Pouya Dini

Background: There is an ethical debate about the application of ovum pick‐up (OPU) and intracytoplasmic sperm injection (ICSI) in mares. Objectives: To survey the opinions of equine veterinarians and breeders of sport horses on the ethics of using OPU–ICSI in mares. Study Design: Cross‐sectional descriptive study. Methods: Three surveys were distributed. The first targeted breeders (‘Breeder Survey’) who were actively involved in sport horse breeding. The second targeted veterinarians performing OPU (‘OPU‐Reprovets Survey’). The third targeted general equine practitioners (‘Practitioner Survey’). There were six questions common to all surveys, and, for analyses, respondents were categorised into four groups: equine veterinarians not performing OPU (VET), equine veterinarians performing OPU (OPUVET), breeders not using OPU in their breeding programme (BREEDER), and those using OPU in their breeding programme (OPUBREEDER). Results: A total of 225, 92 and 112 responses were returned from the Breeders, OPU‐Reprovets and Practitioner surveys, respectively. Differences emerged between groups in how ethically challenging the procedure was perceived to be. A greater proportion of veterinarians not performing OPU (VET; 58.8%) and breeders not using the technology (BREEDER; 55.2%) rated OPU–ICSI as ethically challenging (scores 4–5 on a 5‐point scale), compared with veterinarians performing OPU (OPUVET; 35.2%) and breeders using the technology (OPUBREEDER; 19.6%). Across all groups, mare welfare was the main theme when considering the use of OPU–ICSI, and for BREEDER and OPUBREEDER this was closely followed by Genetic diversity and overuse . Main Limitations: Potential for motivational and response bias. Conclusions: These results demonstrate differences in perception of ethical acceptability of OPU–ICSI between veterinarians and breeders with and without direct experience of OPU–ICSI. Despite these differences, mare welfare was consistently prioritised across all respondent groups, highlighting the importance of further research to better characterise the mare experience during OPU and to support evidence‐based guidance for clinical and breeding practice.

The endocrinology of equine pregnancy is unique, but many of its unusual phenomena closely resemble pregnancy in women. Placental estrone synthesis uses fetal adrenal androgen precursors in women, but from the fetal gonads in mares, peaking midway through the 11-month gestation. Unique to equine pregnancies, other estrogens like equilin, typified by an unsaturated B-ring sterol structure, are synthesized from 7-dehydrocholesterol (7-DHC), the immediate precursor to cholesterol, without cholesterol formation, peaking later in gestation. Currently, the spatial and temporal regulation underlying the redirection of 7-DHC from cholesterol to B-ring unsaturated steroid synthesis in equine fetal gonads remains unknown. Here, we investigate the developmental dynamics of the equine fetal gonads from the fourth to eleventh gestational month using RNA sequencing, immunofluorescence imaging, RNAScope, and gas chromatography–mass spectrometry. Our results suggest that placental estrone synthesis correlates with the expression of the last enzyme in the cholesterol synthesis pathway, 7-dehydrocholesterol reductase (DHCR7) in the fetal gonad, and its downregulation may drive the accumulation of 7-DHC and the synthesis of B-ring unsaturated steroids. B-ring unsaturated steroid secretion is the hallmark of Smith–Lemli–Opitz Syndrome in humans, resulting from mutations in the gene encoding DHCR7 in affected patients. This naturally occurring phenomenon in equine pregnancies may offer a unique comparative model for understanding the metabolic consequences of 7-DHC accumulation in these patients and a platform for drug development.

Testicular seminomas in horses are infrequently reported in the veterinary literature and long-term survival and metastatic potential are not well documented. Describe the clinical, imaging, and histologic features in 9 horses with testicular seminoma. Medical records of 9 horses diagnosed with testicular seminoma at a referral hospital between 2000 and 2024 were reviewed. Horses were included if seminoma was confirmed on histopathology from biopsy or necropsy. Information retrieved included signalment, history, clinical pathology, ultrasonographic findings, treatment, and outcome. Median age was 18 years (range, 8-26). Three of 9 horses presented for systemic disease (colic, 2; ventral edema, 1). All 3 had large abdominal masses palpable on rectal examination and were euthanized after diagnostic evaluation. Six horses presented for unilateral testicular enlargement (5) or cryptorchid castration (1). These horses underwent castration and the removed testes had features of malignancy with lymphatic neoplastic emboli. Five of 6 horses were euthanized between 4 months and 3 years after castration because of acute weight loss and abdominal masses identified on rectal examination or ultrasonographic examination, attributable to metastatic spread. One additional horse was euthanized for unrelated reasons. Horses presented with testicular enlargement and diagnosis of testicular seminoma should undergo diagnostic screening, including transabdominal or transrectal ultrasonographic examination to determine the presence of possible metastasis. Negative screening results at the time of diagnosis do not rule out the possibility of metastasis, and frequent screening at regular intervals is prudent in such cases. Horses with testicular seminoma of retained or descended testicles have a poor long term prognosis because of high potential for metastatic spread.

Background: Ascending placentitis is a leading cause of late‐term pregnancy loss in mares. Although pathogens are presumed to ascend from the caudal reproductive tract, the association between the vaginal microbiome and placentitis has not been systematically examined. Objectives: To characterise microbial and host gene expression in the equine vagina during gestation and to identify taxa or transcripts that were associated with the presence of an abnormal placenta at birth. Study Design: Prospective, paired observational study. Methods: Vaginal wall swabs were collected from 49 multiparous Thoroughbred mares at days 42–46 and 118–133 of gestation (first and second trimester). Pregnancies were monitored to term, and placentas were classified as normal (healthy) or abnormal. Complete data was available for 37 mares, and 13 were included in this study: normal ( n = 6), abnormal ( n = 5), and ascending placentitis ( n = 2). Total RNA underwent deep dual RNA‐sequencing. Alpha‐ and beta‐diversity metrics, differential expression, and microbe–host correlation analyses were performed. Results: Global vaginal microbial diversity did not differ between mares with healthy or abnormal placentas. Several microbes from the phyla Actinomycetota and Pseudomonadota showed altered activity in mares with abnormal placentas. Vaginal transcriptome showed a subtle inflammatory response in the second trimester in the abnormal placenta group in the absence of clinical signs. Correlation analysis suggested an interaction between bacterial survival and virulence genes and host inflammation and apoptosis genes. Main Limitations: Samples were obtained from clinical cases, limiting the availability of a complete history. Conclusions: Although overall vaginal microbial diversity was similar between outcome groups, distinct host vaginal transcriptional and microbial activity signatures distinguished mares with an abnormal placenta at birth. These findings warrant targeted investigation of the vaginal immune response and bacterial virulence factors as early biomarkers and therapeutic targets.

Background: Nocardioform placentitis (NP) is an understudied form of equine placentitis historically attributed to nocardioform bacteria, yet it remains uncertain whether these organisms are the sole pathogens involved. Objectives: To elucidate the pathophysiology of NP and the host–pathogen interaction. Study Design: In vivo clinical multi‐omics study. Methods: Dual RNA sequencing was performed to profile transcriptionally active microbial communities and concurrent placental transcriptome responses in samples from 31 placentas with and without NP. Untargeted metabolomics was performed to study the associated metabolites in the placenta. Results: The most abundant microbial transcripts belonged to Amycolatopsis , Crossiella , Lentzea , Enterococcus , and Mycobacterium. Bacterial gene expression in NP‐affected placentas was enriched in pathways related to ribosomal activity and metabolic processes involving amino acid, carbohydrate, and glycosphingolipid metabolism. Concurrently, placental transcripts demonstrated significant upregulation of inflammatory pathways and downregulation of pathways associated with blood vessel formation. Untargeted metabolomics highlighted an elevated abundance of metabolites such as beta‐D‐fucose, nervonic acid, and zymostenol in the placentitis samples. Significant correlations were found between microbial genes ( mraW , rlmB , amy , afuA , and cysC ) and host inflammation genes ( CXCL14 , IL15RA , TASL , and IFIH1 ). Additionally, elevated beta‐D‐fucose, a microbe‐specific metabolite, showed a strong correlation with microbial genes involved in stress‐adaptive metabolism and DNA repair ( ydhP , ybgC , serC , puuE , and radA ). The bacterial enzymes involved in beta‐D‐fucose were notably upregulated and predominantly expressed by Amycolatopsis and Lentzea . Main Limitations: Classification based on RNA abundance limited the number of Crossiella cases ( n = 3). Conclusions: Both nocardioform and non‐nocardioform bacteria are involved in NP‐diagnosed cases, challenging the current generalisation of the term ‘nocardioform placentitis’ and supporting the need to broaden diagnostic protocols for mucoid placentitis. Multi‐omics profiling revealed potential host–microbe interactions mediated by microbial metabolites, offering mechanistic insights and opportunities for improved diagnostic strategies.

The endometrium is a hormonally responsive tissue that undergoes cyclic remodeling. Although endometrial organoids have been established in several species, detailed characterization remains limited. Here, we assessed the structural and molecular fidelity of equine endometrial organoids across reproductive cycle stages and during extended culture. Organoids were generated from biopsies collected during estrus and diestrus and analyzed using histology, immunohistochemistry, electron microscopy, and bulk- and single-cell RNA sequencing. Organoids formed polarized cystic structures composed of columnar cells with microvilli, tight junctions, and secretory vesicles. Compared to native tissue, organoids showed higher expression of genes involved in proliferation and metabolism, and lower expression of genes related to differentiation, angiogenesis, and immune responses. Single-cell analysis identified diverse epithelial and stromal populations in both tissue and organoids. While most cell types were preserved, organoids were enriched in progenitor-like cells but underrepresented in ciliated, proliferative glandular, endothelial, smooth muscle, and antigen-presenting cells. Cycle-specific differences were observed in morphology, hormone receptor expression, and gene expression profiles. Estrus-derived organoids showed increased proliferation and metabolic activity. Although organoids retained transcriptional signatures reflective of the hormonal cycle stage of the source tissue, these signatures faded with prolonged culture, despite overall transcriptomic stability. In summary, equine endometrial organoids replicate key features of the native tissue, retain reproductive cycle characteristics, and maintain transcriptomic stability over time. Endometrial organoids provide a robust platform to study the equine endometrium, though native tissue differences should be considered in the experimental design and data interpretation.