Search Results (1,102)

This review aims to discuss how heat stress affects ovarian follicles and oocytes, steroidogenesis, and embryo development in ruminants. The literature shows that quiescent primordial follicles appear to be less susceptible to heat stress, but from the primary follicle stage onwards, they begin to suffer the consequences of heat stress. These adverse effects are exacerbated when the follicles are cultured in vitro. In antral follicles, heat stress reduces granulosa cell viability and proliferation in both in vivo and in vitro models. Oocyte maturation, both nuclear and cytoplasmic, is also compromised, and embryo quality declines under elevated thermal conditions. These effects are linked to intracellular disturbances, including oxidative imbalance, mitochondrial dysfunction, and altered hormonal signaling. The differences between in vivo and in vitro responses reflect the complexity of the biological impact of heat stress and emphasize the protective role of the physiological microenvironment. A better understanding of how heat stress alters the function of ovarian follicles, oocytes, and embryos is crucial. This knowledge is critical to devise effective strategies that mitigate damage, support fertility, and improve outcomes in assisted reproduction for livestock exposed to high environmental temperatures. Full article

In remote alpaca breeding regions, access to advanced sperm analysis laboratories is limited. This study validates a practical cytometric method for evaluating sperm viability and acrosomal integrity in epididymal alpaca sperm using early fluorochrome staining, formaldehyde fixation, and intermediate storage. Thirty-two testes were transported at 5 °C, and spermatozoa were collected from the cauda epididymis. After morphometric screening, 26 samples were included. Aliquots were stained with Zombie Green (viability) and FITC–PSA (acrosomal integrity), at time zero. Each aliquot was divided for cytometric analysis at T0 (immediately), T24 (24 h after formaldehyde fixation) and T1w (1 week post-fixation). Fixed samples showed higher viability and acrosomal integrity values (T24: 70.75%, 97.24%; T1w: 71.80%, 97.21%) than T0 (67.63%, 95.89%). This may reflect fluorescence alterations associated with fixation. Strong correlations and Bland–Altman analysis confirmed consistency across time points. This method enables accurate sperm quality evaluation up to one week after collection, offering a useful tool for reproductive monitoring in field conditions without immediate analysis. Further research on ejaculated semen and field protocols is recommended. Full article

This study aimed to determine the factors associated with the occurrence of the first-wave dominant follicle (DF) in the ovary ipsilateral to the corpus luteum (CL) in lactating dairy cows and dairy heifers. A total of 505 estruses were investigated (lactating dairy cows, n = 361; dairy heifers, n = 144). The locations of the preovulatory follicle (PF) and regressed CL were examined at the estrus, and the locations of the first-wave dominant follicle (DF) and newly formed CL were examined seven days after estrus using transrectal ultrasonography. Then, cows were classified into two groups: the first-wave DF in the ovary ipsilateral to the CL (IG) and the first-wave DF in the ovary contralateral to the CL (CG). To evaluate the factors which affect the occurrence of IG and CG, binominal logistic regression analysis was conducted; the location of the PF and regressing CL, season (warm: June–September; cool: October–May), live weight, days in milk at estrus, daily milk production, and body condition score were used as independent variables. The occurrence rate of IG was significantly higher when the PF was located contralateral to the regressing CL (lactating dairy cows, 63.4%; dairy heifers, 58.6%) rather than ipsilateral (lactating dairy cows, 44.9%; dairy heifers, 35.1%). The IG occurrence rate was significantly higher with an increase in daily milk production (<30 kg, 47.3%; 30–40 kg, 55.2%; >40 kg, 60.5%) in lactating dairy cows. In conclusion, the occurrence of IG was associated with relative locations of the PF and regressing CL in lactating dairy cows and dairy heifers and with the level of milk production in lactating dairy cows. Full article

Mitochondria are central to energy production and redox regulation in spermatozoa, supporting key functions such as progressive motility, capacitation, and the acrosome reaction. These processes are essential for successful fertilization and embryo development. However, species-specific differences exist in the reliance on oxidative phosphorylation versus glycolysis. Mitochondria also generate reactive oxygen species, which at physiological levels aid in sperm function but can cause oxidative stress and damage when overproduced. Mitochondrial dysfunction and excessive ROS can impair membrane potential, induce apoptosis, and damage nuclear and mitochondrial DNA, ultimately compromising sperm quality. Sperm mitochondrial DNA is highly susceptible to mutations and deletions, contributing to reduced motility and fertility. Targeted antioxidant strategies have emerged as promising therapeutic interventions to mitigate oxidative damage. This article provides a comprehensive overview of mitochondrial regulation in spermatozoa, the consequences of redox imbalance, and the potential of mitochondria-targeted antioxidants to improve sperm function and male fertility outcomes. The paper aims to deepen our understanding of mitochondrial roles in sperm physiology and contribute to the advancement of strategies for addressing male infertility. Full article

Subfertile boars often go undetected until they cause significant reproductive losses. Current semen quality assessments are limited in their ability to predict fertility, highlighting the need for complementary biomarkers. This study explored whether semen freezability could serve as an indirect indicator of boar fertility. Eighteen boars were classified based on historical fertility records and semen freezability, assessed by post-thaw quality. Fresh and post-thaw semen samples were analyzed using the CASA system and fluorescence microscopy. High-fertility boars showed significantly better motility and functional sperm parameters in fresh semen compared to low-fertility boars. However, these differences were mostly lost after cryopreservation. Conversely, boars with good freezability had consistently better post-thaw semen quality, though this did not correlate directly with higher fertility outcomes. Notably, a combined analysis revealed that boars with both high fertility and poor freezability had the lowest post-thaw semen quality. This suggests that cryopreservation may expose hidden sperm defects not detectable in fresh semen. Total motility was the only parameter associated with both fertility and freezability. In conclusion, while freezability alone may not directly predict fertility, it may help identify low-performing males. The combined assessment of fresh semen motility and freezability could support more effective boar selection strategies. Full article

In the swine industry, artificial insemination (AI) primarily uses chill-stored semen, making sperm preservation crucial for reproductive success. However, sperm quality declines at varying rates during chilled storage at 17 °C, distinguishing high-survival semen from low-survival semen. This study investigates the metabolomic profiles of boar sperm with different abilities to survive liquid storage. We analyzed sperm motility, kinematics, and morphology in freshly extended (Day 0) and 7-day stored AI semen doses. The AI semen doses were classified as high-motile (HM) or low-motile (LM) based on sperm motility after 7 days of storage (Day 7). Metabolomic data were collected in positive (ESI+) and negative (ESI−) ion modes using a Vanquish Flex UPLC coupled with a Q Extractive Plus. We consistently detected 442 metabolites (251 in ESI+, 167 in ESI−, and 24 in both) across samples and storage durations. In freshly extended and 7-day stored AI doses, we identified 42 and 56 differentially expressed metabolites (DEMs), respectively. A clustering analysis showed significant changes in DEMs between the HM and LM samples. These DEMs were mainly enriched in amino acid metabolism, the pentose phosphate pathway, glycerolipid metabolism, glyoxylate and dicarboxylate metabolism, terpenoid backbone biosynthesis, etc. In summary, this study highlights the metabolomic differences between semen doses with varying abilities to survive liquid storage. Glyceric acid and lysoPC(20:3) emerged as potential markers for sperm preservation. Full article

Understanding protein and amino acid deposition in pregnant gilts is important for developing nutritional strategies that meet these demands and enhance reproductive performance. Current models, such as the NRC (2012) gestating sow model, assume a constant proportional protein and amino acid content in tissues throughout pregnancy. However, empirical data suggest that gestational tissue growth and composition change dynamically. In this study, we developed a gestation model that characterizes the dynamic changes in growth, crude protein, and amino acid deposition throughout gestation. Based on a systematized search of published data, mathematical functions were developed to estimate daily protein and amino acid deposition in key tissues, including allantoic and amniotic fluid, uterus, placenta, fetus, mammary gland, and maternal body. Our results suggest that dietary crude protein levels and amino acid profiles should be adjusted to meet metabolic demands, particularly in early gestation, where a potential nutritional deficiency was identified. Additionally, the amino acid profile of deposited protein shifts during late gestation, suggesting a changing demand for specific amino acids. These findings challenge existing models and highlight the need for adaptive dietary strategies that better align with pregnancy’s biological demands. By refining protein and amino acid deposition estimates, this study provides a framework guiding future research on precision feeding, ultimately improving gilt and sow reproductive performance. Full article

Oocytes and cumulus cells undergo meiotic resumption and proliferation via gonadotropins and growth factors during maturation, and various small G proteins are activated when COCs undergo physiological changes. This study investigated the influence of gonadotropins and growth factors on Ras and its GTPases during porcine COC maturation. Unmatured COCs were treated with FSH, LH, or EGF for 44 h. The mRNA expression levels of the Ras subfamily (H-Ras, K-Ras, N-Ras, and R-Ras), its GTPases (RASA1 and SOS1), and proliferation factors (ERK, CCNB1, and Cdc2) were analyzed using RT-PCR. In contrast to other Ras subfamilies, R-Ras expression is upregulated during COC maturation. We evaluated the effects of FSH, LH, and EGF at various concentrations that most effectively regulated the expression of R-Ras and GTPases. The results demonstrated that 0.5 µg/mL FSH, 10 IU/mL human chorionic gonadotropin (hCG), and 10 ng/mL EGF effectively enhanced R-Ras expression and cell proliferation. FSH supplementation during porcine COC maturation significantly upregulated R-Ras and ERK expression, independent of LH and EGF, and downregulated Cdc2 expression. These results indicated that FSH regulates R-Ras expression, thereby promoting cell proliferation during COC maturation. These results provide fundamental knowledge for understanding the role of Ras and its family members in the development of follicular environments in pigs. Full article

The conservation of small and genetically vulnerable brown bear populations, such as the Cantabrian subpopulation in Spain, depends on developing species-specific assisted reproductive technologies and genetic resource banks. However, the lack of standardized sperm collection and cryopreservation protocols hinders their application. This study provides the first comparative analysis of sperm quality and proteomic profiles from three different origins: epididymal, pre-ejaculated, and ejaculated. Sperm quality parameters —motility and kinetic, viability, apoptosis, and oxidative stress— and protein expression were assessed. Although yields were similar, marked differences were observed in sperm quality and protein profiles. Sixty-three proteins involved in metabolism, stress response, and oxidative balance were differentially expressed depending on sperm origin. Epididymal sperm showed the highest viability and motility, lowest apoptosis, and a proteomic profile indicative of active spermatogenesis and enhanced oxidative stress defense. In contrast, ejaculated sperm had increased oxidative stress and reduced expression of metabolic proteins, while pre-ejaculated sperm exhibited lower motility, likely due to urine contamination and mitochondrial protein alterations, despite comparable viability and apoptosis. These findings offer novel insights into brown bear sperm biology and highlight the importance of sperm origin in developing optimized assisted reproduction strategies, ultimately supporting ex situ conservation efforts for this species. Full article

This study aims to analyse the effect of selected variation factors on the ejaculate characteristics of boars and to characterise changes in ejaculate characteristics in Landrace, Large White, Duroc, and Pietrain boars during their use for artificial insemination. The original value of this work lies in the estimation of the percentage share of individual components of variability in shaping the traits of boar ejaculate. A total of 943 ejaculates collected from 77 boars used for artificial insemination were analysed. This study began when the boars were at 8–9 months old. Ejaculates were collected in nine consecutive months from the start of the boars’ use. Immediately after collection, they were analysed for ejaculate volume, sperm concentration, percentage of sperm with progressive motility, total number of spermatozoa, and number of insemination doses per ejaculate. The results were analysed according to three criteria: breed of boar (Landrace, Large White, Duroc, and Pietrain), age of boar (up to 10 months, 11–13 months, 14–17 months, and more than 17 months), and season (spring, summer, autumn, and winter). The analysis of the variation in ejaculate characteristics took into account the share of each factor (boar breed, boar age, and season) in the variation, as well as the interactions between factors. The effects of the three factors and interactions between them were calculated using an ANOVA (analysis of variance). The variation was shown to depend mainly on the breed and age. These two factors and the interaction between them determine about 80% of the variation in ejaculate characteristics. The season also has an effect, but its share in the influence of variation on ejaculate characteristics is relatively small. Ejaculates from Landrace boars are the most favourable for insemination, with a large volume, a relatively high sperm concentration, and the highest number of sperm. The highest number of insemination doses can be prepared from Landrace ejaculates—on average, 2.7–6.7 more doses than from the other breeds. Duroc boar ejaculates are most distinctive, with a very low volume but a very high sperm concentration and the highest sperm motility. The ejaculates of Pietrain boars showed the opposite pattern, with the largest volume but the lowest sperm concentration. The sexual development of young boars, expressed as an increase in ejaculation performance, progresses during their first year of insemination use. Full article

Sexing procedures and subsequent freezing still impact sperm cells, leading to decreased fertility of gender-ablated semen. This study aimed to enhance cryo-survivability and reduce the capacitation-like change rate of gender-ablated semen by adding 2 mg of cholesterol-loaded cyclodextrin (CLC) per mL of extended semen containing 67 × 10⁶ sperm cells. This marks the first use of CLC with gender-ablated semen. Semen from four Jersey bulls was used for this study. Viability, motility, and mitochondrial activity were evaluated and adjusted to account for the inactivation of undesired sex sperm cells during processing. Binding ability to oviduct cells, fertilizing ability, and acrosome status were also evaluated. Adding CLC did not increase sperm motility. The population with intact membranes and acrosomes was significantly increased (p < 0.05) from 28.9 ± 1.2% to 34.1 ± 1.2% in the CLC-treated group. Mitochondrial potential, capacitation status at the membrane, calcium levels, and binding ability to oviduct cells were maintained. CLC treatment did not delay capacitation while significantly improving fertilization rates after 8 and 12 h of co-incubation (77 ± 3% vs. 67 ± 3% and 82 ± 3% vs. 74 ± 3%, respectively; p < 0.05). In conclusion, CLC addition significantly improved gender-ablated post-thaw sperm viability, acrosome integrity, and fertilizing ability while preserving motility, capacitation progress, and binding ability to oviduct cells. Full article

This study examined the impact of heat stress during specific periods before and after insemination to identify the most critical time affecting insemination success in dairy cows. A total of 47,199 insemination records from 6751 Holstein-Friesian cows (2002–2024) were analysed using generalised linear mixed models (GLMMs) with a binomial distribution. The periods assessed were the day of insemination, 21–6 days before (P1), 5–2 days before (P2), 1 day before to 1 day after (P3), 2–7 days after (P4), and 8–30 days after insemination (P5). Mild heat stress significantly reduced insemination success on the day of insemination, with a critical THI threshold of 60. Among all periods, P1 was the most sensitive to heat stress, with even mild stress negatively affecting outcomes. Severe heat stress (THI ≥ 72) also impacted P3 and P5, whereas P2 and P4 showed no significant effect. Heritability estimates increased with heat stress intensity, suggesting genetic variability in resilience. These findings highlight the importance of managing heat stress from three weeks before to one month after insemination and support the potential for genetic selection to improve reproductive performance under heat stress conditions. Full article

Oocyte maturation represents a fundamental biological process in bovine reproduction, establishing the physiological basis for fertilization and early embryonic development while critically determining the propagation of improved varieties and breeding efficiency. The roles of MQC in reproduction have gained substantial scientific attention. The proper maturation of oocytes fundamentally depends on adequate mitochondrial functionality. However, the intrinsic regulatory mechanisms governing MQC during bovine oocyte maturation remain incompletely characterized. Here, we discuss the most recent progress on the molecular mechanisms and roles of mitochondrial fission/fusion, biogenesis, and mitophagy in MQC. Building upon the mechanistic foundations of MQC in bovine oocyte maturation, this review identifies key mitochondrial-targeted supplements with potential applications in enhancing oocyte quality. Furthermore, we evaluate epigenetic influences on mitochondrial regulatory networks through mitochondrial–nuclear communication. Finally, we discuss the challenges in elucidating mitochondrial quality control mechanisms during oocyte maturation and propose corresponding strategies to address these obstacles. Integrating mechanistic insights, this review proposes strategies to enhance in vitro culture systems and identify oocyte quality markers, providing valuable insights for optimizing in vitro production (IVP) of bovine embryos and enhancing reproductive efficiency. Full article

Subclinical mastitis (SCM) is a stealthy but devastating challenge in the dairy industry, leading to economic losses and hindering efforts to achieve milk self-sufficiency. This study investigated the prevalence of SCM, antimicrobial resistance, and virulence profiles of Escherichia coli. A total of 174 milk samples were analyzed using the California mastitis test (CMT), somatic cell counts (SCCs), bacteriological culture, MALDI-TOF MS, and polymerase chain reaction (PCR). The findings revealed that the SCM prevalence was 68/174 (39.08%) based on CMT and SCC. Among SCM-positive samples, 60/68 (88.23%) were identified as E. coli, confirmed by MALDI-TOF MS and PCR assay. The most frequently detected serogroups were 0113 (11.6%) and 0113 (3.3%). Additionally, the genes for Stx1 and Stx2 were also detected in nine (15%) and one (1.7%), respectively. Antimicrobial susceptibility tests showed widespread resistance, with E. coli isolates demonstrating resistance to penicillin in 43 (71.6%), followed by ciprofloxacin in 42 (70%) and gentamicin in 18 (30%). A larger proportion of the E. coli strains (100%) harbored the bla_VIM gene, while 23 (38.3%), 20%, 20%, and 1.47% contained bla_KPC, bla_NMD, suli1, and msrA. Thirty (50%) isolates were considered multidrug-resistant (MDR). These findings underscore the urgent need for enhanced surveillance and antibiotic stewardship in dairy farming. The presence of MDR E. coli in SCM poses a dual threat of potential transmission to humans and treatment failures in mastitis management. This study highlights the importance of proactive control strategies to mitigate the spread of antimicrobial resistance in livestock and beyond. Full article