Impact of oocyte donor age and breed on in vitro embryo production in cattle, and relationship of dairy and beef embryo recipients on pregnancy and the subsequent performance of offspring: A review.

Genomic selection combined with in vitro embryo production (IVEP) with oocytes from heifer calves provides a powerful technology platform to reduce generation interval and significantly increase the rate of genetic gain in cattle. The ability to obtain oocytes with developmental competence from calves has been recognised for more than 60years. There is renewed interest in the potential of this reproductive technology as the beef and dairy industries seek to identify and multiply animals that satisfy consumer demand for efficient utilisation of natural resources, with minimal environmental impact and high product quality. Heifer calves show a good follicular response to FSH stimulation and oocytes can be recovered by ovum pick-up (OPU). However, the efficiency of OPU/IVEP remains lower for calves compared with peripubertal heifers and cows, in both indicus (Zebu, Bos indicus ) and taurus (Bos taurus ) breeds. Indicus cattle generally have more follicles, yield a greater number of oocytes, and have a better IVEP outcome, compared with taurus cattle. The treatment of prepubertal heifers with bovine somatotrophin (bST) and propylene glycol before OPU/IVEP has yet to show a major improvement in embryo production. Holstein (taurus) dairy heifers derived from different reproductive technologies (AI, MOET, OPU/IVEP) showed a similar age at puberty and first-lactation milk production. OPU/IVEP Holstein embryos transferred to beef or dairy cows likewise yielded heifers with the same performance. The gains in productivity that can be achieved with strategic use of OPU/IVEP in heifer calves make this a relevant and highly important reproductive technology in cattle breeding. Ongoing optimisation of the technology is needed for the potential of OPU/IVEP in young donors to be fully realised.

Peripheral action of kisspeptin at reproductive tissues-role in ovarian function and embryo implantation and relevance to assisted reproductive technology in livestock: a review.

Kisspeptin (KISS1) is encoded by the KISS1 gene and was initially found to be a repressor of metastasis. Natural mutations in the KISS1 receptor gene (KISS1R) were subsequently shown to be associated with idiopathic hypothalamic hypogonadism and impaired puberty. This led to interest in the role of KISS1 in reproduction. It was established that KISS1 had a fundamental role in the control of gonadotropin releasing hormone (GnRH) secretion. KISS1 neurons have receptors for leptin and estrogen receptor α (ERα), which places KISS1 at the gateway of metabolic (leptin) and gonadal (ERα) regulation of GnRH secretion. More recently, KISS1 has been shown to act at peripheral reproductive tissues. KISS1 and KISS1R genes are expressed in follicles (granulosa, theca, oocyte), trophoblast, and uterus. KISS1 and KISS1R proteins are found in the same tissues. KISS1 appears to have autocrine and paracrine actions in follicle and oocyte maturation, trophoblast development, and implantation and placentation. In some studies, KISS1 was beneficial to in vitro oocyte maturation and blastocyst development. The next phase of KISS1 research will explore potential benefits on embryo survival and pregnancy. This will likely involve longer-term KISS1 treatments during proestrus, early embryo development, trophoblast attachment, and implantation and pregnancy. A deeper understanding of the direct action of KISS1 at reproductive tissues could help to achieve the next step change in embryo survival and improvement in the efficiency of assisted reproductive technology.

Adhesion molecules in gamete transport, fertilization, early embryonic development, and implantation-role in establishing a pregnancy in cattle: A review.

Cell-cell adhesion molecules have critically important roles in the early events of reproduction including gamete transport, sperm-oocyte interaction, embryonic development, and implantation. Major adhesion molecules involved in reproduction include cadherins, integrins, and disintegrin and metalloprotease domain-containing (ADAM) proteins. ADAMs on the surface of sperm adhere to integrins on the oocyte in the initial stages of sperm-oocyte interaction and fusion. Cadherins act in early embryos to organize the inner cell mass and trophectoderm. The trophoblast and uterine endometrial epithelium variously express cadherins, integrins, trophinin, and selectin, which achieve apposition and attachment between the elongating conceptus and uterine epithelium before implantation. An overview of the major cell-cell adhesion molecules is presented and this is followed by examples of how adhesion molecules help shape early reproductive events. The argument is made that a deeper understanding of adhesion molecules and reproduction will inform new strategies that improve embryo survival and increase the efficiency of natural mating and assisted breeding in cattle.

Transforming growth factor-ß superfamily and interferon-τ in ovarian function and embryo development in female cattle: review of biology and application.

Survival of the embryo and establishment of a pregnancy is a critical period in the reproductive function of female cattle. This review examines how the transforming growth factor-ß (TGFB) superfamily (i.e. bone morphogenetic protein (BMP) 15, growth differentiation factor (GDF) 9, anti-Müllerian hormone (AMH)) and interferon-τ (IFNT) affect ovarian function and embryo development. The oocyte in a primary follicle secretes BMP15 and GDF9, which, together, organise the surrounding granulosa and theca cells into the oocyte-cumulus-follicle complex. At the same time, the granulosa secretes AMH, which affects the oocyte. This autocrine-paracrine dialogue between the oocyte and somatic cells continues throughout follicle development and is fundamental in establishing the fertilisation potential and embryo developmental competency of oocytes. The early bovine embryo secretes IFNT, which acts at the uterine endometrium, corpus luteum and blood leucocytes. IFNT is involved in the maternal recognition of pregnancy and immunomodulation to prevent rejection of the embryo, and supports progesterone secretion. Manipulation of BMP15, GDF9, AMH and IFNT in both invivo and invitro studies has confirmed their importance in reproductive function in female cattle. This review makes the case that a deeper understanding of the biology of BMP15, GDF9, AMH and IFNT will lead to new strategies to increase embryo survival and improve fertility in cattle. The enhancement of oocyte quality, early embryo development and implantation is considered necessary for the next step change in the efficiency of natural and assisted reproduction in cattle.

Short communication: Space allocation in intensive Mediterranean buffalo production influences the profile of functional biomolecules in milk and dairy products.

The aim of the present study was to determine if space allocation influenced the concentration of biomolecules in buffalo milk and dairy products. Intensively housed buffaloes (n = 96) were randomly assigned to 2 groups according to days in milk, parity, and milk yield: group S10 had a space allocation of 10 m2 per buffalo and group S15 had a space allocation of 15 m2 per buffalo. Individual milk yield was recorded daily. Twice a month, a bulk milk sample was collected for each group, as well as whey, ricotta, and mozzarella cheese, to assess cheese yield and to conduct HPLC-electrospray ionization-tandem mass spectrometry, milk antioxidant activity, and cell viability analyses. We tested milk extracts from the 2 groups in vitro to evaluate their efficacy in counteracting endothelial oxidative damage induced by high glucose. We evaluated reproductive function in 28 buffaloes from each group using the Ovsynch-timed artificial insemination program. We observed no differences in milk quantity or quality in terms of fat, protein, or lactose, and reproductive function did not differ between the 2 groups. Compared with group S10, group S15 had higher concentrations of carnitine (56.7 ± 1.1 vs. 39.8 ± 0.7 mg/L in milk and 40.9 ± 0.8 vs. 31.7 ± 0.7 mg/L in whey), acetyl-l-carnitine (51.9 ± 0.3 vs. 39.7 ± 0.7 mg/L in milk and 41.1 ± 1.7 vs. 28.7 ± 2.6 mg/L in whey), propionyl-l-carnitine (34.8 ± 1.0 vs. 21.0 ± 0.9 mg/L in milk and 26.9 ± 0.8 vs. 17.6 ± 1.2 mg/L in whey), glycine betaine (23.1 ± 1.9 vs. 13.5 ± 1.6 mg/L in milk and 10.7 ± 0.4 vs. 7.9 ± 0.5 mg/L in whey), and δ-valerobetaine (24.2 ± 0.5 vs. 16.7 ± 0.5 mg/L in milk and 22.0 ± 0.9 vs. 15.5 ± 0.7 mg/L in whey). Group S15 also had higher total antioxidant activity than group S10 (56.7 ± 1.9 vs. 46.4 ± 1.13 mM Trolox equivalents). Co-incubation of high-glucose-treated endothelial cells with milk extracts from group S15 improved cell viability compared with cells treated with high glucose only; it also reduced intracellular lipid peroxidation (144.3 ± 0.4 vs. 177.5 ± 1.9%), reactive oxygen species (141.3 ± 0.9 vs. 189.3 ± 4.7 optical density units), and cytokine release (tumor necrosis factor-α, IL-1ß, IL-6). Greater space allocation was associated with higher levels of biomolecules in buffalo milk. This could have been the result of improved welfare in buffaloes that were allocated more space.

The role of rumen-protected choline in hepatic function and performance of transition dairy cows.

High-producing dairy cows enter a period of negative energy balance during the first weeks of lactation. Energy intake is usually sufficient to cover the increase in energy requirements for fetal growth during the period before calving, but meeting the demand for energy is often difficult during the early stages of lactation. A catabolic state predominates during the transition period, leading to the mobilisation of energy reserves (NEFA and amino acids) that are utilised mainly by the liver and muscle. Increased uptake of mobilised NEFA by the liver, combined with the limited capacity of hepatocytes to either oxidise fatty acids for energy or to incorporate esterified fatty acids into VLDL results in fatty liver syndrome and ketosis. This metabolic disturbance can affect the general health, and it causes economic losses. Different nutritional strategies have been used to restrict negative effects associated with the energy challenge in transition cows. The provision of choline in the form of rumen-protected choline (RPC) can potentially improve liver function by increasing VLDL exportation from the liver. RPC increases gene expression of microsomal TAG transfer protein and APOB100 that are required for VLDL synthesis and secretion. Studies with RPC have looked at gene expression, metabolic hormones, metabolite profiles, milk production and postpartum reproduction. A reduction in liver fat and enhanced milk production has been observed with RPC supplementation. However, the effects of RPC on health and reproduction are equivocal, which could reflect the lack of sufficient dose-response studies.

Active immunisation of mice with GnRH lipopeptide vaccine candidates: Importance of T helper or multi-dimer GnRH epitope.

Active immunisation against gonadotropin releasing hormone (GnRH) is a potential alternative to surgical castration. This study focused on the development of a GnRH subunit lipopeptide vaccine. A library of vaccine candidates that contained one or more (up to eight) copies of monomeric or dimeric GnRH peptide antigen, an adjuvanting lipidic moiety based on lipoamino acids, and an additional T helper epitope, was synthesised by solid phase peptide synthesis. The candidates were evaluated in vivo in order to determine the minimal components of this vaccine necessary to induce a systemic immune response. BALB/c mice were immunised with GnRH lipopeptide conjugates, co-administered with or without Complete Freund's Adjuvant, followed by two additional immunisations. Significant GnRH-specific IgG titres were detected in sera obtained from mice immunised with four of the seven lipopeptides tested, with an increase in titres observed after successive immunisations. This study highlights the importance of for epitope optimisation and delivery system design when producing anti-hapten antibodies in vivo. The results of this study also contribute to the development of future clinical and veterinary immunocontraceptives.

Pleomorphic adenoma gene1 in reproduction and implication for embryonic survival in cattle: a review.

The pleomorphic adenoma gene1 (PLAG1) encodes a DNA-binding, C2H2 zinc-finger protein which acts as a transcription factor that regulates the expression of diverse genes across different organs and tissues; hence, the name pleomorphic. Rearrangements of the PLAG1 gene, and/or overexpression, are associated with benign tumors and cancers in a variety of tissues. This is best described for pleomorphic adenoma of the salivary glands in humans. The most notable expression of PLAG1 occurs during embryonic and fetal development, with lesser expression after birth. Evidence has accumulated of a role for PLAG1 protein in normal early embryonic development and placentation in mammals. PLAG1 protein influences the expression of the ike growth factor 2 (IGF2) gene and production of IGF2 protein. IGF2 is an important mitogen in ovarian follicles/oocytes, embryos, and fetuses. The PLAG1-IGF2 axis, therefore, provides one pathway whereby PLAG1 protein can influence embryonic survival and pregnancy. PLAG1 also influences over 1,000 other genes in embryos including those associated with ribosomal assembly and proteins. Brahman (Bos indicus) heifers homozygous for the PLAG1 variant, rs109815800 (G > T), show greater fertility than contemporary heifers with either one, or no copy, of the variant. Greater fertility in heifers homozygous for rs109815800 could be the result of early puberty and/or greater embryonic survival. The present review first looks at the broader roles of the PLAG1 gene and PLAG1 protein and then focuses on the emerging role of PLAG1/PLAG1 in embryonic development and pregnancy. A deeper understanding of factors which influence embryonic development is required for the next transformational increase in embryonic survival and successful pregnancy for both in vivo and in vitro derived embryos in cattle.

The pleomorphic adenoma gene1 (PLAG1) produces PLAG1 protein which, by binding to specific regions on DNA, influences the activity of other genes that regulate many body functions. One gene is insulin-like growth factor 2 (IGF2) which controls cell metabolism and growth. The PLAG1 gene is particularly active during embryonic and fetal growth, and through IGF2 determines stature later in life. IGF2 protein is also very important in early embryonic development. This review explores the hypothesis that PLAG1 is an important determinant of embryonic survival and the establishment of pregnancy in mammals.

Genome-wide association and expression quantitative trait loci in cattle reveals common genes regulating mammalian fertility.

Most genetic variants associated with fertility in mammals fall in non-coding regions of the genome and it is unclear how these variants affect fertility. Here we use genome-wide association summary statistics for Heifer puberty (pubertal or not at 600 days) from 27,707 Bos indicus, Bos taurus and crossbred cattle; multi-trait GWAS signals from 2119 indicine cattle for four fertility traits, including days to calving, age at first calving, pregnancy status, and foetus age in weeks (assessed by rectal palpation of the foetus); and expression quantitative trait locus for whole blood from 489 indicine cattle, to identify 87 putatively functional genes affecting cattle fertility. Our analysis reveals a significant overlap between the set of cattle and previously reported human fertility-related genes, impling the existence of a shared pool of genes that regulate fertility in mammals. These findings are crucial for developing approaches to improve fertility in cattle and potentially other mammals.

Proteomic profiles of the embryonic chorioamnion and uterine caruncles in buffaloes (Bubalus bubalis) with normal and retarded embryonic development.

The aim of this study was to compare the proteome profiles of the chorioamnion and corresponding caruncle for buffalo embryos that had either normal or retarded development on Day 25 after artificial insemination (AI). In experiment 1, embryos that were to subsequently undergo late embryonic mortality had a smaller width on Day 25 after AI than embryos associated with pregnancy on Day 45 after AI. In experiment 2, 25 Italian Mediterranean buffaloes underwent transrectal ultrasonography on Day 25 after AI, and pregnant animals were categorized as one of two groups based on embryonic width: normal embryos (embryonic width > 2.7 mm) and retarded embryos (embryonic width < 2.7 mm). Three buffaloes of each group were slaughtered on Day 27 after AI to collect chorioamnion and caruncle tissues for subsequent proteomic analyses. Two-dimensional difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight mass spectrometer analysis were used to ascertain the proteomic profiles. To confirm 2D-DIGE-results, three selected proteins were analyzed by Western blot. The proteomic profiles of the chorioamnion of retarded embryos and the corresponding caruncles showed differences in the expression of several proteins compared to normal embryos. In particular, a down-regulation was observed for proteins involved in protein folding (HSP 90-alpha, calreticulin), calcium binding (annexin A1, annexin A2), and coagulation (fibrinogen alpha-chain) (P < 0.05), whereas proteins involved in protease inhibition (alpha-1-antiproteinase, serpin H1, serpin A3-8), DNA and RNA binding (heterogeneous nuclear ribonucleoproteins A2/B1 and K), chromosome segregation (serine/threonine-protein phosphatase 2A), cytoskeletal organization (ezrin), cell redox homeostasis (amine oxidase-A), and hemoglobin binding (haptoglobin) were up-regulated (P < 0.05).

Review: Development, adoption, and impact of assisted reproduction in domestic buffaloes.

The domestic buffalo (Bubalus bubalis), also known as water buffalo, comprises two sub-species the River buffalo (B. bubalis ssp. bubalis; 50 chromosomes) and the Swamp buffalo (ssp. carabanensis; 48 chromosomes). Domestic buffaloes are a globally significant livestock species. In South Asia, the River buffalo is a primary source of milk and meat and has a very important role in food security. The River buffalo also supports high-value, differentiated food production in Europe and the Americas. The Swamp buffalo is an important draft animal and a source of food in Southeast Asia and East Asia. The growing importance of buffaloes requires that they undergo an accelerated rate of genetic gain for efficiency of production, product quality, and sustainability. This will involve the increased use of assisted reproduction. The initial application of reproductive technology in buffaloes had variable success as it relied on the adoption of procedures developed for cattle. This included artificial insemination (AI), sperm cryopreservation, and embryo technologies such as cloning and in vitro embryo production (IVEP). Reproductive technology has been progressively refined in buffaloes, and today, the success of AI and IVEP is comparable to cattle. Ovarian follicular superstimulation (superovulation) combined with in vivo embryo production results in low embryo recovery in buffaloes and has limited practical application. The contribution of elite female buffaloes to future genetic improvement will therefore rely mainly on oocyte pickup and IVEP. This will include IVEP from females before puberty to reduce generation intervals. This review provides for the first time a clear chronology on the development, adoption, and impact, of assisted reproduction in domestic buffaloes.

Plasma metabolomics reveals major changes in carbohydrate, lipid, and protein metabolism of abruptly weaned beef calves.

1H NMR-based metabolomics was used to study the effect of abrupt weaning on the blood metabolome of beef calves. Twenty Angus calves (258 ± 5 kg BW; 5 to 6 months old) were randomly assigned to a non-weaned (NW) group that remained grazing with their dam or a weaned (W) group that underwent abrupt separation from their dam to a separate paddock on d 0 of the study. Body weight, behaviour, and blood samples for cortisol and metabolomics were measured at d 0, 1, 2, 7, and 14 of the study. On d 1 and 2, W calves spent less time grazing and ruminating, and more time vocalising and walking, had a greater concentration of cortisol, NEFA, 3-hydroxybutyrate, betaine, creatine, and phenylalanine, and lesser abundance of tyrosine (P < 0.05) compared to NW calves. Compared to NW calves at d 14, W calves had greater (P < 0.01) relative abundance of acetate, glucose, allantoin, creatinine, creatine, creatine phosphate, glutamate, 3-hydroxybutyrate, 3-hydroxyisobutyrate, and seven AA (alanine, glutamate, leucine, lysine, phenylalanine, threonine and valine) but lesser (P < 0.05) relative abundance of low density and very low-density lipids, and unsaturated lipids. Both PCA and OPLS-DA showed no clustering or discrimination between groups at d 0 and increasing divergence to d 14. Blood metabolomics is a useful tool to quantify the acute effects of stress in calves during the first 2 days after abrupt weaning, and longer-term changes in carbohydrate, lipid and protein metabolism due to nutritional changes from cessation of milk intake and greater reliance on forage intake.

Incorporation of Testicular Ultrasonography and Hair Steroid Concentrations in Bull Breeding Soundness Evaluation.

Testicular ultrasonography and steroid concentrations (cortisol, dehydroepiandrosterone sulfate (DHEA-S), cortisol/DHEA-S ratio, testosterone) in hair were examined for their utility in the bull breeding soundness evaluation (BBSE). Beef and dairy bulls (n = 16; 2.7 ± 0.4 years old; body condition score 3.2 ± 0.1) of five breeds were maintained under the same conditions at an accredited semen collection center. Bulls underwent routine semen collection twice weekly for 12 weeks and semen was processed and cryopreserved. Ultrasonography and hair sampling were undertaken at the last semen collection. Bulls with homogeneous testicular parenchyma (n = 8) had a higher (p < 0.05) percentage of motile sperm post-thawing compared with bulls with heterogeneous parenchyma (n = 8). There were no differences (p > 0.05) in the hair concentrations of cortisol, DHEA-S, and testosterone between bulls with homogeneous and heterogeneous parenchyma. In bulls with homogeneous parenchyma, hair DHEA-S concentration was positively correlated with percentage motile sperm (R2 = 0.76), progressively motile sperm (R2 = 0.70), and motility yield (R2 = 0.71). The findings indicate that the integration of testicular ultrasonography and hair DHEA-S status in the BBSE could provide a more comprehensive assessment of indicative fertility in bulls. Additionally, ultrasonography can be used in the BBSE when the evaluation of semen parameters is not available.

Plasma fatty acid profile in Italian Holstein-Friesian dairy cows supplemented with natural polyphenols from the olive plant Olea Europaea L.

This study evaluated the effects of supplementing with natural functional feed on the plasma fatty acid profile of lactating Italian Holstein-Friesian dairy cows. Thirty cows in mid-lactation received the natural olive extract PHENOFEED DRY (500 mg/cow/day) which mainly comprises hydroxytyrosol, tyrosol and verbascoside. The total content of polyphenols and the antioxidant power of standard feed, enriched feed and pure extract was evaluated respectively by Folin-Ciocalteu and DPPH assay, and a characterization in HPLC-UV (High-Performance Liquid Chromatography-Ultraviolet) of bioactive molecules present in the extract PHENOFEED DRY was performed. PHENOFEED DRY was provided for 60 days, and the plasma profile of fatty acids was determined by Gas Chromatography. The administration of enriched feed resulted in an increase in the ratio of Omega-6 to Omega-3 polyunsaturated fatty acids from 3:1 to 4:1 (p<0.001). This was not influenced by the calving order. The addition of polyphenols helped to keep monounsaturated (MUFA) and saturated (SFA) levels constant and results in a significant increase in polyunsaturated (PUFA) fatty acid after 15 days of administration. The Omega-6/Omega-3 ratio was in the optimal range. The findings show that inclusion of natural functional food such as plant polyphenols helps to maintain a healthy blood fatty acid profile in lactating dairy cows.

Local action of cytokines and immune cells in communication between the conceptus and uterus during the critical period of early embryo development, attachment and implantation - Implications for embryo survival in cattle: A review.

Early embryo development, implantation and pregnancy involve a complex dialogue between the embryo and mother. In cattle this dialogue starts as early as days 3-4 when the embryo is still in the oviduct, and it continues to implantation. Immunological processes involving cytokines, mast cells and macrophages form an important part of this dialogue. Amongst the cytokines, interleukin-6 (Il-6) and leukemia inhibitory factor (LIF) are secreted by both the embryo and uterine endometrium and form part of an ongoing and reciprocating dialogue. Mast cells and macrophages populate the uterine endometrium during embryo development and are involved in achieving the correct balance between inflammatory and anti-inflammatory reactions at the uterus that are associated with embryo attachment and implantation. Embryo loss is the major cause of reproductive wastage in cattle, and livestock generally. A deeper understanding of immunological processes during early embryo development will help to achieve the next step change in the efficiency of natural and assisted breeding.

Polyacrylate-GnRH Peptide Conjugate as an Oral Contraceptive Vaccine Candidate.

Contraceptive vaccines are designed to elicit immune responses against major components of animal reproductive systems. These vaccines, which are most commonly administered via injection, typically target gonadotropin-releasing hormone (GnRH). However, the need to restrain animals for treatment limits the field applications of injectable vaccines. Oral administration would broaden vaccine applicability. We explored contraceptive vaccine candidates composed of GnRH peptide hormone, universal T helper PADRE (P), and a poly(methylacrylate) (PMA)-based delivery system. When self-assembled into nanoparticles, PMA-P-GnRH induced the production of high IgG titers after subcutaneous and oral administration in mice. PADRE was then replaced with pig T helper derived from the swine flu virus, and the vaccine was tested in pigs. High levels of systemic antibodies were produced in pigs after both injection and oral administration of the vaccine. In conclusion, we developed a simple peptide-polymer conjugate that shows promise as an effective, adjuvant-free, oral GnRH-based contraceptive vaccine.

Estradiol Priming Potentiates the Kisspeptin-Induced Release of LH in Ovariectomized Cows.

The present study examined whether priming with estradiol benzoate (EB) for 12 h increased both the peak and duration of LH release in response to kisspeptin (KISS1, KP) in cows. In a Latin square design, ovariectomized Nelore cows (n = 8) received: Control, i.m. 4 mL of 0.9% saline; KP, i.m. 4 mg murine KISS1-10; EBKP, i.m. 4 mg KISS1-10 + i.m. 2 mg EB simultaneously; EB12KP, i.m. 4 mg KISS1-10 + i.m. 2 mg EB 12 h before KISS1-10. Concentrations of LH were determined in blood samples obtained at time 0 (treatment), 20, 40, 60, 90, 120, 150, 180, 210 and 270 min. Concentrations of LH were analyzed by Proc GLIMMIX for repeated measures. In case of significance, the adjusted Tukey test was used to test for differences among treatments. GraphPad 8.0 PRISM® was used to determine the area under the LH-response curve (AUC) after injection of KISS1-10. Plasma LH remained relatively constant throughout sampling after treatment with saline. The peak in LH after injection of KISS1-10 occurred at 20 min in Groups KP and EBKP and at 40 min in Group EB12KP. The peak LH response (∆LH, ng/mL) was greater (p < 0.01) in Group EB12KP (5.6 ± 0.9) than in Groups KP (2.4 ± 0.9) and EBKP (3.5 ± 0.9), which did not differ. AUC (LH ng/mL*min) was greater (p = 0.02) in Group EB12KP (439 ± 73) than in Groups KP (176 ± 73) and EBKP (241 ± 73), with the latter two groups not differing. The findings indicated that 12 h priming with EB increased both the peak and duration of the LH response to treatment with KISS1. The incorporation of EB priming and KISS1 could improve the efficiency of estrus synchronization with fixed-time AI in cows. This would have an important practical application in assisted breeding in beef and dairy cattle.

Effect of Breeding Techniques and Prolonged Post Dry Aging Maturation Process on Biomolecule Levels in Raw Buffalo Meat.

Recently, several concerns have been expressed on red meat quality and consumption. The aims of this study were to evaluate the influence of different breeding techniques and a prolonged post dry aging (PDA) maturation process on biomolecules level in raw buffalo meat. In the first experiment, two groups of animals were maintained with different space availability (15 vs. 10 m2/animal) for 90 days and biomolecules content was evaluated. In experiment 2, two diets (with or without ryegrass green forage) were used to assess the concentration of these biomolecules. Finally, in experiment 3, the meat of the animals that showed the highest content of biomolecules was chosen to assess the influence of the PDA maturation process. Buffaloes reared at 15 m2 showed a significantly (p < 0.05) higher content of the considered biomolecules compared with their counterparts. Similarly, buffaloes fed green forage showed higher content of biomolecules (p < 0.05) compared with the control group. The meat of the animals bred at 15 m2 and fed green forage showed a significant (p < 0.01) increase of biomolecules content during the PDA maturation process up to 60 days without influence microbiological profile in terms of total aerobic bacterial counts, yeasts, and molds. In conclusion, breeding techniques and PDA maturation system could enhance biomolecules levels in terms of quality, without affect health standards.

Green feed increases antioxidant and antineoplastic activity of buffalo milk: A globally significant livestock.

The effect of green feed on health-promoting biomolecules in milk was examined in dairy buffaloes. Buffaloes received a total mixed ration (TMR) (Control, C; n = 40) or TMR + alfalfa green feed (30% of diet) (Treated, T; n = 40). Biomolecules and functional activity were measured in milk obtained twice-monthly. Treated buffaloes had higher milk l-carnitine, acetyl-l-carnitine, propionyl-l-carnitine and δ-valerobetaine (P < 0.01). They also had higher antioxidant activity (P < 0.01). Compared with C buffaloes, milk of T buffaloes improved the viability of endothelial cells exposed to high-glucose (P < 0.01), and reduced intracellular lipid peroxidation, reactive oxygen species (ROS), and cytokine release (P < 0.01). Milk of T buffaloes inhibited with greater potency the viability of human HCT116 and Cal 27 cancer cells (P < 0.001). The findings show that including green feed in the diet of dairy buffaloes enhances health-promoting biomolecules and the antioxidant and antineoplastic properties of milk.

Changes in the blood metabolome of Wagyu crossbred steers with time in the feedlot and relationships with marbling.

Wagyu crossbred steers (n = 167) were used to (1) compare the metabolome of individual animals at two distant time-points (days 196 and 432) in a feedlot (this corresponded to 272 and 36 days before slaughter); and (2) determine relationships between the metabolome and marbling, and the effect of days in the feedlot (time-points) on these relationships. 1H NMR spectroscopy followed by standard recoupling of variables analysis produced 290 features or 'peaks' from which 38 metabolites were identified. There was a positive correlation between the relative concentration (RC) at days 196 and 432 for 35 of 38 metabolites (P > 0.05). The RC of 21 metabolites mostly involved in muscle energy and glucose metabolism increased (P < 0.05) from day 196 to 432, and the RC of 13 metabolites mostly involved in lipid metabolism decreased (P < 0.05). There were 14 metabolites correlated with marbling including metabolites involved in energy and fat metabolism (glucose, propionate, 3-hydroxybutyrate, lipids). The relationship between marbling and the RC of metabolites was affected by time-point, being positive for 3-hydroxybutyrate and acetate (P < 0.05) at day 432 but not at day 196. The findings indicate that the blood metabolome in Wagyu crossbred steers changes with time in a feedlot. Notwithstanding, the metabolome has potential to predict marbling in Wagyu. The ability to predict marbling from the blood metabolome appears to be influenced by days in a feedlot and presumably the stage of development towards a mature body conformation.