Advancing stem cell technologies for conservation of wildlife biodiversity.

Wildlife biodiversity is essential for healthy, resilient and sustainable ecosystems. For biologists, this diversity also represents a treasure trove of genetic, molecular and developmental mechanisms that deepen our understanding of the origins and rules of life. However, the rapid decline in biodiversity reported recently foreshadows a potentially catastrophic collapse of many important ecosystems and the associated irreversible loss of many forms of life on our planet. Immediate action by conservationists of all stripes is required to avert this disaster. In this Spotlight, we draw together insights and proposals discussed at a recent workshop hosted by Revive & Restore, which gathered experts to discuss how stem cell technologies can support traditional conservation techniques and help protect animal biodiversity. We discuss reprogramming, in vitro gametogenesis, disease modelling and embryo modelling, and we highlight the prospects for leveraging stem cell technologies beyond mammalian species.

In vitro gametogenesis (IVG): reflections from a workshop.

Key ideas from a workshop convened by the National Academies of Sciences, Engineering, and Medicine to discuss developments in IVG (National Academies of Sciences, Engineering, and Medicine 2023) chaired by Dr Eli Y. Adashi (former Dean of Medicine and Biological Sciences at Brown University). The authors are solely responsible for the content of this paper, which does not necessarily represent the views of the National Academies of Sciences, Engineering, and Medicine.

Histological observations and transcriptome analyses reveal the dynamic changes in the gonads of the blotched snakehead (Channa maculata) during sex differentiation and gametogenesis.

BACKGROUND: Blotched snakehead (Channa maculata) displays significant sexual dimorphism, with males exhibiting faster growth rates and larger body sizes compared to females. The cultivation of the all-male population of snakeheads holds substantial economic and ecological value. Nonetheless, the intricate processes governing the development of bipotential gonads into either testis or ovary in C. maculata remain inadequately elucidated. Therefore, it is necessary to determine the critical time window of sex differentiation in C. maculata, providing a theoretical basis for sex control in production practices. METHODS: The body length and weight of male and female C. maculata were measured at different developmental stages to reveal when sexual dimorphism in growth initially appears. Histological observations and spatiotemporal comparative transcriptome analyses were performed on ovaries and testes across various developmental stages to determine the crucial time windows for sex differentiation in each sex and the sex-related genes. Additionally, qPCR and MG2C were utilized to validate and locate sex-related genes, and levels of E2 and T were quantified to understand sex steroid synthesis. RESULTS: Sexual dimorphism in growth became evident starting from 90 dpf. Histological observations revealed that morphological sex differentiation in females and males occurred between 20 and 25 dpf or earlier and 30-35 dpf or earlier, respectively, corresponding to the appearance of the ovarian cavity or efferent duct anlage. Transcriptome analyses revealed divergent gene expression patterns in testes and ovaries after 30 dpf. The periods of 40-60 dpf and 60-90 dpf marked the initiation of molecular sex differentiation in females and males, respectively. Male-biased genes (Sox11a, Dmrt1, Amh, Amhr2, Gsdf, Ar, Cyp17a2) likely play crucial roles in male sex differentiation and spermatogenesis, while female-biased genes (Foxl2, Cyp19a1a, Bmp15, Figla, Er) could be pivotal in ovarian differentiation and development. Numerous biological pathways linked to sex differentiation and gametogenesis were also identified. Additionally, E2 and T exhibited sexual dimorphism during sex differentiation and gonadal development. Based on these results, it is hypothesized that in C. maculata, the potential male sex differentiation pathway, Sox11a-Dmrt1-Sox9b, activates downstream sex-related genes (Amh, Amhr2, Gsdf, Ar, Cyp17a2) for testicular development, while the antagonistic pathway, Foxl2/Cyp19a1a, activates downstream sex-related genes (Bmp15, Figla, Er) for ovarian development. CONCLUSIONS: This study provides a comprehensive overview of gonadal dynamic changes during sex differentiation and gametogenesis in C. maculata, establishing a scientific foundation for sex control in this species.

Blotched snakehead (Channa maculata) exhibits significant sexual dimorphism, as males display faster growth rates and larger body sizes compared to females. The cultivation of the all-male population of snakeheads holds substantial economic and ecological value. However, the mechanisms underlying sex determination and differentiation in C. maculata remain insufficiently elucidated. In this study, sexual dimorphism in growth became evident starting from 90 dpf through the measurement of body length and weight of male and female C. maculata at different developmental stages. Histological observations indicated that morphological sex differentiation in females and males occurred at 20­25 dpf or earlier and 30­35 dpf or earlier, respectively, corresponding to the appearance of the ovarian cavity or efferent duct anlage. Transcriptome analyses revealed divergent gene expression patterns in male and female gonads after 30 dpf, suggesting that the period preceding 30 dpf might be the critical time window for sex control in C. maculata. The periods of 40­60 dpf and 60­90 dpf marked the initiation of molecular sex differentiation in females and males, respectively. Male-biased genes (Sox11a, Dmrt1, Amh, Amhr2, Gsdf, Ar, Cyp17a2) likely play crucial roles in testicular differentiation and spermatogenesis, while female-biased genes (Foxl2, Cyp19a1a, Bmp15, Figla, Er) could be pivotal in ovarian differentiation and oogenesis. Additionally, numerous biological pathways linked to sex differentiation and gametogenesis were identified. Moreover, sexual dimorphism was observed in the levels of E₂ and T during gonadal differentiation and development. Based on these findings, it is hypothesized that in C. maculata, the potential male sex differentiation pathway, Sox11a­Dmrt1­Sox9b, activates downstream sex-related genes (Amh, Amhr2, Gsdf, Ar, Cyp17a2) for testicular development, while the antagonistic pathway, Foxl2/Cyp19a1a, activates downstream sex-related genes (Bmp15, Figla, Er) for ovarian development. This study provides a comprehensive overview of gonadal dynamic changes during sex differentiation and gametogenesis in C. maculata, thereby establishing a scientific foundation for sex control in this species.

Relevance, strategies, and added value of mouse models in androgenetics.

BACKGROUND: Male Infertility is a prevalent condition worldwide, and a substantial fraction of cases are thought to have a genetic basis. Investigations into the responsible genes is limited experimentally, so mice have been used extensively to identify genes required for fertility and to understand their functions. OBJECTIVES: To review the progress made in reproductive genetics based on experiments in mice, the impact upon clinical fertility genetics, and discuss how evolving technologies will continue to advance our understanding of human infertility genes. RESULTS AND DISCUSSION: Gene knockout studies in mice have shown that several hundreds of genes are required for normal fertility and that this number is much higher in males than in females. In addition to gene discovery, the mouse is a powerful platform for functionally dissecting genetic pathways, modeling putative human infertility variants, identifying contraceptive targets, and developing in vitro gametogenesis. CONCLUSION: These ongoing studies in mice have made an enormous contribution to our understanding of the genetics of human reproduction in the sense that the "parts list" of genes for mammalian gametogenesis is being elucidated. This would have been impossible to do in humans, and in vitro systems are not yet adequate to associate genes with andrological phenotypes, especially in the germline.

Identification and Functional Analysis of E3 Ubiquitin Ligase g2e3 in Chinese Tongue Sole, Cynoglossus semilaevis.

Gametogenesis, the intricate developmental process responsible for the generation of germ cells (gametes), serves as a fundamental prerequisite for the perpetuation of the reproductive cycle across diverse organisms. The g2e3 enzyme is a putative ubiquitin E3 ligase implicated in the intricate regulatory mechanisms underlying cellular proliferation and division processes. The present study delves into the function of G2/M phase-specific E3 ubiquitin protein ligase (Cs-g2e3) in gametogenesis in Chinese Tongue Sole (Cynoglossus semilaevis). Sequence analysis shows that the Cs-g2e3 mRNA spans 6479 bp, encoding a 733 amino acid protein characterized by three conserved structural domains: PHD, RING, and HECT-typical of HECT E3 ubiquitin ligases. The predominant expression of Cs-g2e3 in the gonad tissues is further verified by qPCR. The expression profile of Cs-g2e3 in the gonads of the Chinese Tongue Sole is analyzed at different ages, and the results show that its expression peaks at 8 months of age and then begins to decline and stabilize. It is noteworthy that the expression level remains significantly elevated compared to that observed during the juvenile period. In situ hybridization shows that the mRNA of Cs-g2e3 is mainly localized in the germ cells of the ovary and the testis. RNA interference experiments show that the knockdown of Cs-g2e3 in ovarian and testicular germ cell lines significantly downregulates the expression of key genes involved in oogenesis (e.g., sox9 and cyp19a) and spermatogenesis (e.g., tesk1 and piwil2), respectively. Furthermore, the analysis of mutations in the transcription factor binding sites reveals that mutations within the Myogenin, YY1, and JunB binding sites significantly impact the transcriptional activity of the Cs-g2e3 gene, with the mutation in the YY1 binding site exhibiting the most pronounced effect (p < 0.001). This study contributes to a deeper understanding of the tissue-specific expression patterns of Cs-g2e3 across various tissues in Cynoglossus semilaevis, as well as the potential regulatory influences of transcription factors on its promoter activity. These findings may facilitate future research endeavors aimed at elucidating the expression and functional roles of the Cs-g2e3 gene.

From Germ Cells to Implantation: The Role of Extracellular Vesicles.

Extracellular vesicles represent a large heterogeneous class of near and long-distance intercellular communication mediators, released by both prokaryotic and eukaryotic cells. Specifically, the scientific community has shown growing interest in exosomes, which are nano-sized vesicles with an endosomal origin. Not so long ago, the physiological goal of exosome generation was largely unknown and required more investigation; at first, it was hypothesized that exosomes are able to remove excess, reject and unnecessary constituents from cells to preserve cellular homeostasis. However, thanks to recent studies, the central role of exosomes in regulating cellular communication has emerged. Exosomes act as vectors in cell-cell signaling by their cargo, proteins, lipids, and nucleic acids, and influence physiological and pathological processes. The findings on exosomes are widespread in a large spectrum of biomedical applications from diagnosis and prognosis to therapies. In this review, we describe exosome biogenesis and the current methods for their isolation and characterization, emphasizing the role of their cargo in female reproductive processes, from gametogenesis to implantation, and the potential involvement in human female disorders.

Decoding the Epigenetics of Infertility: Mechanisms, Environmental Influences, and Therapeutic Strategies.

Infertility is a complex condition caused by a combination of genetic, environmental, and lifestyle factors. Recent advances in epigenetics have highlighted the importance of epigenetic changes in fertility regulation. This review aims to provide a comprehensive overview of the epigenetic mechanisms involved in infertility, with a focus on DNA methylation, histone modification, and non-coding RNAs. We investigate the specific epigenetic events that occur during gametogenesis, with a focus on spermatogenesis and oogenesis as distinct processes. Furthermore, we investigate how environmental factors such as diet, stress, and toxin exposure can influence these epigenetic changes, potentially leading to infertility. The second part of the review explores epigenetic changes as therapeutic targets for infertility. Emerging therapies that modulate epigenetic marks present promising opportunities for fertility restoration, particularly in spermatogenesis. By summarizing current research findings, this review emphasizes the importance of understanding epigenetic contributions to infertility. Our discussion aims to lay the groundwork for future research directions and clinical applications in reproductive health.

Roles for the canonical polarity machinery in the de novo establishment of polarity in budding yeast spores.

Budding in the yeast Saccharomyces cerevisiae occurs at a single site pre-determined by cortical landmarks deposited during prior budding. During mating between haploid cells in the lab, external pheromone cues override the cortical landmarks to drive polarization and cell fusion. By contrast, in haploid gametes (called spores) produced by meiosis, a pre-determined polarity site drives initial polarized morphogenesis independent of mating partner location. Spore membranes are made de novo so existing cortical landmarks were unknown, as were the mechanisms by which the spore polarity site is made and how it works. We find that the landmark canonically required for distal budding, Bud8, stably marks the spore polarity site along with Bud5, a GEF for the GTPase Rsr1 that canonically links cortical landmarks to the conserved Cdc42 polarity machinery. Cdc42 and other GTPase regulators arrive at the site during its biogenesis, after spore membrane closure but apparently at the site where membrane synthesis began, and then these factors leave, pointing to a discrete "functionalization" step. Filamentous actin may be required for initial establishment of the site, but thereafter Bud8 accumulates independent of actin filaments. These results suggest a distinct polarization mechanism that may provide insights into gamete polarization in other organisms.

Livestock embryonic stem cells for reproductive biotechniques and genetic improvement.

Embryonic stem cells (ESCs) have proven to be a great in vitro model that faithfully recapitulates the events that occur during in vivo embryogenesis, making them a unique tool to study the cellular and molecular mechanisms that define tissue specification during embryonic development. Livestock ESCs are particularly attractive and have broad prospects including drug selection and human disease modeling, improvement of reproductive biotechniques and agriculture-related applications such as production of genetically modified animals. While mice and human ESCs have been established many years ago, no significant advances were made in livestock species until recently. Nowadays, livestock ESCs are available from cattle, pigs, sheep, horses and rabbits with different states of pluripotency. In this review, we summarize the current advances on livestock ESCs establishment and maintenance along with their present and future applications.

Visualization and Quantification of Rapid Chromosome Movements at Early Stages of Mouse Meiosis.

Telomere-led rapid chromosome movements (RPMs) are a conserved characteristic of chromosome dynamics in meiosis. RPMs have been suggested to influence critical meiotic functions such as DNA repair and the association of the homologous chromosomes. Here, we describe a method using 3D time-lapse fluorescence imaging to monitor RPMs in Hoechst-stained mouse seminiferous tubules explants. We supplement visualization with customized quantitative motion analysis and in silico simulation. The ability to carry out live imaging, combined with quantitative image analysis, offers a sensitive tool to investigate the regulation of RPMs, chromosome reorganizations that precede dynamic mid-prophase events, and their contribution to faithful transmission of genetic information.

Generation of Meiotic Mouse Models Using CRISPR/Cas9 Technology.

In recent years, targeted genome editing has emerged as an indispensable tool for creating animal models, facilitating a comprehensive exploration of the molecular mechanisms governing a myriad of biological processes. Within this scientific landscape, the investigation of meiosis in mice has attracted considerable attention across numerous research laboratories. The precision and versatility of the CRISPR/Cas9 genome editing system have revolutionized our ability to generate mice with tailored genetic alterations, including point mutations and null mutations. These genetic modifications have provided invaluable insights into the intricate functionality of various meiotic genes and their associated variants. In this context, we present a detailed state of the art protocol for the creation of novel mouse models, each bearing specific genetic modifications within key meiotic genes, through the application of CRISPR/Cas9 technology. Furthermore, we showcase two distinct genetic modifications, accomplished within our laboratory, that can serve as valuable reference points for researchers seeking to elucidate the molecular intricacies of meiosis in mammals.

Genome-wide analysis of the cytochrome P450 gene family in Pacific oyster Crassostrea gigas and their expression profiles during gonad development.

The cytochrome P450 (CYP) gene superfamily plays a significant role in various physiological processes, producing different compounds such as hormones, fatty acids, and biomolecules. However, little information is known their roles during gonad development in Pacific oyster (Crassostrea gigas). In this study, total of 116 CgCYP (Crassostrea gigas cytochrome P450) genes were identified and their expression pattern was analyzed for the first time. The relative molecular weights of these CgCYP genes ranged from 63.52 to 113.41 kDa, and the length of encoded amino acids ranged from 103 to 993. And total 26 cis-acting elements of these CgCYP genes were identified. GO and KEGG enrichment analysis showed some CgCYP genes are essential for the metabolism of male and female sex hormones. Additionally, expression anslysis showed 69 CgCYP genes were over-expressed in early gonad development and triploid infertile individuals. More importantly, expression levels of CgCYP1, CgCYP15, CgCYP34, CgCYP46, CgCYP69, CgCYP87, CgCYP88, and CgCYP103, were found to be significantly higher in female gonad, suggesting their important roles in female gonad development. The results of this study will provide a better understanding of the CgCYP genes in the gonad development of Pacific oyster.

Transcriptomic dataset of the development and maturation of the Rhipicephalus microplus ovary.

To conduct differential gene expression analysis, ovaries from the cattle tick Rhipicephalus microplus were dissected at three distinct developmental stages (preingurgitated, immature ingurgitated, and mature ingurgitated). Additionally, undissected intact mature males and complete ingurgitated female ticks without ovaries (carcasses) were also collected to serve as reference samples for analysis. To perform total RNA purification, tissue from ten individuals representing each of the five previously described conditions was pooled. mRNA was isolated from the purified total RNA using the oligo (dT) method. Following fragmentation, double stranded cDNA was synthesized and ligated to sequencing adapters. Suitable-sized fragments were subsequently used for PCR amplification. Libraries were analyzed and quantified using an Agilent 2100 Bioanalyzer and an ABI StepOnePlus Real-Time PCR System. A total of 45.64 Gb bases were sequenced using the Illumina HiSeq sequencing platform. After assembling the samples and correcting for abundance, we obtained 82,877 unigenes. The total length, average length, N50, and GC content of the unigenes were 89,754,828 bp,1,082 bp,2,068 bp and 49.04 % respectively. For functional annotation, the unigenes were aligned with 7 functional databases. The number of unigenes identified in the functional databases were as follows: 32,518 (NR:39.24 %), 10,259 (NT:12.38 %), 23,624 (Swissprot:28.50 %), 22,203 (KOG:26.79 %), 25,072 (KEGG:30.25 %), 17,435(GO:21.04 %), and 23,220 (InterPro:28.02 %). Unigene candidate coding regions (CDS) among the unigenes were predicted using TransDecoder software and 42,143 CDS were detected. We also detected 10,522 simple sequence repeats (SSRs) distributed on 8,126 unigenes, and predicted 4,672 transcription factors (TF) coding unigenes. Our data can be used to identify genes that are important for male and female tick and arachnid reproduction and tick general physiology.

Interplay of RNA-binding proteins controls germ cell development in zebrafish.

The specification of germ cells in zebrafish mostly relies on an inherited mechanism by which localized maternal determinants, called germ plasm, confer germline fate in the early embryo. Extensive studies have partially allowed the identification of key regulators governing germ plasm formation and subsequent germ cell development. RNA-binding proteins, acting in concert with other germ plasm components, play essential roles in the organization of the germ plasm and the specification, migration, maintenance, and differentiation of primordial germ cells. The loss of their functions impairs germ cell formation and causes sterility or sexual conversion. Evidence is emerging that they instruct germline development through differential regulation of mRNA fates in somatic and germ cells. However, the challenge remains to decipher the complex interplay of maternal germ plasm components in germ plasm compartmentalization and germ cell specification. Because failure to control the developmental outcome of germ cells disrupts the formation of gametes, it is important to gain a complete picture of regulatory mechanisms operating in the germ cell lineage. This review sheds light on the contributions of RNA-binding proteins to germ cell development in zebrafish and highlights intriguing questions that remain open for future investigation.

RNA HELICASE 32 is essential for female gametophyte development in Arabidopsis.

The normal progression of mitotic cycles and synchronized development within female reproductive organs are pivotal for sexual reproduction in plants. Nevertheless, our understanding of the genetic regulation governing mitotic cycles during the haploid phase of higher plants remains limited. In this study, we characterized RNA HELICASE 32 (RH32), which plays an essential role in female gametogenesis in Arabidopsis. The rh32 heterozygous mutant was semi-sterile, whereas the homozygous mutant was nonviable. The rh32 mutant allele could be transmitted through the male gametophyte, but not the female gametophyte. Phenotypic analysis revealed impaired mitotic progression, synchronization, and cell specification in rh32 female gametophytes, causing the arrest of embryo sacs. In the delayed pollination test, none of the retarded embryo sacs developed into functional female gametophytes, and the vast majority of rh32 female gametophytes were defective in the formation of the large central vacuole. RH32 is strongly expressed in the embryo sac. Knock-down of RH32 resulted in the accumulation of unprocessed 18 S pre-rRNA, implying that RH32 is involved in ribosome synthesis. Based on these findings, we propose that RH32 plays a role in ribosome synthesis, which is critical for multiple processes in female gametophyte development.

GametesOmics: A Comprehensive Multi-omics Database for Exploring the Gametogenesis in Humans and Mice.

Gametogenesis plays an important role in the reproduction and evolution of species. The transcriptomic and epigenetic alterations in this process can influence the reproductive capacity, fertilization, and embryonic development. The rapidly increasing single-cell studies have provided valuable multi-omics resources. However, data from different layers and sequencing platforms have not been uniformed and integrated, which greatly limits their use for exploring the molecular mechanisms that underlie oogenesis and spermatogenesis. Here, we develop GametesOmics, a comprehensive database that integrates the data of gene expression, DNA methylation, and chromatin accessibility during oogenesis and spermatogenesis in humans and mice. GametesOmics provides a user-friendly website and various tools, including Search and Advanced Search for querying the expression and epigenetic modification(s) of each gene; Tools with Differentially expressed gene (DEG) analysis for identifying DEGs, Correlation analysis for demonstrating the genetic and epigenetic changes, Visualization for displaying single-cell clusters and screening marker genes as well as master transcription factors (TFs), and MethylView for studying the genomic distribution of epigenetic modifications. GametesOmics also provides Genome Browser and Ortholog for tracking and comparing gene expression, DNA methylation, and chromatin accessibility between humans and mice. GametesOmics offers a comprehensive resource for biologists and clinicians to decipher the cell fate transition in germ cell development, and can be accessed at http://gametesomics.cn/.

Role of miRNAs in assisted reproductive technology.

Cellular proteins and the mRNAs that encode them are key factors in oocyte and sperm development, and the mechanisms that regulate their translation and degradation play an important role during early embryogenesis. There is abundant evidence that expression of microRNAs (miRNAs) is crucial for embryo development and are highly involved in regulating translation during oocyte and early embryo development. MiRNAs are a group of short (18-24 nucleotides) non-coding RNA molecules that regulate post-transcriptional gene silencing. The miRNAs are secreted outside the cell by embryos during preimplantation embryo development. Understanding regulatory mechanisms involving miRNAs during gametogenesis and embryogenesis will provide insights into molecular pathways active during gamete formation and early embryo development. This review summarizes recent findings regarding multiple roles of miRNAs in molecular signaling, plus their transport during gametogenesis and embryo preimplantation.

Anticipating in vitro gametogenesis: Hopes and concerns for IVG among diverse stakeholders.

In vitro gametogenesis (IVG), the reconstitution of germ cell development in vitro, is an emerging stem cell-based technology with profound implications for reproductive science. Despite researchers' long-term goals for future clinical applications, little is currently known about the views of IVG held by the stakeholders potentially most affected by its introduction in humans. We conducted focus groups and interviews with 80 individuals with lived experience of infertility and/or LGBTQ+ family formation in the US, two intersecting groups of potential IVG users. Respondents expressed hope that IVG would lead to higher reproductive success than current assisted reproductive technology (ART), alleviate suffering associated with ART use, and promote greater social inclusion, while expressing concerns predominantly framed in terms of equity and safety. These findings underscore the importance of sustained engagement with stakeholders with relevant experience to anticipate the implications of IVG for research and clinical translation.

Sex-biased gene expression precedes sexual dimorphism in the agonadal annelid Platynereis dumerilii.

Gametogenesis is the process by which germ cells differentiate into mature sperm and oocytes, cells essential for sexual reproduction. The sex-specific molecular programs that drive spermatogenesis and oogenesis can also serve as sex identification markers. Platynereis dumerilii is a research organism that has been studied in many areas of developmental biology. However investigations often disregard sex, as P. dumerilii juveniles lack sexual dimorphism. The molecular mechanisms of gametogenesis in the segmented worm P. dumerilii are also largely unknown. In this study, we used RNA sequencing to investigate the transcriptomic profiles of gametogenesis in P. dumerilii juveniles. Our analysis revealed that sex-biased gene expression becomes increasingly pronounced during the advanced developmental stages, particularly during the meiotic phases of gametogenesis. We identified conserved genes associated with spermatogenesis, such as dmrt1, and a novel gene psmt, that is associated with oogenesis. Additionally, putative long non-coding RNAs were upregulated in both male and female gametogenic programs. This study provides a foundational resource for germ cell research in P. dumerilii, markers for sex identification, and offers comparative data to enhance our understanding of the evolution of gametogenesis mechanisms across species.

Rapid identification of reproductive toxicants among environmental chemicals using an in vivo evaluation of gametogenesis in budding yeast Saccharomyces cerevisiae.

Infertility affects ∼12 % of couples, with environmental chemical exposure as a potential contributor. Of the chemicals that are actively manufactured, very few are assessed for reproductive health effects. Rodents are commonly used to evaluate reproductive effects, which is both costly and time consuming. Thus, there is a pressing need for rapid methods to test a broader range of chemicals. Here, we developed a strategy to evaluate large numbers of chemicals for reproductive toxicity via a yeast, S. cerevisiae high-throughput assay to assess gametogenesis as a potential new approach method (NAM). By simultaneously assessing chemicals for growth effects, we can distinguish if a chemical affects gametogenesis only, proliferative growth only or both. We identified a well-known mammalian reproductive toxicant, bisphenol A (BPA) and ranked 19 BPA analogs for reproductive harm. By testing mixtures of BPA and its analogs, we found that BPE and 17 ß-estradiol each together with BPA showed synergistic effects that worsened reproductive outcome. We examined an additional 179 environmental chemicals including phthalates, pesticides, quaternary ammonium compounds and per- and polyfluoroalkyl substances and found 57 with reproductive effects. Many of the chemicals were found to be strong reproductive toxicants that have yet to be tested in mammals. Chemicals having affect before meiosis I division vs. meiosis II division were identified for 16 gametogenesis-specific chemicals. Finally, we demonstrate that in general yeast reproductive toxicity correlates well with published reproductive toxicity in mammals illustrating the promise of this NAM to quickly assess chemicals to prioritize the evaluation for human reproductive harm.