Pub Date : 2025-12-25DOI: 10.1016/j.anireprosci.2025.108092
Hao Yu, Xiaotong Li, Xianyi Zhou, Jie Zhao, Wei Wang, Dagan Mao
This study aimed to investigate the effects of microRNA-150 (miR-150) on progesterone (P4) synthesis and apoptosis in goat luteal cells. Corpus luteum (CL) tissues were collected from non-pregnant goats. qPCR and in situ hybridization showed that miR-150 was highly expressed in the early CL but decreased in mid and late CL, and miR-150 was positively localized in luteal cells. Luteal cells from mid-cycle CL were then transfected with Agomir-150 (mimics) or Antagomir-150 (inhibitor). Immunoassays and qPCR results showed that Agomir-150 suppressed P4 levels and down-regulated the mRNA and protein expressions of hormone‑sensitive lipase (HSL), phosphorylated HSL (p-HSL), steroidogenic acute regulatory protein (StAR), and 3β‑hydroxysteroid dehydrogenase (HSD3B), whereas Antagomir-150 had the opposite effects. In addition, flow cytometry, qPCR, and Western blot results showed that Agomir-150 had no significant impact on cell apoptosis, while Antagomir-150 promoted apoptosis, up-regulated the mRNA and protein expressions of BCL2-associated X protein (BAX), Fas cell surface death receptor (FAS), tumor necrosis factor receptor 1 (TNFR1), Caspase-3, cleaved Caspase-3, and down-regulated B-cell lymphoma 2 (BCL-2) expression. Bioinformatic analysis identified nuclear receptor subfamily 4, group A, member 1 (NR4A1) as a potential miR-150 target. Dual-luciferase assays confirmed that miR-150 directly binds to the NR4A1–3′UTR and represses its expression, which was further validated by qPCR and Western blot. A rescue experiment revealed that NR4A1 knockdown partially reversed the Antagomir-150–induced up-regulation of P4 synthesis and apoptosis. Overall, this study demonstrates that miR-150 contributes to P4 synthesis and apoptosis in goat luteal cells by targeting NR4A1.
{"title":"MicroRNA-150 deficiency promotes progesterone synthesis and apoptosis in goat luteal cells by targeting nuclear receptor NR4A1","authors":"Hao Yu, Xiaotong Li, Xianyi Zhou, Jie Zhao, Wei Wang, Dagan Mao","doi":"10.1016/j.anireprosci.2025.108092","DOIUrl":"10.1016/j.anireprosci.2025.108092","url":null,"abstract":"<div><div>This study aimed to investigate the effects of microRNA-150 (miR-150) on progesterone (P4) synthesis and apoptosis in goat luteal cells. Corpus luteum (CL) tissues were collected from non-pregnant goats. qPCR and in situ hybridization showed that miR-150 was highly expressed in the early CL but decreased in mid and late CL, and miR-150 was positively localized in luteal cells. Luteal cells from mid-cycle CL were then transfected with Agomir-150 (mimics) or Antagomir-150 (inhibitor). Immunoassays and qPCR results showed that Agomir-150 suppressed P4 levels and down-regulated the mRNA and protein expressions of hormone‑sensitive lipase (HSL), phosphorylated HSL (p-HSL), steroidogenic acute regulatory protein (StAR), and 3β‑hydroxysteroid dehydrogenase (HSD3B), whereas Antagomir-150 had the opposite effects. In addition, flow cytometry, qPCR, and Western blot results showed that Agomir-150 had no significant impact on cell apoptosis, while Antagomir-150 promoted apoptosis, up-regulated the mRNA and protein expressions of BCL2-associated X protein (BAX), Fas cell surface death receptor (FAS), tumor necrosis factor receptor 1 (TNFR1), Caspase-3, cleaved Caspase-3, and down-regulated B-cell lymphoma 2 (BCL-2) expression. Bioinformatic analysis identified nuclear receptor subfamily 4, group A, member 1 (NR4A1) as a potential miR-150 target. Dual-luciferase assays confirmed that miR-150 directly binds to the NR4A1–3′UTR and represses its expression, which was further validated by qPCR and Western blot. A rescue experiment revealed that NR4A1 knockdown partially reversed the Antagomir-150–induced up-regulation of P4 synthesis and apoptosis. Overall, this study demonstrates that miR-150 contributes to P4 synthesis and apoptosis in goat luteal cells by targeting NR4A1.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"286 ","pages":"Article 108092"},"PeriodicalIF":3.3,"publicationDate":"2025-12-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145838938","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
By utilizing a comprehensive multi-modal approach that combined in silico modeling, in vitro molecular assays, in vivo microscopic observations, and on-farm phenotyping, we aimed to improve comprehension of the genetic architecture related to sheep litter size (LS). Our goal was to assess a T/C polymorphism (rs423810437) in the ovine Estrogen Receptor 2 (ESR2) gene and find potential LS biomarkers by applying network and circuit analysis. The sequencing of this polymorphism involved four populations, specifically the Romanov, Ghezel, Lori Bakhtiari, and their crossbreeds. With strong support for BMPR1B, GDF9, and ESR2, our analysis found more than ten candidate genes linked to LS. In purebred Romanov and Romanov-crossbred sheep, the ESR2 C allele was significantly correlated (p < 0.01) with increased LS. As our research suggests, the ESR2 polymorphism, among the identified candidate genes, is a valuable biomarker for litter size, offering understanding of follicular development in prolific sheep.
{"title":"Combined pathway-based biomarker discovery and ESR2 gene, polymorphism analysis of litter size prediction in sheep using a, multi model bioinformatics toolbox","authors":"Arash Javanmard , Mansour Yazdanyar , Mahdi Ebrahimi , Abbas Rafat , Karim Hasanpur , Babak Ghasemi Panahi , Sadegh Alijani , Habib Cheraghi , Mohammadreza Sheikhlou , Mohammadreza Mohammadabadi , Ali Esmailizadeh","doi":"10.1016/j.anireprosci.2025.108090","DOIUrl":"10.1016/j.anireprosci.2025.108090","url":null,"abstract":"<div><div>By utilizing a comprehensive multi-modal approach that combined <em>in silico</em> modeling, in vitro molecular assays, in vivo microscopic observations, and on-farm phenotyping, we aimed to improve comprehension of the genetic architecture related to sheep litter size (LS). Our goal was to assess a T/C polymorphism (rs423810437) in the ovine Estrogen Receptor 2 (ESR2) gene and find potential LS biomarkers by applying network and circuit analysis. The sequencing of this polymorphism involved four populations, specifically the Romanov, Ghezel, Lori Bakhtiari, and their crossbreeds. With strong support for BMPR1B, GDF9, and ESR2, our analysis found more than ten candidate genes linked to LS. In purebred Romanov and Romanov-crossbred sheep, the ESR2 C allele was significantly correlated (p < 0.01) with increased LS. As our research suggests, the ESR2 polymorphism, among the identified candidate genes, is a valuable biomarker for litter size, offering understanding of follicular development in prolific sheep.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"286 ","pages":"Article 108090"},"PeriodicalIF":3.3,"publicationDate":"2025-12-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145826813","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-20DOI: 10.1016/j.anireprosci.2025.108089
Agus Oman Sudrajat , Shofihar Sinansari , Harton Arfah , Eni Kusrini , Bastiar Nur , Darmawan Setia Budi , Odang Carman , Alimuddin Alimuddin
This study evaluated Synodontis eupterus (squeaker catfish) reproductive responses maintained under blue light exposure and different melatonin dietary levels for 56 days, with dosage: 0 mg/kg (M0), 0.5 mg/kg (M0.5), and 1 mg/kg (M1). Each treatment consisted of 12 individually maintained broodstock (6 individuals of each sex), with 90.15 ± 16.66 g (males) and 89.51 ± 16.65 g (females) initial body weights. Fish were reared individually in 75 × 60 × 60 cm tanks under continuous aeration and blue LED light (550 lux; 465–467 nm). Assessments were conducted using defined subsamples: GSI, HSI, and gonadal histology from 3 fish/sex/treatment; hormone profiles from 4 fish/sex/treatment; and gamete quality, fertilization, and hatching rates from 3 spawning pairs/treatment. Dietary melatonin influenced several reproductive parameters, with the M0.5 treatment showing higher female and male GSI (10.99 ± 4.05 % and 0.51 ± 0.03 %), egg diameter (0.49 ± 0.02 mm), and fecundity (8.73 ± 2.64 eggs/g). Sperm concentration, viability, and motility were also higher in M0.5. Broodstock receiving 0.5 mg/kg melatonin exhibited earlier peaks of estradiol-17β and testosterone. Fertilization, hatching, and early larval survival and growth were highest in M0.5. Responses in the M1 group were generally lower than those in M0.5 across several parameters; however, these differences were not consistently significant, and therefore no definitive inhibitory effect is inferred. Overall, the findings suggest that a moderate melatonin dose (0.5 mg/kg) under blue light supports improvements in gonadal maturation, gamete quality, and early larval performance in S. eupterus, warranting further validation with larger experimental replication.
{"title":"Reproductive responses of Synodontis eupterus to different dietary melatonin levels under blue light conditions","authors":"Agus Oman Sudrajat , Shofihar Sinansari , Harton Arfah , Eni Kusrini , Bastiar Nur , Darmawan Setia Budi , Odang Carman , Alimuddin Alimuddin","doi":"10.1016/j.anireprosci.2025.108089","DOIUrl":"10.1016/j.anireprosci.2025.108089","url":null,"abstract":"<div><div>This study evaluated <em>Synodontis eupterus</em> (squeaker catfish) reproductive responses maintained under blue light exposure and different melatonin dietary levels for 56 days, with dosage: 0 mg/kg (M0), 0.5 mg/kg (M0.5), and 1 mg/kg (M1). Each treatment consisted of 12 individually maintained broodstock (6 individuals of each sex), with 90.15 ± 16.66 g (males) and 89.51 ± 16.65 g (females) initial body weights. Fish were reared individually in 75 × 60 × 60 cm tanks under continuous aeration and blue LED light (550 lux; 465–467 nm). Assessments were conducted using defined subsamples: GSI, HSI, and gonadal histology from 3 fish/sex/treatment; hormone profiles from 4 fish/sex/treatment; and gamete quality, fertilization, and hatching rates from 3 spawning pairs/treatment. Dietary melatonin influenced several reproductive parameters, with the M0.5 treatment showing higher female and male GSI (10.99 ± 4.05 % and 0.51 ± 0.03 %), egg diameter (0.49 ± 0.02 mm), and fecundity (8.73 ± 2.64 eggs/g). Sperm concentration, viability, and motility were also higher in M0.5. Broodstock receiving 0.5 mg/kg melatonin exhibited earlier peaks of estradiol-17β and testosterone. Fertilization, hatching, and early larval survival and growth were highest in M0.5. Responses in the M1 group were generally lower than those in M0.5 across several parameters; however, these differences were not consistently significant, and therefore no definitive inhibitory effect is inferred. Overall, the findings suggest that a moderate melatonin dose (0.5 mg/kg) under blue light supports improvements in gonadal maturation, gamete quality, and early larval performance in <em>S. eupterus</em>, warranting further validation with larger experimental replication.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"286 ","pages":"Article 108089"},"PeriodicalIF":3.3,"publicationDate":"2025-12-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145801625","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-18DOI: 10.1016/j.anireprosci.2025.108088
Paulo Victor dos S. Pereira , Ana Paula da S. Cupello , Lucas Francisco L. Correia , Walter Lilenbaum , Joanna M.G. Souza-Fabjan
Although a few studies have reported Leptospira spp. in cumulus-oocyte complexes (COCs) from experimentally infected cows, it remains unclear whether this bacterium can penetrate COCs in naturally infected animals. This study aimed to detect the presence of Leptospira spp. in COCs from naturally infected cows. Ovaries and uterine body fragments were collected from 40 cows after slaughter. Follicular fluid (FF) and uterine fragments (UF) were screened using conventional PCR (cPCR), and negative results were confirmed by quantitative PCR (qPCR). COCs were immunolabeled using a monoclonal anti-LipL41 antibody, followed by an Alexa Fluor 488-conjugated secondary antibody, and counterstained with propidium iodide. Detection of Leptospira spp. was performed by epifluorescence microscopy. Cows were then classified as positive (POS-FF, POS-UF, or both) or negative (NEG). In POS-FF animals (n = 8), 9.1 COCs/female were recovered, with 51.1 ± 12.7 % showing Leptospira presence (not significant; n.s.). In POS-UF cows (n = 12), 6.6 COCs/female were obtained, and 81.0 ± 5.5 % showed bacterial presence (n.s.). In animals positive for both FF and UF (n = 10), 5.4 COCs/female were collected, with 63.7 ± 12.9 % testing positive (n.s.). In NEG animals (n = 10), which were both cPCR- and qPCR-negative, 69.3 ± 9.7 % of COCs exhibited bacterial labeling, suggesting a low bacterial load (n.s.). These cows yielded 9.7 ± 3.7 COCs/female (n.s.). Although no difference was observed between the groups, this is the first report demonstrating the presence of Leptospira spp. in COCs from naturally infected cows, highlighting a mechanism contributing to reproductive failure in cattle.
{"title":"First detection of Leptospira spp. in cumulus-oocyte complexes from naturally infected cows","authors":"Paulo Victor dos S. Pereira , Ana Paula da S. Cupello , Lucas Francisco L. Correia , Walter Lilenbaum , Joanna M.G. Souza-Fabjan","doi":"10.1016/j.anireprosci.2025.108088","DOIUrl":"10.1016/j.anireprosci.2025.108088","url":null,"abstract":"<div><div>Although a few studies have reported <em>Leptospira</em> spp. in cumulus-oocyte complexes (COCs) from experimentally infected cows, it remains unclear whether this bacterium can penetrate COCs in naturally infected animals. This study aimed to detect the presence of <em>Leptospira</em> spp. in COCs from naturally infected cows. Ovaries and uterine body fragments were collected from 40 cows after slaughter. Follicular fluid (FF) and uterine fragments (UF) were screened using conventional PCR (cPCR), and negative results were confirmed by quantitative PCR (qPCR). COCs were immunolabeled using a monoclonal anti-<em>Lip</em>L41 antibody, followed by an Alexa Fluor 488-conjugated secondary antibody, and counterstained with propidium iodide. Detection of <em>Leptospira</em> spp. was performed by epifluorescence microscopy. Cows were then classified as positive (POS-FF, POS-UF, or both) or negative (NEG). In POS-FF animals (n = 8), 9.1 COCs/female were recovered, with 51.1 ± 12.7 % showing <em>Leptospira</em> presence (not significant; n.s.). In POS-UF cows (n = 12), 6.6 COCs/female were obtained, and 81.0 ± 5.5 % showed bacterial presence (n.s.). In animals positive for both FF and UF (n = 10), 5.4 COCs/female were collected, with 63.7 ± 12.9 % testing positive (n.s.). In NEG animals (n = 10), which were both cPCR- and qPCR-negative, 69.3 ± 9.7 % of COCs exhibited bacterial labeling, suggesting a low bacterial load (n.s.). These cows yielded 9.7 ± 3.7 COCs/female (n.s.). Although no difference was observed between the groups, this is the first report demonstrating the presence of <em>Leptospira</em> spp. in COCs from naturally infected cows, highlighting a mechanism contributing to reproductive failure in cattle.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108088"},"PeriodicalIF":3.3,"publicationDate":"2025-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145788243","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-18DOI: 10.1016/j.anireprosci.2025.108087
Diego A. Galarza , Mauricio Duma , Gissela Sánchez-Pacheco , María García-Pacheco , Camila Ramón-Barrera , Gilda Sanmartín-Ordóñez , Manuel Soria , Antonio J. Vallecillo
HSPA1A is a heat shock protein belonging to the HSP70 family that acts during cryopreservation by mitigating oxidative stress, stabilizing denatured proteins, preventing their aggregation, and maintaining proteostasis. This study evaluated the effect of recombinant HSPA1A protein on the cryosurvival of dog epididymal spermatozoa subjected to slow-freezing (SF) and kinetic vitrification (VIT). Twenty adult dogs were orchiectomized, and epididymal sperm from the left testes were cryopreserved with SF, while the right ones were with VIT. Four treatments were established according to the addition of 10 µg/ml (+HSPA1A) or 0 µg/ml (-Control) of recombinant bovine HSPA1A, produced using Escherichia coli Rosetta 2 (DE3): pET15b-HSPA1A strain and IMAC purified. The different cryopreservation treatments evaluated were SF+HSPA1A, SF-Control, VIT+HSPA1A, and VIT-Control (n = 20 samples per treatment). Samples from SF+HSPA1A and SF-Control treatments were frozen in 0.25 ml straws by exposing them to liquid nitrogen (LN₂) vapors. For VIT, samples were vitrified by direct immersion of 30 µl drops into LN₂ and stored in cryotubes. Results showed that SF+HSPA1A and SF-Control yielded significantly higher (P < 0.05) progressive motility, velocities, linearity and straightness, and beat cross frequency compared to VIT+HSPA1A and VIT-Control. Furthermore, sperm treated with HSPA1A (both SF and VIT) exhibited significantly improved (P < 0.05) motility, viability and DNA integrity compared to their respective controls. In conclusion, the addition of recombinant HSPA1A to both slow-freezing and vitrification media enhanced the post-cryopreserved quality of dog epididymal spermatozoa, demonstrating its protective effect on motility, viability, and integrity of acrosome and DNA.
{"title":"Recombinant heat shock protein HSPA1A enhances the cryosurvival of frozen-thawed or vitrified-warmed dog epididymal spermatozoa","authors":"Diego A. Galarza , Mauricio Duma , Gissela Sánchez-Pacheco , María García-Pacheco , Camila Ramón-Barrera , Gilda Sanmartín-Ordóñez , Manuel Soria , Antonio J. Vallecillo","doi":"10.1016/j.anireprosci.2025.108087","DOIUrl":"10.1016/j.anireprosci.2025.108087","url":null,"abstract":"<div><div>HSPA1A is a heat shock protein belonging to the HSP70 family that acts during cryopreservation by mitigating oxidative stress, stabilizing denatured proteins, preventing their aggregation, and maintaining proteostasis. This study evaluated the effect of recombinant HSPA1A protein on the cryosurvival of dog epididymal spermatozoa subjected to slow-freezing (SF) and kinetic vitrification (VIT). Twenty adult dogs were orchiectomized, and epididymal sperm from the left testes were cryopreserved with SF, while the right ones were with VIT. Four treatments were established according to the addition of 10 µg/ml (+HSPA1A) or 0 µg/ml (-Control) of recombinant bovine HSPA1A, produced using <em>Escherichia coli</em> Rosetta 2 (DE3): pET15b-HSPA1A strain and IMAC purified. The different cryopreservation treatments evaluated were SF+HSPA1A, SF-Control, VIT+HSPA1A, and VIT-Control (n = 20 samples per treatment). Samples from SF+HSPA1A and SF-Control treatments were frozen in 0.25 ml straws by exposing them to liquid nitrogen (LN₂) vapors. For VIT, samples were vitrified by direct immersion of 30 µl drops into LN₂ and stored in cryotubes. Results showed that SF+HSPA1A and SF-Control yielded significantly higher (P < 0.05) progressive motility, velocities, linearity and straightness, and beat cross frequency compared to VIT+HSPA1A and VIT-Control. Furthermore, sperm treated with HSPA1A (both SF and VIT) exhibited significantly improved (P < 0.05) motility, viability and DNA integrity compared to their respective controls. In conclusion, the addition of recombinant HSPA1A to both slow-freezing and vitrification media enhanced the post-cryopreserved quality of dog epididymal spermatozoa, demonstrating its protective effect on motility, viability, and integrity of acrosome and DNA.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108087"},"PeriodicalIF":3.3,"publicationDate":"2025-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145788275","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-15DOI: 10.1016/j.anireprosci.2025.108078
Xiaofeng Li , Min Liu , Hui Xiang , Linsen Gui , Yun Ma , Xingang Dan
Probiotics, particularly composite lactic acid bacteria (CLAB), are emerging as a potential alternative to antibiotics for managing bovine metritis. While prior studies have highlighted CLAB's anti-inflammatory effects, limited data exist on its impact on vaginal and uterine microbiomes and systemic metabolism. In this study, Holstein cows received deep vaginal infusions of CLAB (a blend of Lactobacillus rhamnosus, Pediococcus acidilactici and Lactobacillus reuteri; at a standardized 4.5 × 1010 CFU/dose and a proportion of 25/25/2, respectively) at 20 and 10 days prepartum. We analyzed changes in vaginal and uterine microbiota, plasma metabolomes, and assessed the incidence of metritis, conception rate, and lactation performance postpartum. CLAB administration significantly altered the vaginal microbiota by reducing opportunistic pathogens (Prevotella heparinolytica, Bacteroides and Fusobacteria) and promoting beneficial taxa (Akkermansia and Prevotellaceae). In the uterus, CLAB enriched Rikenellaceae, Christensenellaceae and Lachnospiraceae, while suppressing pathogenic genera such as Cutibacterium and Fournierella. Metabolomic analysis identified pyruvic acid, L-glutamine and L-valine as hub metabolites, with KEGG enrichment revealing involvement in amino acid metabolism and immunomodulatory pathways. Although CLAB infusion did not significantly reduce metritis incidence (5.00 % vs. 5.52 %) or improve conception rate (47.50 % vs. 45.00 %), it showed promising trends without affecting milk production. These findings suggest that prepartum vaginal CLAB infusion modulates reproductive tract microbiota and systemic metabolism, potentially contributing to uterine health maintenance in dairy cows. A key limitation of this study was the absence of significant reductions in metritis incidence and improvements in conception rates, likely attributable to the small sample size and limited study period. Further large-scale studies are warranted to validate its efficacy during high-risk seasons.
益生菌，特别是复合乳酸菌（CLAB），正在成为治疗牛乳酸菌炎的潜在替代抗生素。虽然先前的研究强调了CLAB的抗炎作用，但关于其对阴道和子宫微生物群以及全身代谢的影响的数据有限。在本研究中，荷斯坦奶牛在准备20天和10天阴道深度注射CLAB（鼠李糖乳杆菌、酸乳酸球球菌和罗伊氏乳杆菌的混合物，标准剂量为4.5 × 1010 CFU/，比例分别为25/25/2）。我们分析了阴道和子宫微生物群、血浆代谢组的变化，并评估了子宫炎的发生率、受孕率和产后泌乳表现。CLAB通过减少机会致病菌（溶肝普雷沃氏菌、拟杆菌和梭杆菌）和促进有益类群（Akkermansia和普雷沃氏菌科）显著改变阴道微生物群。CLAB在子宫内富集Rikenellaceae、Christensenellaceae和Lachnospiraceae，同时抑制Cutibacterium和Fournierella等致病属。代谢组学分析发现，丙酮酸、l -谷氨酰胺和l -缬氨酸是枢纽代谢物，KEGG的富集揭示了氨基酸代谢和免疫调节途径的参与。虽然CLAB输注没有显著降低子宫炎发生率（5.00 % vs. 5.52 %）或提高受孕率（47.50 % vs. 45.00 %），但在不影响产奶量的情况下，它显示出很好的趋势。这些结果表明，阴道前注射CLAB可调节生殖道微生物群和全身代谢，可能有助于奶牛子宫健康的维持。本研究的一个关键限制是子宫内膜炎发病率没有显著降低和受孕率的提高，可能是由于样本量小和研究时间有限。需要进一步的大规模研究来验证其在高危季节的有效性。
{"title":"Integrated microbiome and metabolome revealing new insight into the combination of lactic acid bacteria in preventing postpartum metritis of dairy cows","authors":"Xiaofeng Li , Min Liu , Hui Xiang , Linsen Gui , Yun Ma , Xingang Dan","doi":"10.1016/j.anireprosci.2025.108078","DOIUrl":"10.1016/j.anireprosci.2025.108078","url":null,"abstract":"<div><div>Probiotics, particularly composite lactic acid bacteria (CLAB), are emerging as a potential alternative to antibiotics for managing bovine metritis. While prior studies have highlighted CLAB's anti-inflammatory effects, limited data exist on its impact on vaginal and uterine microbiomes and systemic metabolism. In this study, Holstein cows received deep vaginal infusions of CLAB (a blend of <em>Lactobacillus rhamnosus</em>, <em>Pediococcus acidilactici</em> and <em>Lactobacillus reuteri</em>; at a standardized 4.5 × 10<sup>10</sup> CFU/dose and a proportion of 25/25/2, respectively) at 20 and 10 days prepartum. We analyzed changes in vaginal and uterine microbiota, plasma metabolomes, and assessed the incidence of metritis, conception rate, and lactation performance postpartum. CLAB administration significantly altered the vaginal microbiota by reducing opportunistic pathogens (<em>Prevotella heparinolytica</em>, <em>Bacteroides</em> and <em>Fusobacteria</em>) and promoting beneficial taxa (<em>Akkermansia</em> and <em>Prevotellaceae</em>). In the uterus, CLAB enriched <em>Rikenellaceae</em>, <em>Christensenellaceae</em> and <em>Lachnospiraceae</em>, while suppressing pathogenic genera such as <em>Cutibacterium</em> and <em>Fournierella</em>. Metabolomic analysis identified pyruvic acid, <span>L</span>-glutamine and <span>L</span>-valine as hub metabolites, with KEGG enrichment revealing involvement in amino acid metabolism and immunomodulatory pathways. Although CLAB infusion did not significantly reduce metritis incidence (5.00 % vs. 5.52 %) or improve conception rate (47.50 % vs. 45.00 %), it showed promising trends without affecting milk production. These findings suggest that prepartum vaginal CLAB infusion modulates reproductive tract microbiota and systemic metabolism, potentially contributing to uterine health maintenance in dairy cows. A key limitation of this study was the absence of significant reductions in metritis incidence and improvements in conception rates, likely attributable to the small sample size and limited study period. Further large-scale studies are warranted to validate its efficacy during high-risk seasons.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108078"},"PeriodicalIF":3.3,"publicationDate":"2025-12-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145772904","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-13DOI: 10.1016/j.anireprosci.2025.108077
César Augusto Pinzón-Osorio , Thomaz Kranen Cunha , German Holguin-Sanabria , Marco Alves Machado , Favorino José de Freitas Collares , Luís Henrique de Aguiar , Paula Rodriguez-Villamil , Felipe Ledur Ongaratto , Eduardo Pradebom da Silva , Eduardo de Oliveira Sanguinet , Carolina Silveira da Silva , Daiane Mentz , Karine Campagnolo , Paula Viero Marchioretto , José Martin Klafke , Luís Fernando Schutz , Fernando Caetano de Oliveira , José Luiz Rodrigues , Marcelo Bertolini
Timed artificial insemination (TAI) protocols based on estradiol and progesterone (E2/P4) typically yield pregnancy rates (PR) near 50 % in beef cattle, falling short of the 65–70 % expected in fertile females. This study investigated physiological and procedural limitations that may compromise TAI success due to suboptimal synchrony between insemination and ovulation. Three experiments involving 840 suckled crossbred females evaluated follicular responses, semen type, and alternative reproductive strategies to enhance fertility. In Experiment I (n = 28), daily ultrasonography revealed that 46.4 % of females exhibited suboptimal ovarian responses, defined as no emergence/atresia of the new follicular wave, and ovulation delay or failure. In Experiment II (n = 348), cows with small follicles (<8.5 mm) at P4 removal had markedly higher PR when inseminated with cooled versus frozen semen (71.9 % vs 38.0 %), highlighting the relevance of sperm cell viability when ovulation is delayed. In Experiment III (n = 464), strategies combining TAI with estrus detection (ED), timed embryo transfer (TET), or both significantly increased PR compared to TAI alone (TAI: 53.6 %; TAI+TET: 67.9 %; ED-AI/TAI: 70.5 %; ED-AI/TAI+TET: 68.2 %), which was accompanied by increased twinning rates after TET, suggesting that many TAI failures were not due to uterine inadequacy or embryo mortality but to asynchrony in gamete encounter. Collectively, results suggest that most TAI failures stem from impaired synchrony between ovulation and sperm cell availability rather than uterine or embryonic limitations. Incorporating tools such as ultrasonography, semen selection, ED, or TET may enhance reproductive outcomes in commercial beef herds under E2/P4-based TAI protocols.
{"title":"Suboptimal responses to E2/P4-based timed AI protocols compromise pregnancy outcomes in fertile beef cattle: Roles of ovulation timing and semen viability","authors":"César Augusto Pinzón-Osorio , Thomaz Kranen Cunha , German Holguin-Sanabria , Marco Alves Machado , Favorino José de Freitas Collares , Luís Henrique de Aguiar , Paula Rodriguez-Villamil , Felipe Ledur Ongaratto , Eduardo Pradebom da Silva , Eduardo de Oliveira Sanguinet , Carolina Silveira da Silva , Daiane Mentz , Karine Campagnolo , Paula Viero Marchioretto , José Martin Klafke , Luís Fernando Schutz , Fernando Caetano de Oliveira , José Luiz Rodrigues , Marcelo Bertolini","doi":"10.1016/j.anireprosci.2025.108077","DOIUrl":"10.1016/j.anireprosci.2025.108077","url":null,"abstract":"<div><div>Timed artificial insemination (TAI) protocols based on estradiol and progesterone (E2/P4) typically yield pregnancy rates (PR) near 50 % in beef cattle, falling short of the 65–70 % expected in fertile females. This study investigated physiological and procedural limitations that may compromise TAI success due to suboptimal synchrony between insemination and ovulation. Three experiments involving 840 suckled crossbred females evaluated follicular responses, semen type, and alternative reproductive strategies to enhance fertility. In Experiment I (<em>n</em> = 28), daily ultrasonography revealed that 46.4 % of females exhibited suboptimal ovarian responses, defined as no emergence/atresia of the new follicular wave, and ovulation delay or failure. In Experiment II (<em>n</em> = 348), cows with small follicles (<8.5 mm) at P4 removal had markedly higher PR when inseminated with cooled versus frozen semen (71.9 % <em>vs</em> 38.0 %), highlighting the relevance of sperm cell viability when ovulation is delayed. In Experiment III (<em>n</em> = 464), strategies combining TAI with estrus detection (ED), timed embryo transfer (TET), or both significantly increased PR compared to TAI alone (TAI: 53.6 %; TAI+TET: 67.9 %; ED-AI/TAI: 70.5 %; ED-AI/TAI+TET: 68.2 %), which was accompanied by increased twinning rates after TET, suggesting that many TAI failures were not due to uterine inadequacy or embryo mortality but to asynchrony in gamete encounter. Collectively, results suggest that most TAI failures stem from impaired synchrony between ovulation and sperm cell availability rather than uterine or embryonic limitations. Incorporating tools such as ultrasonography, semen selection, ED, or TET may enhance reproductive outcomes in commercial beef herds under E2/P4-based TAI protocols.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108077"},"PeriodicalIF":3.3,"publicationDate":"2025-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145766934","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-13DOI: 10.1016/j.anireprosci.2025.108076
Emily A. Lugo , James K. Graham , James D. Gillis , Jennifer P. Barfield
Domestic cats (Felis catus) serve as a model for studying non-domestic cat reproduction. In particular, assisted reproductive technologies, such as cryopreservation of gametes and embryos, often are developed in domestic species and then applied to non-domestic species. Membrane composition and osmotic tolerance impact a sperm’s ability to withstand the stresses of cryopreservation. In this study, the membrane cholesterol content and osmotic tolerance range of domestic field spermatozoa were investigated and considered when testing alternative cryoprotectants and methods (addition of cholesterol) for felid sperm cryopreservation. The most commonly used cryoprotectant for felid sperm, glycerol, was compared to methylformamide (MF), dimethylformamide (DMF), and combinations of glycerol with both MF and DMF. To verify osmotic tolerance range, domestic cat sperm were subjected to 12 different anisosmotic solutions, ranging from 0 to 1200 mOsm/kg. Using flow cytometry, > 65 % of the sperm maintained intact membranes between 50 and 600 mOsm/kg, verifying that epididymal domestic cat sperm have a wide osmotic tolerance range. In addition, epididymal cat sperm have a relatively high cholesterol:phospholipid ratio (0.70) and attempting to increase the cholesterol content did not benefit cryosurvival. The alternative cryoprotectant DMF, however, did improve post-thaw percentages of motile sperm (43 %) compared to glycerol (24 %; P < 0.05), with 8 % DMF resulting in higher post-thaw motility than sperm frozen with 5 % DMF (33 vs 54 %; P < 0.05). These findings highlight physiological properties of felid sperm that should be considered when updating protocols to improve cat sperm cryopreservation methods.
{"title":"Investigating the plasma membrane composition and osmotic tolerance of domestic felid (Felis catus) spermatozoa as a model for non-domestic felids","authors":"Emily A. Lugo , James K. Graham , James D. Gillis , Jennifer P. Barfield","doi":"10.1016/j.anireprosci.2025.108076","DOIUrl":"10.1016/j.anireprosci.2025.108076","url":null,"abstract":"<div><div>Domestic cats (<em>Felis catus</em>) serve as a model for studying non-domestic cat reproduction. In particular, assisted reproductive technologies, such as cryopreservation of gametes and embryos, often are developed in domestic species and then applied to non-domestic species. Membrane composition and osmotic tolerance impact a sperm’s ability to withstand the stresses of cryopreservation. In this study, the membrane cholesterol content and osmotic tolerance range of domestic field spermatozoa were investigated and considered when testing alternative cryoprotectants and methods (addition of cholesterol) for felid sperm cryopreservation. The most commonly used cryoprotectant for felid sperm, glycerol, was compared to methylformamide (MF), dimethylformamide (DMF), and combinations of glycerol with both MF and DMF. To verify osmotic tolerance range, domestic cat sperm were subjected to 12 different anisosmotic solutions, ranging from 0 to 1200 mOsm/kg. Using flow cytometry, <u>></u> 65 % of the sperm maintained intact membranes between 50 and 600 mOsm/kg, verifying that epididymal domestic cat sperm have a wide osmotic tolerance range. In addition, epididymal cat sperm have a relatively high cholesterol:phospholipid ratio (0.70) and attempting to increase the cholesterol content did not benefit cryosurvival. The alternative cryoprotectant DMF, however, did improve post-thaw percentages of motile sperm (43 %) compared to glycerol (24 %; <em>P</em> < 0.05), with 8 % DMF resulting in higher post-thaw motility than sperm frozen with 5 % DMF (33 vs 54 %; <em>P</em> < 0.05). These findings highlight physiological properties of felid sperm that should be considered when updating protocols to improve cat sperm cryopreservation methods.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108076"},"PeriodicalIF":3.3,"publicationDate":"2025-12-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145766983","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Sperm preservation is a pivotal technique in reproductive science, facilitating the long-term preservation and utilization of valuable genetic material, particularly in animal breeding programs and assisted reproductive technologies (ART). However, the freezing-thawing process imposes significant physiological stress on sperm cells, primarily due to oxidative damage. This oxidative stress disrupts critical cellular functions, leading to reduced sperm motility, viability, DNA integrity, and membrane stability, thereby compromising overall reproductive potential. Among various antioxidant strategies, lycopene, a potent carotenoid, has emerged as a promising candidate for mitigating cryoinjury. This review provides a comprehensive overview of lycopene’s role in sperm cryopreservation and cold storage, emphasizing its effect on post-thaw sperm quality. Experimental evidence from animal studies indicates that lycopene supplementation effectively neutralizes reactive oxygen species, reduces lipid peroxidation, and preserves structural integrity by integrating into cell membranes. These protective effects contribute to enhancing sperm functionality post-thaw, potentially improving fertilization outcomes. Yet, this review critically highlights that the efficacy of lycopene is a double-edged sword: its effects are highly dose- and species-dependent, with excessive concentrations paradoxically impairing sperm performance. We conclude that a 'one-size-fits-all' approach is ineffective, and future investigations must move beyond simple supplementation to focus on optimizing species-specific formulations and validating in vitro benefits with in vivo fertility trials to fully harness lycopene's potential in reproductive applications.
{"title":"Lycopene as a protective antioxidant in sperm preservation","authors":"Mohsen Shayestehyekta , Azita Faramarzi , Zahra Rashidi , Mojtaba Moradi","doi":"10.1016/j.anireprosci.2025.108075","DOIUrl":"10.1016/j.anireprosci.2025.108075","url":null,"abstract":"<div><div>Sperm preservation is a pivotal technique in reproductive science, facilitating the long-term preservation and utilization of valuable genetic material, particularly in animal breeding programs and assisted reproductive technologies (ART). However, the freezing-thawing process imposes significant physiological stress on sperm cells, primarily due to oxidative damage. This oxidative stress disrupts critical cellular functions, leading to reduced sperm motility, viability, DNA integrity, and membrane stability, thereby compromising overall reproductive potential. Among various antioxidant strategies, lycopene, a potent carotenoid, has emerged as a promising candidate for mitigating cryoinjury. This review provides a comprehensive overview of lycopene’s role in sperm cryopreservation and cold storage, emphasizing its effect on post-thaw sperm quality. Experimental evidence from animal studies indicates that lycopene supplementation effectively neutralizes reactive oxygen species, reduces lipid peroxidation, and preserves structural integrity by integrating into cell membranes. These protective effects contribute to enhancing sperm functionality post-thaw, potentially improving fertilization outcomes. Yet, this review critically highlights that the efficacy of lycopene is a double-edged sword: its effects are highly dose- and species-dependent, with excessive concentrations paradoxically impairing sperm performance. We conclude that a 'one-size-fits-all' approach is ineffective, and future investigations must move beyond simple supplementation to focus on optimizing species-specific formulations and validating in vitro benefits with in vivo fertility trials to fully harness lycopene's potential in reproductive applications.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108075"},"PeriodicalIF":3.3,"publicationDate":"2025-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145754729","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-09DOI: 10.1016/j.anireprosci.2025.108074
Darmawan Setia Budi , Agus Priyadi , Asep Permana , Ikhsan Khasani , Tamás Müller , Ahmad Shofy Mubarak , Imam Mustofa
Habitat degradation, overfishing, pollution, and climate change threaten many fish species, necessitating urgent conservation efforts. Captive breeding is essential for fish species recovery and genetic diversity preservation, with maturity determination being crucial for broodstock management, breeding success, and sustainable aquaculture practices. Standardized, species-specific protocols and non-invasive methods for accurately determining fish maturity are currently lacking. This review synthesizes current methodologies and practical applications of maturity determination in fish captive breeding, emphasizing how established knowledge can be applied to improve broodstock management and conservation. Traditional methods, like morphological and gonadal assessments, remain foundational, while histological and molecular techniques offer enhanced precision but are resource-intensive. Non-invasive imaging methods, including ultrasound, provide innovative alternatives for evaluating gonadal development. Behavioral cues, such as aggression, shoaling, and nesting, complement physiological assessments, enhancing accuracy. A multi-method approach is essential to address the diverse reproductive strategies of fish species. Advances in camera-based technologies and diagnostic kits for hormone detection offer novel, non-invasive, and scalable maturity assessments, reducing stress on fish. The integration of maturity determination with aquaculture facilitates broodstock selection, stock enhancement, and conservation efforts, promoting the long-term sustainability of aquatic ecosystems. This review highlights the need for standardized, species-specific protocols and technological advancements to support sustainable fish breeding. Future research should prioritize developing accessible tools and methodologies that integrate traditional and modern approaches, enhancing biodiversity conservation and promoting aquaculture growth.
{"title":"Maturity determination and its potential application for fish captive breeding","authors":"Darmawan Setia Budi , Agus Priyadi , Asep Permana , Ikhsan Khasani , Tamás Müller , Ahmad Shofy Mubarak , Imam Mustofa","doi":"10.1016/j.anireprosci.2025.108074","DOIUrl":"10.1016/j.anireprosci.2025.108074","url":null,"abstract":"<div><div>Habitat degradation, overfishing, pollution, and climate change threaten many fish species, necessitating urgent conservation efforts. Captive breeding is essential for fish species recovery and genetic diversity preservation, with maturity determination being crucial for broodstock management, breeding success, and sustainable aquaculture practices. Standardized, species-specific protocols and non-invasive methods for accurately determining fish maturity are currently lacking. This review synthesizes current methodologies and practical applications of maturity determination in fish captive breeding, emphasizing how established knowledge can be applied to improve broodstock management and conservation. Traditional methods, like morphological and gonadal assessments, remain foundational, while histological and molecular techniques offer enhanced precision but are resource-intensive. Non-invasive imaging methods, including ultrasound, provide innovative alternatives for evaluating gonadal development. Behavioral cues, such as aggression, shoaling, and nesting, complement physiological assessments, enhancing accuracy. A multi-method approach is essential to address the diverse reproductive strategies of fish species. Advances in camera-based technologies and diagnostic kits for hormone detection offer novel, non-invasive, and scalable maturity assessments, reducing stress on fish. The integration of maturity determination with aquaculture facilitates broodstock selection, stock enhancement, and conservation efforts, promoting the long-term sustainability of aquatic ecosystems. This review highlights the need for standardized, species-specific protocols and technological advancements to support sustainable fish breeding. Future research should prioritize developing accessible tools and methodologies that integrate traditional and modern approaches, enhancing biodiversity conservation and promoting aquaculture growth.</div></div>","PeriodicalId":7880,"journal":{"name":"Animal Reproduction Science","volume":"285 ","pages":"Article 108074"},"PeriodicalIF":3.3,"publicationDate":"2025-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145735570","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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