Animal Reproduction最新文献

Due to their self-renewal and differentiation potentials, mesenchymal stem cells (MSCs) may be induced into germ cells (GC) differentiation under in vitro conditions. In veterinary medicine, this technology could provide an alternative method to artificial insemination, as well as potentially useful for the conservation of endangered species. Previous studies have reported the use of SCs and MSCs co-culture systems, as well as SCs conditioned medium (SCCM) to induce GC differentiation of human and murine embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). The objective of this study was to evaluate the effect of SCCM as an inducer of in vitro GC differentiation of MSCs derived from fetal bovine adipose tissue (AT-MSCs). SCCM was collected from bovine SC cultures generated from adult bull testis. The effect of SCCM on MSCs was analyzed using quantitative PCR (Q-PCR) and flow cytometry. CD73 mRNA levels were decreased (P<0.05) in AT-MSC/SCCM at day 14 of culture compared to control. CD90 and CD105 gene expression were detected during the 21 days of culture; however, relative expression levels were not different (P>0.05) between treated and controls cells. DAZL gene expression was detected on day 21 of culture, as well as a proportion of AT-MSC positive for DAZL at day 21 of culture. OCT4, PIWIL2 and DAZL gene expressions were detected from day 0, 7 and 21 of culture, respectively, as well as a proportion of cells positive for each marker were detected at day 21 of culture. However, similar gene and protein expression levels (P>0,05) were detected between AT-MSCs/SCCM and control cultures. DMC1 gene expression levels were detected from day 7 of culture, and expression levels were not different (P>0,05) between treatment and control cells. Expression patterns of MSC, pluripotent, GC and meiotic markers indicate that SCCM did not induce GC differentiation of AT-MSCs.

Leptospirosis is a zoonosis caused by bacteria of the genus Leptospira. In cattle, the infection mainly manifests in the genital form. However, there are still few studies about this manifestation. The aim of this study was to describe the control of an outbreak of leptospirosis in dairy cows with reproductive disorders, through the combination of diagnostic methods and the integration of vaccination with antibiotic therapy. Between December 2022 and April 2023, 17 cows presented reproductive disorders. After the outbreak, two consecutive blood collections and one cervicovaginal mucus (CVM) collection were carried out. The blood samples were tested by the microscopic seroagglutination test (MAT), using a collection of antigens with eight strains of Leptospira and a cutoff point ≥ 1:200. The CVM samples were analyzed by polymerase chain reaction (PCR), with the lipL32 gene as target. The control was carried out with the CattleMaster® GOLD FP 5/L5 vaccine, in addition to the application of streptomycin (25 mg/kg) in positive cows. After one year of sanitary management, the CVM PCR was repeated to evaluate the effectiveness of the integrated control. In serology, 58.8% (10/17) of the cows were reactive, with 100% (10/10) for the serogroup Sejroe. In the molecular analysis, 58.8% (10/17) of the cows were positive. When combining the two methods, the result was 82.3% (14/17) reagent/positive. After the integrated control, 0.0% (0/17) of cows were positive. It was concluded that the outbreak was related to bovine leptospirosis. Furthermore, the combination of diagnostic methods and integrated control proved to be efficient.

Application of assisted reproduction techniques are essential for the preservation of endangered species, and ultrasonography has emerged as an interesting tool in this process, allowing noninvasive assessment of reproductive stages and characterization of male gonads. This review provides a compilation on the applications and perspectives of using ultrasonography for investigation of the morphological and functional aspects of the male reproductive tract in wild species. The technique, which has been improved with the use of vascular doppler, allows detailed analysis of blood flow and aids in the selection of individuals for breeding programs. Although there are challenges, such as physiological variations among species and the difficulty for applying ultrasonography to birds and reptiles, advances in imaging technologies, including elastography and doppler, have expanded the possibilities for diagnosis and monitoring reproductive status in various mammals. Ultrasonographic analysis contributes to the assessment of fertility, detection of testicular diseases and the definition of protocols for reproductive management, becoming an important tool in the conservation of wildlife and in the development of more effective assisted reproductive technologies.

As cattle have not been traditionally considered a model species and the molecular details of germ cell development don't directly inform production practices, the specification of primordial germ cells in the bovine embryo has remained understudied and poorly understood. Recent work by our laboratory builds on previous investigations to establish the molecular profile of primordial germ cells (PGC) at the critical moment when they are being specified in the embryo during the gastrulation stage. Combining advanced immunolocalization, confocal imaging and single-cell RNA sequencing, we identified PGC in the bovine embryo approximately on day 16 of development by co-expression of the core transcription factors OCT4, SOX17, PRDM1, and TFAP2C as demonstrated for several other species in which the embryo develops a bilaminar disc at the onset of gastrulation. Soon after specification, between days 20 and 22 of embryo development, early migratory PGC repress transcripts responsible for the establishment of somatic lineages. Notably, these cells do not seem highly proliferative during the early migratory stage, another aspect of early germ cells that is conserved in cows and other species such as pigs. Advancing the study of germ cell specification and development during bovine embryonic development, particularly at stages when human embryos are unavailable for investigation, places cows as an additional domestic species capable of providing crucial information about events that are paramount for fertility. As the field of in vitro gametogenesis continues to rapidly evolve, the study of bovine PGC and fetal germ cell development will provide invaluable information to facilitate the development and advancement of future assisted reproduction technologies for the improvement of agricultural animals and human reproduction.

Assisted reproductive technologies, particularly sex-sorted semen and in vitro embryo production (IVP) can contribute to accelerating genetic gain in both dairy breeds and beef breeds suitable for mating with dairy cows by increasing the number of offspring produced from genetically elite dams. Use of sexed semen has rapidly increased in recent years, accelerating herd genetic gain through selection of the best genetic merit dams to breed replacements, allowing non-replacement dams to be bred to beef sires or to act as recipients of beef embryos to improve calf marketability. IVP offers significant advantages over traditional multiple ovulation embryo transfer (MOET) including increased flexibility in sire usage allowing multiple pregnancies from elite dam-sire combinations to be generated, the ability to produce more embryos per unit time per genetically elite female, the ability to use oocytes from prepubertal females and the more efficient use of rare or high-cost semen straws. Despite these benefits, significant challenges relating to pregnancy loss after embryo transfer, particularly after cryopreservation of IVP embryos, and issues relating to peri- and postnatal health and development of IVP offspring remain to be resolved and hamper the more widespread application of the technology. Improving our understanding of the underlying physiological and molecular mechanisms that regulate early embryo development, embryo-endometrial interactions and lead to successful pregnancy establishment is necessary to understand and elucidate the causes of pregnancy loss and provide a basis for new strategies to improve pregnancy outcomes and reproductive efficiency.

Reproductive tract inflammatory diseases (RTID) present significant health challenges in domestic animals, impacting welfare, fertility, and productivity. Traditionally, antibiotics have been the primary treatment for these conditions, however, the rise of antimicrobial resistance calls for alternative approaches. The microbiome of the female reproductive tract plays a vital role in maintaining reproductive health, and emerging evidence suggests that microbiome-based therapies, such as 'natural' or 'synthetic' microbiome transplantation, may offer sustainable solutions for RTID management. This review explores the composition and dynamics of the reproductive microbiome in both healthy and diseased states in cows, mares, sows, dogs, and cats. It also examines current treatments and the potential for microbiome-based interventions to replace or complement antibiotic therapies. Although research on microbiome-based therapies for preventing or treating RTID in domestic animals is virtually non-existent, vaginal and uterine microbiomes transplantation in mice and women show promise but require further investigation to evaluate their efficacy and safety across species with varying reproductive physiologies. Additionally, synthetic microbiome therapies present a controlled and reproducible alternative, though they face challenges in design, engraftment, and regulatory approval. The transition from antibiotic dependence to microbiome-based solutions marks a paradigm shift in veterinary medicine, but successful implementation demands a deeper understanding of host-microbiome interactions, rigorous safety protocols, and species-specific research.

Unlocking the developmental potential of oocytes at various stages of folliculogenesis represents a major challenge in reproductive biology and assisted reproductive technologies. While in vitro maturation (IVM) of fully grown oocytes is widely applied, the vast majority of oocytes enclosed within early-stage follicles remain underutilized. This review outlines current advancements in in vitro culture systems designed to support oocyte growth and differentiation, with particular attention to the contributions of the authors. Key developments, mainly encompassing the bovine species, include the use of prematuration strategies to enhance the competence of oocytes retrieved from antral follicles, stepwise in vitro culture protocols for growing oocytes from early antral follicles, and efforts to establish defined systems for preantral follicle culture. Emerging insights into chromatin dynamics, cumulus-oocyte communication, and epigenetic regulation are shaping the design of tailored culture environments. Despite promising progress, significant challenges remain in replicating the complexity of in vivo folliculogenesis, particularly in non-rodent models. Addressing these challenges will be critical to expanding the oocyte pool available for reproductive and biotechnological applications, with broad implications for fertility preservation, livestock breeding, and fundamental research.

In vitro embryo production (IVEP) has become a cornerstone of genetic advancement in cattle, yet its efficiency remains suboptimal and highly variable. This review synthesizes current knowledge on donor selection and management strategies aimed at optimizing IVEP outcomes. Central to IVEP success is the quantity and developmental competence of oocytes, which is influenced by both intrinsic donor characteristics and extrinsic management interventions. Ovarian superstimulation using follicle-stimulating hormone (FSH) has emerged as a key strategy to enhance oocyte yield and quality, with evidence supporting dose-dependent improvements in embryo development and yield. Protocol refinements-including timing, duration, and delivery mode of FSH- can further influence IVEP efficacy. Donor-specific factors such as age, pregnancy status, and size of the ovarian reserve, assessed via antral follicle count (AFC) or anti-Müllerian hormone (AMH) concentrations, significantly affect oocyte competence and/or embryo yield. Additionally, newly developed genomic traits and selection indexes, offer predictive value for donor performance and enable integration of IVEP-specific traits into breeding programs. High AMH donors consistently outperform low AMH counterparts, and emerging evidence suggests that tailoring superstimulation protocols to AMH phenotype can further enhance IVEP outcomes. The integration of physiological and genomic data provides the opportunity for developing targeted, phenotype/genotype-driven superstimulation protocols to maximize IVEP efficiency in a cost-effective and biologically sound manner.

The increase in free fatty acid (FFA) levels in the circulation and follicular fluid in response to the negative energy balance of dairy cows has received significant attention during the last decades. However, until recently the potential effect of FFA on the periovulatory steroid environment has been overlooked. The well-orchestrated luteinizing hormone (LH) peak induces a steroid shift in the periovulatory follicle, from Estradiol-17β (E2) dominance around the LH peak towards progesterone (P4) dominance around ovulation, and is a prerequisite for optimal cytoplasmic and nuclear maturation in the oocyte and oocyte developmental competence. Recent insights in literature demonstrate a link between saturated and mono-unsaturated FFAs and the expression of gonadotrophin receptors, follicle stimulating hormone (FSH)R and LHR, including steroid related enzymes and E2 synthesis by in vitro granulosa cells. The current review will focus on the potential role of mono-unsaturated oleic acid, the most abundant FFA in follicular fluid, on steroidogenesis and its potential effect on the cumulus-oocyte-complex (COC) during final maturation. The data of this review suggest the potential for a regulatory interlinked system, which includes the oocyte secreted factor FGF10 and oleic acid, that modulates the steroidogenic switch from E2 to P4 in the periovulatory follicle, via actions that involve the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway in support of the delicate and well-orchestrated dialogue between the oocyte and cumulus cells during final maturation of COCs.

The acquisition of oocyte competence in cattle, encompassing both cytoplasmic and nuclear maturation, is essential for successful fertilization and embryonic development. This competence is progressively achieved during the latter stages of the oocyte growth phase and completed within the dominant follicle (DF). The unique hormonal and immunological environment of the DF during oestrous supports oocyte "capacitation," a process involving organelle reorganization, mRNA storage and meiosis resumption, which fully prepares the oocyte for fertilization. These changes differentiate oocytes from the DF from those of subordinate follicles, explaining why only oocytes from the DF mature and ovulate successfully. Despite advances in assisted reproductive technologies like in vitro maturation (IVM) and in vitro fertilization (IVF), developmental outcomes remain inferior compared to in vivo matured oocytes, largely due to incomplete or altered oocyte maturation in vitro. Blastocyst rates after IVM/IVF are substantially lower (~35%) than those from in vivo matured oocytes (58-78%). The heterogeneity of oocytes retrieved from antral follicles and the lack of exposure to the natural follicular environment during IVM are key factors limiting developmental competence. Here we describe the molecular changes in bovine oocytes from DFs, collected at 24 and 2 h before ovulation without ovarian stimulation, using single-cell RNA sequencing and bisulfite sequencing to assess gene expression and DNA methylation dynamics. Results revealed significant shifts in transcripts related to oxidative phosphorylation, highlighting the crucial role of energy metabolism during oocyte capacitation. DNA methylation changes were subtle but indicated a more dynamic and less stable epigenome in fully-grown oocytes than previously assumed. Overall, understanding the gene expression and epigenetic landscape during oocyte maturation in the DF offers valuable insights into improving oocyte quality and ART outcomes in cattle. Optimizing the maturation environment to better mimic natural follicular conditions could enhance reproductive efficiency in bovine production systems.