Precision Microbial Therapeutics for Infertility: Next-Generation Probiotics, Engineered Biologics and Translational Pathways.

Translation of microbiome science into reproductive therapeutics is constrained by a mismatch between human reproductive physiology and the models commonly used to test interventions. Murine vaginal and uterine microbial communities differ taxonomically and functionally from human communities, and mouse estrous cycling, glycogen metabolism and mucosal immune architecture create a distinct ecological and immunologic baseline that limits direct extrapolation of colonisation dynamics and host responses observed in mice (Greenberg Jonathan et al.  2022 ; Mejia et al.  2023 ; He et al.  2025 ). To narrow this gap, the field is moving toward humanised models in which human reproductive microbiota and immune elements are engrafted into immunocompromised or antibiotic‐conditioned mice; such humanised‐mouse systems permit testing of colonisation efficiency, interkingdom interactions and microbe driven immune training within a controlled in vivo environment while retaining the capacity for longitudinal sampling and functional readouts (Mejia et al.  2023 ; Rahman et al.  2023 ). Parallel advances in ex vivo systems have made organoids derived from primary human endometrium a practical platform for mechanistic and therapeutic screening because these cultures retain hormone responsiveness, glandular architecture and pregnancy‐associated transcriptional programs (Turco et al.  2017 ). When organoids are combined with stromal, immune and endothelial compartments in microfluidic endometrium‐on‐chip devices, they recapitulate dynamic hormone cycles, perfusion and cell–cell interactions required to assess mucosal colonisation, metabolite diffusion and mucosal immune modulation by candidate live biotherapeutics (Dai et al.  2025 ; Deng et al.  2024 ). Taken together, a preclinical strategy that layers humanised in vivo models with multi‐cellular, perfused in vitro platforms will provide the fidelity required to prioritise strains and formulations for clinical testing while reducing reliance on imperfect conventional mouse models (Mejia et al.  2023 ; Turco et al.  2017 ). Integrating microbiome profiling into fertility workups creates an opportunity to convert descriptive microbiology into actionable patient stratification. Early clinical studies showed that the composition of the endometrial microbiota, and in particular the presence or absence of a Lactobacillus ‐dominated community, predicts implantation and live birth outcomes after embryo transfer, and follow‐up cohorts have reinforced these associations across diverse clinical settings (Moreno et al.  2016 , 2022 ). For therapeutics that seek to modulate the reproductive microbiome, sequencing‐based assays such as 16S rRNA gene amplicon profiling and shotgun metagenomics can operate as companion diagnostics to document baseline dysbiosis, to define microbiome endotypes, and to monitor engraftment and ecological shifts after intervention (Polifke et al.  2024 ; Liu et al.  2025 ). Because sampling depth, amplification targets and contamination control substantially influence result interpretation, diagnostic deployment must adopt standardised sampling protocols and validated, contamination‐aware pipelines for low‐biomass samples, including routine extraction and sampling blanks, quantified biomass (qPCR), mock‐community and dilution series benchmarks and prespecified contaminant‐filtering procedures, and should report thresholds that map microbial states to clinically meaningful endotypes for targeted next‐generation probiotics (Polifke et al.  2024 ; Liu et al.  2025 ; Davis et al.  2018 ; Eisenhofer et al.  2019 ; Salter et al.  2014 ). In practice, a companion diagnostic framework would allow selection of patients most likely to benefit from a specific live biotherapeutic, permit early stopping rules for nonengraftment and create mechanistic biomarkers that bridge microbial modulation with mucosal immunity and reproductive endpoints (Moreno et al.  2022 ; Liu et al.  2025 ). Studies of the endometrium and other upper reproductive tract sites face distinct technical hazards because microbial biomass is low and contaminant DNA from reagents, extraction kits and the laboratory environment can constitute a large fraction of recovered sequences. To ensure diagnostic value, we recommend the minimum methodological elements for any diagnostic pipeline and clinical study that includes low‐biomass sampling: (i) inclusion of multiple negative controls at sampling and extraction (swab blanks and extraction blanks) and sequencing (library blanks); (ii) quantification of biomass (e.g., qPCR or picogreen) to inform downstream filtering and to detect extremely low‐biomass samples; (iii) use of positive controls or mock communities and dilution series to determine limit of detection and to benchmark kit‐to‐kit variability; (iv) randomised processing and explicit tracking of extraction batch, operator and kit lot; (v) orthogonal validation where possible (targeted qPCR, culture, or FISH/imaging) to corroborate sequence‐based findings; and (vi) contamination‐aware bioinformatic workflows (e.g., statistical contaminant identification such as decontam or read‐subtraction approaches such as microDecon) together with transparent reporting of all blanks and filtering rules. These practices (summarised in the RIDE/RID[E] guidance framework) substantially reduce false positives and support reliable companion‐diagnostic calls for patient stratification in trials (Davis et al.  2018 ; Eisenhofer et al.  2019 ; Salter et al.  2014 ; Adam et al.  2019 ). We recommend a three‐stage clinical pathway: (1) early human mechanistic and safety studies focused on engraftment and intermediate biomarkers; (2) phase II randomised, diagnostic‐stratified trials using microbiome conversion and mechanistic endpoints as co‐primary outcomes; and (3) phase III trials powered for live birth only after dosing, population selection and endpoints have been refined. Adaptive designs with prespecified subgroups will improve efficiency and reduce the risk of false negatives (Armstrong et al.  2022 ). Also we recommend that Phase I–IIa studies prioritise safety, manufacturing quality and colonisation efficiency, and incorporate contamination‐aware diagnostic workflows and mechanistic mucosal endpoints (quantified colonisation by qPCR and shotgun/16S with blanks, local cytokines, TLR signalling and epithelial barrier markers) in matched endometrial or cervicovaginal samples to ensure that observed changes reflect true biological engraftment rather than technical artefact (Pot and Vandenplas  2021 ). Small, randomised, placebo controlled phase I designs in healthy volunteers or symptom free women with documented dysbiosis can quantify local tolerability and the kinetics of engraftment; translational immune readouts generate mechanistic proof of principle prior to efficacy testing (Chetty et al.  2025 ; Tseng et al.  2025 ). For efficacy, regulatory and clinical consensus favours live birth or cumulative live birth per woman as the primary outcome in infertility trials because live birth integrates embryologic, endometrial and clinical care factors better than intermediate outcomes. Randomised phase IIb and phase III trials should therefore enrol well phenotyped infertile populations, for example patients with recurrent implantation failure whose endometrial microbiome meets pre specified dysbiosis criteria, and should be powered to detect clinically meaningful differences in live birth while prespecifying secondary mechanistic outcomes such as microbiome conversion rate, implantational biomarkers and adverse pregnancy events (Harbin Consensus Conference Workshop Group et al.  2014 ; Feng et al.  2025 ). Adaptive designs that stratify by microbiome endotype and that include colonisation and functional microbiome endpoints will accelerate go no go decisions and identify subpopulations with the highest benefit to risk ratio (Pot and Vandenplas  2021 ; Bloom and Chung  2025 ). To help readers interpret the literature and to guide new trials, we contrast key features of robust versus problematic study designs in this field: Predefined companion‐diagnostic criteria and validated low‐biomass pipelines; documented negative and positive controls; reporting of extraction and sequencing blanks (Eisenhofer et al.  2019 ; Salter et al.  2014 ). Measurement of biomass (qPCR) and use of orthogonal validation for key taxa (Eisenhofer et al.  2019 ). Randomised, blinded allocation (when ethical/feasible) with pre‐specified microbiome and immunologic mechanistic endpoints (e.g., colonisation/engraftment rate, local cytokine shifts) as well as prespecified clinical endpoints. Power calculations based on realistic effect sizes for both microbiome conversion and clinical outcomes; prespecified stratified analyses by microbiome endotype (Wong  2013 ). Preplanned bioinformatic contaminant‐filtering strategy (e.g., use of decontam and documented blank‐based subtraction) and full reporting of filtering thresholds (Davis et al.  2018 ; Adam et al.  2019 ). Predefined companion‐diagnostic criteria and validated low‐biomass pipelines; documented negative and positive controls; reporting of extraction and sequencing blanks (Eisenhofer et al.  2019 ; Salter et al.  2014 ). Measurement of biomass (qPCR) and use of orthogonal validation for key taxa (Eisenhofer et al.  2019 ). Randomised, blinded allocation (when ethical/feasible) with pre‐specified microbiome and immunologic mechanistic endpoints (e.g., colonisation/engraftment rate, local cytokine shifts) as well as prespecified clinical endpoints. Power calculations based on realistic effect sizes for both microbiome conversion and clinical outcomes; prespecified stratified analyses by microbiome endotype (Wong  2013 ). Preplanned bioinformatic contaminant‐filtering strategy (e.g., use of decontam and documented blank‐based subtraction) and full reporting of filtering thresholds (Davis et al.  2018 ; Adam et al.  2019 ). Absence of negative/extraction controls or failure to report them; no measurement of sample biomass (Salter et al.  2014 ). Small, underpowered ‘pilot’ studies that report clinical outcomes without reporting engraftment, contamination controls or orthogonal validation (Wong  2013 ). Retrospective ad‐hoc filtering of taxa without prespecified rules, or failure to account for kit/reagent batch effects (Davis et al.  2018 ; Adam et al.  2019 ). Lack of randomization, lack of blinding where feasible and selective reporting of only ‘positive’ taxa (Ioannidis  2005 ). Absence of negative/extraction controls or failure to report them; no measurement of sample biomass (Salter et al.  2014 ). Small, underpowered ‘pilot’ studies that report clinical outcomes without reporting engraftment, contamination controls or orthogonal validation (Wong  2013 ). Retrospective ad‐hoc filtering of taxa without prespecified rules, or failure to account for kit/reagent batch effects (Davis et al.  2018 ; Adam et al.  2019 ). Lack of randomization, lack of blinding where feasible and selective reporting of only ‘positive’ taxa (Ioannidis  2005 ). By explicitly adopting the robust features above and by clearly documenting any unavoidable limitations, future trials will be easier to interpret and will better meet regulatory expectations for companion diagnostics and clinical claims. Regulatory expectations vary substantially by therapeutic modality, and a modality‐specific strategy is therefore essential for reproductive microbiome products. In the United States, products intended to prevent, treat or cure disease that contain live microorganisms are typically regulated as live biotherapeutic products (LBPs) and fall under IND/BLA pathways with chemistry, manufacturing and control (CMC) expectations that reflect their biological complexity; the FDA's guidance on Early Clinical Trials With Live Biotherapeutic Products summarises these CMC and safety expectations for IND submissions (FDA  2016 ). Products that use genetically modified bacterial chassis (engineered to secrete cytokines, antimicrobials or sensors) generally face the most stringent oversight: regulators treat them as biologics and (in many jurisdictions) as genetically‐modified organisms (GMOs), requiring both standard medicinal product dossiers (identity, potency, sterility, stability, reproductive toxicology) and environmental risk assessment/GMO authorization processes in Europe. Developers should expect requirements for additional containment/biocontainment evidence (e.g., auxotrophy, kill‐switch validation), detailed characterisation of genetic constructs (stability, absence of mobilizable elements) and expanded nonclinical packages addressing off‐target exposure and reproductive safety. For EU trials this typically invokes EMA GMO/ERA guidance; for the US, early engagement with CBER/OTR (and IND‐level CMC meetings) is recommended (FDA  2016 ; European Medicines Agency  2020 ). Multistrain, non‐genetically modified live consortia that are developed with medical claims (not as dietary supplements) are usually handled as LBPs/drugs rather than foods. Sponsors must provide strain identity (whole‐genome characterisation), potency assays, absence of transferable AMR and validated release criteria; prior trial examples such as LACTIN‐V ( Lactobacillus crispatus CTV‐05) illustrate an IND pathway and the kinds of clinical safety and efficacy endpoints regulators expect in phase II trials (Cohen Craig et al.  2020 ; FDA  2016 ). ‘Postbiotics’ (inactivated microbes or their components) occupy a more variable regulatory space: when labelled/marketed as foods or dietary supplements with general health claims they may be regulated under food law or supplements rules, but if they are intended to treat or prevent disease in a reproductive‐medicine context they will usually be regulated as drugs/biologics and require appropriate preclinical safety and clinical evidence. The ISAPP consensus definition and recent regulatory reviews highlight that regulators will focus on the precise composition (inactivated cells vs. purified metabolites), safety (toxicity, impurities) and the intended claims to determine the pathway. For reproductive uses (e.g., adjunct to embryo transfer) plan drug‐level programs including reproductive toxicology and dose‐finding PK/PD or pharmacodynamic marker studies (Amobonye et al.  2025 ; Salminen et al.  2021 ). Whole‐community transfers (e.g., vaginal microbiota transplantation) have attracted specific regulatory attention because of donor‐derived risks; the FDA and other agencies have issued enforcement policies and donor‐screening expectations for FMT and have emphasised the need for INDs when the product is intended for disease treatment rather than research. For reproductive niche transplantation, rigorous donor screening, long‐term follow‐up and explicit plans for pregnancy and neonatal surveillance will be essential (STI CTG  2016 ; FDA  2022 ). Bacteriophage therapeutics (whole phages or lysins) are regulated as biological medicinal products and the regulatory framework is evolving rapidly; FDA workshops and EMA/Ph.Eur. activities provide emerging quality/CMC expectations (potency, host‐range characterisation, manufacturing host banks, GMP) and often require bespoke assay development for phage potency and resistance monitoring. Sponsors should expect iterative discussions with regulators to define acceptable potency assays and manufacturing controls (Fuerst‐Wilmes et al.  2025 ; FDA  2021 ). For all microbiome modalities, a regulator‐facing dossier should clearly define: (i) product taxonomy and strain/genetic characterisation (WGS and mobile element analysis); (ii) potency assays that reflect mechanism (engraftment metrics, metabolite output or functional activity); (iii) absence/mitigation of transferable AMR; (iv) reproductive and developmental toxicology plan (embryo/gamete compatibility studies where applicable); (v) manufacturing controls including adventitious agent testing and validated low‐biomass contamination controls; and (vi) for GM or transplant products, environmental risk assessment/donor screening and post‐marketing surveillance plans. Early pre‐IND/Scientific Advice meetings will materially reduce uncertainty and help define whether a product is reviewed as an LBP/biologic, a GMO, a drug, or a food/supplement in a specific jurisdiction (FDA  2016 , 2022 ; European Medicines Agency  2020 ).

Conception and design of the research by Huifang Cong, Shumin Liu, acquisition of data by Limiao Li, Lin Feng, Qiang Li, analysis and interpretation of data by: Yue Zhou, Sifan Liu, Limiao Li, Qiang Li, obtaining funding by Heilongjiang Province Postdoctoral Fund (Grant Number: LBH‐ Z24283 ), drafting the manuscript by Limiao Li, Qiang Li, Huimin Song, Shengnan Kang, Huike Chen, revision of manuscript for important intellectual content by Shumin Liu and Huifang Cong.

The authors have nothing to report.

Over the past decade, converging cohort studies and mechanistic experiments have elevated the reproductive microbiome from a descriptive curiosity to a mechanistic and translational priority by linking specific community states with implantation competence, pregnancy outcomes and mucosal immune tone (Moreno et al.  2016 ; Bui et al.  2023 ). Translational successes in related mucosal fields, including randomised evidence that a vaginal live biotherapeutic can reduce recurrence of bacterial vaginosis and more recent reports of successful vaginal microbiota transplantation in refractory cases, provide a proof of principle that directed microbial interventions can durably reconfigure mucosal ecology and downstream host biology (Cohen et al.  2020 ; Wrønding et al.  2023 ; Li et al.  2025 ). Together these observations motivate a focused pipeline to move next generation probiotics and precision microbial therapeutics into infertility care, while recognising that efficacy for reproductive endpoints remains to be proven in large, well controlled trials (Armstrong et al.  2022 ; Bui et al.  2023 ). To realise clinical impact, research must prioritise mechanistic fidelity and patient stratification so that interventions address the biological drivers of infertility rather than an incompletely characterised microbial signal. Advanced human tissue models and organ‐on‐chip systems that recreate endometrial architecture, hormone dynamics and perfusion permit causal interrogation of how candidate strains and metabolites alter epithelial receptivity, decidualization and local immunity, and these platforms are now sufficiently mature to be used for preclinical candidate selection (Dai et al.  2025 ; Ciprietti et al.  2025 ). Complementary humanised in vivo approaches and careful correlative studies in well phenotyped cohorts will be required to understand colonisation barriers, ecological resistance and the kinetics of microbial engraftment that determine whether a strain can functionally reshape the endometrial niche (Armstrong et al.  2024 ; Yang, Sha, et al.  2025 ). A mechanistically driven development path will also exploit metabolite and immune biomarkers as intermediate readouts to accelerate go no go decisions. Recent work implicating lactate signalling pathways at the maternal fetal interface and the protective effects of short chain fatty acids such as butyrate on epithelial integrity suggest that measurement of metabolite profiles and host receptor engagement can serve as proximal pharmacodynamic markers after microbial intervention (Gou et al.  2025 ; Zhu et al.  2025 ). Incorporating these biomarkers into early phase trials will provide biological proof of mechanism even when clinical endpoints require larger, longer studies, and will enable rational selection of strains, dose regimens and formulation strategies that maximise both colonisation and functional impact (Armstrong et al.  2024 ; Gou et al.  2025 ). Rigorous diagnostic integration will be essential for precision deployment. High quality, low biomass metagenomic profiling coupled with standardised sampling and contamination control can convert microbial descriptions into patient endotypes that predict response to specific live biotherapeutics, and companion diagnostic frameworks should be built into trial inclusion criteria so that studies are powered to detect treatment effects in the most relevant subgroups (Bui et al.  2023 ; Kumar et al.  2025 ). Standardisation efforts and consensus on reporting will reduce heterogeneity across studies, enable meta analyses that identify robust responder phenotypes and accelerate clinical translation of strain level therapeutic claims (Bui et al.  2023 ; Ma et al.  2024 ). Clinical development must balance the unique safety considerations of reproductive indications with the operational realities of infertility research. Early phase LBP studies should emphasize product identity, potency, and colonization kinetics alongside rigorous assessment of local immune responses and reproductive toxicology, and later phase randomised controlled trials should use cumulative live birth per woman as a primary endpoint to provide regulators and clinicians with clinically meaningful evidence (Cordaillat‐Simmons et al.  2020 ). Adaptive and stratified designs that prespecify microbiome conversion and mechanistic secondary outcomes will increase efficiency, clarify mechanisms of action and identify subpopulations with the highest likelihood of benefit (Ma et al.  2024 ; Microbiome Therapeutics Innovation Group and Barberio  2024 ). Navigating the regulatory and ethical landscape will require early dialogue with agencies and multidisciplinary consortia to define acceptable evidence for quality, safety and efficacy. Existing FDA guidance for live biotherapeutic products and evolving European regulatory thinking about microbiome interventions provide a framework but also highlight the need for specific reproductive safety packages and post marketing surveillance plans that address pregnancy exposure and potential vertical transmission (Rodriguez et al.  2025 ). Ethical stewardship is equally important because interventions that alter the maternal microbiome may have implications for offspring, and transparent data sharing, donor screening standards for any transplant approaches and robust informed consent processes will be necessary to maintain public trust (Wrønding et al.  2023 ; Maor et al.  2025 ). Looking forward, the field is poised to deliver transformative tools for infertility care if three conditions are met: investment in mechanistic human models and biomarker discovery to narrow candidate selection, integration of diagnostic‐driven patient stratification into clinical development, and early regulatory engagement to align quality and safety expectations (Bui et al.  2023 ; Ciprietti et al.  2025 ). If those conditions are realised, precision microbial therapeutics could shift infertility management from empiric treatments toward interventions that repair mucosal ecology, restore immune and barrier homeostasis, and thereby improve implantation and live birth outcomes for well defined patient subgroups (Armstrong et al.  2022 ; Bui et al.  2023 ). In summary, the reproductive microbiome constitutes both a window into the biology of human fertility and a promising therapeutic target. The coming years should focus on translating mechanistic insights into robust clinical strategies using rigorous models, companion diagnostics and fit for purpose clinical trials so that next generation probiotics become an evidence based option for couples confronting infertility (Moreno et al.  2016 ; Ma et al.  2024 ).

This work was supported by Heilongjiang Province Postdoctoral Fund, LBH‐Z24283.

Engineered live biotherapeutics bring powerful capabilities but also unique reproductive safety and regulatory challenges; as a pragmatic translational hierarchy, postbiotic approaches offer a lower‐risk route to test mechanistic hypotheses in humans, while engineered strains should proceed only after comprehensive preclinical reproductive safety evaluation (Cohen Craig et al.  2020 ; Ma, Lyu, et al.  2022 ; Steidler et al.  2003 ). Recent translational work in other mucosal tissues establishes proof of principle for this strategy: Lactococcus lactis strains engineered to secrete interleukin 10 achieved biological containment and measurable anti‐inflammatory effects in preclinical colitis models and advanced to first‐in‐human phase I testing, demonstrating that live bacterial delivery of cytokines can be both efficacious and clinically tolerable (Braat et al.  2006 ; Steidler et al.  2000 ; He et al.  2019 ). The design principles that enable safe and effective reproductive tract applications are therefore already defined and include choice of a generally recognized as safe chassis, genetic circuits for inducible expression, and multiple biological containment layers such as auxotrophy or kill switches (Ma, Lyu, et al.  2022 ; Steidler et al.  2003 ). Translating this approach to infertility focuses on two complementary functional aims: precise suppression of pathobiont‐driven inflammation and restoration of immunotolerance required for implantation (Hijová  2024 ; Noguès et al.  2022 ). In concept, Lactobacillus or other vaginally adapted species can be engineered to detect pathobiont‐associated molecular patterns or biofilm metabolites and, in response, secrete anti‐inflammatory cytokines such as interleukin 10 or transforming growth factor beta to locally reprogram immune responses without systemic exposure (Steidler et al.  2000 ; Sadhu et al.  2025 ). Preclinical work has already shown that delivery of host defense peptides and peptide precursors by engineered lactic acid bacteria can reduce mucosal inflammation and modulate cytokine profiles, providing a mechanistic basis for using engineered probiotics to bias the uterine immune milieu toward tolerance (Noguès et al.  2022 ; Zeng et al.  2020 ). A parallel, pathogen‐focused engineering strategy is to arm probiotics with antimicrobial peptides or bacteriocins that are expressed only in the presence of pathobionts, thereby combining targeted killing with preservation of beneficial Lactobacillus species; such conditional secretion circuits reduce selection pressure for broad antimicrobial resistance (Garvey  2023 ; Yang, Ma, et al.  2025 ). Finally, because reproductive medicine demands high safety margins, engineered strains for infertility will need rigorous genomic stability testing, containment validation and demonstration of noninterference with sperm, embryo or endometrial function prior to clinical translation (Ma, Lyu, et al.  2022 ; Steidler et al.  2003 ). Postbiotics are defined as preparations of inactivated microbial cells, cell components, or metabolites that deliver health benefits without live organisms, and this modality addresses several safety and regulatory constraints that limit live therapeutics (Hijová  2024 ; Kumar et al.  2024 ; Pei et al.  2024 ). Because they are nonviable, postbiotics offer enhanced stability, reduced risk of horizontal gene transfer, and more straightforward quality control and pharmacokinetic characterisation, which simplifies clinical development for sensitive indications such as assisted reproduction (Hijová  2024 ; Kumar et al.  2024 ). The repertoire of postbiotic modalities relevant to the reproductive tract includes killed bacterial lysates that retain immunomodulatory surface ligands, cell‐free supernatants rich in short‐chain fatty acids and lactic acid isomers, and purified microbial metabolites or peptides with defined molecular targets (Kumar et al.  2024 , 2023 ). Notably, specific microbial metabolites exert stereoisomer‐specific effects in mucosal biology; the D isomer of lactic acid produced by many Lactobacillus strains has been shown to downregulate matrix metalloproteinase activity and to modulate epithelial and immune cell responses in the female reproductive tract, suggesting a concrete molecular candidate for a postbiotic approach that enhances implantation conditions (Grewal et al.  2021 ). Preclinical and early clinical data indicate that postbiotic preparations can induce defensin expression, attenuate proinflammatory cytokine release and promote barrier integrity across mucosal surfaces, which together provide a rationale for testing vaginal or intrauterine postbiotics as adjuncts to embryo transfer (Kumar et al.  2024 ; Nguyen et al.  2023 ). From a translational perspective, manufacturing postbiotics allows precise dosing of active molecules, batch‐to‐batch reproducibility and straightforward stability testing under the cold chain constraints of reproductive medicine, which may accelerate regulatory approval compared with live engineered strains (Hijová  2024 ; Kumar et al.  2024 ). Finally, combining postbiotics with narrowly targeted antimicrobials or phage‐derived enzymes may permit a two‐step therapy in which deleterious biofilms are first disrupted and then the mucosa is conditioned with defined immunomodulatory metabolites to restore receptivity (Hijová  2024 ; Tisakova et al.  2025 ). Bacteriophage based therapeutics provide species specific bactericidal activity and biofilm disruption potential that is particularly attractive for reproductive tract pathobionts such as Gardnerella and Prevotella because phages can spare beneficial lactobacilli (Fujiki and Schnabl  2023 ; Gliźniewicz et al.  2024 ). Recent laboratory studies demonstrate that phage derived enzymes and lytic phages can selectively reduce Gardnerella burden and dismantle biofilms ex vivo, establishing a translational path for phage or endolysin based interventions in bacterial vaginosis associated infertility states (Tisakova et al.  2025 ). However, several technical and biological challenges must be addressed before routine clinical use in infertility: phage host range can be narrow requiring cocktails or engineered broad host range variants, bacterial resistance to phage via receptor mutation or CRISPR mechanisms can emerge, and complex polymicrobial biofilms may shelter target bacteria from phage access (Fujiki and Schnabl  2023 ; Eghbalpoor et al.  2024 ). Formulation is a critical translational variable because phages intended for vaginal or intrauterine delivery must remain viable through manufacturing and administration and must be compatible with existing reproductive procedures such as embryo transfer; encapsulation, mucoadhesive gels and co administration with biofilm degrading enzymes are active areas of formulation research (Gliźniewicz et al.  2024 ; Eghbalpoor et al.  2024 ). Importantly, phage derived lysins and engineered lytic enzymes may offer advantages over whole phages in reproductive medicine because they act rapidly to degrade peptidoglycan or biofilm matrix without requiring bacterial replication, which simplifies dosing and regulatory evaluation (Tisakova et al.  2025 ; Eghbalpoor et al.  2024 ). Finally, the clinical translation of phage therapeutics for infertility will require demonstration of safety for gametes and embryos, standardised assays of phage purity and potency and early phase trials that couple microbiological end points with implantation and live birth outcomes to prove both mechanistic activity and clinical benefit (Fujiki and Schnabl  2023 ; Niazi  2025 ). Table  3 contrasts precision microbial modalities, their opportunities and translational hurdles and lists recommended proximal endpoints for early‐phase testing, while Figure  3 provides a schematic representation of these major therapeutic strategies. Precision microbial therapeutics: Modalities, unique advantages, major translational challenges and recommended early endpoints. Schematic representation of precision microbial therapeutics for reproductive medicine. The diagram illustrates three major therapeutic strategies: engineered live biotherapeutics, postbiotics derived from non‐viable microbial products, and phage‐based approaches. These modalities encompass genetic engineering tools such as plasmids and CRISPR, immunomodulatory effects on host cells, microbiota‐derived metabolites like SCFAs, cell‐free supernatants and targeted bacteriophage or endolysin mechanisms that disrupt pathogens and biofilms. Together, these interventions highlight emerging avenues for modulating the reproductive tract microbiome to improve clinical outcomes. Clinical heterogeneity and ecological endotypes (low‐ Lactobacillus , L. iners ‐predominant, anaerobe/pathobiont‐dominant, chronic endometritis) demand a structured, reproducible approach for choosing between therapeutic modalities (single‐strain NGP, synthetic consortium, engineered live biotherapeutic, postbiotic, phage/endolysin or VMT). Below we propose a pragmatic, diagnostic‐driven decision framework intended for trial design and translational use. The framework integrates: (A) microbial endotype (species/strain and functional markers, such as lactic acid stereoisomers, pH, biofilm markers), (B) clinical urgency and context (e.g., immediate embryo transfer vs. preconception optimization), (C) need for durable engraftment and (D) safety constraints (pregnancy exposure, embryo/gamete safety, risk of horizontal gene transfer). Use a validated low‐biomass pipeline and companion diagnostic thresholds to classify patients into endotypes ( L. crispatus ‐deficient but low inflammation; transitional L. iners ‐dominant; anaerobe/biofilm‐dominant; biopsy‐proven chronic endometritis). Diagnostic gating is required because the choice of modality depends on whether the problem is primarily replacement (lack of protective lactobacilli), eradication (biofilm/pathobiont dominance) or immune‐mediated inflammation (CE) (Abouelela and Helmy  2024 ; Vinderola et al.  2022 ). If immediate embryo transfer is planned or pregnancy is ongoing → prioritise non‐viable modalities (postbiotics, endolysin) or highly characterised, safety‐tested approaches with minimal theoretical embryo exposure; avoid deploying novel engineered live strains during embryo transfer unless reproductive toxicology is fully complete (Vinderola et al.  2022 ; Gliźniewicz et al.  2023 ). If preconception optimization is possible (time available) → consider modalities that require time to engraft or remodel ecology (synthetic consortia, engineered live biotherapeutics or staged consortia + preconditioning). These are appropriate when durable conversion is the goal (Abouelela and Helmy  2024 ; Cruz et al.  2022 ). If immediate embryo transfer is planned or pregnancy is ongoing → prioritise non‐viable modalities (postbiotics, endolysin) or highly characterised, safety‐tested approaches with minimal theoretical embryo exposure; avoid deploying novel engineered live strains during embryo transfer unless reproductive toxicology is fully complete (Vinderola et al.  2022 ; Gliźniewicz et al.  2023 ). If preconception optimization is possible (time available) → consider modalities that require time to engraft or remodel ecology (synthetic consortia, engineered live biotherapeutics or staged consortia + preconditioning). These are appropriate when durable conversion is the goal (Abouelela and Helmy  2024 ; Cruz et al.  2022 ). L. crispatus ‐deficient, low inflammation, no biofilm → single‐strain high‐fitness NGP ( L. crispatus ) or small defined consortium ; aim is functional replacement and acidification; use intravaginal delivery + companion diagnostic to confirm engraftment (Abouelela and Helmy  2024 ). Transitional ( L. iners ‐dominant) or recurrent instability → defined synthetic consortium to provide redundancy and metabolic cooperation (greater resilience than single strain). Consider a staged regimen (antibiotic or biofilm disruptor → consortium) (Jiang et al.  2026 ). Anaerobe/biofilm‐dominant states (Gardnerella/Prevotella/Atopobium) or chronic endometritis → targeted eradication first (phage cocktails/endolysins or narrow‐spectrum antimicrobials combined with biofilm‐degrading enzymes), followed by mucosal conditioning with a postbiotic and then recolonization with a consortium or engineer/NGP as needed. Endolysins hold promise for Gardnerella biofilms ex vivo and are attractive as a targeted preconditioning step (Tisakova et al.  2025 ; Gliźniewicz et al.  2023 ). High safety constraints (immediate embryo transfer/pregnancy/neonatal safety critical)  →  postbiotic formulations (defined metabolites, D‐lactic acid, SCFAs in controlled dose windows) or purified endolysins because they avoid live organism exposure and simplify pharmacology (Vinderola et al.  2022 ). Refractory, recurrent or complex polymicrobial dysbiosis after standard therapies → Vaginal microbiota transplantation (VMT) can be considered in tightly controlled research settings with rigorous donor screening and long‐term follow‐up; reserve for refractory cases where donor engraftment is the most plausible path to durable correction (Tisakova et al.  2025 ; Abouelela and Helmy  2024 ). L. crispatus ‐deficient, low inflammation, no biofilm → single‐strain high‐fitness NGP ( L. crispatus ) or small defined consortium ; aim is functional replacement and acidification; use intravaginal delivery + companion diagnostic to confirm engraftment (Abouelela and Helmy  2024 ). Transitional ( L. iners ‐dominant) or recurrent instability → defined synthetic consortium to provide redundancy and metabolic cooperation (greater resilience than single strain). Consider a staged regimen (antibiotic or biofilm disruptor → consortium) (Jiang et al.  2026 ). Anaerobe/biofilm‐dominant states (Gardnerella/Prevotella/Atopobium) or chronic endometritis → targeted eradication first (phage cocktails/endolysins or narrow‐spectrum antimicrobials combined with biofilm‐degrading enzymes), followed by mucosal conditioning with a postbiotic and then recolonization with a consortium or engineer/NGP as needed. Endolysins hold promise for Gardnerella biofilms ex vivo and are attractive as a targeted preconditioning step (Tisakova et al.  2025 ; Gliźniewicz et al.  2023 ). High safety constraints (immediate embryo transfer/pregnancy/neonatal safety critical)  →  postbiotic formulations (defined metabolites, D‐lactic acid, SCFAs in controlled dose windows) or purified endolysins because they avoid live organism exposure and simplify pharmacology (Vinderola et al.  2022 ). Refractory, recurrent or complex polymicrobial dysbiosis after standard therapies → Vaginal microbiota transplantation (VMT) can be considered in tightly controlled research settings with rigorous donor screening and long‐term follow‐up; reserve for refractory cases where donor engraftment is the most plausible path to durable correction (Tisakova et al.  2025 ; Abouelela and Helmy  2024 ). For early‐phase trials choose proximal, mechanistic endpoints (engraftment/lactic acid production/biofilm biomass reduction/local IL‐10 rise and IL‐6/IL‐1β reduction/epithelial junction markers such as ZO‐1) and prespecify early stopping rules for no engraftment or adverse mucosal reactions. Use adaptive randomization or enrichment by endotype to improve power (Abouelela and Helmy  2024 ).

The authors have nothing to report.

Lactic acid and other microbial metabolites likely modulate barrier integrity and immune tone at the maternal–fetal interface, yet translational readiness requires defining effective local concentrations, stereoisomeric activity and safe delivery strategies so that mechanistic insights can be converted into reproducible therapies (Moreno et al.  2016 ; Delgado‐Diaz et al.  2022 ; Ma et al.  2020 ). Evidence from mechanistic studies shows that trophoblasts both produce and export lactate and that lactate flux influences trophoblast differentiation and invasiveness; inhibition of lactate transport or metabolism impairs decidualization and reduces trophoblast motility in vitro (Gao et al.  2022 ; Xu et al.  2021 ). At least part of this signalling is mediated by the G protein‐coupled receptor HCAR1, also known as GPR81, which senses physiologic lactate concentrations and modulates intracellular signalling pathways that control inflammation, cell migration and survival; uterine expression of GPR81 increases during gestation and is functionally relevant to uterine immune responses (Ma et al.  2020 ; Madaan et al.  2017 ). Mechanistic dissection using primary decidual macrophages and trophoblast models demonstrates that lactate skews macrophage differentiation toward an anti‐inflammatory, tissue‐remodelling phenotype that supports trophoblast invasion and the establishment of immune tolerance, thereby linking microbial and trophoblast metabolism to local immunoregulation required for implantation (Gao et al.  2022 ; Xu et al.  2021 ). Clinically, higher lactate levels both in the reproductive tract and systemically correlate with markers of implantation competence and successful early pregnancy in observational cohorts, suggesting that perturbations of lactate production or signalling may contribute to implantation failure and early pregnancy loss (Abbasi Ranjbar et al.  2023 ; Gurner and Gardner  2025 ). Together, these data recast lactic acid as a bona fide signalling metabolite in reproduction: it enforces a regulatory, low‐inflammation niche that promotes trophoblast invasion while simultaneously enhancing epithelial barrier function and antimicrobial defence in the cervicovaginal milieu (Delgado‐Diaz et al.  2022 ; Ma et al.  2020 ). Short‐chain fatty acids, particularly butyrate, produced by gut microbes and potentially by translocated or local anaerobic taxa, act at distant mucosal sites through circulation and locally via paracrine mechanisms to influence epithelial cell biology and immune tone in the female reproductive tract (Chadchan et al.  2021 ; Liu, Peng, et al.  2024 ). Experimental models show that butyrate enhances epithelial barrier assembly by promoting the transcriptional upregulation and correct assembly of tight junction components, increasing transepithelial resistance in diverse epithelial lines and primary tissues; these effects are mediated by several complementary mechanisms including histone deacetylase inhibition, activation of AMPK and HIF‐1α stabilization, and signaling through hydroxycarboxylic acid receptor 2 (McArthur  2023 ; Peng et al.  2009 ). In reproductive tissues, preclinical studies indicate that butyrate reduces pathological epithelial remodeling and suppresses inflammatory programs implicated in endometriosis and other disorders linked to infertility, in part by restoring epithelial integrity and by potent epigenetic reprogramming of stromal and immune cell transcriptional networks (Chadchan et al.  2021 ; Kim and Yang  2024 ). Importantly, while moderate concentrations of butyrate strengthen tight junction expression and barrier function, several human tissue studies caution that supraphysiologic doses may provoke epithelial stress or apoptosis and transiently disrupt barrier proteins, highlighting a dose dependent window for therapeutic exploitation (Pérez‐Reytor et al.  2021 ; Tabat et al.  2020 ). Collectively, these mechanistic and translational data imply that targeted modulation of SCFA production or delivery could fortify endometrial epithelial integrity and simultaneously engage anti‐inflammatory pathways supportive of implantation and early placentation (Liu, Peng, et al.  2024 ; Zhang et al.  2023 ). The reproductive tract epithelium and resident immune cells express an array of pattern recognition receptors including Toll‐like receptors that continuously sample microbial and metabolite cues; commensal organisms and their molecular products sculpt basal TLR expression and downstream signalling thresholds, thereby setting the responsiveness to pathogens versus tolerance to conceptus antigens (Gryaznova et al.  2024 ; Yang et al.  2022 ). Molecular and transcriptional profiling studies show correlations between community composition and epithelial TLR and NOD‐like receptor expression across the cervix and endometrium, and mechanistic experiments indicate that Lactobacillus cell surface structures and secreted metabolites attenuate proinflammatory TLR signalling while favouring regulatory cascades (Decout et al.  2024 ; Vinitha et al.  2025 ). This microbiome‐dependent priming results in a more restrained innate response to microbial ligands and a bias toward tolerogenic cytokine profiles that promote regulatory T cell and M2 macrophage phenotypes essential for implantation (Gao et al.  2022 ; Xu et al.  2021 ). Pathologic shifts toward dysbiosis increase exposure to potent TLR agonists, such as lipopolysaccharide and peptidoglycan fragments, provoking exaggerated TLR activation that impairs decidual receptivity, disrupts vascular remodelling and correlates with adverse reproductive outcomes including implantation failure and preterm birth (Fasoulakis et al.  2025 ; Zou et al.  2022 ). Therefore, mechanistic modulation of microbiome–TLR interactions represents a tractable axis for precision microbial therapeutics aimed at restoring a receptive, low‐inflammation milieu (Gryaznova et al.  2024 ; Valeriano et al.  2024 ). A central, indirect route by which the microbiome governs fertility is via preservation of mucosal barrier integrity through regulation of tight junction proteins. Studies in human cervicovaginal epithelial cells and in vivo models demonstrate that lactic acid and lactobacilli enhance expression and localization of ZO‐1 and occludin and reduce paracellular permeability, thereby limiting pathogen translocation and inflammatory exposure of the endometrium (Delgado‐Diaz et al.  2022 ; Schwecht et al.  2023 ). Parallel work with SCFAs, and butyrate in particular, shows upregulation and reorganisation of multiple tight junction components including claudins, occludin and ZO‐1 via transcriptional activation and posttranslational assembly pathways; these effects have been validated across intestinal and reproductive epithelial models, linking microbial metabolites with the physical competence of mucosal barriers (Peng et al.  2009 ; Pérez‐Reytor et al.  2021 ). The clinical relevance of barrier modulation is underscored by findings that dysbiotic communities and pathogen‐associated molecular patterns downregulate ZO‐1 and occludin and associate with epithelial disruption, inflammatory cytokine release and impaired implantation (Zou et al.  2022 ; Huang et al.  2024 ). Taken together, these mechanistic layers, metabolic signalling through lactate and SCFAs, microbiome‐tuned TLR responsiveness, and reinforcement of epithelial junctions, form an integrated circuit by which commensal microbes sustain a receptive uterine environment, as summarised in Figure  2 . Interventions that precisely restore metabolite profiles or deliver defined probiotic strains that reinforce these pathways therefore hold promise as next‐generation therapeutics to rescue barrier and immune homeostasis in infertility (Gao et al.  2022 ; McArthur  2023 ). The gut and lower genital tract microbiota influence reproductive success through four major mechanisms. Lactic acid produced by Lactobacillus and trophoblasts activates GPR81 and promotes anti‐inflammatory M2 macrophage polarization. Short‐chain fatty acids (SCFAs), particularly butyrate, modulate epithelial and immune function through HDAC inhibition, AMPK activation, and HIF‐1α stabilization. Pattern recognition receptor (PRR) signaling induced by commensal microbes regulates TLR‐mediated immune responses and supports regulatory immune cell phenotypes. Regulation of tight junctions induced by SCFAs and lactate enhances epithelial barrier integrity. Together, these pathways protect against dysbiosis‐related infertility, enhance immune tolerance, strengthen epithelial barriers and ultimately enhance implantation. A useful way to present microbiota → infertility mechanisms is to follow the immune cascade from innate sensing to adaptive polarization and to name the key downstream molecular effectors. Below we summarise the sequence and the evidence linking each step to implantation failure. Dysbiotic reproductive communities (especially Gram‐negative and biofilm‐forming taxa) expose the epithelium and resident macrophages/dendritic cells to high levels of LPS and other PAMPs, which activate TLRs and canonical NF‐κB signaling. Increased NF‐κB activity has been documented in pathological endometrium and is associated with thin or non‐receptive endometrium and reduced live‐birth rates in RIF cohorts (Kalı et al.  2025 ; Blazheva et al.  2024 ). NF‐κB drives transcription of IL‐1β, IL‐6, TNF‐α and chemokines that recruit and activate neutrophils and monocytes, and this inflammatory milieu perturbs decidualization and vascular remodelling. IL‐6 in particular engages STAT3 signalling in stromal and epithelial cells and acts as a bridge to adaptive T‐cell differentiation (Blazheva et al.  2024 ; Li et al.  2024 ; Zhou et al.  2021 ). High IL‐6 and IL‐1β concentrations favour STAT3‐dependent induction of Th17 cells (RORγt‐driven) while simultaneously inhibiting Treg induction/maintenance (FOXP3), shifting the local Th17/Treg ratio toward inflammation; several human studies show elevated Th17/Treg ratios in RIF and chronic endometritis samples. At the same time, a Th1 bias (elevated IFN‐γ and TNF‐α) has been repeatedly linked to implantation failure in other cohorts; therefore, the dominant immune pattern can be context dependent (Th17 predominance is common in CE and some RIF cohorts; Th1 predominance is reported in other infertility/early‐loss cohorts). Clinically, patients with increased Th17/Treg ratios show higher IL‐6/IL‐17 expression and worse implantation outcomes (Berdiaki et al.  2024 ; Garmendia et al.  2025 ). NF‐κB output amplifies and synergizes with STAT3 and MAPK pathways; IL‐6/STAT3 signalling can reinforce pro‐inflammatory transcription and cell survival programs that favour Th17 maintenance. NLRP3 inflammasome activation (downstream of TLR/NF‐κB priming and a second activation signal) increases IL‐1β/IL‐18 maturation and can induce pyroptosis or local tissue remodelling detrimental to implantation. These convergent pathways are mechanistically plausible targets for interventions (e.g., probiotics/postbiotics that reduce LPS/TLR stimulation, or engineered strains that locally deliver IL‐10/TGF‐β to re‐establish a Treg‐favouring milieu) (Balci and Acar  2024 ; Cheng et al.  2021 ; Li et al.  2024 ; Yeşilyurt et al.  2021 ).

Infertility affects millions worldwide and remains a major clinical and public‐health challenge: contemporary estimates indicate that around one in six people of reproductive age experience infertility during their lifetime (pooled lifetime prevalence ≈17.5%), though estimates vary by method and population (Cox et al.  2022 ; World Health Organization  2023 ). The pathogenesis of infertility is heterogeneous, major categories include ovulatory dysfunction (including PCOS), tubal and structural factors, uterine/endometrial disorders (including chronic endometritis), male factor infertility (sperm quantity/quality) and unexplained or idiopathic infertility, with immunologic, endocrine, anatomic, genetic and environmental/lifestyle contributors interacting in many patients (Carson and Kallen  2021 ; Kicińska et al.  2025 ; Luo et al.  2024 ). Current problems in infertility care include gaps in etiologic diagnosis (a nontrivial fraction of couples remain ‘unexplained’), variable access to and high cost of assisted reproductive technologies (ART) and treatments that often address downstream reproductive steps rather than root causes. These limitations motivate complementary therapeutic paradigms that target modifiable upstream factors (e.g., local mucosal ecology and inflammation) to improve implantation and live‐birth outcomes (Mesfin et al.  2025 ; Stern et al.  2022 ). For most of the twentieth century the uterus and upper reproductive tract were considered sterile, except in overt infection. Advances in sequencing and improved sampling now reveal reproducible microbial signals in the endometrium and adjacent niches, requiring a reassessment of that view (Moreno et al.  2016 ; Winters et al.  2019 ). Recent high‐resolution studies using contamination‐aware workflows and deep sequencing have shown that the endometrial cavity can harbour low‐biomass but biologically meaningful communities whose composition correlates with key reproductive endpoints, a finding that has shifted discussion from absence to ecological nuance (Franasiak et al.  2016 ; Moreno et al.  2022 ). At the same time, rigorous reappraisals of other low‐biomass tissues such as the placenta have modelled the methodological pitfalls and urged caution, reminding the field that detection of microbial DNA is not synonymous with established resident ecosystems and that rigorous negative controls and orthogonal validation are essential (Blaser et al.  2021 ; de Goffau et al.  2019 ). This dialectic between discovery and skepticism has been productive: methodological skepticism sharpened experimental design while reproducible clinical associations between endometrial or vaginal community states and implantation or pregnancy outcomes generated a new, testable clinical hypothesis (Franasiak et al.  2016 ; Moreno et al.  2022 ). To define the terrain for intervention we must distinguish the major ecological players while acknowledging their connectivity. The vagina is characterised by well described community state types dominated in health by Lactobacillus spp., which maintain low pH and produce antimicrobial compounds that discourage overgrowth of anaerobic pathobionts; these canonical community configurations were first and most comprehensively described in population‐scale sequencing efforts (De Seta et al.  2019 ; Ravel et al.  2011 ). By contrast the endometrial microbiome is a lower‐biomass, compositionally distinct ecosystem that nonetheless frequently shows Lactobacillus predominance in many women but also exhibits patterns of non‐ Lactobacillus dominance that have been associated with implantation failure, recurrent pregnancy loss and other adverse reproductive outcomes (Moreno et al.  2016 , 2022 ). Comparative studies reveal both overlap and divergence between vaginal and endometrial communities, some taxa transit or seed adjacent niches while mucosal immune conditioning, hormonal milieu and local anatomical barriers shape niche‐specific assembly, so the two habitats must be conceptualised as distinct but linked ecologies with different therapeutic access points and risk–benefit profiles (Miyagi et al.  2023 ; Polifke et al.  2024 ). Importantly, longitudinal and intervention studies indicate that manipulating the vaginal microbiome can change local inflammation and community structure and that endometrial compositions measured at the time of embryo transfer may predict outcome, thereby providing both mechanistic plausibility and a potential biomarker‐guided framework for therapeutic development (Franasiak et al.  2016 ; Cohen et al.  2020 ). Together, these developments create a clear translational path from descriptive ecology to precision therapeutics: rigorous, niche‐specific microbial characterisation; recognition of low‐biomass sampling pitfalls; and early clinical signals from live biotherapeutic products (LBPs) and microbiota transplantation that justify development of next‐generation probiotics for infertility (Blaser et al.  2021 ; Cohen et al.  2020 ; Lev‐Sagie et al.  2019 ). Importantly, the clinical link between reproductive‐tract microbial state and fertility is beginning to be demonstrated: observational IVF cohorts show that Lactobacillus ‐dominant profiles at the time of embryo transfer associate with higher implantation and live‐birth rates, and early intervention studies indicate that manipulating the vaginal microbiome can change local ecology and clinical outcomes. The most direct clinical evidence to date comes from vaginal microbiota transplantation (VMT) case series and proof‐of‐concept reports: Lev‐Sagie et al. reported feasibility of donor VMT to treat intractable bacterial vaginosis with durable engraftment in several cases, and more recently a proof‐of‐concept antibiotic‐free VMT case study documented donor engraftment, dysbiosis resolution and a subsequent live birth in a patient with prior recurrent pregnancy loss. These clinical signals remain preliminary but provide the most direct human evidence that microbiome reconstitution can favourably alter reproductive outcomes and therefore motivate hypothesis‐driven trials of VMT, defined live biotherapeutics and postbiotic strategies in infertility (Lev‐Sagie et al.  2019 ; Bosma et al.  2024 ; Wrønding et al.  2023 ).

The authors declare no conflicts of interest.

Bacterial interactions and the case for multi‐strain approaches. Bacteria in the reproductive tract act as an interconnected community where interspecies interactions, cross‐feeding and functional redundancy determine ecological stability and host effects; consequently, the effect of a single strain is frequently limited and multi‐strain or defined consortia approaches are increasingly seen as a promising future direction for durable modulation of dysbiosis. Synthetic consortia and multi‐strain products are conceptually appealing for functional redundancy and cooperative metabolism, but head‐to‐head comparative data versus single‐strain approaches are sparse; priorities should include controlled ecological comparisons, manufacturability and safety assessments, and early human trials emphasising microbiome conversion as a proximal endpoint (Armstrong et al.  2022 ). Preclinical and early translational studies support this concept: defined consortia reduce vaginal inflammation and restore healthy community states more effectively than monotherapies in animal models, and early human formulations have shown higher conversion rates toward Lactobacillus ‐dominant community types. Designing and deploying such consortia requires (i) strain‐level genomic and phenotypic selection, (ii) metabolic modelling to avoid antagonism and (iii) iterative in vitro/ex vivo mucosal validation prior to clinical testing. The synthetic consortia approach thus frames the development of next‐generation probiotics as community engineering rather than single‐strain replacement, and it motivates the use of combination strategies (live consortia, synbiotics or staged regimens) in infertility trials (Li et al.  2023 ; Liu, He, et al.  2024 ; Tanoue et al.  2019 ; van der Lelie et al.  2021 ). Clinical trials show mixed results for generic, food‐derived probiotics in the reproductive tract, highlighting the need for strain‐level selection: colonisation and efficacy vary by strain rather than by genus (Cohen et al.  2020 ; Romeo et al.  2024 ). Genomic and phenotypic surveys of vaginal Lactobacillus populations demonstrate that strains differ in key functional traits, such as glycogen utilisation, pullulanase gene variants, bacteriocin repertoires and adhesins, and these differences map to ecological fitness in the vaginal niche, explaining why some isolates persist after administration while others are transient (van der Veer et al.  2019 ; Santarelli et al.  2025 ). Experimental and translational work furthermore shows that certain vaginal isolates display host and site adaptation signatures absent from gut or food strains, implying that candidate NGPs should be selected from clinical vaginal collections or engineered to possess vaginal‐adaptive traits (Mancabelli et al.  2021 ; Pan et al.  2020 ). Finally, contemporary trials of live biotherapeutics in the urogenital tract reveal that formulation, dosing schedules and host preconditioning influence engraftment, reinforcing that strain choice must be integrated with a product‐level strategy rather than assumed sufficient on its own (Cohen et al.  2020 ; Armstrong et al.  2024 ). An ideal next‐generation probiotic for the reproductive tract should satisfy a short list of mechanistic, safety and translational criteria beginning with demonstrable niche fitness characterised by sustained acidification through lactic acid production, efficient glycogen or mucin‐utilisation pathways, and adhesins that promote mucosal retention without provoking deleterious inflammation (van der Veer et al.  2019 ; Clabaut et al.  2021 ). Antagonistic potential against pathobionts mediated by hydrogen peroxide, bacteriocins or competitive nutrient capture is a second essential property because suppression of anaerobic overgrowth is a primary mechanism by which lactobacilli reduce inflammation and maintain epithelial integrity (Santarelli et al.  2025 ; Mancabelli et al.  2021 ). Third, robust safety and manufacturing attributes are required: absence of transferable antibiotic resistance, low proinflammatory potential in ex vivo mucosal models, amenability to high‐titre cultivation and to stabilisation processes such as lyophilization or encapsulation, and genomic stability under production conditions (Agriopoulou et al.  2023 ; D'Amico et al.  2025 ). Fourth, the ideal candidate should either show evidence of clinical association with favourable reproductive outcomes or possess mechanistic plausibility for improving implantation milieu, for example by modulating local cytokine profiles or barrier function in endometrial models (Armstrong et al.  2024 ; Takada et al.  2023 ). Finally, for precision applications in infertility the ability to be deployed in companion diagnostics is valuable so that strain selection or multi‐strain prescriptions can be matched to a patient's baseline community state type (Ravel et al.  2025 ; Chowdhury et al.  2024 ). To operationalise NGP discovery we recommend the candidate‐level criteria and associated assays summarised in Table  2 . Ideal next‐generation probiotic (NGP) candidate: functional, safety and assayable criteria. Lactobacillus crispatus is widely regarded as the prototypical vaginal health species because populations dominated by L. crispatus exhibit the lowest vaginal pH, the lowest proinflammatory cytokine signature and the most stable community state types in longitudinal cohorts (van der Veer et al.  2019 ; Vieira‐Baptista et al.  2022 ). Clinical trials of a defined L. crispatus live biotherapeutic, CTV‐05 (commercially Lactin‐V), have shown reduced recurrence of bacterial vaginosis and partial but sometimes transient colonisation after dosing, thereby establishing both clinical proof of concept and the practical challenge of achieving durable engraftment (Cohen et al.  2020 ; Armstrong et al.  2024 ). Genomic analyses of L. crispatus isolates reveal strain‐level differences in glycogen metabolism, adhesins and bacteriocin content that predict growth on vaginal substrates and antagonism toward common pathobionts, supporting strain selection criteria grounded in functional genomics rather than species label alone (van der Veer et al.  2019 ; Santarelli et al.  2025 ). Recent formulation advances including mucoadhesive slow‐release vaginal tablets and multi‐strain synbiotic approaches have improved conversion of nonoptimal vaginal community types to L. crispatus dominance in early trials, indicating that pairing high‐fitness strains with delivery technologies can markedly increase ecological resilience (Armstrong et al.  2024 ; Ravel et al.  2025 ). Ligilactobacillus salivarius and several other noncanonical vaginal lactobacilli are emerging as promising NGP candidates because clinical and mechanistic studies suggest they can modulate urogenital dysbiosis and in some trials increase term pregnancy rates in selected infertility cohorts (Huerga López et al.  2025 ; Iniesta et al.  2022 ). These species frequently encode complementary functional repertoires, such as distinct bacteriocins, bile salt tolerance that facilitates oral delivery, and immunomodulatory surface molecules, which broaden the mechanistic toolbox beyond lactic acid production alone (Santarelli et al.  2025 ; Wu et al.  2024 ). Importantly, positive infertility signals reported with some L. salivarius preparations emphasize the need for randomised, mechanistically informed trials that measure colonisation, local immune modulation and endometrial receptivity rather than pregnancy endpoints alone in small open‐label cohorts (Huerga López et al.  2025 ; Thanaboonyawat et al.  2023 ). Viability and stability are the twin practical constraints for any live biotherapeutic, and techniques such as optimised lyophilization, protective excipients, and microencapsulation materially increase shelf‐life and survival during storage and delivery while also enabling controlled release at the mucosa (D'Amico et al.  2025 ; Vivek et al.  2023 ). Microencapsulation methods including alginate matrices, double‐coating, and mucoadhesive polymers preserve cell viability and can be tuned to resist ambient humidity or to dissolve slowly in the vaginal lumen, allowing lower dosing frequencies and improved colonisation probability (Agriopoulou et al.  2023 ; Sin et al.  2025 ). Route of administration remains a fundamental choice that shapes both pharmacokinetics and mechanistic reach; intravaginal delivery maximises local inoculum and immediate mucosal contact and has shown superior direct conversion to Lactobacillus ‐dominant states in several studies, while oral administration benefits from ease of use and potential engagement of the gut–vagina immunologic axis but typically results in lower rates of direct vaginal engraftment (Ravel et al.  2025 ; Rezazadeh et al.  2024 ). An emerging pragmatic strategy is dual‐channel administration or staged regimens that combine oral preconditioning of systemic immunity and mucosal seeding by intravaginal formulations, a design that may be especially relevant in infertility protocols where endometrial conditioning and synchronised mucosal states are necessary for implantation (Takada et al.  2023 ; Wang et al.  2024 ). For completeness we briefly summarise the principal clinical routes used to deliver microbiota to the gastrointestinal tract, because many of these approaches (colonoscopic infusion, enemas, nasoenteric delivery, oral capsules and transendoscopic enteral tubing) are directly relevant to how a microbial product might be formulated or targeted to influence distant mucosal sites (e.g., via gut–vagina or gut–endometrium axes). Lower‐GI delivery methods include colonoscopy (direct infusion to the colon) and retention enema; these methods place microbiota immediately into the large bowel and are commonly used in clinical FMT practice because of good local engraftment but require procedural resources and can be invasive. Upper‐GI/mid‐gut routes include nasogastric, nasojejunal or nasoduodenal tubes and upper endoscopic (OGD) infusion, which may be chosen when small‐bowel delivery is desired but carry theoretical aspiration risks and generally require careful patient selection and preparation. Oral encapsulated formulations (freeze‐dried stool in acid‐resistant capsules) have been developed as a noninvasive alternative and were shown to be non‐inferior to colonoscopic delivery for recurrent C. difficile in a randomised non‐inferiority trial, supporting capsule use where convenience and repeated dosing are priorities. Importantly, transendoscopic enteral tubing (TET), a transendoscopically placed tube that provides repeated, controlled access to the colon (colonic‐TET) or mid‐gut (mid‐gut TET), has been adopted in several centers to enable multiple or staged microbiota administrations without repeated full endoscopic procedures; prospective series report feasibility, safety and high patient acceptability, making TET especially practical when repeated dosing is planned. Each route has tradeoffs (invasiveness, reach, engraftment, safety and patient acceptability) and therefore the choice should be tailored to indication, product formulation (e.g., liquid vs. capsule vs. encapsulated probiotics) and the need for repeated dosing or local biofilm disruption (Brusnic et al.  2024 ; Kao et al.  2017 ; Wang et al.  2023 ; Zhang et al.  2020 ). Practical implications for reproductive‐tract microbial therapeutics: for vaginal or intrauterine goals, intravaginal formulations remain the most direct route to the target niche (see above). However, if an intervention seeks to act via gut→reproductive tract axes (metabolite delivery, systemic immune modulation, or engraftment of bile/orally‐tolerant strains), oral capsules or mid‐gut delivery may be preferred because they are less invasive and support repeated dosing; conversely, lower‐GI infusion methods (colonoscopy, colonic‐TET) are advantageous when high colon inoculum and rapid engraftment are required. When repeated administrations are expected (e.g., serial dosing during an IVF cycle), TET provides a practical compromise: it limits repeated sedation/endoscopy while enabling controlled instillation of a liquid microbiota product. These method‐level considerations should be reported and justified in clinical protocols, and product development should explicitly match formulation (capsule, lyophilized powder, mucoadhesive gel, liquid) to the chosen delivery route and safety package (Kao et al.  2017 ; Wang et al.  2023 ). Collectively, these advances point to an NGP paradigm that is strain‐conscious, mechanism‐driven, and formulation‐aware and that prizes robust preclinical validation and targeted clinical designs which measure ecological engraftment, mucosal immunology and reproductive physiology as proximal endpoints on the path to fertility outcomes (Cohen et al.  2020 ; D'Amico et al.  2025 ; Ravel et al.  2025 ).