The epididymis is a highly specialized organ essential for promoting the post-testicular functional maturation of spermatozoa, a process underpinning male fertility. This review examines the latest proteomics advances that have been used to unravel the complex molecular landscape of the epididymis, revealing the dynamic protein networks that shape sperm function beyond their genomic and transcriptomic blueprints. Here, we highlight how high-resolution mass spectrometry has helped to map the proteomic signatures of epididymal tissue, luminal extracellular vesicles (epididymosomes), and spermatozoa at different maturation stages, pinpointing key regulators of motility, capacitation, fertilization competence, and immune regulation. However, critical knowledge gaps remain, including deep protein characterization of the cytoplasmic droplet, epididymal fluid, and relatively underexplored anatomical tissue segments such as the corpus and cauda epididymis. We discuss how integrating global proteomic insights with complementary omics, single-cell proteomics, and advanced imaging is poised to reveal the spatial and temporal refinement of the sperm proteome, providing insights into how its disruption may contribute to idiopathic infertility. To promote data accessibility and accelerate discovery in epididymal biology, we introduce ShinyEpididymis (https://reproproteomics.shinyapps.io/ShinyEpididymis/), an interactive, web-based resource integrating publicly available proteomic datasets from spermatozoa, epididymosomes, and epididymal tissue. This platform enables researchers to rapidly query proteins of interest, explore spatial patterns of expression, and identify potential biomarkers or therapeutic targets. By consolidating current knowledge and defining future priorities, this review positions proteomics at the forefront of understanding epididymal biology, emphasising its clinical relevance and untapped potential for diagnosing and treating male infertility.

Protein phosphorylation is critical to successful sperm cell maturation and oocyte interaction. However, despite longstanding and largely global use of protein tyrosine phosphorylation as a robust marker of "readiness for fertilization," a deeper understanding of sperm-specific phosphorylation events and phosphoproteins is still lacking. To address this critical knowledge gap, here we present a complete guide to produce and analyze highly comprehensive phosphoproteomes from human spermatozoa. This chapter provides a bespoke guide to in vitro human sperm capacitation, phosphopeptide enrichment, and high-throughput sample preparation for mass spectrometry based on the renowned EasyPhos method. Replete with hints for successful application, we intend this guide to be of equal use in improving our understanding of male infertility, as well as establishing new insight for the development of non-hormonal, sperm-targeted contraceptives.

Capacitation, the process whereby sperm gain the functional competence to fertilize an egg in the absence of de novo transcription and translation, is orchestrated by a hierarchy of kinases driving the phosphorylation of sperm proteins. While increased phosphorylation, in particular tyrosine phosphorylation, is a revered hallmark of fertilization competency in our species, only a limited repository of phosphorylated substrates and kinases have ever been reported from human sperm. To broaden therapeutic targets for sperm targeted contraceptives and infertility therapies, we adapted a contemporary phosphoproteomic technique termed EasyPhos to generate bespoke methodology for the investigation of human sperm signalling. This approach yielded high depth phosphoproteomes of non-capacitated and capacitated human spermatozoa with in silico investigation of the phosphosites revealing 52 kinases with previously uncharacterized roles in sperm capacitation. Investigating the function of the putative sperm capacitation kinases identified yielded several kinases with novel roles in the regulation of sperm function. Of particular interest, polo like kinase 1 (PLK1) inhibition significantly reduced progressive sperm motility, attenuated capacitation-associated tyrosine phosphorylation and reduced the sperm acrosome reaction, an essential step to achieve fertilization. These findings reveal extensive phosphoproteome remodelling during human sperm capacitation, expanding the landscape of molecular targets for fertility control.

Abstract The manchette is a transient microtubule (MT)-based structure that is vital for the correct shaping of sperm during spermiogenesis. Throughout spermiogenesis, the manchette retains structural integrity for several days, raising the question of how its MTs are regulated. Here, using cryo-electron tomography of manchettes isolated from rat testes, we find that manchette MT ends are structurally diverse. We show that the MT-binding protein CLASP2 is present throughout the manchette and likely regulates both MT ends. Using cryo-electron microscopy single particle analysis and super-resolution microscopy, we reveal that SPACA9 and MNMIP1 (SH3D21) bind to the seam of manchette MTs from the luminal side. SPACA9 binds to both α- and β-tubulin of protofilament 1 but does not interact directly with protofilament 13, while MNMIP1 binds directly to protofilament 13. MNMIP1 further extends and threads through the MT lattice at the seam. Our study reveals a novel seam MT inner protein complex with a unique binding mode, providing a plausible explanation for MT regulation that maintains manchette integrity over an extended period.

Oxidative stress, characterized by an imbalance between prooxidants and antioxidants in favor of the former, can lead to cellular damage due to the accumulation of reactive oxygen species (ROS) and reactive nitrogen species (RNS). In spermatozoa, oxidative stress plays a crucial role in signaling pathways associated with capacitation, a process essential for fertilization. However, the limited capacity of spermatozoa to buffer ROS makes them susceptible to oxidative stress. Measurement of oxidative stress in spermatozoa is important as it precedes or directly causes the loss of basic sperm functions-such as motility and the membrane changes required for zona binding and gamete fusion-associated with fertility. This chapter aims to describe protocols for assessing oxidative damage in spermatozoa, encompassing the generation of ROS in mitochondria to the manifestation of oxidative damage in DNA, proteins, and lipids. In addition to the detailed instructions provided within this chapter, we have sought to provide background information on the development and significance of the assays which we most commonly utilize in our laboratory.

The functional maturation of mammalian spermatozoa is driven by modification of their intrinsic proteome as the cells transit the male (epididymal sperm maturation) and female reproductive tracts (capacitation). Here, high-resolution mass spectrometry was used to interrogate the central role that phosphoproteomic changes play in the functional remodeling of mouse spermatozoa. This strategy identified 14,586 site-specific phosphorylation events, including the phosphorylation of 573 proteins and dephosphorylation of 426 during epididymal maturation and additional phosphorylation changes in 211 proteins linked to capacitation. We identified over 300 kinases that putatively govern these events, including three novel kinases (STK33, HIPK4, and PAK1) implicated in acrosomal exocytosis. The functional relevance of these data was confirmed via the use of knockout mouse models, which demonstrated several phosphoproteins as being essential for sperm motility and fertilization capacity. These findings illustrate that large-scale phosphorylation remodeling occurs during sperm maturation with implications extending to novel means of fertility regulation. HIGHLIGHTS ### Competing Interest Statement The authors have declared no competing interest. National Health and Medical Research Council of Australia, APP1147932, APP2034392

Seminal extracellular vesicles (SEVs) carry a diverse array of bioactive molecules, including proteins, lipids, and nucleic acids, which influence sperm function and have potential to modulate the female reproductive tract immune response after intromission. However, the full spectrum of SEV cargo involved in these processes remains incompletely defined. Here, we employed label-free quantitative high-resolution mass spectrometry to characterize the human SEV proteome, identifying 5079 associated proteins. These proteins were shown to likely originate from multiple regions of the male reproductive tract, notably the seminal vesicles and prostate, providing evidence for heterogeneous tissue origins of SEVs. Bioinformatic analysis revealed enrichment in sperm- and immune-related functions, as well as functions linked to protein translation. Notably, we identified several proteins with established roles in sperm physiology and immune signaling that had not previously been linked with SEV function. These included; Adenylate kinase isoenzyme (AK)2/9, and Calcium-binding tyrosine-phosphorylation regulated protein (CABYR), implicated in sperm motility, and immune regulators such as the Toll-like receptor 4 ligand, High mobility group protein B1 (HMGB1), and the Nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) inhibitor epsilon (NFκBIE). Altogether, these findings expand the known SEV proteome and highlight proteins that may influence both male and female reproductive capacity.

Sperm capacitation describes a series of biochemical and biophysical changes that collectively render the cell competent to fertilize an ovum. This phase of posttesticular maturation is initiated soon after ejaculation in response to physiological cues arising from within the female reproductive tract and encompasses major remodeling of the plasma membrane, activation of complex signaling cascades, expression of hyperactivated motility and the ability to complete an acrosome reaction. This article discusses the biological significance of capacitation in terms of enhancing synchrony between sperm activation and ovulation and our current mechanistic understanding of how this is achieved.

BACKGROUND:Bisphenol A (BPA) is an endocrine disrupting chemical released from plastic materials, including food packaging and dental sealants, persisting in the environment and ubiquitously contaminating ecosystems and human populations. BPA can elicit an array of damaging health effects and, alarmingly, 'BPA-free' alternatives mirror these harmful effects. Bisphenol exposure can negatively impact female fertility, damaging both the ovary and oocytes therein. Such damage can diminish reproductive capacity, pregnancy success, and offspring health. Despite global government regulations in place to indicate 'safe' BPA exposure levels, these policies have not considered the effects of bisphenols on oocyte health. OBJECTIVE AND RATIONALE:This scoping review was conducted to evaluate evidence on the effects of BPA and BPA alternatives on standardized parameters of oocyte health. In doing so, this review addresses a critical gap in the literature providing a comprehensive, up-to-date synthesis of the effects of bisphenols on oocyte health. SEARCH METHODS:This scoping review was conducted in accordance with PRISMA guidelines. Four databases, Medline, Embase, Scopus, and Web of Science, were searched twice (23 February 2022 and 1 August 2023) to capture studies assessing mammalian oocyte health post-bisphenol exposure. Search terms regarding oocytes, ovarian follicles, and bisphenols were utilized to identify relevant studies. Manuscripts written in English and reporting the effect of any bisphenol on mammalian oocyte health from all years were included. Parameters for toxicological studies were evaluated, including the number of bisphenol concentrations/doses tested, dosing regimen, biological replicates and/or animal numbers, and statistical information (for human studies). Standardized parameters of oocyte health including follicle counts, oocyte yield, oocyte meiotic capacity, morphology of oocyte and cumulus cells, and oocyte meiotic spindle integrity were extracted across the studies. OUTCOMES:After screening 3147 studies, 107 studies of either humans or mammalian animal models or humans were included. Of the in vitro exposure studies, 96.3% (26/27) and 94.1% (16/17) found at least one adverse effect on oocyte health using BPA or BPA alternatives (including BHPF, BPAF, BPB, BPF, and BPS), respectively. These included increased meiotic cell cycle arrest, altered morphology, and abnormal meiotic spindle/chromosomal alignment. In vivo, 85.7% (30/35) of studies on BPA and 92.3% (12/13) on BPA alternatives documented adverse effects on follicle development, morphology, or spindle/chromosome alignment. Importantly, these effects were recorded using levels below those deemed 'safe' for human exposure. Over half (11/21) of all human observational studies showed associations between higher urinary BPA levels and reduced antral follicle counts or oocyte yield in IVF patients. Recommendations are presented based on the identified shortcomings of the current evidence, incorporating elements of FDA requirements for future research in the field. WIDER IMPLICATIONS:These data highlight the detrimental impacts of low-level BPA and BPA alternative exposure, contributing to poor oocyte quality and reduced fertility. These outcomes are valuable in promoting the revision of current policies and guidelines pertaining to BPA exposure internationally. This study serves as a valuable resource to scientists, providing key recommendations on study design, reporting elements, and endpoint measures to strengthen future studies. Ultimately, this review highlights oocyte health as a fundamentally important endpoint in reproductive toxicological studies, indicating an important direction for future research into endocrine disrupting chemicals to improve fertility outcomes.

Forecasted increases in the prevalence and severity of extreme weather events accompanying changes in climatic behavior pose potential risk to the reproductive capacity of humans and animals of ecological and agricultural significance. While several studies have revealed that heat stress induced by challenges such as testicular insulation can elicit a marked negative effect on the male reproductive system, and particularly the production of spermatozoa, less is known about the immediate impact on male reproductive function following subchronic whole-body exposure to elevated ambient temperature. To address this knowledge gap, we exposed unrestrained male mice to heat stress conditions that emulate a heat wave (daily cycle of 8 h at 35 °C followed by 16 h at 25 °C) for a period of 7 d. Neither the testes or epididymides of heat-exposed male mice exhibited evidence of gross histological change, and similarly, spermatozoa of exposed males retained their functionality and ability to support embryonic development. However, the embryos generated from heat-exposed spermatozoa experienced pronounced changes in gene expression linked to acceleration of early embryo development, aberrant blastocyst hatching, and increased fetal:placental weight ratio. Such changes were causally associated with an altered sperm small noncoding RNA (sncRNA) profile, such that these developmental phenotypes were recapitulated by microinjection of wild-type embryos sired by control spermatozoa with RNAs extracted from heat-exposed spermatozoa. Such data highlight that even relatively modest excursions in ambient temperature can affect male reproductive function and identify the sperm sncRNA profile as a particular point of vulnerability to this imposed environmental stress.

Abstract Seminal fluid extracellular vesicles (SFEVs) have previously been shown to interact with spermatozoa and influence their fertilisation capacity. Here, we sought to extend these studies by exploring the functional consequences of SFEV interactions with human spermatozoa. SFEVs were isolated from the seminal fluid of normozoospermic donors prior to assessing the kinetics of sperm-SFEV binding in vitro, as well as the effects of these interactions on sperm capacitation, acrosomal exocytosis, and motility profile. Biotin-labelled SFEV proteins were transferred primarily to the flagellum of spermatozoa within minutes of co-incubation, although additional foci of SFEV biotinylated proteins also labelled the mid-piece and head domain. Functional analyses of high-quality spermatozoa collected following liquefaction revealed that SFEVs did not influence sperm motility during incubation at pH 5, yet SFEVs induced subtle increases in total and progressive motility in sperm incubated with SFEVs at pH 7. Additional investigation of sperm motility kinematic parameters revealed that SFEVs significantly decreased beat cross frequency and increased distance straight line, linearity, straightness, straight line velocity, and wobble. SFEVs did not influence sperm capacitation status or the ability of sperm to undergo acrosomal exocytosis. Functional assessment of both high- and low-quality spermatozoa collected prior to liquefaction showed limited SFEV influence, with these vesicles inducing only subtle decreases in beat cross frequency in spermatozoa of both groups. These findings raise the prospect that, aside from subtle effects on sperm motility, the encapsulated SFEV cargo may be destined for physiological targets other than the male germline, notably the female reproductive tract. Lay Summary A male’s influence over the biological processes of pregnancy extends beyond the provision of sperm. Molecular signals present in the ejaculate can influence the likelihood of pregnancy and healthy pregnancy progression, but the identity and function of these signals remain unclear. In this study, we wanted to understand if nano-sized particles present in the male ejaculate, called seminal fluid extracellular vesicles, can assist sperm in traversing the female reproductive tract to access the egg. To explore this, we isolated seminal fluid extracellular vesicles from human semen and incubated them with sperm. Our data showed that seminal fluid extracellular vesicles act to transfer molecular information to sperm, but this resulted in only subtle changes to the movement of sperm. Graphical abstract

Understanding the systemic biochemistry of early pregnancy in the mare is essential for developing new diagnostics and identifying causes for pregnancy loss. This study aimed to elucidate the dynamic lipidomic changes occurring during the initial stages of equine pregnancy, with a specific focus on days 7 and 14 post-ovulation. By analysing and comparing the plasma lipid profiles of pregnant and non-pregnant mares, the objective of this study was to identify potential biomarkers for pregnancy and gain insights into the biochemical adaptations essential for supporting maternal recognition of pregnancy and early embryonic development. Employing discovery lipidomics, we analysed plasma samples from pregnant and non-pregnant mares on days 7 and 14 post-conception using the SCIEX ZenoTOF 7600 system. This high-resolution mass spectrometry approach enabled us to comprehensively profile and compare the lipidomes across these critical early gestational timepoints. Our analysis revealed significant lipidomic alterations between pregnant and non-pregnant mares and between days 7 and 14 of pregnancy. Key findings include the upregulation of bile acids, sphingomyelins, phosphatidylinositols, and triglycerides in pregnant mares. These changes suggest enhanced lipid synthesis and mobilization, likely associated with the embryo's nutritional requirements and the establishment of embryo-maternal interactions. There were significant differences in lipid metabolism between pregnant and non-pregnant mares, with a notable increase in the sterol lipid BA 24:1;O5 in pregnant mares as early as day 7 of gestation, suggesting it as a sensitive biomarker for early pregnancy detection. Notably, the transition from day 7 to day 14 in pregnant mares is characterized by a shift towards lipids indicative of membrane biosynthesis, signalling activity, and preparation for implantation. The study demonstrates the profound lipidomic shifts that occur in early equine pregnancy, highlighting the critical role of lipid metabolism in supporting embryonic development. These findings provide valuable insights into the metabolic adaptations during these period and potential biomarkers for early pregnancy detection in mares.

Significance: Stringent regulation of protein homeostasis pathways, under both physiological and pathological conditions, is necessary for the maintenance of proteome fidelity and optimal cell functioning. However, when challenged by endogenous or exogenous stressors, these proteostasis pathways can become dysregulated with detrimental consequences for protein fate, cell survival, and overall organism health. Most notably, there are numerous somatic pathologies associated with a loss of proteostatic regulation, including neurodegenerative disorders, type 2 diabetes, and some cancers.Recent Advances: Lipid oxidation-derived reactive carbonyl species (RCS), such as 4-hydroxynonenal (4HNE) and malondialdehyde, are relatively underappreciated purveyors of proteostatic dysregulation, which elicit their effects via the nonenzymatic post-translational modification of proteins. Emerging evidence suggests that a subset of germline proteins can serve as substrates for 4HNE modification. Among these, prevalent targets include succinate dehydrogenase, heat shock protein A2 and A-kinase anchor protein 4, all of which are intrinsically associated with fertility.Critical Issues: Despite growing knowledge in this field, the RCS adductomes of spermatozoa and oocytes are yet to be comprehensively investigated. Furthermore, the manner by which RCS-mediated adduction impacts protein fate and drives cellular responses, such as protein aggregation, requires further examination in the germline. Given that RCS-protein adduction has been attributed a role in infertility, there has been sparked research investment into strategies to prevent lipid peroxidation in germ cells.Future Directions: An increased depth of knowledge regarding the mechanisms and substrates of RCS-mediated protein modification in reproductive cells may reveal important targets for the development of novel therapies to improve fertility and pregnancy outcomes for future generations.

Sperm cells are terminally differentiated cells that are essential for reproduction in sexually reproducing species. Consistent with their highly specialized function, sperm cells harbor a unique proteome containing many proteins not expressed in somatic cells. In contrast, the post-translational landscape of the sperm proteome remains largely unexplored, limiting our understanding of how modifications such as glycosylation impact sperm function and sperm-egg interactions. Here, we used glycopeptide-centric glycoproteomics to comprehensively characterize protein N-glycosylation in sperm from three mammalian species, revealing clear conservation of glycosylation profiles. We find that glycosylation patterns in sperm proteins are distinct from those in plasma, with as clear distinctive features less sialyation and more paucimannosylation in sperm. Moreover, based on their subcellular location, sperm protein glycosylation varies, with paucimannose species enriched in the acrosomal vesicle, oligomannose species in the sperm head membrane, and complex glycan species in the acrosomal membrane.

BACKGROUND:Previously, we reported that cysteine-rich secretory protein 2 is involved in high molecular weight complexes in boar spermatozoa. These cysteine-rich secretory protein 2protein complexes are formed at the last phase of sperm formation in the testis and play a role in sperm shaping and functioning.OBJECTIVES:This study aimed to identify cysteine-rich secretory protein 2 interacting partners. These binding partner interactions were investigated under different conditions, namely, non-capacitating conditions, after the induction of in vitro sperm capacitation and subsequently during an ionophore A23187-induced acrosome reaction.MATERIALS AND METHODS:The incubated pig sperm samples were subjected to protein extraction. Extracted proteins were subjected to blue native gel electrophoresis and native immunoblots. Immunoreactive gel bands were excised and subjected to liquid chromatography-mass spectrometry (LC-MS) analysis for protein identification. Protein extracts were also subjected to CRISP2 immunoprecipitation and analyzed by LC-MS for protein identification. The most prominent cystein-rich secretory protein 2 interacting proteins that appeared in both independent LC-MS analyses were studied with a functional in situ proximity interaction assay to validate their property to interact with cystein-rich secretory protein 2 in pig sperm.RESULTS:Blue native gel electrophoresis and native immunoblots revealed that cystein-rich secretory protein 2 was present within a ∼150 kDa protein complex under all three conditions. Interrogation of cystein-rich secretory-protein 2-immunoreactive bands from blue native gels as well as cystein-rich secretory protein 2 immunoprecipitated products using mass spectrometry consistently revealed that, beyond cystein-rich secretory protein 2, acrosin and acrosin binding protein were among the most abundant interacting proteins and did interact under all three conditions. Co-immunoprecipitation and immunoblotting indicated that cystein-rich secretory protein 2 interacted with pro-acrosin (∼53 kDa) and Aacrosin binding protein under all three conditions and additionally to acrosin (∼35 kDa) after capacitation and the acrosome reaction. The colocalization of these interacting proteins with cystein-rich secretory protein 2 was assessed via in situ proximity ligation assays. The colocalization signal of cystein-rich secretory protein 2 and acrosin in the acrosome seemed dispersed after capacitation but was consistently present in the sperm tail under all conditions. The fluorescent foci of cystein-rich secretory protein 2 and acrsin binding protein colocalization appeared to be redistributed within the sperm head from the anterior acrosome to the post-acrosomal sheath region upon capacitation.DISCUSSION AND CONCLUSION:These results suggest that CRISP2 may act as a scaffold for protein complex formation and dissociation to ensure the correct positioning of proteins required for the acrosome reaction and zona pellucida penetration.

Early pregnancy in the mare presents a risk of embryo loss, with little known of the systemic lipid dynamics that contribute to a successful or failed pregnancy. Observations in horses and other species suggest that pregnancy is characterized by an anabolic (i.e. fat deposition) lipid profile initially, followed by a catabolic (lipid mobilization) profile toward the end of gestation. There have been no investigations of lipid metabolism in the very early stages of pregnancy, leading up to and during the time of maternal recognition. High resolution lipidomics presents an unprecedented opportunity to map the changes in lipid profiles of pregnant mares, to examine metabolic pathways contributing to establishment of pregnancy and to identify biomarkers for detection and monitoring of early pregnancy and pregnancy loss. Blood samples were collected from Thoroughbred mares in a commercial breeding operation at day 7 and day 14 following ovulation. Mares were live-covered, and pregnancy tested by transrectal ultrasound at day 14, hereby designated pregnant or non-pregnant. High resolution lipidomics analysis was performed on isolated plasma to compare profiles of pregnant (7P, 14P) and non-pregnant (7NP, 14NP) mares at day 7 and day 14 (total n = 72, split evenly in groups). Lipids were extracted using ice-cold butanol/methanol (1:1 v/v) with 10 mM ammonium acetate containing internal standards.LC separation was performed on a Phenomenex Kinetex C18 Column (2.6 µm, 100 Å, 100 × 2.1 mm) using 5 µL injection volume. The samples were analyzed in data dependent mode using Zeno EAD IDA on the ZenoTOF 7600 system. All data were analyzed using MS-DIAL 5.1. The analysis yielded a total of 10034 unique lipid species across all samples. A total of 277 and 1722 lipids were significantly altered (p<0.05) in pregnant vs non-pregnant animals at day 7 and day 14, respectively. Phosphatidylcholines (PC) were significantly increased in 7P vs 7NP, and phosphatidylethanolamines (PE) were significantly decreased in 7P vs 7NP, with similar trends observed at day 14. Lysophosphatidylcholines (LPC) were increased in 14P vs 14NP. PC and LPC concentrations also increased from day 7 to day 14, but only in pregnant animals. Collectively, these observations point to a major role for phospholipid metabolism in early pregnancy and are consistent with PE being a substrate for methylation to PC, which is a key source of choline crucial for early embryonic development. The roles of PC biosynthesis and supplementation in early equine pregnancy warrant further investigation. Furthermore, alterations in the lipid profile of mares at day 7 post-ovulation challenge the current paradigm that no systemic changes occur in the pregnant mare prior to classical maternal recognition and luteal maintenance.

Male infertility is a commonly encountered pathology that is estimated to be a contributory factor in approximately 50% of couples seeking recourse to assisted reproductive technologies. Upon clinical presentation, such males are commonly subjected to conventional diagnostic andrological practices that rely on descriptive criteria to define their fertility based on the number of morphologically normal, motile spermatozoa encountered within their ejaculate. Despite the virtual ubiquitous adoption of such diagnostic practices, they are not without their limitations and accordingly, there is now increasing awareness of the importance of assessing sperm quality in order to more accurately predict a male’s fertility status. This realization raises the important question of which characteristics signify a high-quality, fertilization competent sperm cell. In this review, we reflect on recent advances in our mechanistic understanding of sperm biology and function, which are contributing to a growing armory of innovative approaches to diagnose and treat male infertility. In particular we review progress toward the implementation of precision medicine; the robust clinical adoption of which in the setting of fertility, currently lags well behind that of other fields of medicine. Despite this, research shows that the application of advanced technology platforms such as whole exome sequencing and proteomic analyses hold considerable promise in optimizing outcomes for the management of male infertility by uncovering and expanding our inventory of candidate infertility biomarkers, as well as those associated with recurrent pregnancy loss. Similarly, the development of advanced imaging technologies in tandem with machine learning artificial intelligence are poised to disrupt the fertility care paradigm by advancing our understanding of the molecular and biological causes of infertility to provide novel avenues for future diagnostics and treatments.