Effects of Mixed Pasture Legume Phytoestrogens on Superovulatory Response and Embryo Quality in Angus Cows.

Ovulation and artificial insemination rates have been observed to decrease in sheep and cows when exposed to dietary phytoestrogens at concentrations greater than 25 mg/kg DM. A grazing trial was undertaken to investigate the effects of coumestrol and other key phytoestrogens on the superovulatory response, embryo numbers and quality in beef cows grazing legume pastures. A 7-week controlled grazing trial was conducted with legume and ryegrass pasture treatments, with cows exposed to legumes at two timed treatments, 4 and 7 weeks. Twenty Angus cows were subjected to a conventional estrus synchronization and superovulation protocol. Embryos were recovered via conventional uterine body flushing 7 days post artificial insemination (AI). Numerous phytoestrogens were identified in both pasture and plasma samples, including coumestrol and formononetin. Concentrations of phytoestrogens in the pasture ranged from 0.001 to 47.5 mg/kg DM and 0 to 2.6 ng/mL in plasma. Approximately 50% of cows produced viable embryos 7 days post AI. A significant interaction between the effect of treatment groups on the embryo stage was observed (p < 0.05). The results suggest that concentrations of >25 mg/kg DM of phytoestrogens less than 20 days preceding AI may negatively affect oocyte developmental competence, reduce progesterone production and thus contribute to early embryonic loss.

Effect of Spirulina platensis Supplementation on Reproductive Parameters of Sahrawi and Jabbali Goat Bucks.

Spirulina platensis (SP) is a protein-rich dietary supplement that improves animal reproductive traits. This study investigated the effect of SP supplementation on puberty onset, semen characteristics, scrotal circumference (SC), libido, and hormone concentrations in Sahrawi and Jabbali bucks. The study was conducted in 36 bucks, divided into three groups (n = 6/group), for 70 days. The rations included the following: (1) Control feed (Con) with 14% crude protein and 11.97% MJ/kg DM energy; (2) Con with 2 g SP/head/day SP treatment (T1) and (3) Con with 4 g SP/head/day treatment (T2). The mean (±SEM) SC of both SP groups in the Sahrawi breed was significantly higher (p ≤ 0.05) compared to the Con. The mean of the semen volume significantly increased (p ≤ 0.05) in the SP group than in the Con group in both breeds. SP groups vs. Con groups had increased sperm concentration in Sahrawi bucks than Jabbali bucks. Mean serum luteinizing hormone (LH) and testosterone (Tes) concentrations in Jabbali bucks were significantly higher (p ≤ 0.05) in the SP groups compared to Sahrawi bucks. SP improved the SC, semen quality, libido, sperm concentration, and LH and Tes concentrations in both breeds. The results of the current study suggest that adding SP to the diet may have the ability to improve the semen quality of the local Omani bucks.

QTLs and Candidate Genes Associated with Semen Traits in Merino Sheep.

Ram semen traits play a significant role in conception outcomes, which in turn may influence reproductive efficiency and the overall productivity and profitability of sheep enterprises. Since hundreds of ewes may be inseminated from a single ejaculate, it is important to evaluate semen quality prior to use in sheep breeding programs. Given that semen traits have been found to be heritable, genetic variation likely contributes to the variability observed in these traits. Identifying such genetic variants could provide novel insights into the molecular mechanisms underlying variability in semen traits. Therefore, this study aimed to identify quantitative trait loci (QTLs) associated with semen traits in Merino sheep. A genome-wide association study (GWAS) was undertaken using 4506 semen collection records from 246 Merino rams collected between January 2002 and May 2021. The R package RepeatABEL was used to perform a GWAS for semen volume, gross motility, concentration, and percent post-thaw motility. A total of 35 QTLs, located on 16 Ovis aries autosomes (OARs), were significantly associated with either of the four semen traits in this study. A total of 89, 95, 33, and 73 candidate genes were identified, via modified Bonferroni, within the QTLs significantly associated with volume, gross motility, concentration, and percent post-thaw motility, respectively. Among the candidate genes identified, SORD, SH2B1, and NT5E have been previously described to significantly influence spermatogenesis, spermatozoal motility, and high percent post-thaw motility, respectively. Several candidate genes identified could potentially influence ram semen traits based on existing evidence in the literature. As such, validation of these putative candidates may offer the potential to develop future strategies to improve sheep reproductive efficiency. Furthermore, Merino ram semen traits are lowly heritable (0.071-0.139), and thus may be improved by selective breeding.

The GLP-1 agonist, exendin-4, stimulates LH secretion in female sheep.

Our previous studies showed that microinjection into the median eminence of the sheep of glucagon-like peptide- 1 (GLP-1) or its receptor agonist exendin-4 stimulates luteinising hormone (LH) secretion, but it is unknown whether the same effect may be obtained by systemic administration of the same. The present study measured the response in terms of plasma LH concentrations to intravenous (iv) infusion of exendin-4. A preliminary study showed that infusion of 2 mg exendin-4 into ewes produced a greater LH response in the follicular phase of the oestrous cycle than the luteal phase. Accordingly, the main study monitored plasma LH levels in response to either 0.5 mg or 2 mg exendin-4 or vehicle (normal saline) delivered by jugular infusion for 1 h in the follicular phase of the oestrous cycle. Blood samples were collected at 10 min intervals before, during and after infusion. Both doses of exendin-4 increased mean plasma LH concentrations and increased LH peripheral pulse amplitude. There was no effect on inter-pulse interval or timing of the preovulatory LH surge. These doses of exendin-4 did not alter plasma insulin or glucose concentrations. Quantitative PCR of the gastrointestinal tract samples from a population of ewes confirmed the expression of the preproglucagon gene (GCG). Expression increased aborally and was greatest in the rectum. It is concluded that endogenous GLP-1, most likely derived from the hindgut, may act systemically to stimulate LH secretion. The present data suggest that this effect may be obtained with levels of agonist that are lower than those functioning as an incretin.

Comparison of PCR-HRM, colorimetric LAMP and culture based diagnostic assays in the detection of endometritis caused by Streptococcus equi subsp. zooepidemicus in mares.

Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is one of the causative agents of equine endometritis. In this study, a panel of different bacterial species, and colonies derived from bacteriological cultures of 38 clinical samples, were subjected to Loop-Mediated Isothermal Amplification (LAMP) assay and PCR, followed by high-resolution melt (HRM) curve analysis. All clinical samples were genotyped into three distinct groups based on HRM curve analysis. Differences in melting curve profiles were a reflection of DNA variation in sorD gene which was confirmed by DNA sequencing. A mathematical model based on Genetic Confidence Percentage (GCP) was used in HRM curve analysis and a cut-off point value was established which differentiated S. zooepidemicus isolates without requiring visual interpretation of curve profiles. The accuracy of PCR-HRM and bacterial culture in detection of S. zooepidemicus were identical with 100% sensitivity and specificity, while LAMP assay had similar specificity but a lower sensitivity (89.5%). PCR-HRM and LAMP assay provided an effective detection method with a turn-around time of six hours for PCR-HRM and 120 min for LAMP assay, compared to a minimum three days that was required when routine bacteriological culture method was used. In summary, results indicate that LAMP had the quickest turnaround, and HRM curve analysis could potentially be used for genotyping without DNA sequencing. Any mare suspected of endometritis will benefit from developed rapid diagnostic tests for detection of S. zooepidemicus and proper treatment prior to being bred and will mitigate unnecessary treatment and antibiotic resistance.

Heritability and Genetic Parameters for Semen Traits in Australian Sheep.

Semen characteristics including volume, gross motility, spermatozoal concentration, and percent post-thaw motility are routinely assessed to determine the quality and quantity of an ejaculate prior to use in artificial breeding programs. Currently, artificial breeding programs in sheep place relatively little emphasis on ram-side factors, such as the fertilising potential of an ejaculate, which may contribute to variability in conception outcomes. Estimating genetic parameters for ejaculate quality and quantity traits could provide insights into whether selective breeding can be used to improve such ram-side traits, improving ovine reproductive performance and farm profitability. Therefore, in this study, a total of 11,470 ejaculate records, including data for ejaculate volume, gross motility, spermatozoal concentration, and percent post-thaw motility, collected over a 20-year period was used to estimate genetic parameters in sheep. Univariate and bivariate mixed model analysis was performed including a variety of fixed effects such as breed, age at collection, centre of collection, collection number, season of collection, and method of collection; and the permanent environmental effects associated with each ram, stud and year of collection, and the breeding value of rams included as random effects. The heritability for ejaculate volume, gross motility, concentration, and percent post-thaw motility was estimated to be 0.161, 0.170, 0.089, and 0.081. Repeatability estimates were moderate, ranging between 0.4126 and 0.5265. Overall, results indicate that semen traits are lowly heritable and moderately repeatable, indicating that these traits are significantly influenced by environmental variables.

Subtotal Ovariohysterectomy Following Fetal Maceration and Pyometra in a Maiden Welsh Pony Mare.

Fetal maceration and pyometra were diagnosed in a 16-year-old maiden Welsh pony mare. Due to anatomical and surgical difficulties encountered throughout treatment, previously reported techniques including both per-vaginum removal of fetal remnants and ovariohysterectomy were attempted and resulted in novel complications. Fetal bones were unable to be removed completely transcervically due to failure of cervical dilation despite repeated attempts. Subsequent surgical complications such as inadequate exposure of the uterus and incorporation of fetal bones into the uterine stump following ovariohysterectomy also occurred. This case highlights some of the difficulties in managing rare cases of mid-term fetal loss and retention that result in maceration. Ultimately, this may provide useful insights to practitioners when managing similar cases in future.

Phytoestrogens: A Review of Their Impacts on Reproductive Physiology and Other Effects upon Grazing Livestock.

Legume crops and pastures have a high economic value in Australia. However, legume species commonly used for grazing enterprises have been identified to produce high concentrations of phytoestrogens. These compounds are heterocyclic phenolic, and are similar in structure to the mammalian estrogen, 17ß-estradiol. The biological activity of the various phytoestrogen types; isoflavones, lignans and coumestans, are species-specific, although at concentrations of 25 mg/kg of dry matter each of the phytoestrogen types affect reproductive functions in grazing livestock. The impacts upon fertility in grazing livestock such as cattle and sheep, vary greatly over length of exposure time, age and health of animal and the stress stimuli the plant is exposed to. More recently, research into the other effects that phytoestrogens may have upon metabolism, immune capacity and growth and performance of grazing livestock has been conducted. Potential new benefits for using these phytoestrogens, such as daidzein and genistein, have been identified by observing the stimulation of production in lymphocytes and other antibody cells. Numerous isoflavones have also been recognized to promote protein synthesis, increase the lean meat ratio, and increase weight gain in cattle and sheep. In Australia, the high economic value of legumes as pasture crops in sheep and cattle production enterprises requires proactive management strategies to mitigate risk associated with potential loss of fertility associated with inclusion of pasture legumes as forages for grazing livestock.

The future of assessing bull fertility: Can the 'omics fields identify usable biomarkers?†.

Breeding soundness examinations for bulls rely heavily on the subjective, visual assessment of sperm motility and morphology. Although these criteria have the potential to identify infertile males, they cannot be used to guarantee fertility or provide information about varying degrees of bull fertility. Male factor fertility is complex, and the success of the male gamete is not necessarily realized until well after the spermatozoon enters the oocyte. This paper reviews our existing knowledge of the bull's contribution from a standpoint of the sperm's cargo and the impact that this can have on fertilization and the development of the embryo. There has been a plethora of recent research characterizing the many molecular attributes that can affect the functional competence of a spermatozoon. A better understanding of the molecular factors influencing fertilization and embryo development in cattle will lead to the identification of biomarkers for the selection of bulls of superior fertility, which will have major implications for livestock production. To see this improvement in reproductive performance, we believe incorporation of modern technology into breeding soundness examinations will be necessary-although many of the discussed technologies are not ready for large-scale field application. Each of the 'omics fields discussed in this review have shown promise for the identification of biomarkers of fertility, with certain families of biomarkers appearing to be better suited to different evaluations throughout a bull's lifetime. Further research is needed for the proposed biomarkers to be of diagnostic or predictive value.

Maternal Behavior in Beef Cattle: The Physiology, Assessment and Future Directions-A Review.

Bovine maternal behavior is known to be influenced by a variety of factors including hormonal mediation, breed, age, parity, host genetics and general management practices. Following centuries of varying levels of domestication processes, the behavior of the bovine cow has altered from that of her original wild ungulate ancestors, although many maternal instincts have remained unchanged. The influence of maternal behavior on calf health and performance is of interest to cow-calf beef production operations, as in most instances, the cow is solely responsible for rearing the calf until weaning. However, investigating the magnitude of this influence is challenging, in part because objective measurement of behavioral traits is difficult, particularly in extensive settings. In recent years, while a number of remote monitoring devices have been developed that afford opportunities for objective measurement of behavioral traits in livestock, characterization of physiological mechanisms that underlie superior maternal behavior, including identification of potential biomarkers remains elusive in cattle. Hormonal profiles during the periparturient period have been shown to influence behavioral patterns in both current and future generations in other mammalian species and may provide insights into the physiology of bovine maternal behavior. Therefore, the aim of this review is to describe general characteristics of bovine maternal behavior and the factors known to influence it, including hormonal drivers, through which cross-reference to other species is made. Current methods of measuring and assessing behavior that may also be applicable to most production settings have also been reviewed. At present, there is no known hormonal assay that can be used to measure and/or reliably predict bovine maternal behavior post-calving or across generations. Being able to objectively assess superior maternal behavior, whether that be through remote monitoring, hormonal profiling or indirectly through measuring calf performance will be beneficial to livestock industries in the future.

Characterization of Phytoestrogens in Medicago sativa L. and Grazing Beef Cattle.

Phytoestrogens are plant-produced bioactive secondary metabolites known to play an integral role in plant defense that frequently accumulate in times of stress and/or microbial infection. Phytoestrogens typically belong to two distinct chemical classes; flavonoids (isoflavones) and non-flavonoids (lignans and coumestans). Upon consumption by livestock, high concentrations of phytoestrogens can cause long-term disruption in reproduction due to structural similarities with mammalian estrogens and their tendency to bind estrogen receptors. Wide variation in phytoestrogen concentration has been reported in pasture legumes and corresponding silage or hay. Lucerne is a common perennial pasture legume in temperate climates, but information on phytoestrogen production or accumulation in grazing livestock is currently limited. Therefore, metabolic profiling using UHPLC-MS-QToF was performed to identify and quantitate key phytoestrogens in both fresh and dried lucerne fodder from replicated field or controlled glasshouse environments. Phytoestrogens were also profiled in the blood plasma of Angus cattle grazing field-grown lucerne. Results revealed that phytoestrogens varied quantitatively and qualitatively among selected lucerne cultivars grown under glasshouse conditions. Fresh lucerne samples contained higher concentrations of coumestans and other phytoestrogenic isoflavones than did dried samples for all cultivars profiled, with several exceeding desirable threshold levels for grazing cattle. Coumestans and isoflavones profiled in plasma of Angus heifers grazing lucerne increased significantly over a 21-day sampling period following experimental initiation. Currently, threshold concentrations for phytoestrogens in plasma are unreported. However, total phytoestrogen concentration exceeded 300 mg·kg-1 in fresh and 180 mg·kg-1 in dried samples of selected cultivars, suggesting that certain genotypes may upregulate phytoestrogen production, while others may prove suitable sources of fodder for grazing livestock.

Characterization of Breed Specific Differences in Spermatozoal Transcriptomes of Sheep in Australia.

Reduced reproductive efficiency results in economic losses to the Australian sheep industry. Reproductive success, particularly after artificial insemination, is dependent on a number of contributing factors on both ewe and ram sides. Despite considerable emphasis placed on characterising ewe side contributions, little emphasis has been placed on characterising ram side contributions to conception success. Over 14,000 transcripts are in spermatozoa of other species, which are transferred to the ova on fertilisation. These transcripts conceivably influence early embryonic development and whether conception is successful. Semen was collected (n = 45) across three breeds; Merino, Dohne, and Poll Dorset. Following collection, each ejaculate was split in two; an aliquot was assessed utilising Computer Assisted Semen Analysis (CASA) and the remaining was utilised for RNA extraction and subsequent next-generation sequencing. Overall, 754 differentially expressed genes were identified in breed contrasts and contrast between ejaculates of different quality. Downstream analysis indicated that these genes could play significant roles in a broad range of physiological functions, including maintenance of spermatogenesis, fertilisation, conception, embryonic development, and offspring production performance. Overall results provide evidence that the spermatozoal transcriptome could be a crucial contributing factor in improving reproductive performance as well as in the overall productivity and profitability of sheep industries.

The effect of false mount on quality of frozen-thawed semen in Bos indicus beef bulls.

This study evaluated the effect of false mounts (FM) on quality of frozen-thawed semen in beef bulls. Five mature Brahman bulls at the Standard Semen Production Center in Thailand were subjected to semen collection. Overall, 60 ejaculates were collected over a 12-week period of which 30 were collected without, and another 30 were collected using FM. A range of quality parameters of fresh, pre-frozen, and frozen-thawed semen were evaluated. Use of FM resulted in significant improvement of most parameters of fresh semen except progressive motility and sperm viability. The quality parameters of pre-frozen and frozen-thawed semen also significantly improved with FM. Use of FM significantly improved the viability of frozen-thawed semen, possibly indicating the protective effect of seminal plasma proteins.

The impact of ecbolic therapy in the early postpartum period on uterine involution and reproductive health in dairy cows.

Postpartum uterine disease due to poor uterine involution continues to be a significant factor that contributes to poor reproductive efficiency in dairy cattle. Therapy that increases the frequency, duration and strength of uterine contractions in the postpartum period might enhance uterine involution, resulting in improved reproductive performance. The objective of this clinical trial was to study the effect of two uterine ecbolic therapies, oxytocin and prostaglandinF2α on uterine involution, postpartum endometritis, and reproductive performance. A randomized double-blinded clinical trial was conducted in 118 dairy cows from two research herds that had normal parturition and expulsion of the fetal membranes. Within 24 hr after calving, cows were randomly assigned to receive intramuscular injections twice a day of 50 IU of oxytocin, or 25 mg of dinoprost (PGF2α) or saline (control) for 7 days. Cows were monitored from day 3 to day 63 ± 3 days postpartum by transrectal palpation of the uterus, vaginoscopy, Metricheck® examination and by endometrial cytology. Blood samples were collected for measurement of progesterone in weeks 3 (21 ± 3 days), 5 (35 ± 3 days), 7 (49 ± 3 days), and 9 (63 ± 3 days) postpartum. Herd breeding records were obtained to determine reproductive performance in all cows. Neither oxytocin nor prostaglandin F2α therapy during the first week postpartum had any significant effect on the rate of uterine involution, prevalence of endometritis or reproductive performance, compared to untreated controls. Ecbolic drugs, as used here, are not recommended for use in clinical practice to improve involution or reproductive tract health in normal cows.

Assessment of uterine luminal pH in mares and the effect of dilute vinegar lavage on uterine luminal pH and endometrial health.

Uterine luminal pH has been demonstrated to be a valid indicator of uterine health in species such as cattle and sheep. However, research regarding uterine luminal pH in equines is lacking. The objectives of this study were to assess uterine luminal pH in mares during the estrous cycle, and evaluate the effect of dilute vinegar lavage on both uterine luminal pH and endometrial health. The study was conducted using a randomized block design in which eight mares (four Thoroughbred and four Standardbred) were aged matched then randomly assigned to two groups. Endometrial biopsies were taken from each mare prior to trial commencement. The treatment group (n = 4; 1 Thoroughbred, 3 Standardbreds) received a uterine lavage of one liter dilute vinegar (20 mL of vinegar in 1 L saline) every second day during each estrus period throughout the trial. Control group mares did not receive a uterine lavage. Uterine luminal pH measurements were recorded in all mares in both groups for a period of up to 10 min immediately prior to lavage (0 h), one hour and 24 h post lavage (same time points in control group mares as if they had been treated). Diestrus uterine luminal pH measurements were recorded once between days 6-10 post-ovulation. Endometrial biopsies were repeated from all mares at trial completion. Mean uterine luminal pH ranged from pH 5.3 to 7.6 and was significantly lower during diestrus compared to estrus (P < 0.001). Regression analysis indicated that this variation in pH was best explained by estrous cycle day, with uterine luminal pH increasing by a mean of 0.03 units each day (P < 0.001) from 6 to 10 days post-ovulation through to ovulation. Uterine lavage with dilute vinegar did not significantly affect uterine luminal pH (P > 0.05). A scoring system to quantify the abundance of cell types in the endometrial biopsies showed that mares in the treatment group had a significant decrease in polymorphonuclear cell abundance between pre- and post-trial biopsies (P = 0.03). Mares in the treatment group also had a significant decrease in lymphocyte, plasma cell and eosinophil cell abundance (P = 0.05). Although dilute vinegar lavage was not associated with a significant change in uterine luminal pH, it was associated with a significant improvement in endometrial biopsy scores. Because the control group did not receive a uterine lavage, further research is required to determine if this significant improvement results from the addition of dilute vinegar, or the uterine lavage itself.