Preantral follicle numbers and size in heifers carrying Trio, a bovine high fecundity allele.

In brief: The bovine high fecundity allele, Trio, results in the occurrence of multiple ovulations and is characterized by antral follicles that develop slower and acquire ovulatory capacity at smaller sizes. This study provides novel information on the effect of the Trio allele on early folliculogenesis. Abstract: The bovine high fecundity allele, Trio, causes overexpression in granulosa cells (GCs) of SMAD6, an inhibitor of BMP15-activated SMAD signalling. Furthermore, the Trio allele results in antral follicles that develop slower, acquire ovulatory capacity at smaller sizes, and have three-fold greater ovulation rate compared to half-sib non-carriers. The present study was designed to determine preantral follicle numbers and size in Trio carrier and non-carrier cattle testing the hypothesis that inhibition of SMAD signalling would alter preantral follicle activation and/or growth. Ovarian tissues from Trio carrier (n = 12) and non-carrier (n = 12) heifers were obtained by laparotomy after follicle wave synchronization. Follicle numbers and dimensions were determined for each stage of development (primordial, transitional, primary, and secondary) from paraffin-embedded sections. There were no differences in the number of primordial, transitional, or secondary follicles or in antral follicle count, circulating AMH, or ovarian volume between carriers and non-carriers. Trio carriers had ~2.5-fold greater (P < 0.01) number of primary follicles than non-carriers, and transitional and primary follicles were larger (~1.2-fold; P < 0.1) in Trio carriers. Oocyte volume of primordial and transitional follicles was not different between genotypes; however, oocytes were larger (P < 0.05) in primary (~1.3-fold) and secondary (~1.8-fold) follicles for Trio carriers. Granulosa cell numbers were not different (P > 0.3) between carriers and non-carriers, irrespective of the stage of development. These results suggest that, after primordial follicle activation, follicles in Trio carrier cattle have slower progression through the primary stage, hence the larger oocyte and greater number of primary follicles.

Effect of use and dosage of p-follicle-stimulating hormone for ovarian superstimulation before ovum pick-up and in vitro embryo production in pregnant Holstein heifers.

The benefit of ovarian superstimulation using exogenous FSH before ovum pick-up (OPU) and in vitro embryo production (IVEP) has been the subject of conflicting results. The objective of the present study, therefore, was to evaluate the effect of use and dose of porcine FSH (p-FSH) before OPU/IVEP on ovarian response and embryo production in pregnant heifers. Pregnant Holstein heifers (n = 48) were randomly assigned to receive 0, 160, or 300 mg NIH-FSH-P1 in a crossover design. Ovum pick-up was performed at 49, 63, and 77 d of gestation with a 14 d "washout" between OPU sessions. Follicle ablation was performed on D 0 (p.m.) and p-FSH treatments, consisting of 4 decreasing dose injections administered 12 h apart, were initiated 36 h after follicle ablation (d 2 a.m.). Heifers underwent OPU on D 5 (a.m.), 40 h after the last p-FSH treatment, and cumulus oocyte complexes (COC) were subjected to IVEP procedures. Differences between treatment groups were evaluated using generalized linear mixed models. There were quadratic effects of treatment on both number and percentage of small (<6 mm), medium (6-10 mm), and large (>10 mm) follicles. Number and percentage of medium follicles increased with increasing p-FSH dosages, although the magnitude of the change was greater between 0 and 160 mg, than between 160 and 300 mg of p-FSH. Total number of follicles, number of COC recovered and number of viable COC increased linearly with increasing p-FSH dose. Conversely, there was no evidence for an effect of p-FSH dose on COC recovery percentage nor the percentage of viable COC. Cleavage percentage, number of cleaved oocytes, blastocyst percentage, and number of blastocysts increased linearly with increasing p-FSH dose. In conclusion, use of p-FSH before OPU resulted in greater superstimulatory response and oocyte competence which in turn increased IVEP. Furthermore, these effects were dose dependent such that use of a greater dose of p-FSH up to 300 mg progressively increased embryo yield.

Selection of fewer dominant follicles in Trio carriers given GnRH antagonist and luteinizing hormone action replaced by nonpulsatile human chorionic gonadotropin†.

Studying selection of multiple dominant follicles (DFs) in monovulatory species can advance our understanding of mechanisms regulating selection of single or multiple DFs. Carriers of the bovine high fecundity Trio allele select multiple DFs, whereas half-sib noncarriers select a single DF. This study compared follicle selection during endogenous gonadotropin pulses versus during ablation of pulses with Acyline (GnRH antagonist) and luteinizing hormone (LH) action replaced with nonpulsatile human chorionic gonadotropin (hCG) treatment in Trio carriers (n = 28) versus noncarriers (n = 32). On Day 1.5 (Day 0 = ovulation), heifers were randomized: (1) Control, untreated; (2) Acyline, two i.m. doses (Days 1.5 and D3) of 3 µg/kg; (3) hCG, single i.m. dose of 50 IU hCG on Day 1.5 followed by daily doses of 100 IU; and (4) Acyline + hCG. Treatments with nonpulsatile hCG were designed to replace LH action in heifers treated with Acyline. Acyline treatment resulted in cessation of follicle growth on Day 3 with smaller (P < 0.0001) maximum follicle diameter in Trio carriers (6.6 ± 0.2 mm) than noncarriers (8.7 ± 0.4 mm). Replacement of LH action (hCG) reestablished follicle diameter deviation and maximum diameter of DFs in both genotypes (8.9 ± 0.3 mm and 13.1 ± 0.5 mm; P < 0.0001). Circulating follicle stimulating hormone (FSH) was greater in Acyline-treated than in controls. Finally, Acyline + hCG decreased (P < 0.0001) the number of DFs from 2.7 ± 0.2 to 1.3 ± 0.2 in Trio carriers, with most heifers having only one DF. This demonstrates the necessity for LH in acquisition of dominance in Trio carriers (~6.5 mm) and noncarriers (~8.5 mm) and provides evidence for a role of GnRH-induced FSH/LH pulses in selection of multiple DFs in Trio carriers and possibly other physiologic situations with increased ovulation rate.

Trio a novel bovine high-fecundity allele: II. Hormonal profile and follicular dynamics underlying the high ovulation rate.

The newly discovered Trio high-fecundity allele produces multiple ovulations in cattle. This study evaluated (1) size and growth rates of follicles in Trio carriers during a synchronized follicular wave, induced by follicle aspiration; (2) follicle-stimulating hormone (FSH) patterns associated with the follicular wave; (3) size of corpora lutea (CL) and circulating progesterone; and (4) intrafollicular estradiol concentrations prior to normal deviation. Trio carriers had mean dominant follicles that were significantly smaller in diameter and volume than noncarriers. Onset of diameter deviation occurred at ∼3 days after the last follicle aspiration in both genotypes despite Trio carriers having much smaller individual follicles. Follicles of Trio carriers grew at a slower rate than noncarrier follicles (∼65% in mm/day or ∼30% in mm3/day) resulting in much smaller individual dominant follicles (∼25% volume). However, total dominant follicle volume, calculated as the sum of all dominant follicles in each animal, was similar in carriers and noncarriers of Trio throughout the entire follicular wave. Circulating FSH was greater in Trio carriers during the 24 h encompassing deviation. Trio carriers had significantly more ovulations than noncarriers, and individual CL volume was smaller, although total luteal tissue volume and circulating P4 were not different. Thus, increased ovulation rate in Trio carriers relates to smaller individual follicles (one-third the volume) near the time of deviation due to slower follicle growth rate, although time of deviation is similar, with increased circulating FSH near deviation leading to selection of multiple dominant follicles in Trio carriers with similar total follicle volume.

Trio, a novel high fecundity allele: I. Transcriptome analysis of granulosa cells from carriers and noncarriers of a major gene for bovine ovulation rate.

A major gene for bovine ovulation rate has been mapped to a 1.2 Mb region of chromosome 10. Screening of coding regions of positional candidate genes within this region failed to reveal a causative polymorphism, leading to the hypothesis that the phenotype results from differences in candidate gene expression rather than alteration of gene structure. This study tested differences in expression of positional candidate genes in granulosa cells between carriers and noncarriers of the high fecundity allele, as well as characterizing differences in the transcriptomic profile between genotypes. Five carriers and five noncarriers, female descendants of "Trio," a carrier of the high fecundity allele were initially used in an RNA-seq analysis of gene expression. Four of ten samples were contaminated with theca cells, so that six samples were used in the final analysis (three of each genotype). Of 14 973 genes expressed, 143 were differentially expressed (false discovery rate P < 0.05) in carriers versus noncarriers. Among the positional candidate genes, SMAD6 was 6.6-fold overexpressed in the carriers compared to noncarriers (P < 5 × 10-5). This result was replicated in an independent group of 12 females (7 carriers and 5 noncarriers) using quantitative real-time PCR; SMAD6 was 9.3-fold overexpressed in carriers versus noncarriers (P = 1.17 × 10-6). Association of overexpression of SMAD6, an inhibitor of the BMP/SMAD signaling pathway, with high ovulation rate corresponds well with disabling mutations in ligands (BMP15 and GDF9) and a receptor (BMPR1B) of this pathway that cause increased ovulation rate in sheep.

Mechanisms for rescue of corpus luteum during pregnancy: gene expression in bovine corpus luteum following intrauterine pulses of prostaglandins E1 and F2α.

In ruminants, uterine pulses of prostaglandin (PG) F2α characterize luteolysis, while increased PGE2/PGE1 distinguish early pregnancy. This study evaluated intrauterine (IU) infusions of PGF2α and PGE1 pulses on corpus luteum (CL) function and gene expression. Cows on day 10 of estrous cycle received 4 IU infusions (every 6 h; n = 5/treatment) of saline, PGE1 (2 mg PGE1), PGF2α (0.25 mg PGF2α), or PGE1 + PGF2α. A luteal biopsy was collected at 30 min after third infusion for determination of gene expression by RNA-Seq. As expected, IU pulses of PGF2α decreased (P < 0.01) P4 luteal volume. However, there were no differences in circulating P4 or luteal volume between saline, PGE1, and PGE1 + PGF2α, indicating inhibition of PGF2α-induced luteolysis by IU pulses of PGE1. After third pulse of PGF2α, luteal expression of 955 genes were altered (false discovery rate [FDR] < 0.01), representing both typical and novel luteolytic transcriptomic changes. Surprisingly, after third pulse of PGE1 or PGE1 + PGF2α, there were no significant changes in luteal gene expression (FDR > 0.10) compared to saline cows. Increased circulating concentrations of the metabolite of PGF2α (PGFM; after PGF2α and PGE1 + PGF2α) and the metabolite PGE (PGEM; after PGE1 and PGE1 + PGF2α) demonstrated that PGF2α and PGE1 are entering bloodstream after IU infusions. Thus, IU pulses of PGF2α and PGE1 allow determination of changes in luteal gene expression that could be relevant to understanding luteolysis and pregnancy. Unexpectedly, by third pulse of PGE1, there is complete blockade of either PGF2α transport to the CL or PGF2α action by PGE1 resulting in complete inhibition of transcriptomic changes following IU PGF2α pulses.

Trio, a novel bovine high fecundity allele: III. Acquisition of dominance and ovulatory capacity at a smaller follicle size.

The acquisition of dominance and ovulatory capacity was evaluated in follicles from cows that were carriers or half-sibling noncarriers of the Trio allele. Follicle size at acquisition of follicular dominance was determined by evaluating whether follicles ovulate after GnRH challenge (ovulatory capacity-experiment 1) and by determination of intrafollicular concentrations of estradiol and free insulin like growth factor 1 (IGF1) and relative mRNA expression of cytochrome P450 family 19 subfamily A member 1 (CYP19A1), luteinizing hormone/choriogonadotropin receptor (LHCGR), and pappalysin 1 (PAPPA, previously known as pregnancy-associated plasma protein A, pappalysin 1) in granulosa cells from follicles of different sizes (experiment 2). Ovulatory capacity developed in follicles at 8.3 mm (50% ovulatory capacity) in noncarriers but at smaller sizes (5.5 mm) in Trio carriers. Similarly, in experiment 2, follicles of Trio carriers acquired a dominant phenotype, as determined by intrafollicular estradiol and CYP19A1, LHCGR, and PAPPA mRNA expression in granulosa cells, at significantly smaller sizes but at a similar time after wave emergence. Overall, dominance/ovulatory capacity was acquired when follicles of Trio carriers were ∼30% the size (volume basis) of follicles in noncarriers. In addition, follicles in Trio carriers appear to acquire dominance in a hierarchal manner, as demonstrated by the progressively greater number of follicles with a dominant phenotype between days 2 and 4 after wave emergence. Thus, results from this study provide further support for a physiological model in which selection of multiple follicles in Trio allele carriers is characterized by acquisition of dominance at a smaller follicle size but at a similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence.

Proteomic analysis of follicular fluid in carriers and non-carriers of the Trio allele for high ovulation rate in cattle.

This study was conducted to characterise differences in follicular fluid proteins between carriers and non-carriers of a bovine allele for high ovulation rate. A total of four non-carrier and five carrier females were used in an initial study with four and six additional non-carriers and carriers respectively used in a validation study. Emergence of the follicular wave was synchronised and the ovaries containing the dominant follicle(s) were extracted by ovariectomy for follicular fluid collection. A hexapeptide ligand library was used to overcome the masking effect of high-abundance proteins and to increase detection of low-abundance proteins in tandem mass spectrometry. After correcting for multiple comparisons, only two proteins, glia-derived nexin precursor (SERPINE2) and inhibin ß B chain precursor (INHBB), were significantly differentially expressed (false-discovery rate <0.05). In a replicate study of analogous design differential expression was confirmed (P<0.05). Joint analysis of results from the two studies indicated that three additional proteins were consistently differentially expressed between genotypes. For three of these five, previous studies have indicated that expression is increased by transforming growth factor-ß-bone morphogenetic protein signalling; their reduction in follicular fluid from carrier animals is consistent with the ~9-fold overexpression of SMAD family member 6 (SMAD6) in carriers that is inhibitory to this pathway.

Increase in average testis size of Canadian beef bulls.

Selection for adequate testis size in beef bulls is an important part of bull breeding soundness evaluation. Scrotal circumference (SC) is highly correlated with paired testis weight and is a practical method for estimating testis weight in the live animal. Most bulls presented for sale in Canada have SC included in the presale information. Scrotal circumference varies by age and breed, and may change over time due to selection for larger testis size. Therefore, it is important to periodically review the mean SC of various cattle breeds to provide valid bull selection criteria. Scrotal circumference data were obtained from bulls sold in western Canada from 2008 to 2011 and in Quebec from 2006 to 2010. Average scrotal circumferences for the most common beef breeds in Canada have increased significantly in the last 25 years. Differences between breeds have remained unchanged and Simmental bulls still have the largest SC at 1 year of age. Data provided here could aid in the establishment of new suggested minimum SC measurements for beef bulls.

Augmentation de la taille moyenne des testicules chez les taureaux de boucherie canadiens. La sélection de la taille adéquate des testicules chez les taureaux de boucherie représente une partie importante de l'évaluation de l'aptitude à l'utilisation comme reproducteur du taureau. La circonférence scrotale (CS) présente une corrélation élevée avec le poids des paires de testicules et est une méthode pratique d'estimer le poids des testicules chez l'animal vivant. La CS de la plupart des taureaux mis en vente au Canada est incluse dans les renseignements de prévente. La circonférence scrotale varie selon l'âge et la race et peut changer au fil du temps en raison de la sélection pour une taille supérieure de testicules. Par conséquent, il est important d'examiner périodiquement la CS moyenne des diverses races bovines afin de fournir des critères de sélection valides pour les taureaux. Les données sur la circonférence scrotale ont été obtenues de taureaux vendus dans l'Ouest canadien de 2008 à 2011 et au Québec de 2006 à 2010. Les circonférences scrotales moyennes pour les races bovines les plus communes ont augmenté significativement au cours des 25 dernières années. Les différences entre les races sont demeurées inchangées et les taureaux Simmental possèdent toujours la CS la plus grande à l'âge de 1 an. Les données fournies ici pourraient faciliter l'établissement de nouvelles mesures minimales suggérées pour la CS des taureaux de boucherie.(Traduit par Isabelle Vallières).

Application of a new diagnostic approach to a bovine genital campylobacteriosis outbreak in a Saskatchewan beef herd.

A new real-time quantitative polymerase chain reaction (qPCR) test was used to diagnose Campylobacter fetus subsp. venerealis infection associated with dramatic reproductive losses in a commercial cow-calf herd. The results were verified with repeated culture, phenotypic characterization of the organism and DNA sequencing. This case demonstrates the need for a practical field test for C. fetus subsp. venerealis and the importance of considering this organism as a potential cause of pregnancy failure in beef herds.

Application d'une nouvelle approche diagnostique lors d'une éclosion de campylobactériose génitale bovine dans un troupeau bovin de la Saskatchewan. Un nouveau test quantitatif d'amplification en chaîne par la polymérase en temps réel (qACP) a été utilisé pour diagnostiquer une infection par Campylobacter fetus sous-espèce venerealis associée à une baisse spectaculaire de la reproduction dans un troupeau commercial de vaches-veaux. Les résultats ont été vérifiés à l'aide de cultures répétées, d'une caractérisation phénotypique de l'organisme et du séquençage de l'ADN. Ce cas démontre le besoin d'un test sur le terrain pratique pour C. fetus sous-espèce venerealis et l'importance de considérer cet organisme comme une cause potentielle d'échec de la gestation dans les troupeaux bovins.(Traduit par Isabelle Vallières).

Effects of polyunsaturated fatty acid supplementation on plasma and follicular fluid resolvin D1 concentration and mRNA abundance in granulosa cells in ewes.

The aim of this experiment was to evaluate the effect of increasing dietary omega-3 (n-3) polyunsaturated fatty acid (PUFA) supplementation on plasma and follicular fluid resolvin D1 (RvD1) concentration and the mRNA expression of genes related to RvD1 production, inflammatory response, oxidative stress, hormone receptors and production, and free fatty acid receptors in the granulosa cells of ewes. Dorset × Hampshire ewes (n = 24) aged 2 to 4 yr and with an initial body weight (BW) of 84.08 ±â€…13.18 kg were blocked by body condition score (BCS) and BW, and randomly assigned to 12 pens. Each pen within each block was randomly assigned to one of three treatments: 1) diet without fatty acid supplementation (control), 2) diet with 0.5% n-3 PUFA supplementation (PUFA0.5), and 3) diet with 1% n-3 PUFA supplementation (PUFA1). BW, BCS, and blood samples were obtained on day 1 and every 21 d for 3 mo. Ewes were then synchronized, superstimulated, and ovariectomized. Antral follicles were aspirated to evaluate RvD1 concentration in follicular fluid, and granulosa cells were used to determine mRNA abundance. Data were analyzed as a randomized complete block design using a mixed model (MIXED or GLIMMIX with log as a link function when data presented a nonnormal distribution). A polynomial effect of treatments was used to analyze RvD1 concentration and mRNA expression when there was no interaction. In addition, the correlation between plasma and follicular fluid RvD1 concentration was evaluated. We found no differences in BW (P = 0.28) and BCS (P = 0.29) between treatments. The concentration of RvD1 in plasma and follicular fluid linearly increased (P = 0.03) and tended to increase (P = 0.06) concomitantly to increasing PUFA supplementation. Plasma and follicular fluid RvD1 concentrations were positively correlated (r = 0.61; P < 0.01). The abundance of GPX1 and GPR32 mRNA tended to increase linearly with increasing PUFA supplementation (P = 0.06). In addition, PUFA supplementation linearly decreased and tended to decrease IL-1ß and COX-2 mRNA abundance (P = 0.01 and P = 0.06, respectively). In conclusion, the correlation between plasma and follicular fluid RvD1 concentration indicates a relationship between both compartments. Also, the decrease of IL-1ß and the increase of GPX1 mRNA abundance after PUFA supplementation could have beneficial effects on follicle development.

The aim of this experiment was to evaluate the effects of increasing dietary omega-3 (n-3) polyunsaturated fatty acid (PUFA) supplementation on plasma and follicular fluid resolvin D1 (RvD1) concentration, and the mRNA expression of genes related to RvD1 synthesis, inflammatory response, oxidative stress, reproductive hormone receptors and production, and free fatty acid receptors in ewes. Twenty-four ewes aged 2 to 4 yr were assigned to 12 pens and randomly allocated to one of three treatments: 1) diet without fatty acid supplementation (control), 2) diet with 0.5% n-3 PUFA (PUFA0.5), and 3) diet with 1% n-3 PUFA (PUFA1). Body weight (BW), body condition score (BCS), and blood samples were obtained on day 1 and every 21 d for 3 mo. Ewes were then synchronized, superstimulated, and ovariectomized. Antral follicles were aspirated to evaluate follicular fluid RvD1 concentration, and granulosa cells were used to analyze mRNA concentration. We found no differences in BW and BCS between treatments. Plasma and follicular fluid RvD1 concentration increased concomitantly to increasing dietary PUFA supplementation. There was a positive correlation between plasma and follicular fluid RvD1 concentrations. Omega-3 PUFA increased the mRNA abundance of genes associated with RvD1 synthesis and oxidative damage response. In addition, PUFA supplementation linearly decreased the mRNA abundance of genes associated with inflammatory response. In conclusion, the positive correlation between plasma and follicular fluid RvD1 concentrations demonstrates a relationship between both compartments. Also, changes in gene expression after PUFA supplementation may have a beneficial effect on the follicle and, in turn, on reproduction.

Environmental stress during the last trimester of gestation in pregnant cows and its effect on offspring growth performance and response to glucose and adrenocorticotropic hormone.

Winter and spring precipitation are predicted to increase in the Midwest region of the United States, causing muddy conditions. In a previous experiment, Angus cows (8 per treatment) were paired based on initial body weight (BW) and one cow from each pair was randomly allocated to either the mud or control treatment. Though cows consumed the same amount of dry matter, cows in the mud treatment weighed 37.4 kg less than cows in the control treatment by day 269 of gestation. The objective of this experiment was to evaluate developmental programming effects of steers born to cows in the mud treatment (MUD; n = 7) or the control treatment (CON; n = 6). Steers were weighed at birth and then weekly from approximately 56 d of age until weaning and were subjected to a glucose tolerance test (GTT) and adrenocorticotropic hormone (ACTH) challenge after weaning. Steers were then placed in the feedlot for an 84-d growing phase and were weighed weekly and 12th rib back fat (BF) and ribeye area (REA) were imaged every 28 d using ultrasonography. Data were analyzed as a randomized complete block design with repeated measurements when appropriate (SAS 9.4). Although there was a 37.4 kg decrease in BW of cows by the end of gestation, there was no evidence of a pen treatment effect on calf birth weight (P = 0.60) or weaning weight (P = 0.99). Additionally, there was no evidence of a pen treatment × day effect for steer BW from birth to weaning (P = 0.67) or growing phase BW (P = 0.60). There was evidence of a treatment × day of growing phase effect (P = 0.02) for BF, such that CON steers had greater BF on day 28 of the growing phase; however, there was no evidence of a treatment × day effect for REA (P = 0.20). Furthermore, there was no evidence of a pen treatment effect for the growing phase average daily gain (P = 0.74), dry matter intake (P = 0.65), gain:feed (P = 0.48), plasma glucose concentration (P = 0.67) or plasma insulin concentration (P = 0.61) in response to the GTT, or plasma cortisol concentration in response to the ACTH challenge (P = 0.51). These results indicate that while mud increased net energy requirements for cows in the MUD treatment, there were no subsequent effects observed for steer BW, gain:feed, or response to glucose and ACTH during the growing phase.

Predictions for the Midwest U.S. indicate that both winter and spring temperatures and precipitation will increase. These climatic changes could result in muddier conditions during winter and spring in the Midwest. It has previously been demonstrated that a muddy environment increases the net energy requirements of mature cow's by approximately 3.9 Mcal/d. If cows are not provided this extra energy and are exposed to mud during late gestation, it is likely that this will cause an adverse environment for the conceptus that could result in compromised growth and metabolism later in life. This study evaluated the developmental programming effects after birth and into the growing phase with beef steers born to cows that were housed in a muddy environment during late gestation compared with steers born to cows that were housed in pens bedded with wood chips during late gestation. Based on the present results, the mature cows housed in the muddy conditions weighed approximately 37.4 kg less than cows housed in pens bedded with wood chips during late gestation, however, calf birth weight, postnatal growth, postnatal feed intake and gain:feed, and postnatal response to glucose and adrenocorticotropic hormone was not affected. This indicates that the mature cows were able to mobilize body stores and supplied the fetus with adequate nutrients during gestation without impairment of growth or postnatal response to glucose and adrenocorticotropic hormone.

Research on timed AI in beef cattle: Past, present and future, a 27-year perspective.

This review aimed to (1) summarize the results from fixed-timed artificial insemination (TAI) fertility studies performed during the last 27 years; (2) compile and evaluate, as examples from the literature base, the direct comparisons made of specific manipulations to synchronization protocols; (3) evaluate the impact of the TAI programs on the reproductive performance during the breeding season, and (4) provide perspective on the future of TAI programs in beef cattle. A search of the literature published from 1995 to 2021 was conducted to identify experiments in which synchronization of ovulation and TAI in beef cattle was performed. The primary outcome of interest was fertility expressed as pregnancies per TAI. The literature included two search engines, the SIS Web of Science and the US National Library of Medicine Institutes of Health through PubMed. After the initial search and screening, a total of 228 manuscripts were selected containing a total of 272,668 TAI. A dramatic increase in the number of publications and TAIs occurred throughout the years. Most of them were from Brazil and United States, followed by Canada, Argentina, Uruguay, and Australia. Two main types of TAI programs were identified: GnRH-based and E2/P4-based protocols. In terms of GnRH-based programs, two variations were evaluated in the present manuscript. First, we evaluated the effect of the progesterone implant during the protocol. The progesterone implant increased pregnancy/TAI (P/TAI) from 44.3 to 54.3%. Second, the use of a second prostaglandin F2α treatment in 5-d CO-synch program increased the P/TAI from 53.2 to 60.9%. In E2/P4-based programs, use of GnRH at TAI increased P/TAI from 54.7 to 59.2% in cows. However, no increase was detected in heifers. Other research showed that use of TAI can increase the overall proportion of the cows pregnant at end of the breeding season and produce earlier calvings compared with bulls. In conclusion, there have been a large number of excellent research studies that have been performed during the last 27 years on TAI in beef cattle. This technology is being utilized successfully in the beef cattle industry. This success is largely because of the valid research that underlies the application of the technology and the economic value of the technology.

Short communication: A comparison between two glucose measurement methods in beef steers during a glucose tolerance test.

Glucose tolerance tests (GTT) are commonly performed in beef cattle to evaluate the glucose-insulin signaling pathway. Blood samples are obtained via a catheter and then transferred back to the laboratory for further analysis. A hand-held glucometer used chute-side can make performing GTT's and quantifying blood glucose concentration much easier and faster for research purposes. The purpose of this study was to evaluate the agreement between a hand-held electronic glucometer (Precision Xtra; Abbott Diabetes Care Inc., Mississauga, ON, Canada) for chute-side use in beef cattle compared with a colorimetric assay in the laboratory (Stanbio Glucose LiquiColor; Stanbio Laboratory, Boerne, TX, USA). A GTT was performed on 13 Simmental × Angus steers during the growing phase. Blood samples were obtained via a jugular catheter. Glucometer readings were taken immediately after blood was sampled from the jugular with no preservative, and laboratory measurements were conducted on plasma preserved with sodium fluoride. A paired t-test (P = 0.40), Pearson's correlation (P < 0.001; r = 0.95), Bland-Altman plot, and Lin's concordance correlation coefficient (LCCC = 0.90) were completed to evaluate the performance of the glucometer relative to the results from the laboratory assay. Based on the results, we conclude that the glucometer is an acceptable method for measuring blood glucose concentration in beef cattle under field conditions.

Beef cows housed in mud during late gestation have greater net energy requirements compared with cows housed on wood chip bedding.

Mud increases net energy requirements for cattle because mud and precipitation compromise the ability of the hair coat to insulate and maintain core body temperature of the cow. The increase in energy required for a gestating cow to compensate for a muddy environment is unknown. The objective of this study was to evaluate effects of muddy conditions on cow body weight (BW) and fetal growth during late gestation. Sixteen multiparous Angus cows (n = 8/treatment) were paired based on initial BW and one cow from each pair was randomly allocated to either the mud (MUD) or control (CON) treatment on day 213 of gestation. Pens in the CON group were bedded with wood chips, while pens in the MUD group were designed to create a muddy lot (average depth of 23.6 ± 5.8 cm). Cows were housed outdoors individually and fed the same diet that consisted of a limit-fed total mixed ration. Each pair was fed to meet energy and protein requirements for maintenance and gestation. From day 213 to 269 of gestation, cows were weighed and sampled for blood metabolites weekly. Data were analyzed as a randomized complete block design with repeated measurements (SAS 9.4). Though cows consumed the same amount of dry matter, cows in the MUD treatment weighed 37.4 kg less than cows in the CON treatment (P < 0.01) by day 269 of gestation. Cows in the MUD treatment decreased approximately half a body condition score (BCS), while cows in the CON treatment gained approximately 1 BCS during the treatment period (P < 0.01). There was no evidence of a treatment × day of gestation effect for 12th rib back fat (P = 0.85), rump fat (P = 0.48), total plasma protein concentrations (P = 0.85), or plasma 3-methylhistidine (P = 0.84); however, there was a marginally significant treatment × day of gestation effect for plasma non-esterified fatty acid concentration (P = 0.09). Despite differences in cow BW at the end of the treatment period, calf birth weight (P = 0.66) and calf total plasma protein (P = 0.27) were not different; however, the divergence in cow BW remained marginally significant at parturition (P = 0.06). These results indicate that mud increased net energy requirements for cows in the MUD treatment, as calf birth weight was not different but maternal BW was decreased compared with cows in the CON treatment.

Synchronization of follicle wave emergence before ovarian superstimulation with FSH and ovum pick-up improves in vitro embryo production in pregnant heifers.

The objective of the present study was to evaluate effects of synchronization of timing of follicle wave emergence, before ovarian superstimulation and ovum pick-up (OPU), on ovarian response and embryo production in pregnant heifers. Pregnant (47-69 days of gestation) Holstein heifers (n = 64), 19.0 ± 0.3 months of age, were assigned in a completely randomized design to one of two groups: synchronization of follicular wave emergence using follicle ablation (Synchronized) or untreated control (Non-synchronized). Superstimulatory treatments consisting of 160 mg (280 IU) of porcine follicle stimulating hormone (p-FSH), administered in four decreasing dose treatments 12 h apart, were initiated 36 h after follicle ablation or at random stages of the follicular wave in heifers of the Synchronized and Non-synchronized group, respectively. Ovum pick-up was performed in all heifers 40 h after the last p-FSH administration and retrieved cumulus oocyte complexes (COCs) were subjected to in vitro embryo production (IVEP) procedures. Ultrasonography was performed immediately before OPU to determine number and size of ovarian follicles. Differences in treatment responses between groups were evaluated using generalized linear mixed models. Total number of follicles at the time of OPU was not different between treatment groups (P = 0.61), however, the number of small follicles (<6 mm) was greater (P = 0.05) in heifers of the Non-synchronized group, whereas number of medium size follicles (6-10 mm) tended (P = 0.09) to be greater in heifers of the Synchronized group. Heifers in the Synchronized group had a greater (P = 0.01) percentage of medium sized follicles and a lesser (P = 0.01) percentage of smaller sized follicles than heifers in the Non-synchronized group. There were no differences (P > 0.15) in total number of recovered COCs, or number of viable COCs between groups. Cleavage percentage (84.5% and 72.8%) and blastocyst percentage (48.2% and 33.4%) were greater (P < 0.01) in heifers of the Synchronized than Non-synchronized group, respectively. As a result, mean number of blastocysts per OPU/heifer was greater (P = 0.006) in the heifers of the Synchronized (8.9 ± 1.0) than the Non-synchronized (5.5 ± 0.9) group. In conclusion, synchronizing the time of follicle wave emergence in pregnant heifers, prior to ovarian superstimulation with FSH and OPU results in a greater superstimulatory response and oocyte competence leading to greater embryo production.

Energy restriction and housing of pregnant beef heifers in mud decreases body weight and conceptus free live weight.

Average temperatures in the Midwest, USA are predicted to increase 2-9°C by the end of the century; resulting in muddy pastures for spring calving beef heifers as they enter late gestation. The objective of this study was to evaluate the effects of muddy conditions on heifer body weight (BW), body condition score (BCS), conceptus free live weight (CFLW), and fetal growth when heifers were energy restricted during late gestation. Eighteen Angus heifers (n = 9/treatment) were paired based on initial BW. One heifer from each BW pair was randomly allocated to either the mud (MUD) or control (CON) treatment on day 196 of gestation. Pens in the CON treatment were bedded with wood chips, while pens in the MUD treatment were filled with mud (average depth of 19.5 ±â€…7.9 cm). Heifers were housed individually and fed the same diet that consisted of a limit-fed total mixed ration from day 196 to 266 of gestation that was formulated to meet 66% of the net energy for maintenance, growth, and gestation requirements. Requirements and the amount of the diet offered were adjusted weekly, and heifers were weighed and sampled for blood metabolites weekly. Data were analyzed as a randomized complete block design with repeated measurements. There was a treatment × day of gestation interaction, such that heifers had similar BW, BCS, and CFLW on day 196 of gestation. By day 266 of gestation; however, heifers in the MUD treatment weighed 43.5 kg less (P < 0.01) and were 1.8 BCS units less (P < 0.01) than heifers in the CON treatment. This is further supported by the treatment × day effects we observed for back fat (BF) and rump fat (RF) thickness, such that the MUD heifers had less BF (P = 0.02) and RF (P < 0.01) by day 266 of gestation. There was a marginally significant difference for gestation length (P = 0.06), such that heifers in the MUD treatment calved approximately 3.1 days before the heifers in the CON treatment. Though heifers in the MUD treatment decreased their BW and CFLW during the treatment period, we did not observe a difference in calf birth weight (P = 0.34), calf plasma IgG concentration (P = 0.37), or calf weaning weight (P = 0.63). Despite heifers in the MUD treatment having greater BW, CFLW, and BCS losses compared with the heifers in the CON treatment, the heifers in the MUD treatment seemed to prioritize fetal growth, as they mobilized their body tissues to meet the energetic demands of pregnancy.

Postovulatory treatment with GnRH on day 5 reduces pregnancy loss in recipients receiving an in vitro produced expanded blastocyst.

The present study tested the hypothesis that administration of GnRH on day 5 of the estrous cycle in embryo transfer (ET) recipients would increase progesterone (P4) concentrations, embryo size, and improve fertility. Holstein and cross-bred Holstein heifers (n = 1562) were synchronized using a modified 5-day CIDR-Synch protocol as follows (All AM treatments): D-8, CIDR inserted; D-3, CIDR removed and PGF2α (500 µg cloprostenol) treatment; D-2, second PGF2α; D0, GnRH (G1, 100 µg gonadorelin acetate) to induce ovulation. On D5 in the afternoon, heifers were assigned in a completely randomized design to one of two treatments: Control (untreated) or GnRH (200 µg). Transfer of day 7 fresh IVP embryos was performed between D6 and D8 after G1. Data collected from each heifer included: embryo stage and quality, body condition score, technician performing ET, interval from G1 to ET, and number of previous transfers. All heifers were evaluated by transrectal ultrasonography on D5, D33, and D60 and a subset of heifers was scanned on D12 (n = 718; to determine ovulation to treatment) and another subset on D33 (n = 295; 16 s video to determine embryo and amniotic vesicle size). Serum P4 was determined from a subset of heifers on D12 (n = 467) and on D21 (n = 837) and pregnancy specific protein B (PSPB) on D28 (n = 843). Pregnancies per ET (P/ET) were analyzed by logistic regression and continuous outcomes by ANOVA. Ovulation to D5 GnRH, defined by the presence of an accessory CL on D12, was 83.9% (302/360) in GnRH-treated heifers vs. 3.3% (12/358) in Controls (P < 0.001). On D12, P4 was greater (P < 0.001) in GnRH-treated heifers (7.2 ±â€¯0.1 ng/ml) vs Controls (6.0 ±â€¯0.1 ng/ml). There was greater P/ET at D33 and D60 of pregnancy for Stage 7 than Stage 6 embryos. Treatment with GnRH did not alter P/ET with either embryo stage but decreased pregnancy loss between D33 and D60 in heifers receiving Stage 7 embryos. Presence of an accessory CL at the D33 pregnancy diagnosis was associated with a larger reduction in pregnancy loss from D33 to D60 in recipients of Stage 7 embryos (11.6 vs 27.6%). Although there was no GnRH effect on embryo size, the presence of an accessory CL was associated (P < 0.05) with larger amniotic vesicle volume in recipients of Stage 7 embryos. In addition, greater PSPB was linked to greater amniotic vesicle volume (P = 0.01) and to reduced pregnancy loss (P < 0.0001). In conclusion, treatment with GnRH on D5 caused ovulation and formation of an accessory CL, increased circulating P4, and reduced pregnancy loss in heifers receiving a Stage 7 but not a Stage 6 IVP embryo.

Mechanisms regulating follicle selection in ruminants: lessons learned from multiple ovulation models.

Selection of a single dominant follicle from a cohort of growing follicles is a unique biological process, a key step in female reproductive function in monovular species, and lies at the core of reproductive technologies in cattle. Follicle growth and the number of follicles that ovulate are regulated by precise endocrine, paracrine, and autocrine mechanisms. Most of our current understanding about follicle selection focuses on the role of FSH, LH, and the IGF family in follicle growth and selection of the dominant follicle. However, more recently the role of members of the TGF-ß family has been highlighted, particularly in high fecundity genotypes in sheep. Intercellular signaling between the oocyte and granulosa cells (GC) regulates proliferation and differentiation due to actions of bone morphogenetic protein 15 (BMP15) and growth and differentiation factor 9 (GDF9) within the follicle. Mutations that either knockout or reduce the activity of BMP15 or GDF9 have been found to increase ovulation rate in heterozygotes and generally cause severe follicle abnormalities in homozygotes. A mutation in the intracellular kinase domain of the BMPR1B receptor (Booroola fecundity gene) increases ovulation rate in heterozygotes with further increases in ovulation in homozygotes. The physiological mechanisms linking these mutations to increased ovulation rates are still not well defined. A recently identified high fecundity bovine genotype, Trio, causes increased expression of SMAD6, an intracellular inhibitor of the BMP15/GDF9 signalling pathways. This bovine model has provided insights into the mechanisms associated with selection of multiple dominant follicles and multiple ovulations in carriers of fecundity alleles. The present review focuses on the mechanisms involved in follicle selection in ruminants with a special emphasis on the contribution made by multiple ovulation models in both cattle and sheep. The evaluation of multiple ovulation models in ruminants has allowed us to construct a new physiological model that relates changes in the BMP15/GDF9 signalling pathways to the physiological changes that result in selection of multiple dominant follicles. This model is characterized by acquisition of dominance at a smaller follicle size but at a similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence, delaying FSH suppression and, thus allowing additional follicles to continue to grow and acquire dominance.

Physiological mechanisms involved in maintaining the corpus luteum during the first two months of pregnancy.

Maintenance of the corpus luteum (CL) during pregnancy is essential for continuing the elevated circulating progesterone (P4) that is required to maintain pregnancy. The mechanisms that protect the CL during early pregnancy when the non-pregnant animal would typically undergo CL regression have been extensively investigated. It is clear uterine prostaglandin F2α (PGF) causes regression of the CL in non-pregnant ruminants and that maintenance of the CL during early pregnancy is dependent upon secretion of interferon-tau (IFNT) from the elongating embryo. A number of specific mechanisms appear to be activated by IFNT. Most studies indicate that there is an inhibition of oxytocin-induced secretion of uterine PGF. There is also evidence for increased resistance to PGF action, perhaps due to secretion of PGE2 and PGE1 or direct endocrine actions of circulating IFNT. These mechanisms occur concurrently and each may help to maintain the CL during the first month of pregnancy. However, during the second month of pregnancy, IFNT is no longer secreted by the embryo. Attachment of the embryo to the uterus and subsequent placentome development have been linked to silencing of expression from the IFNT gene. In addition, there is some evidence that oxytocin responsiveness of the uterus returns during the second month of pregnancy leading to substantial basal secretion of PGF and perhaps PGF pulses. There is also no evidence that the CL during the second month of pregnancy is resistant to the actions of PGF as observed during the first month. Thus, this manuscript attempts to compare the mechanisms that maintain the CL during the first and second months of pregnancy in ruminants and provides a new, speculative, physiological model for maintenance of the CL during month two of pregnancy that is distinct from the previously-described mechanisms that maintain the CL during the first month of pregnancy.