Comparison of Gonadotropin-Releasing Hormone versus Estrogen-Based Fixed-Time Artificial Insemination Protocols in Grazing Bos taurus Suckled Beef Cows.

Fixed-timed artificial insemination (FTAI) protocols for beef cattle in South America are primarily based on estradiol esters and intravaginal progesterone-releasing devices (IVPD). The objective of this study was to determine the optimal gonadotropin-releasing hormone (GnRH)-based protocol as an alternative to the use of estrogen-based protocols in grazing Bos taurus suckling beef cows. All cows received an IVPD on the day of protocol initiation and prostaglandin F2α (PG) plus equine chorionic gonadotropin (eCG) treatments at the time of IVPD removal. In Experiment 1, cows (n = 235) were randomly assigned to one of four treatments: (i) 7-day estradiol = 2 mg of estradiol benzoate (EB) at IVPD insertion on Day 9 and 1 mg of estradiol cypionate (ECP) at IVPD removal on Day 2; (ii) 7-day GnRH = 10 µg of GnRH at IVPD insertion on Day 10, IVPD removal on Day 3 and GnRH at FTAI; (iii) 7 & 7 estradiol = PG at IVPD insertion on Day 16, EB on Day 9 and ECP at IVPD removal on Day 2; (iv) 7 & 7 GnRH = PG at IVPD insertion on Day 17, GnRH on Day 10, IVPD removal on Day 3 and GnRH at FTAI. In Experiment 2, cows (n = 462) were randomly assigned to one of four treatments: (i) 6-day estradiol = EB at IVPD insertion on Day 9, IVPD removal on Day 3 and GnRH at FTAI; (ii) 7-day estradiol; (iii) 7-day GnRH; (iv) 7 & 7 GnRH. In Experiment 1, plasma progesterone concentrations and percentage of cows with a corpus luteum (CL) at IVPD removal, and pregnancy per AI (P/AI) were greater for cows subjected to GnRH-based protocols compared with cows subjected to estrogen-based protocols (p < 0.01). In Experiment 2, cows subjected to the 7 & 7 GnRH protocol had the greatest P/AI (p < 0.01). In summary, GnRH-based FTAI protocols resulted in similar or greater P/AI compared to estrogen-based FTAI protocols in grazing postpartum Bos taurus suckled beef cows. The greatest P/AI was attained with the 7 & 7 GnRH protocol.

Morphologic, Steroidogenic, and Transcriptomic Assessment of the Corpus Luteum in Holstein Cows after Spontaneous or Hormone-Induced Ovulation.

There is evidence that replacing the gonadotropin-releasing hormone (GnRH) with porcine luteinizing hormone (pLH) to synchronize ovulation prior to artificial insemination (AI) increased pregnancy per AI in dairy cows without affecting blood progesterone (P4) concentrations. Whether morphologic, steroidogenic, and transcriptomic differences exist among corpora lutea (CL) formed after ovulation induced by GnRH and pLH is unclear. Our main objective, therefore, was to compare CL characteristics between GnRH- and pLH-induced CL. In 24 non-lactating Holstein cows, ovulations were spontaneous (Spont-Ov) or induced with 100 µg GnRH, 25 mg pLH, or 1 mg estradiol benzoate (EB), with CL excised 12 d after ovulation. In pLH- versus GnRH-treated cows, the duration of elevated LH (above baseline) was prolonged (10 versus 6 h, respectively, p < 0.01), but CL dimensions, pixel intensity of CL images, proportions of steroidogenic and non-steroidogenic luteal cells, and mean plasma LH did not significantly differ. Post-ovulation mean plasma P4 (ng/mL) did not differ among Spont-Ov (3.0) pLH (3.1) or GnRH (3.0) cows but were lower in EB cows (2.0). In vitro P4 concentration was greater in luteal explants of pLH-treated cows than in all other groups (combined means, 16.0 vs. 12.3 µg/mL, p < 0.02). Relative abundance of mRNA for oxytocin receptor (OXTR) was 2-fold higher (p < 0.01) in CL of pLH vs. GnRH cows and highest in Spont-Ov CL. In summary, pLH-treated cows had a longer LH peak, and greatest luteal tissue concentrations and in vitro production of P4. We inferred that increased P4 concentrations at the ovarian-uterine level in pLH-treated cows could have promoted embryo development and increased pregnancy per AI.

Development of a blood calcium test for hypocalcemia diagnosis in dairy cows.

Hypocalcemia, defined as total blood calcium concentrations below 2.1 mM, has detrimental impacts on welfare, production and reproduction in dairy cattle. Yet, no cow-side test exists for testing total bovine blood calcium. Here, we modified the split trehalase complementation assay to detect total calcium in serum by incorporating calmodulin and the M13 peptide as fusion partners to the trehalase fragments. In the presence of calcium, calmodulin undergoes a conformation change and gains strong affinity for M13 peptide. A high reactive assay for calcium was developed with detection threshold of 10 uM and dynamic range between 1 uM and 1 mM. The addition of a specific concentration of calcium chelator, EDTA, in mild acidic conditions, shifted the dynamic range to physiological calcium concentrations and transformed the sensor from ionized calcium sensor to total calcium sensor. The sensor was validated on a collection of 213 bovine serum samples by comparison with quantitative colorimetric calcium test. A correlation coefficient of 0.81 was achieved and the accuracy of detecting subclinical hypocalcemia was 0.86 and specificity of 100%. The area under the Receiver Operating Characteristic curve was 0.93. The concordance correlation coefficient (0.80), Bland-Altman plot and weighted Kappa coefficient (0.71) demonstrated a substantial agreement between both methods. In conclusion, a novel total calcium test was developed that can be used as a convenient high throughput laboratory test and with potential to be incorporated into a version compatible with on-farm testing.

Genetic potential for residual feed intake and diet fed during early- to mid-gestation influences post-natal DNA methylation of imprinted genes in muscle and liver tissues in beef cattle.

Discovery of epigenetic modifications associated with feed efficiency or other economically important traits would increase our understanding of the molecular mechanisms underlying these traits. In combination with known genetic markers, this would provide opportunity to improve genomic selection accuracy in cattle breeding programs. It would also allow cattle to be managed to improve favorable gene expression. The objective of this study was to identify variation in DNA methylation between beef cattle of differential pre-natal nutrition and divergent genetic potential for residual feed intake (RFI). Purebred Angus offspring with the genetic potential for either high (HRFI) or low (LRFI) RFI were prenatally exposed to either a restricted maternal diet of 0.5 kg/d average daily gain (ADG) or a moderate maternal diet of 0.7 kg/d ADG from 30 to 150 d of gestation. We performed DNA methylation analysis of differentially methylated regions (DMR) of imprinted genes (Insulin-like growth factor 2 (IGF2) DMR2, IGF2/H19 imprinting control region (ICR) and IGF2 receptor (IGF2R) DMR2) using post-natal samples of longissimus dorsi (LD) muscle taken from male and female calves at birth and weaning, and of LD muscle, semimembranosus (SM) muscle, and liver samples collected from steers at slaughter (17 months of age). Interestingly, for all three DMR investigated in liver, LRFI steers had higher levels of methylation than HRFI steers. In LD muscle, IGF2/H19 ICR methylation differences for heifers at birth were due to pre-natal diet, while for steers at birth they were mostly the result of genetic potential for RFI with LRFI steers again having higher levels of methylation than HRFI steers. While results from repeated measures analysis of DNA methylation in steers grouped by RFI revealed few differences, in steers grouped by diet, we found higher methylation levels of IGF2 DMR2 and IGF2R DMR2 in LD muscle of restricted diet steers at weaning and slaughter than at birth, as well as increased methylation in LD muscle of restricted diet steers compared with moderate diet steers at weaning and/or slaughter. Our results suggest that differential pre-natal nutrition, and divergent genetic potential for RFI, induces tissue- and sex-specific alterations in post-natal IGF2 and IGF2R methylation patterns and that these patterns can vary with age in Angus beef cattle.

Prevalence of early postpartum health disorders in Holstein cows and associations with production, reproduction, and survival outcomes on Alberta dairy farms.

The objective of this study was to determine the prevalence and associations of early postpartum health disorders with production, reproduction, and survivability in dairy cows in Alberta. Holstein cattle (N = 1096) from 11 commercial dairy farms in Alberta, Canada were enrolled. Cows were evaluated daily for clinical postpartum health disorders and a blood sample was collected once within 2 wk after calving to diagnose subclinical disorders. Overall, 61% of cows were diagnosed with at least 1 postpartum health disorder, with 25% of cows having multiple disorders. Of the cows with only 1 health disorder, 71% were classified as inflammatory and 29% as metabolic disorders. Cows with multiple disorders were at the greatest risk of reduced milk production, impaired fertility, and leaving the herd. Cows with an inflammatory disorder had reduced productive and reproductive performance, whereas cows with a metabolic disorder were at the greatest risk of mortality.

Prévalence des troubles de santé post-partum précoces chez les vaches Holstein et associations avec les résultats de production, de reproduction et de survie dans les fermes laitières de l'Alberta. L'objectif de la présente étude était de déterminer la prévalence et les associations des troubles de santé post-partum précoces avec la production, la reproduction et la capacité de survie chez les vaches laitières en Alberta. Des bovins Holstein (N = 1096) de 11 fermes laitières commerciales en Alberta, au Canada, ont été recrutés. Les vaches ont été évaluées quotidiennement pour des problèmes cliniques de santé post-partum et un échantillon de sang a été prélevé une fois dans les 2 semaines après le vêlage pour diagnostiquer les problèmes subcliniques. Dans l'ensemble, 61 % des vaches ont reçu un diagnostic d'au moins un problème de santé post-partum, 25 % des vaches ayant des problèmes multiples. Parmi les vaches présentant un seul problème de santé, 71 % ont été classées comme inflammatoires et 29 % comme des problèmes métaboliques. Les vaches souffrant de problèmes multiples couraient le plus grand risque de réduction de la production de lait, d'impact sur la fertilité et d'élimination du troupeau. Les vaches présentant un problème inflammatoire avaient des performances de production et de reproduction réduites, tandis que les vaches présentant un problème métabolique étaient les plus exposées au risque de mortalité.(Traduit par Dr Serge Messier).

Effect of types of breeding on embryo survival following first AI in lactating Holstein cows.

The main objective was to investigate the effects of timed-AI protocols versus AI following oestrus detection on circulating progesterone (P4) and embryo survival after first service in Holstein cows. Cycling status was determined by ultrasonography and by plasma P4 concentrations 14 and 26 days after calving, and only cows with a corpus luteum and/or P4 ≥ 1 ng/ml were used. Cows were randomly allocated to one of three types of breeding: DO (n = 80), received GnRH-7d-PGF2α-3d-GnRH and Ovsynch56 was initiated 7 days later; G7G (n = 70), received PGF2α-2d-GnRH and Ovsynch56 (GnRH-7d-PGF2α-56h-GnRH-16h-AI) was initiated 7 days later; or AI based on oestrus detection, EDAI (n = 60). Progesterone was also determined at AI and 8, 16, 18 and 20 days after AI; ISG15 and MX2 mRNA abundance were determined 16 days after AI. Mean plasma P4 at AI was greater in the EDAI group compared with DO and G7G groups, while after AI, P4 was greater in DO and G7G groups compared with EDAI group. However, the percentage of cows with a concentration of P4 < 0.8 ng/ml at AI did not differ among groups. Relative mRNA abundance of ISG15 and MX2 was greater in the DO and G7G groups compared to those in EDAI group. Pregnancy per AI 16, 32 and 60 days after AI was greater (p < .05) in cows in the DO group compared with those in EDAI group (47.5%, 38.8% and 36.3% vs. 30.0%, 21.7% and 15.0%). Pregnancy losses between 16 and 60 days after AI were greater (p < .05) in cows in the EDAI (50.0%) group compared to those subjected to DO (23.7%) or G7G (24.1%). In conclusion, the use of timed-AI synchronization protocols resulted in greater circulating P4 concentrations post-AI and greater embryo survival following first service in lactating Holstein cows.

Common and specific mineral and metabolic features in dairy cows with clinical metritis, hypocalcaemia or ketosis.

The objectives were to evaluate differences in serum concentration of metabolites, macro minerals and hepatic enzymes at pre and postpartum time-points in dairy cows diagnosed with clinical metritis, hypocalcaemia or ketosis postpartum. A total of 144 Holstein cows from 11 commercial dairy herds in Alberta, (Western Canada) were enrolled in this study. Cows with clinical metritis had lower serum concentrations of glutamate dehydrogenase (GLDH) at pre and postpartum and lower total Ca, albumin, urea, and cholesterol at postpartum when compared to control cows. Cows with hypocalcaemia had greater serum concentrations of Na, Cl, and calculated osmolarity (CalOsmo) at prepartum and lower concentration of total serum Ca, glucose, cholesterol, gamma-glutamyl transpeptidase (GGT), GLDH, total protein and albumin at postpartum. Prepartum serum concentrations of non-esterified fatty acids (NEFA), beta hydroxybutyrate (BHB), Cl, albumin/globulin ratio (A/G), Na, K and sum of Na and K were greater in ketotic cows when compared with control cows. Cows with ketosis had also greater postpartum serum concentrations of NEFA, BHB, GGT and aspartate transaminase (AST) when compared with control cows. Prepartum serum Na and Cl concentrations and CalOsmo were greater in cows diagnosed with hypocalcaemia or ketosis when compared with control cows. Furthermore, postpartum serum concentrations of total Ca, cholesterol, albumin and GLDH were significantly affected by hypocalcaemia or clinical metritis and concentrations of GGT by hypocalcaemia or ketosis. Finally, postpartum serum concentrations of haptoglobin increased in all disease groups when compared with control cows. These results suggest common metabolic features for clinical metritis, hypocalcaemia and ketosis in dairy cows in addition to the specific ones.

Poly(N-isopropylacrylamide) microgel-based etalons for the label-free quantitation of estradiol-17ß in aqueous solutions and milk samples.

A novel estradiol-17ß (E2) biosensor was constructed from poly(N-isopropylacrylamide) (pNIPAm) microgel-based etalons by modification of their outermost Au layer with an E2 binding 75-mer DNA aptamer. When E2 is not present in the solution, the aptamer forms a loose/linear structure that allows ions to pass through and into the microgel layer. The ions can change the solvation state of the microgels, which changes the optical properties of the etalon. When E2 is present in the solution, the aptamer binds the E2 and undergoes a conformational change to a form that can block the diffusion of salt ions into the microgel layer. This blocking decreases the response of the device to salt exposure, which can be related to the concentration of E2 in solution. Using this approach, E2 sensor showed a dynamic range of 0.9-200 pg/mL with a calculated detection limit of 0.9 pg/mL (3.2 pM) E2, and the lowest measured concentration of E2 is 5.0 pg/mL. This sensor also showed low cross reactivity with progesterone, a similar steroid hormone. Moreover, this sensor could be regenerated five times without losing its sensitivity. Finally, we demonstrated that the sensor could also be used to quantify E2 in commercial skim and 2% milk, as well as farm milk directly without any pre-treatment. The successful quantitation of E2 in unprocessed milk demonstrates its potential use as a "cow-side" testing device for the dairy industry. Graphical abstract ᅟ.

Update on Multiple Ovulations in Dairy Cattle.

This review updates the causal mechanisms and risk factors for multiple ovulations (MOV) in cattle. Clearly, MOV can lead to twin pregnancies, which negatively affects the health, production, and reproduction of cows. Therefore, a better understanding of the factors causing MOV may help to reduce twinning. Multiple ovulations occur after two or more follicles deviate and achieve codominance. The MOV rate is influenced by a complex network of hormones. For example, MOV is more common during periods of low progesterone (P4), that is, in anovulatory cattle or when luteolysis coincides with the selection of the future ovulatory follicle. There is also strong evidence for the luteinizing hormone (LH) being the primary factor leading to codominance, as high P4 concentrations suppress the transient LH surges and can reduce the ovulation rate in cattle or even inhibit deviation. Rates of MOV are increased in older and higher-producing dairy cows. Increased milk production and dry matter intake (DMI) increases hormone clearance, including P4; however, the association between milk yield and MOV has not been consistent. Additional risk factors for MOV include ovarian cysts, diet, season, and genetics.

Effect of Exogenous Progesterone or Flunixin Meglumine After AI on Serum Progesterone Concentration and Pregnancy per AI in Lactating Dairy Cows.

The objective of this study was to determine the effect of post AI administration of exogenous progesterone (P4) or a prostaglandin F2α (PGF2α) synthesis inhibitor agent on serum P4 concentrations and pregnancy per AI (P/AI) in lactating dairy cows. Eighty lactating cows were randomly allocated to one of four treatment groups: 1) CON (control), received 5 mL of saline solution on d 6 and 14 post AI; 2) IP4 (injection of P 4), received 125 mg of P4 im on d 6 and 14 post AI; 3) CIDR, received a controlled internal drug release insert containing 1.38g of P4 from d 6 to 20 post AI; and 4) FM (Flunixin Meglumine), received 0.625 g of Flunixin Meglumine, a nonsteroidal anti-inflammatory drug, twice daily on d 15 and 19 post AI. Blood samples were taken on d 0, 6, 14, 17 and 20 post AI to determine P4 concentrations. Transrectal palpation was performed between 40 and 45 d post AI to determine pregnancy status. All treatment groups (i.e. IP4, CIDR and FM) resulted in greater serum P4 concentration on d 17 and 20 post AI compared to CON (P < 0.05). Cows given a CIDR insert had greater concentrations of P 4 on d 17 and 20 than IP4 and FM cows (P < 0.05). However, no significant difference was found between IP4 and FM groups for serum P4 concentrations. The P/AI was greater (P < 0.05) in CIDR-treated cows (55%, 11/20) than CON (25%, 5/20), and intermediate in IP4 (40%, 8/20) and FM (35%, 7/20) cows. In summary, treatment with exogenous P4 (i.e. CIDR and IP4) or FM increased serum P4 concentrations in lactating dairy cows. However, results suggest that only CIDR administration would improve P/AI.

Dynamics of pre- and post-insemination progesterone profiles and insemination outcomes determined by an in-line milk analysis system in primiparous and multiparous Canadian Holstein cows.

The objective was to evaluate in-line milk progesterone (mP4) data to determine dynamics of pre- and post-insemination mP4 profiles and their associations with parity and outcomes of artificial insemination (AI) in Holstein cows. Milk progesterone (ng/mL) was quantified at pre-determined time points before and after AI through an automated in-line milk analysis system (Herd Navigator™, DeLaval International, Tumba, Sweden). Only AI (∼d0; n = 605) preceded by an mP4-decline (at least two samples of mP4 ≥5 ng/mL followed by at least one sample <5 ng/mL; d-2) were evaluated. Maximum mP4 attained between d-15 and d-2 (PrePeak), d-2, d5, d10, d14, maximum mP4 attained within 21d post-AI (PostPeak), and rate-of-daily-change between mP4 time points (ng/mL/d) were analyzed. Primiparous and multiparous cows were classified by AI outcomes based on post-AI mP4 profiles into three groups: (1) non-pregnant (OPEN; mP4-decline ≤ 30d post-AI), (2) presumed-pregnant (PREG; no mP4-decline until 55d post-AI), and (3) presumed-pregnancy loss (P-LOSS; mP4-decline between 31 and 55d post-AI). For profile comparisons, smoothed mP4 data were analyzed using mixed linear models. Primiparous cows had greater (P < 0.01) mP4 than multiparous cows at d5 (4.6 ± 0.2 vs. 2.8 ± 0.1), 10 (11.1 ± 0.4 vs. 7.6 ± 0.2), 14 (19.7 ± 0.4 vs. 16.1 ± 0.3) and PostPeak (23.5 ± 0.3 vs. 21.7 ± 0.2). The rate-of-daily-change was greater (P < 0.01) in primiparous than in multiparous cows from d-2 to 5 (+0.2 ± 0.03 vs. -0.1 ± 0.02) and from d5 to 10 (+1.2 ± 0.05 vs. +0.9 ± 0.03), but lesser (P < 0.01) from d14 to PostPeak (+0.9 ± 0.09 vs. +1.3 ± 0.06). In primiparous cows, mP4 in PREG was greater at d10 and PostPeak than OPEN (11.1 ± 0.5 and 24.2 ± 0.5 vs. 9.6 ± 0.4 and 22.3 ± 0.4, respectively, P < 0.04), but lesser at d5 than P-LOSS (4.4 ± 0.3 vs. 5.7 ± 0.4, P = 0.04). In multiparous cows, mP4 at d-2 was lesser in PREG than OPEN and P-LOSS (3.2 ± 0.1 vs. 3.4 ± 0.04 and 3.5 ± 0.1, respectively, P ≤ 0.03), but greater at d10, d14 and PostPeak in PREG than in OPEN (8.2 ± 0.4, 16.8 ± 0.5 and 22.7 ± 0.4 vs. 6.9 ± 0.3, 14.8 ± 0.3 and 19.7 ± 0.2, respectively, P < 0.01). Multiparous PREG cows had greater rate-of-daily-change in mP4 than OPEN cows from d5 to 10 and from d10 to 14 (+1.0 ± 0.06 and +2.2 ± 0.11 vs. +0.8 ± 0.04 and +1.9 ± 0.08, respectively, P < 0.03). Overall post-AI mP4 increased faster and were greater in primiparous than in multiparous cows. Based on in-line mP4 profiles, greater mP4 levels near time of AI (d-2 in multiparous and d5 in primiparous cows) and lesser mP4 beyond d10 were negatively associated with a successful pregnancy.

Relationships among early postpartum luteal activity, parity, and insemination outcomes based on in-line milk progesterone profiles in Canadian Holstein cows.

The objectives of this retrospective study were to use in-line milk progesterone (mP4) data to investigate relationships of (1) commencement of luteal activity (CLA), and (2) luteal phase (LP) length and frequency preceding first postpartum AI, with parity and AI outcomes in Canadian Holstein cows. Starting 21 ± 1 days postpartum (DPP), levels of mP4 were assessed every 2.2 ± 2.0 d through an automated in-line milk analysis system (Herd Navigator™, DeLaval International, Tumba, Sweden) until ∼55 d after first or second AI in 748 Holstein cows from two herds. The CLA was defined as the DPP of the first of at least two consecutive samples with mP4 ≥5 ng/mL, and the period with elevated mP4 (≥5 ng/mL) was defined as the LP. Cows were categorized by CLA [earlier (≤) or later (>) than 28, 35, 42, 49, 56, and 63 DPP], and by the pattern of LP frequency preceding first AI as having or not: (1) one or more normal LP (LP length ≥7 and ≤19 d); (2) one or more abnormal LP (LP length <7 or >19 d, or interluteal period ≥12 d); and (3) two or more LP (either normal or abnormal). Outcomes of first or second AI were determined by the interval between AI and cessation of the ensuing LP as: non-pregnant (mP4-decline ≤30 d), presumed-pregnant (no mP4-decline until 55 d), or presumed-pregnancy loss (mP4-decline between 31 and ≤55 d). The odds of pregnancy per AI (P/AI) at 55 d and pregnancy loss were evaluated using generalized linear mixed models. Primiparous cows had lower odds of having CLA ≤28 DPP [Odds ratio (OR) = 0.58, P = 0.002] and one or more abnormal LP (OR = 0.73, P = 0.04) than multiparous cows. In multiparous cows, CLA ≤28 DPP decreased pregnancy loss (OR = 0.48, P = 0.05) and CLA ≤56 DPP increased P/AI (OR = 4.69, P < 0.01) compared to a later CLA. Primiparous and multiparous cows that had one or more normal LP before first AI had increased P/AI (OR = 3.85 and 3.45, respectively, P < 0.01) and reduced pregnancy loss (OR = 0.26 and 0.27, respectively, P < 0.01) than cows without a normal LP. Primiparous cows that had one or more abnormal LP had decreased P/AI (OR = 0.62, P = 0.04) and increased pregnancy loss (OR = 1.64, P = 0.04) compared to those without an abnormal LP. In summary, AI outcomes were improved in multiparous cows that had early CLA and in cows of both parity groups that had at least one normal LP before first AI. However, primiparous cows that had at least one abnormal LP had reduced AI outcomes. Relationships between early postpartum luteal activity and AI outcomes were inconsistent between primiparous and multiparous cows.

Pregnancy per AI in Holstein heifers inseminated with sex-selected or conventional semen after estrus detection or timed-AI.

This study compared pregnancy per AI (P/AI) of heifers inseminated with sex-selected or conventional semen after estrus detection (ED) or timed-AI (TAI). Heifers in the ED group received 2 treatments with prostaglandin F2α 14 d apart and those in the TAI group received a modified 5-day Co-synch protocol plus an intravaginal progesterone releasing insert device (PRID) and were inseminated 72 h after PRID removal. Overall P/AI were 69.2% (74/107) and 64.1% (75/117) for conventional and sex-selected semen (P > 0.05). Although P/AI in ED heifers following the use of conventional semen were numerically higher (75.0% versus 63.6%), P/AI with sex-selected semen were almost identical (65.0% versus 63.2%) for ED and TAI heifers. Pregnancy losses from first pregnancy diagnosis to term did not differ between ED and TAI heifers (6.0% versus 11.3%). In summary, heifers subjected to TAI with sex-selected semen following the application of a modified 5-day Co-synch plus PRID protocol had acceptable P/AI.

Gestation par IA chez les génisses Holstein inséminées avec du sperme déterminant le sexe ou un sperme conventionnel après la détection de l'oestrus ou l'IA à temps prédéterminé. Cette étude a comparé la gestation par IA (G/IA) des génisses inséminées avec du sperme déterminant le sexe ou du sperme conventionnel après la détection de l'oestrus (DOE) ou l'IA à temps déterminé (IATD). Les génisses dans le groupe DOE ont reçu 2 PGF à un intervalle de 14 jours et celles dans le groupe IATD ont reçu un protocole modifié 5-d Co-synch et PRID et ont été inséminées 72 heures après l'enlèvement de PRID. Dans l'ensemble, le taux de G/IA était de 69,2 % (74/107) et de 64,1 % (75/117) pour le sperme conventionnel et celui déterminant le sexe (P > 0,05). Même si le taux de G/IA chez les génisses DOE après l'utilisation de sperme conventionnel était numériquement supérieur (75,0 % c. 63,6 %), le taux de G/IA avec le sperme déterminant le sexe était presque identique (65,0 % c. 63,2 %) pour les génisses DOE et IATD. La perte de gestations entre le premier diagnostic de gestation jusqu'à la parturition ne différait pas entre les génisses DOE et TAI (6,0 c. 11,3 %). En résumé, les génisses soumises à l'IATD avec du sperme déterminant le sexe après l'application d'un protocole modifié 5-d Co-synch et PRID avaient une G/IA acceptable qui confirmait notre hypothèse.(Traduit par Isabelle Vallières).

Transcriptome Profiling of In-Vivo Produced Bovine Pre-implantation Embryos Using Two-color Microarray Platform.

Early embryonic loss is a large contributor to infertility in cattle. Moreover, bovine becomes an interesting model to study human preimplantation embryo development due to their similar developmental process. Although genetic factors are known to affect early embryonic development, the discovery of such factors has been a serious challenge. Microarray technology allows quantitative measurement and gene expression profiling of transcript levels on a genome-wide basis. One of the main decisions that have to be made when planning a microarray experiment is whether to use a one- or two-color approach. Two-color design increases technical replication, minimizes variability, improves sensitivity and accuracy as well as allows having loop designs, defining the common reference samples. Although microarray is a powerful biological tool, there are potential pitfalls that can attenuate its power. Hence, in this technical paper we demonstrate an optimized protocol for RNA extraction, amplification, labeling, hybridization of the labeled amplified RNA to the array, array scanning and data analysis using the two-color analysis strategy.

Effects of prepartum oilseed supplements on subclinical endometritis, pro- and anti-inflammatory cytokine transcripts in endometrial cells and postpartum ovarian function in dairy cows.

Postpartum uterine infections affect ovarian function and delay ovulation in cattle. As dietary fats can affect immune cell function, we investigated the influence of prepartum diets on postpartum uterine inflammatory status (UIS) as assessed 25±1 days postpartum by endometrial cytology (normal: ≤8% polymorphonuclear cells (PMN) vs subclinical endometritis (SCE): >8% PMN) and associations between SCE, pro- and anti-inflammatory cytokine gene expression and ovarian function. During the last 5 weeks of gestation, dairy cows received a diet supplemented with 8% rolled sunflower (n=10) or canola seed (n=9) or no oilseed (n=9). Ovaries were scanned until 35 days postpartum. Prepartum diets did not influence SCE, but a preovulatory-size follicle developed sooner (P≤0.05), the interval to first ovulation was shorter and the proportion of cows ovulating within 35 days postpartum was greater in the sunflower seed group. Although mRNA expression of cytokines was not affected by diet, cows with SCE had higher (P≤0.05) expression of interleukin-1ß (IL1B), interleukin-8 (CXCL8), IL10 and tumour necrosis factor-α (TNF) than normal cows. The interval (mean ± s.e.m.) from calving to preovulatory-size follicle was shorter (P≤0.05) in normal (13.2±0.9 days) than SCE cows (18.7±1.4 days). In summary, a prepartum diet supplemented with sunflower seed positively influenced postpartum ovarian function without affecting UIS or pro- and anti-inflammatory cytokine gene expression in endometrial cells.

Morphologic and transcriptomic assessment of bovine embryos exposed to dietary long-chain fatty acids.

The main objectives of this study were to determine the influence of diets enriched in α-linolenic, linoleic or oleic acid on the development and transcriptomic profile of embryos collected from dairy cattle. Non-lactating Holstein cows received one of the three diets supplemented with 8% rolled oilseeds: flax (FLX, n = 8), sunflower (SUN, n = 7) or canola (CAN, n = 8). After a minimum 35-day diet adaptation, cows were superovulated, artificially inseminated and ova/embryos recovered non-surgically after 7.5 days. Cows fed FLX had less degenerated embryos and more viable embryos than those fed CAN or SUN. In total, 175 genes were differentially expressed in blastocysts from cows fed FLX than in cows fed CAN or SUN. These differentially expressed genes were mainly involved in cellular growth and proliferation, cellular development, and cell survival and viability. In conclusion, dietary n-3 polyunsaturated fatty acids reduced early embryonic degeneration possibly through improving embryonic cell survival and viability.

Alterations in bone morphogenetic protein 15, growth differentiation factor 9, and gene expression in granulosa cells in preovulatory follicles of dairy cows given porcine LH.

In a previous work, using porcine LH (pLH) in lieu of GnRH for synchronizing ovulation in dairy cows improved pregnancy rates without increasing plasma progesterone concentrations after ovulation. The LH profile is known to remain elevated above basal concentrations (≥1 ng/mL) for up to 20 hours in pLH-treated cows compared to less than 6 hours in GnRH-treated cows. Because LH triggers a cascade of signaling networks in the preovulatory follicle to promote final maturation and support oocyte competence, we hypothesized that dissimilar LH profiles will differentially regulate the intrafollicular factors and expression of downstream genes associated with improved oocyte competence. Specific objectives were to determine differences in the abundance of oocyte-secreted factors in the preovulatory follicular fluid and target genes in granulosa cells associated with oocyte competence, in response to exogenous porcine LH or GnRH-induced endogenous bovine LH exposure, in dairy cows. Follicular contents were aspirated by a transvaginal ultrasound-guided procedure from the preovulatory follicle of cyclic, nonlactating Holstein cows 21 ± 1 hour after administration of either pLH (25-mg) or GnRH (100-µg). Mature forms of bone morphogenetic protein 15, growth differentiation factor 9, and transforming growth factorß1 were approximately 2-fold more abundant in pLH-treated cows which were exposed to an extended, low LH profile, than in GnRH-treated cows that had a short, high LH profile. The relative abundance of messenger RNA for cyclooxygenase-2, LH receptor, and progesterone receptor in granulosa cells, was about two-, eight-, and two-fold higher, respectively, in cows subjected to pLH than GnRH treatment. We infer that the improved pregnancy rate after pLH-induced ovulation reported previously, occurred through greater activation of intrafollicular transforming growth factor-ß1 superfamily members, as these proteins promote cumulus expansion and oocyte competence.

A prepartum diet supplemented with oilseeds high in oleic or linoleic acid reduced GnRH-induced LH release in dairy cows during second week postpartum.

BACKGROUND: The objective was to determine the effect of prepartum diets supplemented with rolled canola seed (high in oleic acid) or sunflower seed (high in linoleic acid) on luteinizing hormone (LH) pulsatility and gonadotropin releasing hormone (GnRH)-induced LH release during early postpartum. METHODS: Thirty-one pregnant Holstein cows, blocked by body condition score, parity and expected calving date, were assigned to 1 of 3 prepartum diets supplemented with 8 % rolled canola or sunflower seed, or no oilseed (control) during the last 35 d of gestation. Blood samples were collected at Weeks (wk)-3, 0, +1 and +2, relative to calving, to determine non-esterified fatty acids (NEFA), Beta-hydroxy butyric acid (BHBA) and glucose. Additional blood samples were collected during wk1 (n = 5 per treatment) or wk2 (n = 5 or 6 per treatment), for 6 h, to measure LH pulsatility; thereafter, 100 mcg GnRH was administrated i.m., and blood was sampled for 4 h more, to measure GnRH-induced LH release. RESULTS: Dietary treatment did not affect prepartum energy balance, but cows fed the control diet were in a deeper state of negative energy balance during wk2, than those fed canola (P = 0.03) or sunflower (P = 0.01). Prepartum diets did not influence the mean plasma concentration of BHBA and glucose. However, NEFA concentration during wk2 was greater in control cows than those fed sunflower (P = 0.03) or canola (P = 0.07). Prepartum diets did not affect LH pulsatility (i.e. mean, minimum, maximum concentration, pulse frequency, and amplitude during wk1 and 2). GnRH-induced LH release did not differ among dietary treatments during wk1 but the mean GnRH-induced LH release during wk2 was either greater (P = 0.02) and tended to be greater (P = 0.09) in control cows than in those fed canola and sunflower, respectively. CONCLUSIONS: Prepartum diets did not affect LH pulsatility and GnRH-induced LH release during the first week postpartum, but cows fed a diet supplemented with oilseeds high in oleic or linoleic acid released less LH than control cows, in response to an exogenous GnRH challenge during the second week postpartum.

A review of current timed-AI (TAI) programs for beef and dairy cattle.

This is a review of the physiology and endocrinology of the estrous cycle and how ovarian physiology can be manipulated and controlled for timed artificial insemination (TAI) in beef and dairy cattle. Estrus detection is required for artificial insemination (AI), but it is done poorly in dairy cattle and it is difficult in beef cattle. Protocols that synchronize follicle growth, corpus luteum regression and ovulation, allowing for TAI, result in improved reproductive performance, because all animals are inseminated whether they show estrus or not. As result, TAI programs have become an integral part of reproductive management in many dairy herds and offer beef producers the opportunity to incorporate AI into their herds. Gonadotropin-releasing hormone-based protocols are commonly used in North America for estrus synchronization as part of a TAI program. Protocols that increase pregnancy rates in lactating dairy cows and suckling beef cows have been developed. Protocols that improve pregnancy rates in heifers, acyclic beef cows, and resynchronized lactating dairy cows are also discussed.

Examen des programmes d'insémination artificielle à temps prédéterminé (IA à temps prédéterminé) pour les bovins de boucherie et les bovins laitiers. La physiologie et l'endocrinologie du cycle œstral et la façon dont la physiologie ovarienne peut être manipulée et contrôlée pour l'insémination artificielle à temps prédéterminé (IA à temps prédéterminé) chez les bovins de boucherie et les bovins laitiers ont été examinées. Même si la détection de l'œstrus est requise pour l'insémination artificielle (IA), elle est réalisée maladroitement chez les bovins laitiers et elle est difficile chez les bovins de boucherie. Les protocoles qui synchronisent la croissance des follicules, la régression du corps jaune et l'ovulation, permettant l'IA à temps prédéterminé, se traduisent par une performance reproductive améliorée parce que tous les animaux sont inséminés, qu'ils manifestent l'œstrus ou non. Par conséquent, les programmes d'IA à temps prédéterminé font maintenant partie intégrante de la gestion de la reproduction dans beaucoup de troupeaux laitiers et offrent aux producteurs bovins l'occasion d'intégrer l'IA dans leurs troupeaux. Les protocoles à gonadolibérine (GnRH) sont couramment utilisés en Amérique du Nord pour la synchronisation de l'œstrus dans le cadre d'un programme d'IA à temps prédéterminé. Des protocoles qui augmentent les taux de gestation chez les vaches laitières et les vaches de boucherie allaitantes ont été mis au point. Des protocoles qui améliorent les taux de gestation chez les taures, les vaches de boucherie acycliques et les vaches laitières allaitantes resynchronisées sont aussi discutés.(Traduit par Isabelle Vallières).

Progesterone supplementation before timed AI increased ovulation synchrony and pregnancy per AI, and supplementation after timed AI reduced pregnancy losses in lactating dairy cows.

One of the causes of poor fertility in high producing dairy cows is inadequate progesterone. Therefore, we determined the efficacy of an intravaginal insert containing 1.55 g of progesterone (PRID) given before and/or after timed AI (TAI) on ovarian response, plasma progesterone concentrations, pregnancy per AI (P/AI) and pregnancy losses. Lactating dairy cows at three locations were assigned (Day 0) to an Ovsynch protocol with (N = 294) or without (N = 314) a PRID. The Ovsynch protocol consisted of two injections of 100 µg gonadorelin (GnRH) 9 days apart and one injection of 500 µg cloprostenol (PG) 7 days after the first GnRH treatment. Insertion and removal of PRID occurred concurrent with the first GnRH and PG treatments, respectively. Timed AI was carried out 12 to 16 hours after the second GnRH. Ovarian status of a subset of 217 first service cows had been presynchronized with 2 treatments of PG 14 days apart with the last PG given 12 days before the first GnRH of the Ovsynch protocol. Body condition score (scale of one to five) was recorded at TAI. Ultrasonographic examinations were done in all cows at first GnRH, at PG, at TAI, and 24 hours after TAI for response to treatment and at 32 and 60 days after TAI for confirmation of pregnancy. At 4.5 days after TAI (Day 14), cows that responded to PG and ovulated after the second GnRH treatment were reassigned to receive (N = 223) or not receive (N = 229) a PRID for 7 days. Blood samples were taken for progesterone determination at PG treatment, at TAI, and post TAI on Days 14 and 21. The PRID treatment pre-TAI reduced the percentage of cows ovulating before TAI (5.8% vs. 11.1%), and significantly increased P/AI in nonpresynchronized cows (41.3% vs. 25.1%). Cows ovulating in response to the first GnRH treatment, cyclic cows, and cows with body condition score of 2.75 or more had increased P/AI, but the addition of a PRID pre-TAI to these cows did not increase P/AI. The PRID treatment post TAI did not affect P/AI, but reduced pregnancy losses (6.1% vs. 11.4%) between 32 and 60 days of gestation. The reduction in pregnancy losses tended (P = 0.10) to be significant in acyclic cows receiving a PRID than in those not receiving a PRID (5.6% vs. 33.3%). Plasma progesterone concentrations at PG treatment and on Day 21 (11.5 days after TAI) were linearly associated with P/AI. In conclusion, progesterone supplementation pre-TAI increased P/AI in nonpresynchronized cows. Progesterone supplementation post TAI reduced pregnancy losses, particularly in acyclic cows.