Impact of gonadotropin-releasing hormone administration at the time of artificial insemination on conception risk and its association with estrous expression.

Cows with reduced estrous expression have compromised fertility. The aim of this study was to determine whether the administration of GnRH at the time of artificial insemination (AI) would affect ovulation rates and the fertility of animals expressing estrous behavior of lesser intensity. Cows were enrolled at the time of estrus from 3 farms (n = 2,607 estrus events; farm A: 1,507, farm B: 429, farm C: 671) and randomly assigned to receive GnRH at AI or not (control). The intensity of estrous expression, monitored through leg-mounted activity monitors, was determined using the maximum activity during estrus; estrous expression was categorized as greater or lower relative to the farm median. On farm A, cows were assessed at alert, and 24 h, 48 h, and 7 d post-alert for ovulation using ultrasonography. Pregnancy per AI was confirmed at 35 ± 7 d post-estrus for cows that were inseminated. Differences between treatments were tested using the GLIMMIX procedure of SAS. Treatment with GnRH at the time of AI increased pregnancy per AI (41.3 ± 1.6 vs. 35.7 ± 1.7%). An interaction between treatment and estrous expression on pregnancy per AI was found. Control cows with greater estrous expression had greater pregnancy per AI than those with lesser expression, whereas GnRH administration increased pregnancy per AI for cows with lesser estrous expression but not those with greater expression (GnRH, greater intensity: 43.5 ± 2.1; GnRH, lesser intensity: 37.8 ± 2.2; control, greater intensity: 42.6 ± 2.2; control, lesser intensity: 31.0 ± 2.2%). A higher proportion of cows with greater estrous expression that were administered GnRH at AI were found to ovulate by 48 h and 7 d post-estrus; however, ovulation of cows with lesser estrous expression was unaffected by GnRH administration. In conclusion, fertility of cows with reduced estrous expression may be increased using GnRH at the time of AI; however, increased ovulation rates do not seem to be the direct mechanism behind this relationship.

Plasma concentrations of progesterone in the preceding estrous cycle are associated with the intensity of estrus and fertility of Holstein cows.

The aim of this study was to determine the association between concentrations of progesterone (P4) during previous the estrous cycle with the intensity of spontaneous or estrogen-induced estrous expression and pregnancy per artificial insemination (P/AI). A total of 1,953 AI events from lactating Holstein cows were used, consisting of 1,289 timed AI events from experiment 1 (Exp. 1) and 664 AI events from experiment 2 (Exp. 2). In Exp. 1, cows were bred after a timed AI protocol based on estradiol and P4. In Exp. 2 animals were bred upon spontaneous estrus detection. In both experiments cows were continuously monitored by an automated activity monitor (AAM), in Exp.1 a relative increase of activity was calculated (i.e., percentage of increase activity at estrus compared to cow's baseline activity) and in Exp.2, activity data from each cow were computed into an index value that ranged from 0 to 100. In Exp.2 duration (hours) of estrus were calculated and defined as the total time above the threshold (35 index). The intensity of estrous expression was determined for each event and classified as either high or low intensity using the median of each experiment. Blood samples were collected for P4 analysis in Exp. 1 at -4 d, 0 d, and 7 d relative to timed AI, and in Exp. 2 immediately following estrus (0 d), 7 d, 14 d, and 21 d post-AI. Concentration of P4 was classified as greater or lower according to the median value in each experiment. Cows with lower concentrations of P4 at AI had greater estrous expression in Exp. 1 (363.6 ± 5.2 vs. 275.9 ± 8.0% relative increase) and Exp. 2 (76.7 ± 1.9 vs. 67.4 ± 4.7 index; and 12.5 ± 0.5 vs. 9.3 ± 1.8 hours). Cows with a greater intensity of estrous expression at timed AI had greater concentrations of P4 at -4 d than cows with lower intensity estrus or no estrous expression (4.6 ± 0.2 vs. 3.6 ± 0.2 vs. 3.7 ± 0.2 ng/mL). Cows with greater concentrations of P4 at -4 d had greater P/AI (32.8 ± 4.4 vs. 22.4 ± 4.5%), whereas cows with lesser concentrations of P4 at d0 for either timed AI (35.2 ± 3.4 vs. 19.6 ± 4.6%) or spontaneous estrus (31.8 ± 2.8 vs. 23.4 ± 3.2%) had greater P/AI. Cows with greater concentrations of P4 7 d post-timed AI had greater P/AI compared with cows that had lower concentration of P4 (39.1 ± 2.9 vs. 24.7 ± 2.6%). Similarly, cows that had lower concentrations of P4 at 7 d, 14 d and 21 d post-spontaneous estrus tended to have lower P/AI when compared with cows with greater concentrations of P4. Overall, concentrations of P4 prior to and at AI were associated with greater estrous intensity and P/AI at both spontaneous and timed AI events.

Rumen-Reticular Temperature During Estrus and Ovulation Using Automated Activity Monitors in Dairy Cows.

This study aimed to determine changes in rumen-reticular temperature (RRT) at estrus or ovulation and determine if these changes are associated with the intensity of estrous expression. Cows were equipped with an automated activity monitor (AAM) and a rumen-reticular bolus thermometer. A total of 190 estrus episodes were used where physical activity data was recorded using the AAM and ovulation was determined via ultrasonography of the ovaries at alert and twice daily, for a maximum of 60 h. Estrous expression was assessed using the maximum activity and duration in which activity remained above the AAM threshold; both characteristics were categorized using the median. Temperature data was collected for the duration of estrus, as well as for the interval of time where ovulation was determined to occur. Three measures of temperature were calculated: (1) positive area under the curve (AUC), (2) maximum positive temperature change (PTC), (3) maximum negative temperature change (NTC) at estrus (AUCE/ PTCE/ NTCE) and around ovulation (AUCO/PTCO/ NTCO). Both AUC and PTC were greater during estrus than around ovulation (2.7 ± 0.2 and 1.1 ± 0.3°C2 for AUCE and AUCO; 0.55 ± 0.03 and 0.26 ± 0.04°C for PTCE and PTCO, respectively). In contrast, NTC was lower around ovulation than estrus (-0.28 ± 0.05 and -0.60 ± 0.06°C for NTCE and NTCO). Cows with greater estrous expression had greater AUC and PTC during estrus than around ovulation, but cows with lesser estrous expression had similar AUC and PTC. Increases in AUC [High THI (Temperature Humidity Index): High activity: 4.7 ± 0.5, Low activity: 1.5 ± 0.4; Low THI: High activity: 3.1 ± 0.2, Low activity: 1.4 ± 0.2 °C2] and PTC (High THI: High activity: 0.79 ± 0.08, Low activity: 0.36 ± 0.07; Low THI: High activity: 0.60 ± 0.04, Low activity: 0.47 ± 0.04°C) associated with estrous expression were found to be greater on days with higher THI. Alerts created using standard deviations from the mean were unable to detect estrus or ovulation with acceptable precision. Further research is required to determine how changes in RRT can be used successfully to predict estrus and ovulation.

Rumen-protected B vitamin complex supplementation during the transition period and early lactation alters endometrium mRNA expression on day 14 of gestation in lactating dairy cows.

Greater metabolic demands in high-producing dairy cows are believed to be a cause of sub-fertility in these animals. Previously, supplementation with vitamin B complex molecules has shown benefits in improving milk production, health, and reproductive efficiency of dairy cows. The primary aim of this project was to determine the effects of rumen-protected vitamin B complex supplementation of 100 g of Transition VB (Jefo, St. Hyacinthe, QC, Canada) and 4 g of Lactation VB (VB; Jefo), during the transition and early lactation periods, respectively, compared with a control diet containing no supplementation on d 14 endometrial outcomes of pregnancy. In the vitamin B supplemented cows, we expect to see a change in the mark-up of endometrial genes important for embryo survival before implantation. Multiparous Holstein cows were enrolled into the study 3 wk before parturition and were randomly assigned to either the VB or control treatment. Twice-a-week blood samples, weekly milk samples, and daily feed intake were collected. Cows were enrolled onto a double-ovsynch protocol at 33 ± 3 d postpartum and inseminated by timed artificial insemination. Milk production and components, concentrations of BHB, haptoglobin, and progesterone in serum, and ovarian dynamics were also measured, but no treatment effect was observed. The uterus was flushed on d 14 after artificial insemination (around 72 DIM) for conceptus collection, and endometrial samples were collected at the same time. Overall, 42 cows were flushed and 13 embryos were collected. Analysis of mRNA expression of genes related to embryo development, immune system, adhesion, and regulation of vitamin B molecules showed that OXTR, MUC5B, MUC1, IL1B, SPP, TRD, FZD8, and FOLR1 genes were significantly upregulated in the VB group. Vitamin B supplementation had no effect on the size of the embryo and ovulatory follicle or corpus luteum diameter at embryo collection. In conclusion, the benefits of strategic dietary VB supplementation during the transition and early lactation might be directly linked to endometrial functions required for embryo survival during the peri-implantation period.

Effect of estrous expression on timing and failure of ovulation of Holstein dairy cows using automated activity monitors.

The aim of this study was to determine if estrous expression, as measured by an automated activity monitor (AAM), affects timing and failure of ovulation of lactating Holstein dairy cows. Cows were equipped with 2 AAM, 1 neck-mounted (AAMC) and 1 leg-mounted (AAML), by 10 d postpartum and enrolled into the trial when their activity crossed the alert threshold on the AAMC. A total of 850 episodes of estrus from 293 different cows were used for this study. When cows were enrolled, their ovaries were scanned by transrectal ultrasonography and gait and body condition scored. Ovaries of cows detected in estrus were scanned twice daily for a maximum of 3 d to determine the disappearance of the preovulatory follicle (ovulation) and the interval from estrus to ovulation was calculated. Physical activity data recorded from the AAM were used to determine estrus behavior using 2 traits: (1) peak activity and (2) duration. Peak activity was only available for the AAML. Peak activity was defined as the maximum activity during an estrus episode. Duration of estrus was defined as the time the activity of the cow exceeded threshold values set by the AAM software. The AAMC correctly identified 87.8% of the estrus alerts, with 12.2% false positives. The average (±standard deviation) intervals from activity alert to ovulation were 25.8 ± 10.2 and 24.7 ± 9.3 h for the AAMC and AAML, respectively. Changes in estrous expression were associated with differences in the interval from alert to ovulation. Cows with short intervals to ovulation were found to have less intense estrous expression than cows with medium and long length intervals to ovulation using the AAMC, whereas using the AAML, cows with short intervals to ovulation exhibited less intense estrous expression than cows with medium but the same as those with long intervals to ovulation. Furthermore, irrespective of the AAM, estrus events with less estrous expression had increased odds of having a short interval to ovulation (below the median of 20 h) when compared with those having greater estrous expression (2.6 and 1.9 increased odds for the AAMC and AAML, respectively). Ovulation failure was affected by estrous expression because estrus events with greater peak activity or longer duration had reduced ovulation failure compared with those with less estrous expression (AAMC peak activity: 1.9 ± 1.4 vs. 9.5 ± 1.7%; AAML peak activity: 2.3 ± 1.4 vs. 6.2 ± 1.5%; AAMC duration: 2.1 ± 1.4 vs. 8.9 ± 1.7%). In addition, cows with more estrous expression had greater pregnancy per artificial insemination than those with less estrous expression with both the AAMC (42.3 ± 0.4 vs. 31.7 ± 0.4%) and the AAML (43.1 ± 0.4 vs. 36.3 ± 0.4%). Pregnancy per artificial insemination results were consistent even when removing cows that failed to ovulate. In conclusion, expression of estrus was highly associated with ovulation timing, ovulation failure, and fertility when using 2 different AAM. Cows with greater estrous expression have longer intervals from activity alert to ovulation, experience less ovulation failure, and have greater pregnancy per artificial insemination.

Prevalence of lameness and leg lesions of lactating dairy cows housed in southern Brazil: Effects of housing systems.

Within the last few decades, the North American and European dairy industries have been collecting information about lameness and leg injury prevalence on dairy farms and have tried to develop solutions to mitigate these ailments. Few published articles report the prevalence of lameness and leg lesions in areas outside of those 2 regions, or how alternative housing systems, such as compost-bedded packs, affect the prevalence of these maladies. The objectives of this study were to compare the prevalence of lameness and leg lesions on confined dairies that used freestall, compost-bedded packs, or a combination of these 2 systems in Brazil. Data were collected in the autumn and winter of 2016 from 50 dairy farms located in Paraná state, including 12 compost-bedded pack dairies (CB), 23 freestall dairies (FS), and 15 freestall dairies that used compost-bedded packs for vulnerable cows (FS+C). A visit to the farm consisted of a management questionnaire, an inspection of the housing areas as well as the milking parlor, and an evaluation of all lactating cows as they exited the parlor for lameness (score 1-5), hygiene (score 0-2), body condition score (score 1-5), and hock and knee lesions (score 0-1). Median 1-way chi-squared test was used to compare production systems. We found no difference between farm types in management practices related to hoof health management or average daily milk production per cow [31 (29-33.9) kg/d; median (quartile 1-3)], percentage of Holstein cattle in the herd [100% (90-100%)], conception rate [35.8% (30.2-38%)], or pregnancy rate [15% (13.7-18%)]. The CB farms were smaller [85 (49.5-146.5) milking cows] than both the FS [270 (178-327.5) milking cows] and FS+C farms [360 (150-541.5) milking cows). The overall prevalence of severe lameness (score 4 and 5) across all farms was 21.2% (15.2-28.5%) but was lower on the CB farms [14.2% (8.45-15.5%)] in comparison to the FS [22.2% (16.8-26.7%)] and the FS+C farms [22.2% (17.4-32.8%)]. Less than 1% of all cows scored on CB farms were observed with swollen or wounded knees (or both), which was lower than either the FS or FS+C farms [7.4% (3.6-11.9%) and 6.4% (2.6-11.8%) of all cows scored, respectively]. The same pattern was found for hock lesions, where the farm-level prevalence within the 3 different housing types was 0.5% (0-0.9%), 9.9% (0.8-15.3%), and 5.7% (2.6-10.9%) for CB, FS, and FS+C farms, respectively. No differences between farm systems were observed for hygiene or body condition score. On average, 2.7% (0.8-10.9%) of lactating cows had a soiled side, 15.4% (2.1-37.4%) had dirty legs and 1.7% (0-9.3%) had dirty udders. The average herd-level body condition score across farms was 2.9 (2.9-3), with 0.86% of the all cows scored having a body condition score <2.5. These results indicate that lameness prevalence on confined dairies in Brazil is high and highlight the need for remedial changes in environmental design and management practices. We found that CB farms in this region had reduced lameness and lesions in relation to FS or FS+C dairies.

Integrating an automated activity monitor into an artificial insemination program and the associated risk factors affecting reproductive performance of dairy cows.

The aim of this study was to compare 2 reproductive programs for the management of first postpartum artificial insemination (AI) based on activity monitors and timed AI, as well as to determine the effect of health-related factors on detection and expression of estrus. Lactating Holstein cows (n = 918) from 2 commercial farms were enrolled. Estrous cycles of all cows were presynchronized with 2 injections of PGF2α administered 2 wk apart. Treatments were (1) first insemination performed by timed AI (TAI) and (2) first insemination based upon the detection of estrus by activity monitors (ACT; Heatime, SCR Engineering, Netanya, Israel) after the presynchronization, whereas cows not inseminated by the detection of estrus were enrolled in the Ovsynch protocol. Body condition score (BCS; scale 1 to 5), hock score (scale: 1 to 4), gait score (scale: 1 to 4), and corpus luteum presence detected by ovarian ultrasonography were recorded twice during the presynchronization. On the ACT treatment, 50.5% of cows were inseminated based on detected estrus, whereas 83.2% of the cows on the TAI treatment were inseminated appropriately after the timed AI protocol. Pregnancy per AI did not differ by treatment (30.8 vs. 33.5% for ACT and TAI, respectively). Success of pregnancy was affected by parity, cyclicity, BCS, milk production, and a tendency for leg health. In addition, treatment × cyclicity and treatment × parity interactions were found to affect pregnancy success, where anovulatory cows and older cows had compromised pregnancy outcomes on the ACT treatment but not on the TAI treatment. Factors affecting pregnancy outcomes varied among farms. Hazard of pregnancy by 300 DIM was affected by farm, parity, BCS, a treatment × cyclicity interaction, and a tendency for an interaction between leg health and farm. Detection of estrus was affected by farm, parity, cyclicity, and leg health, but not BCS or milk production. Expression of estrus was compromised in anovular and older cows, and by the timing of the estrus event, but not by gait score, BCS, or milk production. Increased duration of estrus, but not intensity of estrus, improved pregnancy per AI. In conclusion, using an automated activity monitor for the detection of estrus within a Presynch-Ovsynch program resulted in similar pregnancy per AI and days open compared with a reproduction program that was strictly based on timed AI for first postpartum AI. In contrast, notable variations in reproductive outcomes were detected between farms, suggesting that the use of automated activity monitors is prone to individual farm management.

Relationship of concentrations of cortisol in hair with health, biomarkers in blood, and reproductive status in dairy cows.

Hair cortisol has been used to measure chronic stress in dairy cows as it offers the advantage of being noninvasive, fast, and able to indicate levels of cortisol over long periods. The aim of this study was to determine the associations between hair cortisol with clinical disorders, reproductive status, and the development of subclinical endometritis in dairy cows. Furthermore, we aimed to determine the association between hair cortisol concentrations and blood markers associated with metabolic status and acute inflammation. In experiment 1, cows (n=64) were hair sampled every 3wk from the tail switch beginning at calving (d 0) until d 126 for cortisol analysis; blood samples were collected every 3wk from d 0 until 42 for ß-hydroxybutyrate and glucose analysis. In experiment 2, cows (n=54) were chosen retrospectively by diagnosis of subclinical endometritis (END), subclinical endometritis and at least 1 clinical disease (END+CLIN), or as healthy (control) using a cytobrush and ultrasonography at 30±3d in milk. At the same time, animals were hair sampled for cortisol analysis and blood sampled for haptoglobin and ceruloplasmin analysis. Health records were recorded throughout both experimental periods. Animals with clinical disease presented higher cortisol concentrations than clinically healthy animals in experiment 1 [geometric mean (95% confidence interval); 8.8 (7.8, 9.9) vs. 10.7 (9.6, 12.0) pg/mg]; however, animals diagnosed with subclinical endometritis in experiment 2 did not differ in hair cortisol concentrations [11.7 (9.8, 14.0), 12.2 (9.3, 15.9), 10.5 (8.1, 13.6) pg/mg for control, END, and END+CLIN, respectively]. In experiment 1, an effect of sample day was noted, where d 21 had higher cortisol concentrations than d 42, 84, and 126, but not from d 0 for both parities. Within both experiments, a parity effect was present where multiparous animals consistently had higher cortisol concentrations than primiparous animals. Multiparous cows that became pregnant by 100d postpartum had lower concentrations of hair cortisol at d 42 and 84 in milk. Lastly, other biomarkers associated with metabolic status and acute inflammation, such as glucose, ß-hydroxybutyrate, haptoglobin, and ceruloplasmin, were not strongly correlated with measurements of cortisol in hair. Overall, hair cortisol measurements appear to be associated with clinical disorders and have a direct association with pregnancy status; however, concentrations of hair cortisol may not be suited to differentiate situations of stress with lower magnitudes, such as the development of subclinical disease.

Automated and visual measurements of estrous behavior and their sources of variation in Holstein heifers. II: Standing and lying patterns.

Lying (LY) and standing (ST) behavior of Holstein heifers (n = 57) was studied from Day -7 to Day +2 relative to estrus (Day 0) using leg-mounted accelerometers. A total of 269 estrus episodes were studied. The objectives were to quantify the effects of estrus on ST and LY patterns, identify sources of variation, and provide insight into new approaches for the use of sensors in estrus detection. The sensors were used to obtain ST bout frequency (bouts per day), total daily ST time, and mean bout duration and duration of the longest bout for ST and LY on a daily basis. Estrus episodes were identified using walking activity peaks and validated by ovarian ultrasonography. Repeated-measures ANOVA was used to analyze the effects of estrus order (pubertal vs. second and greater), season, time of estrus onset, category of baseline walking activity, and number of heifers simultaneously in estrus on ST and LY variables. Overall, ST bout frequency was lower, whereas mean ST bout duration, duration of the longest ST bout, and total daily ST time were longer on Day 0 (P < 0.05). The longest ST bout increased from 232 ± 5 minutes on Day -7 to 488 ± 16 minutes on Day 0 (P < 0.05), and on Day 0, it started within -2 to 4 hours of time of estrus onset. Measurements of ST bouts varied the most on Day 0 (coefficient of variation = 21.5%-52.8%). The pubertal estrus was characterized by smaller increase in mean ST bout duration, longest ST bout duration, and total daily ST time (P < 0.05) compared to the second and greater episodes. Second and greater episodes had greater longest LY bout on Day +1, but on Day 0 it was not different from other nonestrus days (P < 0.01). The longest ST bout on Day 0 was longer for heifers with high baseline walking activity and for episodes occurring during the cold season (P < 0.05). The effect of estrus on ST variables for morning-onset episodes was observed only on Day 0. Episodes with afternoon and night onset showed the effect of estrus spread over Days -1 and 0, resulting in a smaller change between Day -7 and Day 0. The number of heifers simultaneously in estrus did not affect the studied variables (P > 0.05). Measurements of ST and LY changed during estrus and were correlated to walking activity. The large increase in duration of the longest ST bout and its occurrence in proximity of estrus onset suggest potential for automation. Incorporating measurements of ST and LY corrected for sources of variation could improve estrus detection systems.

Short communication: Factors affecting hair cortisol concentrations in lactating dairy cows.

Cortisol has long been used as a marker of the stress response in animals. Cortisol can be analyzed from different media, most notably from the blood, saliva, and feces; however, the collection of cortisol from some of these media requires invasive procedures or excessive handling of the animals. Furthermore, it is not possible to capture long-term increases in circulating concentrations of cortisol from the blood, saliva, or feces. Hair cortisol has been found to be a reliable alternative for measuring chronic stress. With this emerging measure, appropriate sampling methodology must be developed and validated. The aim of this study was to determine the effects of hair color, sampling location, and processing method on cortisol concentrations in hair from lactating black and white Holstein cows (n=18). Furthermore, we aimed to measure the hair growth rates at different body locations (n=12) and test hair cortisol levels when resampled over short intervals (n=37). Both black- and white-colored hair was collected from the shoulder, top line, hip, and tail switch of Holsteins; due to breed characteristics only white hair was harvested from the tail switch. All samples were cleaned with water and isopropanol, and then ground in a ball mill or finely cut with scissors once dry. Cortisol was extracted with methanol before being measured using a commercially available ELISA kit. Concentrations of cortisol were greater in white than in black hair (7.8 ± 1.1 vs. 3.8 ± 1.1 pg/mg). When only white samples were analyzed, hair from the tail switch had more cortisol than hair from the shoulder (11.0 ± 1.2 vs. 6.2 ± 1.2 pg/mg), whereas no difference was found when compared with the hip and top line. Samples ground with a ball mill had greater concentrations of cortisol extracted than those minced with scissors (10.4 ± 1.2 vs. 4.7 ± 1.2 pg/mg). The growth rate of hair was significantly greater at the tail switch compared with the hip and shoulder (0.51 ± 0.05 vs. 0.04 ± 0.05 vs. 0.03 ± 0.05 mm/d). When hair was collected every 3 wk after calving, a tendency was detected for multiparous cows to have greater concentrations of hair cortisol and significantly greater concentrations of cortisol on d 0 and 21 after calving compared with d 42, 84, and 126. In Holsteins, the hair on the tail switch is always white, grows more rapidly than other sites, and is sensitive enough to capture changes in cortisol over intervals as short as 3 wk, making it the ideal location for measuring hair cortisol.