Is prolonged luteal phase a problem in lactating Holstein cows?

In this study, the main objective was to assess if long luteal phases could have other causes than pregnancy losses. We enrolled Holstein dairy cows ≥50 d in milk (DIM) from a commercial herd in Brazil from October 2016 to August 2017. All cows received an estradiol-based synchronization protocol, and, on the day of insemination (d 0), were randomly assigned either an artificial insemination (AI) or a placebo insemination (PBO) in a 3:1 ratio. An ultrasound was used to assess the presence of a CL on d17, 24, and 31, which, combined to the information from patches for the detection of estrus, was used to determine the length of the luteal phase following AI or PBO. Pregnancy was assessed by ultrasound on d 31 and cows that were pregnant were excluded from the analyses. The length of the estrous cycles was categorized as short (<17 d), normal (17-23 d), long (24-30 d), and very long (≥31 d). We compared the proportion of cows in each category between the AI and PBO groups using a cumulative ordinal mixed model. We define prolonged luteal phase as estrous cycles ≥24 d and tested its association with potential risk factors (parity, season, DIM, uterine size and position score, milk production, body condition score, and the presence of a corpus luteum (CL) at enrollment to the synchronization protocol) using mixed logistic regression models. Results are presented as odds ratio (OR) and 95% credible intervals (BCI). Data from 876 inseminations (AI: n = 616, PBO: n = 260) was collected. Overall, 12% of estrous cycles were short, 31% were normal, 19% were long, and 38% were very long. There was no difference in the odds of being in longer estrous cycle categories for the AI compared with the PBO group (OR = 0.92, 95% BCI = 0.76-1.10). Season and presence of a CL at enrollment were associated with prolonged luteal phase. In the AI group, there was a possible effect of early pregnancy losses on the lifespan of the CL, but not the PBO group, which led us to conclude that long and very long estrous cycles were not all caused by the embryonic loss. In fact, the high prevalence of cows with an extended CL lifespan in the present study suggests this could be an under- or miss-reported characteristic of high-producing lactating Holstein cows. This finding may have important repercussions in the understanding of the CL function physiology of lactating Holstein cows.

Validity of luminometry and bacteriological tests for diagnosing intramammary infection at dry-off in dairy cows.

The objective of this cross-sectional study was to estimate the validity of laboratory culture, Petrifilm and Tri-Plate on-farm culture systems, and luminometry to correctly identify IMI at dry-off in dairy cows, considering all tests as imperfect. From September 2020 until December 2021, we collected composite milk samples from cows before dry-off and divided them into 4 aliquots for the luminometry test, the Petrifilm (aerobic count), the Tri-Plate, and the laboratory culture. While we assessed multiple thresholds of relative light units (RLU) for the luminometry, we used thresholds of ≥100 cfu/mL for the laboratory culture, ≥ 50 cfu/mL for the Petrifilm, and ≥1 cfu for the Tri-Plate. We fitted Bayesian latent class analysis (LCA) models to estimate the sensitivity (Se) and specificity (Sp) for each test to identify IMI, with 95% credibility interval (BCI). Using different prevalence measures (0.30, 0.50, and 0.70), we calculated the predictive values (PV) and misclassification cost terms (MCT) at different false-negative to false-positive ratios (FN:FP). A total of 333 cows were enrolled in the study from one commercial Holstein herd. The validity of the luminometry was poor for all thresholds, with Se of 0.51 (95% BCI = 0.43-0.59) and Sp of 0.38 (95% BCI = 0.26-0.50) when using a threshold of ≥150 RLU. The laboratory culture had Se of 0.93 (95% BCI = 0.85-0.98) and Sp of 0.69 (95% BCI = 0.49-0.89), the Petrifilm had Se of 0.91 (95% BCI = 0.80-0.98) and Sp of 0.71 (95% BCI = 0.51-0.90), and the Tri-Plate had Se of 0.65 (95% BCI = 0.53-0.82) and Sp of 0.85 (95% BCI = 0.66-0.97). Bacteriological tests had good PVs, with comparable positive PV for all 3 tests, but lower negative PV for the Tri-Plate compared with the laboratory culture and the Petrifilm. For a prevalence of IMI of 0.30, all 3 tests had similar MCT, but for prevalence of 0.50 and 0.70, the Tri-Plate had higher MCT in scenarios where leaving a cow with IMI untreated is considered to have greater detrimental impacts than treating a healthy cow (i.e., FN:FP of 3:1). Our results showed that the bacteriological tests have adequate validity to diagnose IMI at dry-off, but the luminometry does not. We concluded that, while luminometry is not useful to identify IMI at dry-off, the Petrifilm and Tri-Plate tests performed similarly to the laboratory culture, depending on the prevalence and the importance of the FP and FN results.

Herd-level associations between the proportion of elevated prepartum nonesterified fatty acid concentrations and postpartum diseases, reproduction, or culling on dairy farms.

The objectives of this herd-level prospective observational cohort study were to describe the proportion of cows with elevated prepartum nonesterified fatty acid concentrations (PropElevNEFA) in dairy herds and to assess the herd-level associations between PropElevNEFA and postpartum diseases, reproductive performance, and culling. From November 2018 to December 2020, a convenience sample of 49 herds was enrolled in this study. Blood sampling (16 to 29 cows per herd) was performed during the week before and during the 2 wk following calving to quantify the concentration of nonesterified fatty acids (NEFA) and ß-hydroxybutyrate acids (BHBA), respectively. Elevated NEFA was defined as ≥280 µmol/L and hyperketonemia as BHBA ≥1.4 mmol/L. Retained placenta, metritis, purulent vaginal discharge, endometritis, and mastitis were diagnosed on-farm following standardized definitions, and success at first artificial insemination (AI) and culling events were recorded. The associations between PropElevNEFA and each individual disease, success at first AI, and culling were evaluated using Bayesian aggregated binomial regression models with weakly informative priors, from the which odds ratio (OR) and the 95% credible intervals (BCI) were obtained. A total of 981 cows were included in the statistical analyses representing 16 to 29 (median = 19) cows per herd. Cows were enrolled in the prepartum period of their first to tenth (median = third) lactation, and 41% of them had an elevated prepartum NEFA concentration. At the herd level, PropElevNEFA varied between 11% and 78% (median = 39%). The odds of metritis (OR = 1.37, 95% BCI = 1.13-1.67) increased for every 10-point increase in PropElevNEFA, whereas the odds of success at first AI decreased (OR = 0.69, 95% BCI = 0.59-0.80). The PropElevNEFA was not associated with the other tested diseases or culling. Our results suggest that the herd-level proportion of cows having elevated prepartum NEFA concentrations is associated with metritis and poor success at first AI in dairy herds.

Validation of a leg-mounted pedometer for the measurement of steps in lactating Holstein cows.

The aim of this study was to validate the pedometer AfiAct II (AfiMilk) for the measurement of steps in lactating Holstein cows housed in a freestall design by assessing its agreement with visual observation of step counts. A total of 41 primiparous (n = 12) and multiparous (n = 29) cows were enrolled in the study between August and September 2018. Steps were monitored continuously by the pedometer and visually assessed for a 24-h period using video recordings. Visually observed steps were categorized as walking and stationary steps. The total number of steps taken per cow was calculated using the sum of walking and stationary steps. Unprocessed step count data from the study day were retrieved from the AfiMilk system in time-blocks of approximately 15 min. Repeated measures correlation was used to quantify the association between the pedometer measurements and visual observation of step counts. Nonindependence among observations were accounted adjusting for interindividual (cow) variability with an analysis of covariance. Pearson correlation coefficients (r) were categorized from negligible (0.00-0.30) to very high (0.90-1.00). Bland-Altman plots were created to evaluate the bias between the pedometer and visual observations. A total of 2,261 time-blocks were used in this study with an average (mean ± standard deviation) of 55.14 ± 8.1 time-blocks per cow. A high correlation was found for the evaluation between the pedometer and observed walking steps (r = 0.74; 95% confidence interval [CI] = 0.73-0.76), stationary steps (r = 0.71; 95% CI = 0.69-0.63), and total steps (r = 0.88; 95% CI = 0.87-0.89). The results of the Bland-Altman plot suggested limited bias between the pedometer step counts and visual observation of steps, independent of the type of steps. Numerical differences and several time-block differences outside of the 95% interval of agreement suggested an overestimation of step counts by the pedometer, which increased as the number of steps increased. The pedometer measured, on average, 97.6 ± 118.5 (28%), 249.2 ± 126.2 (125%), and 297.2 ± 205.4 (196%) steps/day more than the visual observed total steps, stationary steps, and walking steps, respectively. Our findings indicate that the pedometer counts all movement in which the pedometer leg is lifted off the floor without distinguishing if there was body movement of the animal.

The effect of progesterone concentrations during superovulation of Holstein heifers in a randomized trial.

The aim of this study was to evaluate the effect of different progesterone (P4) concentrations during the follicular growth on the intensity of estrous expression, ovarian response to the superovulatory treatment, and embryo production and quality in superovulated heifers. A total of 63 Holstein heifers were randomly assigned into 2 experimental groups: Low P4 (n = 31) and High P4 (n = 32). Animals received a pre-synchronization protocol followed by a protocol of superovulation that included the allocated P4 treatment. Activity was monitored continuously by an automated activity monitor, and estrus characteristics (maximum intensity and duration) were recorded. Embryo collection was performed 7 d post artificial insemination (AI). Embryos were counted and graded from good or excellent (1) to degenerated (4). The outcomes of interest were: number and diameter of follicles at the time of AI, ovulation success (confirmed 7 d post-AI), time to estrus event, maximum intensity and duration of estrus, number and quality of embryos. Data were analyzed according to the type of outcome variable using logistic, linear, or Poisson regression models. A total of 105 embryos (High P4: n = 42; Low P4: n = 63) were graded for quality. Different P4 levels did not affect the maximum intensity (High P4 = 497.8 ± 23.9%; Low P4 = 542.2 ± 23.5%) or the duration (High P4 = 13.5 ± 1.5 h; Low P4 = 14.3 ± 1.4 h) of estrus. Heifers in the High P4 treatment had greater number of follicles at time of AI (High P4 = 16.6 ± 1.6 follicles; Low P4 = 13.9 ± 1.2 follicles), but with smaller diameter (High P4 = 11.3 ± 0.1 mm; Low P4 = 12.0 ± 0.1 mm) compared with Low P4. High P4 heifers tended to have better embryo quality compared with Low P4 heifers (odds ratio = 1.98; 95% CI = 0.90-4.35). High P4 heifers had less embryos than Low P4 heifers, but this was modified by the CIDR (intravaginal implant of P4) removal to estrus interval (interval 0-21 h: mean ratio = 1.15, 95% CI = 0.42-1.87; interval 22-46 h: mean ratio = 0.58, 95% CI = 0.27-0.96). Although estrous expression was not associated with embryo quality, as the duration and the maximum intensity of estrous expression increased, the number of embryos recovered 7 d post-AI increased (duration: mean ratio = 1.04; 95% CI = 1.03-1.05; maximum intensity: mean ratio = 1.50; 95% CI = 1.42-1.58). In conclusion, P4 during the follicular growth, and intensity of estrus, are playing a role in regulating the quality and the number of embryos produced by superovulated heifers. This study was supported by contributions from Resilient Dairy Genome Project and the Natural Sciences and Engineering Research Council.

Methods used by Canadian dairy farmers for on-farm euthanasia and the emotions associated with the decision and the practice of euthanasia.

On-farm euthanasia is complicated in that farmers have to make the decision to end the life of an animal they are caring for. Studies have shown that some culling decisions are made too late, which results in animal welfare issues. However, information about on-farm euthanasia methods and the decision process leading to the euthanasia of an animal is limited. Additionally, emotions and feelings are involved in the decision and the act of euthanasia. The objectives of this study were to describe current practices related to on-farm euthanasia performed by Canadian dairy farmers and to assess the emotions and complex feelings surrounding the act. A cross-sectional study design was used to gather this information through an online questionnaire completed exclusively by Canadian dairy farmers. Data were analyzed descriptively using frequencies, and associations between the participants' characteristics and their practices and emotions regarding on-farm euthanasia were assessed using logistic regression models. A total of 479 dairy farmers from 5 provinces answered the survey. The most commonly used primary method of on-farm euthanasia was a firearm (>70%). Unacceptable euthanasia methods, such as standard 0.22-caliber long rifle for adult cows or no adjunct method following the use of captive bolt, were used by 25%, 18%, and 58% of the participants for calves, replacement animals, and dairy cows, respectively. Ninety-four percent of the participants reported that the farm owner was the person who always or often made the euthanasia decisions. In addition, 32% of the participants reported that the veterinarian always examined the animal before performing euthanasia, 51% reported at least one person received training to perform euthanasia, and 16% reported having a decision tree for euthanasia. Some participants (17%) were troubled with the responsibility for ending the life of their animals, and 19% perceived other people on the farm to be uncomfortable with performing euthanasia. Half of the participants reported that people performing euthanasia on the farm felt at least one related painful emotion or complex feeling, and half reported they had at least one reason to feel anxiety or unease concerning euthanasia. The results of this study showed gaps such as the limited use of acceptable euthanasia methods and decision trees by Canadian dairy farmers, low availability of on-farm training for euthanasia, and variable involvement of veterinarians.

Randomized controlled trial assessing the impact of a combined treatment of insulin glargine and propylene glycol on the resolution of hyperketonemia and milk production in postpartum dairy cows.

Multiple treatments for hyperketonemia have been studied, and although propylene glycol has been the most constantly reported as successful in mitigating the effects of this condition, a recent study showed that a subpopulation of cows affected by hyperketonemia and hypoglycemia could benefit from a combined treatment of propylene glycol and insulin glargine. The objective of the present study was to assess the effect of this combination on the time-to-hyperketonemia resolution, and on milk production following the initial treatment. A randomized controlled trial was conducted on 2 commercial farms on which postpartum exams were conducted twice a week. Cows naturally affected by both hyperketonemia [ß-hydroxybutyrate (BHB) ≥ 1.4 mmol/L] and hypoglycemia (glucose <2.2 mmol/L) were enrolled and randomly assigned to receive 2 mL of placebo (physiological saline) subcutaneously (PBO), or to receive 2 mL of insulin glargine (200 IU) subcutaneously (INS). Cows in both groups also received 300 g of propylene glycol orally for 3 d. Time-to-hyperketonemia resolution (return to BHB <1.4 mmol/L) and daily milk production following the initial treatment were compared between groups using Cox proportional hazard and mixed linear regression models, respectively. A total of 248 cows between 1 and 30 d in milk (median = 16) were enrolled in the trial from January 2018 to February 2022. Cows were of first (32.3%; n = 80/248), second (32.3%; n = 80/248), or third or greater (35.4%; n = 88/248) parity. The overall time-to-hyperketonemia resolution was not different between treatments, but there was a 2-way interaction between treatment group and parity. Primiparous cows had a shorter time-to-resolution in the INS group than in the PBO group; no difference was observed between treatment groups for older cows. The overall milk production of cows in the INS group was 3.4 kg higher than cows in the PBO group during the 14 d after enrollment. There was a 2-way interaction between treatment group and parity. Primiparous cows in the INS group produced 5.3 kg more milk daily than primiparous cows in the PBO group; no effect of treatment was found for older cows. These results suggest that cows affected simultaneously by hyperketonemia and hypoglycemia benefited from a combined treatment of propylene glycol and insulin glargine to improve subsequent milk production and that the effect primarily came from the positive effect in primiparous cows. Time-to-hyperketonemia resolution was also improved in primiparous cows but not in older cows.

Association between genomic daughter pregnancy rates and reproductive parameters in Holstein dairy cattle.

The objective of this observational study was to evaluate the relationship between genomic daughter pregnancy rate (GDPR) with reproduction parameters such as pregnancy at first artificial insemination (AI), pregnancy per AI, and pregnancy losses (PL). A total of 12,949 events from 3,499 Holstein cows were included. Cows were enrolled as nulliparous (n = 1,220), primiparous (n = 1,314), or multiparous (n = 965). Cows were bred either after a timed AI protocol, timed embryo transfer (ET), or spontaneous estrus. Most lactating cows were bred following a timed AI protocol based on estradiol and progesterone, and most nulliparous were artificially inseminated following estrus detection. Hair samples were collected from the tail switch and cows were genotyped using a SNP platform (Clarifide, Zoetis). Cows that were bred by timed AI were evaluated for estrous behavior using tail chalk. Tail chalk was applied on the head of the tail 2 d before timed AI and the chalk was evaluated at AI (no estrus: 100% of chalk remaining or ≥50% of chalk remaining; Estrus: <50% of chalk remaining). Pregnancy diagnosis was performed at d 32 and 60 after AI using ultrasonography, and the presence of a heartbeat was considered a positive diagnosis. Pregnancy loss was defined as a pregnant cow on d 32 that was nonpregnant on d 60. As GDPR increased, the odds of pregnancy at first AI increased [odds ratio (OR) = 1.28, 95% CI = 1.20-1.35], the odds of pregnancy per AI increased (OR = 1.31, 95% CI = 1.25-1.36), and the odds of PL decreased (OR = 0.66, 95% CI = 0.60-0.72). Most cows that were bred on the day of the timed AI demonstrated estrus (n = 6,075; 92.9%). The odds of demonstrating estrus on the day of timed AI increased as GDPR increased (OR = 1.31, 95% CI = 1.17-1.48). There was no interaction between GDPR and parity or breeding management for pregnancy at first AI, pregnancy per AI, and PL. In conclusion, the odds of pregnancy at first AI and pregnancy per AI increased as GDPR increased. Moreover, the odds of PL increased as GDPR decreased. Greater GDPR was also associated with greater occurrence of estrus on the day of timed AI. These results suggest that selecting for higher GDPR could result in better reproductive performance, but this would need to be assessed with additional research.

Randomized controlled trial of pegbovigrastim as an adjunct therapy for naturally occurring severe clinical mastitis cases in dairy cows.

The main objective of this study was to quantify the effect of pegbovigrastim (PEG) as an adjunct therapy for naturally occurring severe mastitis cases on survival, intramammary bacteriological cure, and subsequent milk production. A double-blinded, randomized controlled trial was conducted on a single commercial dairy farm. During the study period, all cows having a case of severe clinical mastitis, defined as the presence of abnormal milk and inflammation in one quarter or more combined with the presence of systemic signs (pyrexia, dehydration, or recumbency), were enrolled in the study. In addition to a standardized therapy combining systemic and intramammary antimicrobials as well as systemic anti-inflammatory drugs, cows received a subcutaneous injection of either 2.7 mL of 0.9% sterile saline (control group; CON) or 2.7 mL of PEG (PEG group). A milk sample for bacteriological analysis was taken before treatment was administered and a second sample was taken 14 d after enrollment. Survival (30 d post-treatment) and bacteriological cure (14 d post-treatment) were analyzed by survival analysis and chi-squared tests, respectively, whereas daily milk production was treated as a repeated measure in mixed regression models. The 77 cows enrolled in the study were of parity 2 to 4 (mean = 3) and between 3 and 302 d in milk (mean = 154). The bacteria identified in milk at enrollment were Klebsiella spp. (n = 48; 62%), Escherichia coli (n = 16; 21%), Enterobacter spp. (n = 10; 13%), or no growth (n = 3; 4%). The probability of survival during the first 30 d after treatment was higher in the PEG group (84%) than in the control group (46%). Daily milk production over the 30-d period following treatment and bacteriological cure 14 d post-treatment did not differ between groups. Overall, cows treated with PEG as an adjunct therapy for naturally occurring cases of severe clinical mastitis had a better survival than cows treated with saline.

Randomized controlled trial of reinsemination strategies in dairy cows diagnosed nonpregnant using color flow Doppler ultrasonography on d 21 after insemination.

The objective of this study was to quantify the reproductive performance of 4 reinsemination strategies in cows diagnosed nonpregnant using corpus luteum color flow Doppler ultrasonography on d 21 after last insemination. A total of 2,140 color flow Doppler ultrasonography exams from 845 Holstein cows from 10 commercial dairy herds were used in this study. Farms were visited every 2 wk by the research team. On d 7 after insemination, cows were enrolled in a randomized controlled trial to be assigned 1 of 4 treatments if they were subsequently diagnosed nonpregnant on d 21. All cows were then examined on d 21 using Doppler ultrasonography to diagnose nonpregnancy. Treatment A (CON) was the control group: nonpregnant cows were programmed to receive a standard Ovsynch protocol starting on d 32. Nonpregnant cows in treatment B (GnRH) were injected i.m. with GnRH on d 21 after insemination and reinseminated immediately. Cows in treatment C (2×GnRH) received an i.m. injection of GnRH on d 11 after insemination. If diagnosed nonpregnant on d 21 after insemination, they were injected i.m. with GnRH on d 21 after insemination and inseminated immediately. Cows in treatment D (Resynch) received an i.m. injection of GnRH on d 14 after insemination. If diagnosed nonpregnant on d 21 after insemination, they were injected i.m. with PGF2α on d 21 after insemination and injected i.m. with GnRH on d 24. Then, a standard Ovsynch protocol was started on d 32. Statistical analyses were performed using mixed logistic regression models accounting for cow clustering and herd effect. A theoretical simulation was performed for each treatment to quantify the proportion of cows remaining nonpregnant 42 d after initial insemination. Of the 2,140 color flow Doppler ultrasonography exams in the study, 870 (40.6%) had a Doppler score of D0 (n = 444) or D1 (n = 426), which were indicative of nonpregnancy and used for data analysis. Overall, the number of exams assigned to each treatment was as follows: CON = 223 (25.6%), GnRH = 214 (24.6%), 2×GnRH = 220 (25.3%), and Resynch = 213 (24.5%). The final mixed multivariable logistic regression model included treatment, parity, days in milk at enrollment, and herd. Conception risk at first insemination following enrollment was 31.4, 20.6, 31.9, and 48.7% for treatments CON, GnRH, 2×GnRH, and Resynch, respectively. Based on the simulation, if 100 cows were diagnosed nonpregnant at their color flow Doppler ultrasonography exam on d 21 after last insemination, the total number of cows remaining nonpregnant 42 d after the initial insemination would be 69, 72, 58, and 51 for treatments CON, GnRH, 2×GnRH, and Resynch, respectively. Thus, the treatments used in groups 2×GnRH and Resynch for cows diagnosed nonpregnant on d 21 after insemination yielded better results than those used in the other groups.

Technical note: Validation of an in-house bovine serum enzyme immunoassay for progesterone measurement.

Measuring circulating progesterone (P4) of dairy cows is a key component of many research studies dealing with basic and applied reproduction physiology. The gold standard in dairy cows for the measurement of P4 in serum is radioimmunoassay (RIA), but it generates radioactive waste and requires licensed facilities. The purpose of this study was to develop and validate an in-house competitive enzyme immunoassay (EIA) to measure the P4 concentration in serum of dairy cattle. The secondary objective was to validate a commercial EIA. In the present study, a competitive EIA was developed using commercially available antibodies and conjugates. Ninety-six well microtiter plates were coated with the secondary antibody and incubated overnight. Following a washing step, the wells were blocked using the primary antibody. Serum samples were prepared by first extracting P4 using petroleum ether, then diluted in working conjugate solution. Samples were pipetted into the coated and blocked plates, then the matching HRP conjugate label (P4-3-HRP, East Coast Bio, North Berwick, ME) was added. The plates were incubated for 2 h, then washed. The substrate solution was added, and the plate was incubated up to 1 h at room temperature in the dark until a blue color had developed. A stop solution was added, and the optical density measured on a microplate reader was set at 450 nm. The binding proportion was calculated by a visible spectrum absorbance reader, and the amount of P4 was calculated using a log-logit regression line. The commercial EIA was executed as suggested by the manufacturer. The validation of the in-house EIA was done by calculating the inter- and intraassay coefficients of variation (CV) and evaluating the parallelism of diluted samples. The results from the in-house and commercial EIA were also compared with the ones from the RIA graphically (scatterplots and Bland-Altman plots) and statistically, using the Spearman correlation coefficient (r) and the Cohen's kappa statistics using a threshold of 1.0 ng/mL (κ). For the in-house EIA, the intraassay CV were all <10%, but the interassay for samples with small and large P4 concentration had CV of 12.5 and 11.0%, respectively. The correlations between the results from the EIA and the RIA were strong (in-house: r = 0.90; commercial: r = 0.83). At small concentrations (<1.0 ng/mL), however, the correlation with the gold standard was weak (in-house: r = 0.27; commercial: r = 0.14). This was likely due to the lack of accuracy at small concentrations, also shown by the absence of parallelism in samples ≤0.4 ng/mL. In conclusion, results from both the in-house and commercial EIA strongly correlated with the gold standard, but less so at smaller concentrations. The in-house EIA offers good accuracy to measure P4 in samples with a concentration >0.4 ng/mL, and a perfect agreement with RIA using a threshold of 1.0 ng/mL.

Symposium review: Linking activity-sensor data and physiology to improve dairy cow fertility.

Several studies have demonstrated that the intensity of estrous expression is associated with ovulation, ovarian and uterine function, and fertility, and is dependent on social hierarchy and the housing system used. Data from recent studies involving spontaneous and induced estrus have shown that a greater relative increase and longer estrus (captured by different automated activity monitors; AAM) are both associated with improved pregnancy per artificial insemination (AI; around 10 to 14% increase) and decreased pregnancy losses. Intensity and duration of estrus were surprisingly weakly associated with preovulatory follicle diameter and concentrations of plasma estradiol at estrus, whereas ovulation failure was associated with low estrus intensity. Studies have also shown that the display of estrous behavior near AI was associated with the modification of expression of genes related to the immune system, adhesion molecules, and prostaglandin synthesis in the endometrium. Transcripts in leukocytes and in the conceptus tissue associated with maternal recognition of pregnancy as well as conceptus elongation were all associated with differences in the intensity of estrous expression. Most recently, studies from the United States and Canada have demonstrated that reproductive programs emphasizing detection of estrus using AAM can be successful and comparable to intensive timed AI protocol-based programs that incorporate GnRH and PGF2α treatments. Further, one study concluded that the administration of GnRH at AI for spontaneous estrus events greatly improved pregnancy per AI, but only for cows with reduced intensity of estrous expression, showing the potential to use AAM data as a tool in targeted reproductive programs. Quantitative information from estrus events could be used to improve estrus detection and develop decision-making strategies at the farm level. Future studies in this field should aim to better understand ovarian, conceptus, and endometrial mechanisms associated with either the expression or the intensity of estrus, and to refine the identification of phenotypes related to estrus (relative increase, absolute increase, baseline levels, duration, and repeatability within cow) to improve data usage, estrus detection, and possibly genetic selection.

Short communication: Accuracy of estimation of lameness, injury, and cleanliness prevalence by dairy farmers and veterinarians.

Lameness, injuries, and cleanliness are considered important indicators of dairy cow welfare, milk production, and milk quality. Previous research has identified that farmers globally underestimate the prevalence of these cow-based measurements, but no information on the perceptions of veterinarians is available. Because veterinarians are often perceived as the main providers of health advice on farms, the objective of the present study was to evaluate the relationship between the true prevalence of lameness, injury (hock, knee, neck), and cleanliness (udder, legs, flanks), and the estimated prevalence of these issues by farmers and their herd veterinarians. A cross-sectional study was conducted between February 2016 and July 2017. First, the farm owner and the herd veterinarian were asked to estimate the prevalence of lameness, of neck, knee and hock injuries, and of udder, leg, and flank cleanliness on the farm. The research team then visited the farm and scored all lactating cows in the herd for each measurement. Linear regression models were used to assess the relationship between the prevalence estimated by the veterinarians and the farmers, of each cow-based measurement, and the true prevalence on the farm. The 93 herds enrolled had a median of 55 milking cows and were housed in tiestall (90%) and freestall (10%) barns. Ten herd veterinarians participated and were involved with 2 to 22 enrolled farms each. A wide variation was detected in the true prevalence of the different cow-based measurements among herds (lameness: range = 19-72%, median = 36%; neck injuries: range = 0-65%, median = 14%; knee injuries: range = 0-44%, median = 12%; hock injuries: range = 0-57%, median = 25%; dirty udder: range = 0-55%, median 13%; dirty legs: range = 0-91%, median = 18%; and dirty flanks: range = 0-82%, median = 20%). For both veterinarians and farmers, the perception of each cow-based measurement prevalence increased incrementally as the herd's true prevalence increased. Overall, farmers and veterinarians underestimated cow-based measurements. Farmers and veterinarians more accurately estimated lameness prevalence in herds with higher prevalence than in herds with low prevalence, suggesting a better awareness of the issue on farms with lameness problems. Injuries were less accurately estimated in herds with higher injury prevalence compared with herds with lower prevalence, suggesting an opportunity for better knowledge transfer in this area.

Canadian dairy farmers' perception of the efficacy of biosecurity practices.

The adoption of biosecurity practices on dairy farms is limited worldwide. Multiple aspects, one of which is the perception of the effectiveness of these practices, influence the adoption of preventive behavior. The objectives of the present study were to investigate the general understanding of biosecurity by Canadian dairy farmers and to describe their perception of the effectiveness of specific biosecurity practices. In 2015, 368 Canadian dairy farmers were selected to participate in an on-farm national cross-sectional benchmarking study during which they were asked about their general understanding of biosecurity and their perception of the effectiveness of 17 practices to minimize biosecurity risk. For these same biosecurity practices, respondents were asked whether they discussed that particular practice with a veterinarian and whether they implemented it on their farm. Herd and respondent characteristics were also recorded. Associations between the perception of effectiveness of each biosecurity practice and the discussion with a veterinarian, its implementation on farm, and the herd and respondent characteristics were examined using ordered regression models. Most respondents thought the purpose of on-farm biosecurity was to prevent both entry of a new pathogen and spread of an existing pathogen (73%) and considered general biosecurity to be effective (92%) and important (58%). When asked about specific biosecurity practices, respondents considered most of them to be effective (60-94%). Practices related to direct animal-to-animal contact were perceived as effective by more respondents than biosecurity practices related to fomites and visitors. Less than 20% of the respondents reported discussing the different biosecurity practices with a veterinarian, and less than 60% reported implementing these practices on their farm. Geographical region, involvement of a veterinarian, and implementation of the practice on farm were associated with the perception of effectiveness for multiple practices. Finally, the reasons chosen by most respondents as motivation to implement biosecurity practices were (1) if the practice helped to regionally prevent the introduction or the spread of a disease and (2) if it was proven to be beneficial to the health and welfare of animals. These results show that there is a positive perception of the effectiveness of specific biosecurity practices, that there might be a lack of understanding of the practices involving transmission of diseases via indirect contact, and that the perceived threat associated with not adopting practices is minimal.

Biosecurity and herd health management practices on Canadian dairy farms.

The Canadian dairy industry has created national standards to support the adoption of biosecurity practices and to reduce disease risks across the country. There is, however, very little information on the adoption of these practices. The present study aimed to describe the adoption of biosecurity practices on Canadian dairy farms shortly after the creation of the national standards and to identify demographic factors associated with practice adoption. In 2015, 2 questionnaires (phase 1 and 2) were administered to Canadian dairy farmers during an extensive cross-sectional study. Associations between adoption of biosecurity practices as well as associations between adoption of these practices and demographic variables were tested using multiple correspondence analysis. A total of 1,157 questionnaires were completed in phase 1, and a subsample of 368 respondents was selected using stratified random sampling to complete phase 2 during visits to the farms. There was a lack of investigation into general disease syndromes such as a high prevalence of abortion or an unexplained death (38 and 22% of respondents, respectively). Biosecurity measures within herds and between herds to minimize the spread of infection were not widely adopted (e.g., 27% of the respondents never housed sick or lame animals in their calving pen, 41% had closed herds, and 25 and 48% of the open herds had no strategy for introducing new additions and reintroducing returning animals, respectively). Cleanliness of the cows before calving was always ensured by 29% of the respondents, and 27% of the respondents reported always sanitizing the calving pen after each calving. Less than 15% of the respondents had measures in place to limit or control visitors coming on their farm. Moreover, less than half of the respondents reported requiring family members, employees, and visitors to wear farm-designated or clean boots and coveralls. From the multiple correspondence analysis, 2 dimensions were retained and were summarized as "animal movement, calving area, and visitor biosecurity" and "employee biosecurity." Geographical region, type of housing, and milk production were associated with the "employee biosecurity" dimension. The present study demonstrates that many important biosecurity practices are not implemented on Canadian dairy farms; therefore, efforts to promote the adoption of these practices will be necessary to improve biosecurity in dairy herds.

Dairy producers' attitudes toward reproductive management and performance on Canadian dairy farms.

The objectives of this study were to explore Canadian dairy producers' attitudes toward reproductive performance and challenges they perceive to be related to reproduction and reproductive management practices. A survey in both English and French was developed, validated, and administered to Canadian dairy farmers between March and May 2014 to collect general farm, reproduction management, and reproductive performance data, as well as opinions and perceptions about different facets of reproduction. Associations between management practices and the perceived importance of reproduction were tested using a logistic regression model. Thematic network analysis was used to identify themes from the open-ended survey questions about challenges concerning reproduction. Finally, questions that were answered on a Likert scale were graphically represented using diverging stacked bar charts. A total of 832 questionnaires were completed online and by mail, which represents approximately 7% of all dairy farms in Canada. Respondents that ranked reproduction in lactating dairy cows as 1 of the 3 most important challenges faced on their farm (66%) were more likely to house their lactating cows in a tiestall and to have a lower herd annual 21-d pregnancy rate. Estrus detection and conception risk were 2 major themes raised and discussed by the respondents. Other concepts, including housing and milk production, were also perceived to affect estrus detection and conception risk. Whereas analysis of open-ended survey questions does not allow for quantification of the importance of different themes in the sample as a whole, it does show that respondents are aware of the multifactorial complexity of reproductive challenges on dairy farms. Improving performance was the main factor influencing decisions concerning reproduction for 80% of the respondents, and they adopted tools and technologies such as synchronization programs and automated activity monitoring systems to improve herd reproductive performance. More research is required to describe how this performance is defined and perceived by the respondents, and how it relates to the actual variability of performance (i.e., pregnancy rate) among farms.

Performance of automated activity monitoring systems used in combination with timed artificial insemination compared to timed artificial insemination only in early lactation in dairy cows.

Identifying cows in estrus remains a challenge on dairy cattle farms, and tools and technologies have been developed and used to complement or replace visual detection of estrus. Automated activity monitoring (AAM) systems and timed artificial insemination (TAI) are technologies available to dairy farmers, but many factors can influence their relative performance. The objective of the present study was to compare reproductive performance of cows managed with an AAM system combined with TAI, or with a TAI program (Double Ovsynch) for insemination before 88 DIM. From April 2014 to April 2015, 998 cows from 2 herds were randomly assigned either to be inseminated at 85 ± 3 DIM exclusively using the Double Ovsynch protocol for TAI, or to be inseminated based on estrus detection by AAM without hormonal intervention between 50 and 75 DIM; if no alarm was detected by 75 DIM, cows were inseminated following the single Ovsynch protocol (AAM + Ovsynch). The herds used different AAM systems. Parity, hyperketonemia at wk 1 and 2 postpartum (PP), purulent vaginal discharge at wk 5 PP, body condition score at wk 7 PP, and anovulation to wk 9 PP were recorded. These health indicators did not significantly differ between treatments, but did between herds. The effect of treatment on pregnancy at first insemination and by 88 DIM were assessed using logistic regression models. Time to pregnancy was assessed using survival analysis. Results are reported from intention-to-treat analyses. Treatment did not affect pregnancy at first insemination or pregnancy by 88 DIM, but we found significant interactions between treatment and herd for both outcomes. In herd 2, marginal mean pregnancy at first AI was greater with Double Ovsynch (38%) than AAM + Ovsynch (31%), but no difference was observed in herd 1 (Double Ovsynch = 31%; AAM + Ovsynch = 34%). By 88 DIM, a smaller proportion of cows in herd 1 were pregnant in Double Ovsynch (31%) than AAM + Ovsynch (49%), but there was no difference in herd 2 (Double Ovsynch = 38%; AAM + Ovsynch = 38%). We observed a treatment by herd interaction for median (95% confidence interval) time to pregnancy, which were, in herd 1, 110 (106 to 129) and 98 (88 to 113) d, and, in herd 2, 126 (113 to 139) and 116 (105 to 131) d for the Double Ovsynch and AAM + Ovsynch treatments, respectively. The relative performance of AAM-based reproductive management compared with TAI only is likely influenced by herd-specific variables, in particular related to insemination rate when estrus detection is employed.

A dairy herd-level study of postpartum diseases and their association with reproductive performance and culling.

The objectives of this study were to quantify the herd-level prevalence of postpartum diseases in a large number of dairy farms, and to identify prevalence alarm levels of these diseases based on association with a low prevalence of success at first service, with a high prevalence of pregnancy loss following pregnancy diagnosis at first service, and with a high prevalence of postpartum culling. A total of 126 commercial dairy herds were enrolled in this cohort study, and the herd was the unit of interest. Twenty cows from every herd were enrolled during the study period (a total of 2,520 lactating cows in the study). Cows were diagnosed with hyperketonemia, retained placenta, displaced abomasum, purulent vaginal discharge, cytological endometritis, leukocyte esterase endometritis, and prolonged anovulation. The prevalence of each of these diseases was computed for every herd. The study outcomes were the prevalence of success at first service, the prevalence of pregnancy loss following pregnancy diagnosis at first service, and the prevalence of postpartum culling (≤60 d in milk). Descriptive statistics of disease and outcome prevalence were computed. Logistic regression models were used to identify prevalence alarm levels associated with poor outcome prevalence. Median herd prevalence for hyperketonemia, retained placenta, displaced abomasum, purulent vaginal discharge, cytological endometritis, leukocyte esterase endometritis, and prolonged anovulation were 18.8, 4.9, 4.0, 5.0, 29.4, 43.8, and 35.2%, respectively. Herds were defined as having low prevalence of success at first service if <40.0%, as having a high prevalence of pregnancy loss if ≥6.3%, and as having a high prevalence of postpartum culling if ≥13.3%. Risk factors for herds having a low prevalence of success at first service were ≥11.8% hyperketonemia, ≥5.0% purulent vaginal discharge, ≥18.8% cytological endometritis, ≥35.3% leukocyte esterase endometritis, ≥21.0% prolonged anovulation, and ≥4.0% of displaced abomasum. Risk factors for herds having a high prevalence of pregnancy loss were ≥5.0% purulent vaginal discharge and ≥4.9% retained placenta. Risk factors for herds having a high prevalence of postpartum culling were ≥23.1% hyperketonemia, ≥4.9% retained placenta, and ≥4.0% displaced abomasum. Overall, postpartum diseases were prevalent in these dairy herds and alarm levels were identified as risk factors for poor reproductive performance and increased culling.

Survey of reproduction management on Canadian dairy farms.

The objectives of this study were to (1) quantify current reproduction management practices, and (2) assess the association between these practices and herd reproductive performance on dairy farms in Canada. A bilingual survey was developed, validated, and administered from March to May 2014 to collect general and reproduction management and performance measures [annual 21-d pregnancy rate (PR), 21-d insemination rate (IR), and conception risk (CR)]. Associations between management practices and reproductive performance measures were tested using linear regression models. A total of 832 questionnaires were completed online and by mail, representing a response rate of 9%. On average, farms had 77 lactating cows (median=50) and 13 dry cows (median=10), and Holstein was the most common breed (92% of herds). Lactating cow housing was tiestall on 61% of the farms, freestall on 37%, and bedded pack on 2%. The average voluntary waiting period was 58 d in milk (DIM). The main reproduction management practice per farm was defined as the means employed for >50% of inseminations. Farms reported their main reproduction management practice for first and subsequent inseminations, respectively, as visual estrus detection (51 and 44% of herds), timed AI (21 and 23% of herds), automated activity monitoring (AAM; 10 and 10% of herds), other management practice (bulls; 2 and 2% of herds), and a combination of management practices (16 and 21% of herds). On farms using visual estrus detection, cows were observed for signs of estrus on average 3.5 times per day, for an average total of 36 min/d. The most common use of reproductive hormones was to synchronize ovulation using Ovsynch (58% of the farms). Average PR, IR, and CR were 17.6, 44.1, and 40.5%, respectively. In linear regression analyses adjusted for confounders, pregnancy rate was significantly associated with geographic region, housing (tiestall: PR=15.4%, freestall: PR=17.6%), herd size (<50 lactating cows: PR=16.2%, 50-100 cows: PR=16.5%, >100 cows: PR=17.8%), voluntary waiting period (≤60 DIM: PR=17.6%, >60 DIM: PR=15.9%), and frequency of insemination per day (once daily: PR=16.6%, twice or more daily: PR=18.1%). The main reproduction management practice at first and subsequent inseminations was divergently associated with IR and CR, but not with PR (visual heat detection: PR=17.4%, timed AI: PR=18.4%, AAM: PR=17.1%, combined practices: PR=18.2%).

Randomized clinical trial of intrauterine cephapirin infusion in dairy cows for the treatment of purulent vaginal discharge and cytological endometritis.

The objectives of this study were to quantify the effect of an intrauterine infusion of cephapirin on reproductive performance at first service of postpartum dairy cows affected by purulent vaginal discharge (PVD) and cytological endometritis (ENDO) using different diagnostic strategies, and to determine if the presence of prolonged anovulation would influence the magnitude of treatment benefit. In total, 2,259 Holstein cows in 28 herds were enrolled in a randomized clinical trial. At 35 (± 7) days in milk (DIM), cows were diagnosed with PVD using the Metricheck device (Simcro, Hamilton, New Zealand), with cytological endometritis using endometrial cytology (ENDO-CYTO), and with cytological endometritis using leukocyte esterase (ENDO-LE). Regardless of reproductive tract disease status, cows were randomly assigned to receive an intrauterine cephapirin infusion or to not be treated. Serum progesterone was measured at 35 and 49 (± 7) DIM (14 d apart); cows were considered to have prolonged anovulation if progesterone was <1 ng/mL at both times. Reproductive events of cows were collected until 200 DIM. Statistical analyses were conducted using multivariable mixed logistic regression models. Intrauterine cephapirin treatment was associated with an increased first-service pregnancy risk in cows diagnosed with PVD (no treatment: 15.4%; treatment: 31.4%), ENDO-CYTO (no treatment: 16.2%, treatment: 24.4%), and ENDO-LE (no treatment: 15.8%; treatment: 25.1%), but not in cows unaffected by any form of reproductive tract disease (no treatment: 34.8%; treatment: 32.6%). Cephapirin treatment was also associated with an increased first-service reproductive performance in cows affected simultaneously by both PVD and ENDO-CYTO (no treatment: 8.7%; treatment: 23.4%). The effect of cephapirin treatment in anovular cows (no treatment: 21.0%; treatment: 26.4%) was numerically lower than in cyclic cows (no treatment: 22.7%; treatment: 34.1%). Overall, an intrauterine infusion of cephapirin improved first-service pregnancy risk in cows with postpartum reproductive tract disease and this effect was influenced by postpartum anovulation status.