate their strategy for achieving live birth in the shortest timespan possible in the individual patient/ couple.

Vaginal, Cervical and Uterine pH in Women with Normal and Abnormal Vaginal Microbiota.

INTRODUCTION: Healthy women of reproductive age have a vaginal pH around 4.5, whereas little is known about pH in the upper genital tract. A shift in the vaginal microbiota may result in an elevated pH in the upper genital tract. This might contribute to decreased fertility and increased risk of preterm birth. Therefore, we aimed to measure pH in different compartments of the female genital tract in both nonpregnant and pregnant women, stratifying into a normal and abnormal vaginal microbiota. MATERIAL AND METHODS: In this descriptive study, we included 6 nonpregnant, 12 early-pregnant, and 8 term-pregnant women. A pH gradient was recorded with a flexible pH probe. An abnormal vaginal microbiota was diagnosed by a quantitative polymerase chain reaction technique for Atopobium vaginae; Sneathia sanguinegens; Leptotrichia amnionii; bacterial vaginosis-associated bacterium 1, 2, 3, and TM7; and Prevotella spp. among others. RESULTS: In all participants we found the pH gradient in the lower reproductive canal to be most acidic in the lower vagina and most alkaline in the upper uterine cavity. Women with an abnormal vaginal microbiota had an increased pH in the lower vagina compared to the other groups. CONCLUSIONS: There is a pronounced pH gradient within the female genital tract. This gradient is not disrupted in women with an abnormal vaginal microbiota.

Effect of clindamycin and a live biotherapeutic on the reproductive outcomes of IVF patients with abnormal vaginal microbiota: protocol for a double-blind, placebo-controlled multicentre trial.

INTRODUCTION: Recent studies in in vitro fertilisation (IVF) patients have associated abnormal vaginal microbiota (AVM) with poor clinical pregnancy rates of 6%-9% per embryo transfer. The biological plausibility for this finding is hypothesised to be ascending infection to the endometrium which in turn hampers embryo implantation. New molecular based diagnosis may offer advantages compared to microscopical diagnosis of AVM which has huge inter-study variability ranging from 4 to 38%; however, the important question is whether screening and treatment of AVM would improve reproductive outcomes in IVF patients. Herein, we describe a protocol for an ongoing double-blind, placebo-controlled multicentre trial of IVF patients diagnosed with AVM and randomised in three parallel groups 1:1:1. METHODS AND ANALYSIS: This is a drug intervention study where IVF patients will be screened for AVM, using a qPCR assay targeting Atopobium vaginae and Gardnerella vaginalis. If positive, patients will be randomised to one of the three study arms. The first arm consists of clindamycin 300 mg ×2 daily for 7 days followed by vaginal Lactobacillus crispatus CTV-05 until clinical pregnancy scan week 7-9. The second arm consists of clindamycin and placebo L. crispatus CTV-05, whereas patients in the third arm will be treated with placebo/placebo. We used a superiority design to estimate that active treatment in both arms will increase the primary outcome, clinical pregnancy rate per embryo transfer, from 20% to 40%. A potential difference between the two active arms was considered exploratory. With a power of 80% and an alpha at 5%, the sample size is estimated to be 333 patients randomised. A pre-planned interim analysis is scheduled at 167 patients randomised. ETHICS AND DISSEMINATION: All patients have to give informed consent. Dissemination of results is ensured in clinical trial agreements whether they be positive or not. Ethics committee, Central Denmark Region approved this protocol. TRIAL REGISTRATION NUMBER: ICH-GCP monitored trial, EudraCT 2016-002385-31; Pre-results.

Characterization of the Vaginal DNA Virome in Health and Dysbiosis.

Bacterial vaginosis (BV) is characterized by a reduction in Lactobacillus (L.) spp. abundance and increased abundance of facultative anaerobes, such as Gardnerella spp. BV aetiology is not fully understood; however, bacteriophages could play a pivotal role in the perturbation of the vaginal bacterial community. We investigated the vaginal viral community, including bacteriophages and the association to the bacterial community and BV-status. Vaginal samples from 48 patients undergoing IVF treatment for non-female factor infertility were subjected to metagenomic sequencing of purified virus-like particles. The vaginal viral community was characterized and correlated with the BV-status by Nugent score, bacterial community, structure, and the presence of key vaginal bacterial species. The majority of identified vaginal viruses belonged to the class of double-stranded DNA bacteriophages, with eukaryotic viruses constituting 4% of the total reads. Clear links between the viral community composition and BV (q = 0.006, R = 0.26) as well as the presence of L. crispatus (q = 0.001, R = 0.43), L. iners, Gardnerella spp., and Atopobium vaginae were found (q < 0.002, R > 0.15). The eukaryotic viral community also correlated with BV-status (q = 0.018, R = 0.20). In conclusion, the vaginal virome was clearly linked with bacterial community structure and BV-status.

Gonadotropin-releasing hormone agonist for ovulation trigger - OHSS prevention and use of modified luteal phase support for fresh embryo transfer.

The introduction of gonadotrophin-releasing hormone agonist (GnRHa) trigger greatly impacted modern IVF treatment. Patients at low risk of ovarian hyperstimulation syndrome (OHSS) development, undergoing fresh embryo transfer and GnRHa trigger can be offered a virtually OHSS-free treatment with non-inferior reproductive outcomes by using a modified luteal phase support in terms of small boluses of human chorionic gonadotrophin (hCG), daily recombinant luteinizing hormone LH (rLH) or GnRHa. In the OHSS risk patient, GnRHa trigger can safely be performed, followed by a 'freeze-all' policy with a minimal risk of OHSS development and high live birth rates in the subsequent frozen embryo transfer cycle. Importantly, GnRHa trigger opened the 'black box' of the luteal phase, promoting research in the most optimal steroid levels during the luteal phase. GnRHa trigger allows high-dose gonadotropin stimulation to achieve the optimal number of oocytes and embryos needed to ensure the highest chance of live birth. This review thoroughly discusses how the GnRHa trigger concept adds safety and efficacy to modern IVF in terms of OHSS prevention. Furthermore, the optimal luteal phase management after GnRHa trigger in fresh embryo transfer cycles is discussed.

Research and business - the yin and yang in modern medicine.

Yin and yang is a concept of dualism in Chinese philosophy, describing how opposite or contrary forces may be complementary, interconnected and interdependent, and how they may give rise to each other as they interrelate with one another. In line with this, modern clinical research and business can definitely be described as yin and yang. With the increasing need for funding, researchers at a very early stage during the development of a new concept may be forced or tempted to enter the business world. Furthermore, researchers are encouraged and supported by their own universities to collaborate with possible future business partners, not only to acquire funding, but also to explore potential patenting. This collaboration between the business world and research can definitely be very fruitful and provide benefit for both parties, patients and society as a whole, but it may also introduce the risk of premature materialization.

Novel Physiology and Definition of Poor Ovarian Response; Clinical Recommendations.

Poor ovarian response (POR) to controlled ovarian stimulation (OS) presents a major challenge in assisted reproduction. The Bologna criteria represented the first serious attempt to set clear criteria for the definition of POR. However, the Bologna criteria were questioned because of the persistent heterogeneity among POR patients and the inability to provide management strategies. Based on these facts, a more recent classification, the POSEIDON (Patient-Oriented Strategies Encompassing IndividualizeD Oocyte Number) classification, was developed to provide a homogeneous and refined definition of POR that significantly reduces the heterogeneity of the Bologna criteria definition of POR and helps in the clinical handling and counseling of patients. In this review, we discuss the impact of the POSEIDON classification on the clinical management of patients with POR.

Gonadotropin-releasing hormone agonist ovulation trigger-beyond OHSS prevention.

In this review the advantages of the gonadotropin-releasing hormone agonist (GnRHa) trigger are discussed beyond those immediately associated with ovarian hyperstimulation syndrome (OHSS) prevention. The GnRHa trigger concept has sparked the development of novel protocols, enriching the assisted reproductive technology (ART) armamentarium for the benefit of present and future patients. Thus, GnRHa trigger already has a pivotal role, not only for the standard in vitro fertilisation (IVF) patient, but also for patient groups like oocyte donors, cancer patients, patients with poor ovarian reserve, and patients with immature oocyte syndrome and empty follicle syndrome. Herein, we discuss the importance of the GnRHa-elicited midcycle FSH surge and the potential improvement in oocyte yield and embryo competence.