A randomized, controlled trial examining quarter-level somatic cell count and culture-based selective dry cow therapy against blanket dry cow therapy on early lactation production outcomes.

The objective of this study was to determine quarters requiring antimicrobial treatment using either a benchtop somatic cell counter or culture with gram-positive selective media and compare the outcomes in these cows to those receiving blanket dry cow therapy (BDCT) in a randomized, controlled trial. We evaluated 2 novel methods of identifying cows with intramammary infections followed by selective antimicrobial treatment at a commercial dairy farm to determine their usefulness in decreasing antibiotic usage during the dry period without significant detrimental effects on milk quality and production. Cows (n = 840) were randomly allocated to one of 3 groups (BDCT, gram-positive selective media culture-based selective dry cow therapy [C-SDCT], and somatic cell count-based SDCT [S-SDCT]) the day before dry-off, and quarter-level milk samples (QLMS) were collected. The QLMS from cows in the S-SDCT group were evaluated using the cell counter, and quarters were treated if SCC was ≥200,000 cells/mL, whereas the QLMS from cows in the C-SDCT group were cultured, and quarters were treated if the culture showed growth. All cows in the BDCT received antimicrobial therapy, and all cows received an internal teat sealant regardless of treatment group. Outcomes measured were first and second DHIA test SCC, milk production through 60 DIM, cows leaving the farm, clinical mastitis, and bacteriologic new infections in a subset of quarters. Cows in both SDCT groups had fewer antimicrobial treatments than cows in the BDCT group as was expected, and cows in the C-SDCT group had fewer treatments than those in the S-SDCT group. Cows in both SDCT groups had a higher linear score at the first DHIA test (BDCT: 1.8, S-SDCT: 2.2, C-SDCT: 2.2); however, we found no other differences between groups regarding any other outcomes measured. Although antimicrobial use was significantly reduced, farms should use caution in adopting the benchtop analyzer and the selective media described in this study as ways to identify infected cows for dry cow therapy because they may result in increased linear scores early in lactation.

Evaluating the performance of herd-specific Long Short-Term Memory models to identify automated health alerts associated with a ketosis diagnosis in early lactation cows.

The growing use of automated systems in the dairy industry generates a vast amount of cow-level data daily, creating opportunities for using these data to support real-time decision-making. Currently, various commercial systems offer built-in alert algorithms to identify cows requiring attention. To our knowledge, no work has been done to compare the use of models accounting for herd-level variability on their predictive ability against automated systems. Long Short-Term Memory (LSTM) models are machine learning models capable of learning temporal patterns and making predictions based on time series data. The objective of our study was to evaluate the ability of LSTM models to identify a health alert associated with a ketosis diagnosis (HAK) using deviations of daily milk yield, milk FPR, number of successful milkings, rumination time, and activity index from the herd median by parity and DIM, considering various time series lengths and numbers of d before HAK. Additionally, we aimed to use Explainable Artificial Intelligence method to understand the relationships between input variables and model outputs. Data on daily milk yield, milk fat-to-protein ratio (FPR), number of successful milkings, rumination time, activity, and health events during 0 to 21 d in milk (DIM) were retrospectively obtained from a commercial Holstein dairy farm in northern Indiana from February 2020 to January 2023. A total of 1,743 cows were included in the analysis (non-HAK = 1,550; HAK = 193). Variables were transformed based on deviations from the herd median by parity and DIM. Six LSTM models were developed to identify HAK 1, 2, and 3 d before farm diagnosis using historic cow-level data with varying time series lengths. Model performance was assessed using repeated stratified 10-fold cross-validation for 20 repeats. The Shapley additive explanations framework (SHAP) was used for model explanation. Model accuracy was 83, 74, and 70%, balanced error rate was 17 to 18, 26 to 28, and 34%, sensitivity was 81 to 83, 71 to 74, and 62%, specificity was 83, 74, and 71%, positive predictive value was 38, 25 to 27, and 21%, negative predictive value was 97 to 98, 95 to 96, and 94%, and area under the curve was 0.89 to 0.90, 0.80 to 0.81, and 0.72 for models identifying HAK 1, 2, and 3 d before diagnosis, respectively. Performance declined as the time interval between identification and farm diagnosis increased, and extending the time series length did not improve model performance. Model explanation revealed that cows with lower milk yield, number of successful milkings, rumination time, and activity, and higher milk FPR compared with herdmates of the same parity and DIM were more likely to be classified as HAK. Our results demonstrate the potential of LSTM models in identifying HAK using deviations of daily milk production variables, rumination time, and activity index from the herd median by parity and DIM. Future studies are needed to evaluate the performance of health alerts using LSTM models controlling for herd-specific metrics against commercial built-in algorithms in multiple farms and for other disorders.

Association of time to metritis diagnosis with circulating concentration of metabolites, minerals, and haptoglobin in Jersey cows.

Our objective was to investigate the association of early metritis [EMET, diagnosed at <5 d in milk (DIM)] and late metritis (LMET, diagnosed at ≥5 DIM) with circulating concentrations of energy metabolites, minerals, and haptoglobin (Hp) throughout the first 14 d postpartum. A total of 379 purebred Jersey cows were enrolled in a prospective cohort study from a single herd in west Texas. Cows were examined for metritis using the Metricheck device (Simcro Ltd.) at 4, 7, and 10 DIM. Cows identified by farm employees as possible metritis cases were also evaluated for metritis. Blood samples were collected for analysis of concentrations of Ca, Mg, and glucose at DIM 1 through 5, 7, 10, and 14. Albumin, urea, fructosamine, free fatty acids (FFA), creatinine, and ß-hydroxybutyrate (BHB) were analyzed at DIM 3, 5, 7, 10, and 14, and Hp at DIM 1 through 5 and 7. Data were analyzed using the MIXED and PHREG procedures of SAS (SAS Institute Inc.). A series of mixed general linear models accounting for repeated measures were fitted to the data. The independent variables metritis [no metritis (NMET), EMET, and LMET], DIM of analyte assessment, and parity were forced in all models. Multivariable Cox proportional hazard models were built to assess the risk of pregnancy and culling within 150 DIM. The overall metritis incidence was 26.9% (EMET = 49; LMET = 53; NMET = 277). Average concentrations of glucose, Mg, and urea were not associated with metritis. The associations of Ca, creatinine, BHB, and fructosamine with metritis were dependent on the DIM of analyte assessment. Cows categorized as EMET and LMET had, on average, lower albumin and fructosamine compared with NMET cows. Both EMET and LMET cows had, on average, greater BHB than NMET cows. A greater FFA concentration was only observed in cows diagnosed with EMET compared with NMET cows (EMET = 0.58, LMET = 0.52, NMET = 0.48 mmol/L). Additionally, circulating Hp concentration was greater for LMET and EMET compared with NMET cows, and EMET cows had greater Hp compared with LMET cows (EMET = 1.15; LMET = 1.00; NMET = 0.84). In conclusion, several blood biomarkers were temporally associated with early versus late metritis diagnosis in postpartum Jersey cows. No meaningful differences were observed in production, reproduction, or culling between EMET and LMET cows. These results suggest that cows with EMET undergo a more severe degree of inflammation and negative energy balance compared with NMET cows.

Prolonged, low-grade inflammation in the first week of lactation: Associations with mineral, protein, and energy balance markers, and milk yield, in a clinically healthy Jersey cow cohort.

Our objectives were to perform a proof-of-concept study to assess the association of a prolonged inflammatory state (based on a continually elevated haptoglobin concentration at the end of the first week after parturition) with mineral, protein, and energy balance markers in the first 2 wk after parturition, and milk production in the first 60 d of lactation in clinically healthy cows. We conducted a cohort study in 1 herd in west Texas that was milking Jersey and Jersey-Holstein crosses. Only multiparous Jersey cows were eligible for enrollment. Cows were classified as having or not having elevated haptoglobin concentrations based on plasma concentrations evaluated on d 4 and 7 postpartum. We used median concentrations of haptoglobin in the reference population (i.e., before the exclusion of cows diagnosed with clinical diseases) as the limits for categorizing cows into 2 groups: cows with plasma haptoglobin concentrations greater than the median values on both d 4 (0.49 g/L) and 7 (0.35 g/L) had continually elevated haptoglobin (with eHp); and cows with plasma haptoglobin concentrations lower than or equal to the median values of the reference population on d 4 or 7 did not have continually elevated haptoglobin (without eHp). Next, cows with clinical diseases in the first 2 wk of the postpartum period were excluded, so that 233 cows remained for the final analyses. Evaluated outcomes were average daily milk production across the first 60 d of lactation, plasma Ca, Mg, and glucose concentrations on d 1, 2, 3, 4, 5, 7, 10, and 14 postpartum, and ß-hydroxybutyrate (BHB), nonesterified fatty acids (NEFA), fructosamine, albumin, urea, and creatinine concentrations on d 3, 5, 7, 10, and 14 postpartum. Rectal temperatures measured on d 4, 7, and 10 postpartum were also analyzed. We performed statistical analyses using linear mixed models while accounting for the repeated effect of sampling time (plasma analytes and rectal temperature models) and weekly milk test (milk production model). Cows with eHp had lower plasma Ca concentrations in the first 2 wk after calving, but no differences in Mg, BHB, NEFA, glucose, or rectal temperatures compared to cows without eHp. Cows with eHp had lower plasma fructosamine, albumin, and urea concentrations in a time-dependent manner. They also had lower milk production (2.3 kg/d less than cows without eHp in the first 60 DIM). Our study demonstrated that 25% of cows without clinical disorders in the first 2 wk after parturition had continually elevated haptoglobin concentrations at d 7 after parturition relative to d 4, suggestive of a prolonged, low-grade systemic inflammatory state. More observational studies are needed to more fully characterize the duration of prolonged postpartum subclinical inflammation in cows without clinical diseases, as well as its long-term effects, and to evaluate the use of other potential markers of systemic inflammation to describe this disorder.

Short communication: Associations of serum biomarkers of stress and inflammation measured at arrival with health, mortality, and growth of calves transported within the first 4 days of life.

The objective of this study was to assess the association between serum concentrations of haptoglobin (Hp), cortisol, and l-lactate following transport of dairy heifer calves within their first week of life with health, mortality, and growth during the preweaning period. Blood samples were collected at arrival and included a cohort of 168 Jersey and Jersey-cross calves from 8 different sources located in Minnesota. All calves were raised at a single facility in New Mexico. Incidence of respiratory disease (BRD) and mortality during the preweaning period (60 d of life) were extracted from the farm's software database. Individual body weight was measured at birth and at weaning to estimate average daily gain. No simple linear correlations were found between the biomarkers. The incidence of BRD during the preweaning period was 7.7%. While cortisol and l-lactate serum concentrations were not associated with BRD, Hp was negatively associated with BRD. Receiving operating characteristic curve analysis was performed to determine the optimal threshold value for Hp [threshold ≤0.63 µmol/L (63 µg/mL), area under the curve = 0.65]. A Cox's proportional hazards model revealed that calves with Hp concentration ≤0.63 µmol/L were more likely to be diagnosed with BRD (hazard ratio = 5.0, 95% confidence interval = 1.3-19.0). A mixed linear model showed that calves with Hp concentration ≤0.63 µmol/L tended to have lower average daily gain (454.4 vs. 479.9 g/d) during the preweaning period than calves with Hp >0.63 µmol/L at arrival. Overall mortality of the cohort was 3.5%, and Hp was not associated with mortality. Although circulating concentrations of l-lactate and cortisol measured at arrival were not associated with BRD incidence during the preweaning period of heifer calves transported within the first 4 d of life, calves with serum Hp concentration >0.63 µmol/L were less likely to be diagnosed with BRD and tended to grow more than calves with Hp ≤0.63 µmol/L. Further research is needed to replicate these results in a larger cohort and to better understand the possible influence of greater inflammatory status at arrival on health and growth of calves transported to a calf-raising facility.

Association of blood calcium concentration in the first 3 days after parturition and energy balance metabolites at day 3 in milk with disease and production outcomes in multiparous Jersey cows.

Research exploring specific associations of markers of negative energy balance and Ca in postpartum Jersey cows with lactation performance is lacking. Our objectives were to evaluate the associations of total Ca concentration (tCa) measured at 1 through 3 d in milk (DIM) and free fatty acids (FFA), ß-hydroxybutyrate (BHB), and glucose measured at 3 DIM with (1) the risk of multiparous Jersey cows being diagnosed with early-lactation diseases and culling, (2) milk production in the first 9 wk of lactation, and (3) the risk of pregnancy in the first 150 DIM. A cohort study was performed in 1 dairy herd in Texas. Multivariable Poisson regression models were built to evaluate the association of the analytes of interest with the risks of early-lactation diseases and culling in the first 60 DIM (i.e., binary outcomes). Linear mixed models were used to evaluate the association of the analytes of interest with milk production within the first 9 wk of lactation, and a Cox proportional hazard model was built to assess the risk of pregnancy within 150 DIM. A total of 380 cows were used in the final analyses. Total Ca measured at 1 through 3 DIM was not associated with the risk of metritis. Cows with increased FFA and BHB had an increased risk of being diagnosed with metritis and clinical mastitis, respectively. Increased concentrations of glucose and FFA and decreased tCa at 3 DIM were associated with an increased risk of culling. Reduced tCa concentrations at 1 DIM (≤1.84 mmol/L) and 2 DIM (≤2.04 mmol/L) were associated with increased milk production across the first 9 wk of lactation compared with tCa concentrations above those thresholds. Total Ca was not associated with milk production when assessed at 3 DIM, whereas increased FFA (≥0.37 mmol/L) and decreased glucose (≤2.96 mmol/L) at 3 DIM were associated with increased milk production. None of the metabolites measured were associated with the risk of pregnancy in the first 150 DIM. Our results demonstrate that tCa concentration assessed in the first 3 DIM show temporary associations with milk production and culling in multiparous Jersey cows. Although increased concentration of FFA assessed at 3 DIM was associated with greater milk yield, it was a detrimental factor for the risk of metritis. This study attempted to better elucidate the relationship of tCa, FFA, BHB, and glucose assessed in early postpartum with health and performance of Jersey cows. Based on this study, assessments performed at 3 DIM using tCa concentration ≤1.99 mmol/L for increased risk of early-lactation culling and FFA ≥0.43 mmol/L for increased risk of metritis could be used as starting points. More studies evaluating the dynamics of energy balance markers and tCa in postpartum Jersey cows using a greater number of herds are needed to better inform dairy consultants on critical levels for exacerbated postpartum negative energy balance and subclinical hypocalcemia for the Jersey breed.

Association of transient, persistent, or delayed subclinical hypocalcemia with early lactation disease, removal, and milk yield in Holstein cows.

Our objectives were to evaluate the association of subclinical hypocalcemia (SCH) dynamics with the risk of early lactation disease, removal, and milk production. We conducted a prospective observational cohort study in 407 Holstein cows in 2 dairy herds in New York. Cows were stratified by parity group (144 primiparous, 263 multiparous) and classified into 1 of 4 groups based on postpartum plasma Ca concentrations previously associated with improved milk production or increased risk of disease: normocalcemic (NC; primiparous [Ca] >2.15 mmol/L at 1 and 2 d in milk, n = 67; multiparous [Ca] >1.77 at 1 d in milk and 2.20 mmol/L at 4 d in milk, n = 109); transient SCH (tSCH; primiparous [Ca] ≤2.15 at 1 d in milk and >2.15 mmol/L at 2 d in milk, n = 25; multiparous [Ca] ≤1.77 at 1 d in milk and >2.20 mmol/L at 4 d in milk, n = 50); persistent SCH (pSCH; primiparous [Ca] ≤2.15 mmol/L at 1 and 2 d in milk, n = 33; multiparous [Ca] ≤1.77 at 1 d in milk and ≤2.20 mmol/L at 4 d in milk, n = 34); or delayed SCH (dSCH; primiparous [Ca] >2.15 at 1 d in milk and ≤2.15 mmol/L at 2 d in milk, n = 19; multiparous [Ca] >1.77 at 1 d in milk and ≤2.20 mmol/L at 4 d in milk, n = 70). Evaluated outcomes were development of an adverse event [hyperketonemia (blood ß-hydroxybutyrate concentration ≥1.2 mmol/L at 3, 5, 7, or 10 d in milk), metritis, displaced abomasum, or herd removal in the first 60 d in milk] and average milk yield per day across the first 10 wk of lactation. Multivariable Poisson regression was used to analyze the adverse event outcome and generalized linear mixed models for milk yield analysis. Primiparous cows with tSCH were no more likely to have an adverse event than NC cows [risk ratio = 1.3; 95% confidence interval (CI) = 0.5 to 3.2], whereas multiparous cows tended to have a higher risk for an adverse event than NC cows (risk ratio = 1.4; 95% CI = 0.9 to 2.1). However, pSCH cows were 4.1 (95% CI = 2.1 to 7.9, primiparous) and 1.8 (95% CI = 1.2 to 2.7, multiparous) times more likely, and dSCH cows 3.2 (95% CI = 1.5 to 7.0, primiparous) and 1.9 (95% CI = 1.3 to 2.6, multiparous) times more likely, to have an adverse event than NC cows. Primiparous and multiparous cows with tSCH made more milk per day than NC, pSCH, or dSCH cows across the first 10 wk of lactation. Primiparous cows averaged 28.5 ± 0.7, 31.9 ± 1.1, 29.7 ± 0.9, and 28.7 ± 1.2 kg/d, and multiparous cows averaged 44.6 ± 0.7, 49.1 ± 1.1, 46.4 ± 1.3, and 41.4 ± 0.9 kg/d for NC, tSCH, pSCH, and dSCH cows, respectively. Our results suggest that cows with tSCH adapt well to early lactation, develop fewer disease or removal events than pSCH or dSCH cows, and make more milk than NC, pSCH, or dSCH cows. Cows with pSCH or dSCH, regardless of parity group, are at an increased risk for early lactation disease or removal events.

Technical note: Effect of storage time and temperature on total calcium concentrations in bovine blood.

Our objectives were to examine the stability of (1) serum and plasma total calcium (tCa) after delayed separation of bovine whole blood stored at 4°C, and (2) frozen serum and plasma tCa stored at -80°C. Whole blood was collected from 19 cows into vacutainer tubes (Becton Dickinson, Franklin Lakes, NJ) containing no additives (serum) or lithium heparin (plasma). Baseline tCa concentrations (0-h tCa) were determined on serum and plasma harvested within 2 h of blood collection. To evaluate the effect of prolonged whole blood storage at 4°C, serum and plasma were harvested from samples after 6, 24, 48, 72 h, and 7 and 14 d of storage. For evaluation of the stability of frozen serum and plasma stored at -80°C, samples were stored for 7 d and 1, 3, 6, and 12 mo. The median 0-h tCa concentration for serum and plasma was 2.25 mmol/L (range: 1.58 to 2.60 mmol/L) and 2.28 mmol/L (range: 1.60 to 2.65 mmol/L), respectively. The average difference in tCa from 0-h samples ranged from -0.02 to 0.03 mmol/L for all samples stored at either 4 or -80°C. The median within-cow variability for whole blood samples stored at 4°C was 1.1 and 1.0% for serum and plasma, respectively, and for serum and plasma stored at -80°C was 1.1 and 1.3%, respectively. When whole blood was stored at 4°C, no differences in serum or plasma tCa concentrations were observed across the evaluated time points as compared with 0-h concentrations. Similarly, frozen serum and plasma stored at -80°C remained stable up to 12 mo. Therefore, our findings show that whole blood samples may be stored for at least 14 d at 4°C in vacutainer tubes containing no additives or lithium heparin without biologically important changes in tCa concentrations beyond expected analytical variation. Additionally, serum or plasma may be stored at -80°C for up to 12 mo with no effect on tCa concentrations.

Technical note: Effect of delayed analysis of cooled lithium-heparinized whole blood on the stability of ionized calcium, ionized magnesium, sodium, potassium, chloride, glucose, and lactate in samples from dairy cows.

The objectives of this study were to describe the stability of bovine whole-blood electrolytes, glucose, and lactate in samples collected in lithium heparin tubes and stored in thermoconductive modules immersed in ice water. A total of 99 Jersey cows (40 first-parity, 18 second-parity, and 41 third-parity or greater cows) from a commercial dairy farm in West Texas were enrolled between June and July 2018. Blood was collected from the jugular vein using a 60-mL polypropylene syringe and equally distributed into 5 spray-dried evacuated lithium heparin tubes. Baseline samples were analyzed within 90 s of collection using a benchtop blood gas analyzer. The remaining 4 tubes were stored in a thermoconductive, passive-temperature-regulating module inside a cooler with ice water. At 30 min and 2, 4, and 8 h post-collection, samples were removed from the temperature-regulating module, gently inverted for 10 s, and analyzed. Repeated-measures models were built to evaluate the effect of time on the stability of ionized Ca (iCa), ionized Mg (iMg), Na, K, Cl, glucose, and lactate. Most of the analytes investigated remained stable up to 8 h under ice water storage conditions before analysis, including iCa, iMg, Cl, glucose, and lactate. However, Na and K were significantly affected by delayed analysis: Na remained stable up to 4 h post-collection, but K was not stable starting at 2 h post-collection. The results of this study are useful in helping future researchers and consultants to recognize acceptable time delays between whole blood collection and processing or analysis for electrolytes, glucose, and lactate.

Effects of oral calcium bolus supplementation on intracellular polymorphonuclear leukocyte calcium levels and functionality in primiparous and multiparous dairy cows.

The objectives of this study were (1) to characterize Ca levels and polymorphonuclear leukocyte (PMN) function in primiparous and multiparous animals following oral Ca bolus supplementation, and (2) to determine differential responses of boluses containing a lower dose of Ca than traditionally used in primiparous animals on Ca levels and PMN function. Jersey × Holstein crossbred animals (n = 104) were enrolled within 24 h of parturition. All animals were blocked by time relative to calving and randomly assigned to treatment. The Ca boluses were composed of a mixture of Ca chloride, Ca sulfate, and Ca propionate. For objective 1, animals were assigned to control (CON; no Ca supplementation), or a series of 2 Ca boluses given 24 h apart for a total of 50 g of Ca. Objective 2 treatments included control (CON; no Ca supplementation), a series of 2 Ca boluses given 24 h apart containing 50 g of Ca, or a series of 2 Ca boluses given 24 h apart containing 25 g of Ca. Blood samples were collected on d 1 (<24 h), 2, 3, 5, and 7 relative to parturition. Total serum Ca, serum haptoglobin, PMN intracellular Ca, PMN intracellular Ca after stimulation with an environmental Escherichia coli, PMN L-selectin surface expression, and PMN phagocytic and oxidative burst activities were analyzed. For objective 1 a tendency was detected for a treatment difference on basal intracellular PMN Ca and a treatment difference on E. coli-stimulated intracellular PMN Ca. We detected a parity × DIM effect for PMN oxidative burst intensity. However, no other interactions or parity effects on other functional PMN variables were detectable. In primiparous animals, we found a treatment difference for E. coli-stimulated intracellular PMN Ca among animals given 50 g of Ca but no treatment difference on basal intracellular PMN Ca. The 50 g of Ca treatment increased both PMN phagocytosis and oxidative burst intensities. Supplementing animals with 50 g of oral Ca increased intracellular PMN Ca and influenced PMN function.

Method comparison and validation of a prototype device for measurement of ionized calcium concentrations cow-side against a point-of-care instrument and a benchtop blood-gas analyzer reference method.

The objective of this study was to assess an optimized ion-selective electrode Ca-module prototype as a potential cow-side device for ionized Ca (iCa) measurements in bovine blood. A linearity experiment showed no deviation from linearity over a range of iCa concentrations compared with a commercial point-of-care (POC) device commonly used in the field (POCVS; VetScan i-STAT, Abaxis North America, Union City, CA) and a laboratory gold standard benchtop blood-gas analyzer [reference analyzer (RA); ABL-800 FLEX, Radiometer Medical, Copenhagen, Denmark]. Coefficient of variation on 3 samples with high, within-range, and low iCa concentrations ranged from 1.0 to 3.9% for the prototype. A follow-up validation experiment was performed, in which our objectives were to (1) assess the performance of the prototype cow-side against the POCVS (farm gold-standard) using fresh non-anticoagulated whole-blood samples; (2) assess the performance of the prototype and the POCVS against the RA in a diagnostic laboratory using blood collected in a heparin-balanced syringe; and (3) assess the agreement of the prototype and POCVS on-farm (fresh non-anticoagulated whole blood) against the RA on heparin-balanced blood. Finally, sensitivity and specificity of the results obtained by the prototype and the POCVS cow-side compared with the results obtained by the laboratory RA using 3 different iCa cut points for classification of subclinical hypocalcemia were calculated. A total of 101 periparturient Holstein cows from 3 dairy farms in New York State were used for the second experiment. Ionized Ca results from the prototype cow-side were, on average, 0.06 mmol/L higher than the POCVS. With heparin-balanced samples under laboratory conditions, the prototype and POCVS measured an average 0.04 mmol/L higher and lower, respectively, compared with the RA. Results from the prototype and POCVS cow-side were 0.01 mmol/L higher and 0.05 mmol/L lower, respectively, compared with results from the laboratory RA on heparinized blood. Sensitivity and specificity for the prototype and the POCVS under farm conditions at 3 potential subclinical hypocalcemia cut points were 100 and ≥93.5%, respectively. This novel ion-selective electrode Ca-module could become a rapid low-cost tool for assessing iCa cow-side, while qualitatively allowing classification of subclinical hypocalcemia on-farm.

Epidemiology of subclinical hypocalcemia in early-lactation Holstein dairy cows: The temporal associations of plasma calcium concentration in the first 4 days in milk with disease and milk production.

The objectives of this study were to characterize the epidemiology of subclinical hypocalcemia (SCH) in Holstein dairy cows by assessing the temporal associations of plasma Ca concentrations in the first 4 d in milk (DIM) with the risk of cows being diagnosed with metritis or displaced abomasum (or both), and milk production across the first 15 wk of lactation. A prospective cohort study was conducted in 2 dairy herds in New York State, in which cows had a blood sample collected daily for the first 4 DIM. A total of 396 Holstein cows (137 primiparous and 259 multiparous) were enrolled. Multivariable Poisson regression models were built to evaluate the associations of plasma Ca concentration at each of the 4 d following parturition with the risk of primiparous cows being diagnosed with metritis and multiparous cows being diagnosed with metritis, displaced abomasum, or both. Similarly, generalized linear mixed models were built to evaluate the associations of plasma Ca concentration with milk production across the first 15 wk of lactation. Plasma Ca concentration was assessed on a continuous scale in all models; dichotomization and SCH classification only occurred in the final models if the Ca concentration variable was meaningful by creating an optimized threshold based on receiver operating characteristic curve analysis. Plasma Ca concentration assessed at 1 DIM was not associated with the risk of metritis in primiparous cows, but an association was observed at 2, 3, and 4 DIM (critical thresholds were plasma Ca concentration ≤2.15, 2.10, and 2.15 mmol/L, respectively). Plasma Ca concentration was associated with the risk of metritis or displaced abomasum diagnosis (or both) for 2nd parity animals at 2 DIM (threshold ≤1.97 mmol/L), and at 4 DIM for 3rd and greater lactations (threshold ≤2.20 mmol/L). Reduced plasma Ca concentration was associated with higher milk production when assessed at 1 DIM in primiparous and multiparous cows, and lower milk production when assessed at 4 DIM in multiparous cows only. For primiparous cows, plasma Ca concentration was not associated with lower milk production at any of the DIM assessed. In conclusion, assessments of SCH at the individual cow level must take into account the DIM of Ca concentration measurement and parity of the cow, as the epidemiology of the disorder was demonstrated to be highly dependent on these variables. This study advances the knowledge of the epidemiology of SCH and better establishes thresholds for optimizing SCH diagnosis.

Association of immediate postpartum plasma calcium concentration with early-lactation clinical diseases, culling, reproduction, and milk production in Holstein cows.

The objective of this study was to evaluate the association of postpartum plasma Ca concentration with early-lactation disease outcomes, culling within 60 d in milk, pregnancy to first service, and milk production. A total of 1,453 cows from 5 commercial dairy farms in New York State were enrolled in a prospective cohort study from February to November 2015. Blood samples were collected within 12 h of parturition, and plasma was submitted to a diagnostic laboratory for total Ca measurement. Early-lactation disease, reproductive performance, and milk production from Dairy Herd Improvement Association (DHIA) test-day data were compiled from each farm's management software. Multivariable Poisson regression models were built to evaluate the association of plasma Ca with the risks of retained placenta (RP), metritis, displaced abomasum (DA), clinical mastitis, culling within 60 d in milk, and pregnancy to first service. Repeated-measures ANOVA were used to evaluate the association of Ca at parturition with milk production across the first 9 DHIA tests. Herd was considered a random effect in all models. Primiparous cows were modeled separately from multiparous cows if differential responses were observed. Calcium was not associated with the risk of RP, metritis, clinical mastitis, or pregnancy to first service in primiparous or multiparous cows. For multiparous cows only, higher Ca concentration tended to be associated with increased culling within the first 60 d in milk. Multiparous cows with Ca ≤1.85 mmol/L had an increased risk of being diagnosed with a DA compared with cows with Ca >1.85 mmol/L. For the milk production models, Ca was not associated with the amount of milk produced within the first 9 DHIA tests in primiparous cows; however, multiparous cows with Ca ≤1.95 mmol/L produced, on average, 1.1 kg more milk per day across the 9 DHIA tests than their multiparous counterparts with Ca >1.95 mmol/L. Our results indicate that plasma Ca concentration measured within 12 h of parturition is a poor predictor of early-lactation health outcomes. Reduced Ca concentration in the immediate postpartum period was associated with higher milk production in multiparous cows. From these results, we caution that studies attempting to categorize subclinical hypocalcemia based on a single sample in the immediate postpartum period could misclassify the disorder.

Differential effects of a single dose of oral calcium based on postpartum plasma calcium concentration in Holstein cows.

Our objectives were to determine (1) the effect of a single dose of an oral Ca bolus within 24 h after parturition on plasma Ca concentration, (2) the response of primiparous (PP) and multiparous (MP) cows to this supplementation strategy, and (3) differential responses based on plasma Ca at enrollment. For objective 1, cows from 1 commercial dairy in New York State were enrolled within 19 h after parturition (mean ± standard deviation = 8.3 ± 5.3 h) and randomized within parity group (first, second, and ≥third) to control [CON (n = 25); no placebo] or a single dose bolus treatment [BOL (n = 25); 3 oral Ca boluses supplying 54 to 64 g of Ca]. Plasma Ca was measured repeatedly between 1 and 24 h following treatment. For objectives 2 and 3, cows on 6 commercial farms in New York State were assigned to treatment as for objective 1 (CON, n = 1,973; BOL, n = 1,976). Herd records for health, reproduction, and Dairy Herd Improvement Association test day milk production were collected. Mixed effect multivariable models were developed using repeated measures ANOVA, Poisson regression, or proportional hazard models. Objective 2 analyses considered treatment with periparturient risk factors, whereas objective 3 analyses also considered Ca status. No difference was observed for plasma Ca between 1 and 24 h after treatment. Primiparous cows assigned to BOL calving at >712 d old had decreased risk of one or more health disorders [≤30 d in milk; risk ratio (RR) = 0.65, 95% confidence interval (CI) = 0.51 to 0.84] and those with body condition score >3.5 responded to BOL with increased milk production (CON = 31.7 ± 1.1, BOL = 35.1 ± 1.1 kg/d), as did those with days carried calf >277 (CON = 31.9 ± 1.0, BOL = 34.7 ± 1.0 kg/d). Reduced risk of one or more health disorders was observed in parity ≥3 (RR = 0.85, 95% CI = 0.81 to 0.89) and MP cows with body condition score >3.5 (retained placenta; RR = 0.70, 95% CI = 0.58 to 0.84) or that were lame (displaced abomasum; RR = 0.49, 95% CI = 0.32 to 0.75). Differential responses for PP cows by Ca status were minimal. For MP cows with low plasma Ca, BOL decreased risk of additional Ca treatment (≤1.8 mmol/L; RR = 0.57, 95% CI = 0.40 to 0.80) as well as risk of one or more health disorders (≤2.15 mmol/L; RR = 0.90, 95% CI = 0.85 to 0.95). Supplementation with a single oral dose of Ca could be targeted to periparturient risk groups for improved health. Calcium status did not differentiate responses of PP cows, but MP cows with low Ca at parturition had improved health status when supplemented.

Risk factors associated with postpartum subclinical hypocalcemia in dairy cows.

Our study objectives were to evaluate the association of prepartum plasma Mg concentrations with subclinical hypocalcemia (SCH) classification at parturition and to evaluate the association of other cow-level risk factors with SCH classification at calving or at 2 d in milk (DIM). A total of 301 animals from 2 dairy herds located in New York were enrolled in a cohort study. Blood samples were collected at approximately 1 wk before the expected calving date, within 4 h of calving, and at 2 DIM. Prepartum samples had plasma macromineral concentrations (Ca, K, Mg, P), albumin, and ß-hydroxybutyrate analyzed. Samples collected at calving were analyzed for Ca only, and samples from 2 DIM had macromineral and albumin concentrations determined. Postpartum SCH was defined as Ca concentrations ≤2.1 mmol/L. The prevalence of SCH at calving was 2, 40, and 66% for first, second, and third or greater parities, respectively. Only 4% of cows could be classified with prepartum subclinical hypomagnesemia (Mg concentrations <0.8 mmol/L), which did not provide enough power to appropriately determine the association of plasma Mg with postpartum Ca concentrations and its effect on SCH classification. Multiparous cows with Ca concentrations ≤2.4 mmol/L in the prepartum period and third or greater parity cows had a higher risk of being categorized as SCH at calving [relative risk (RR) = 1.4 and 1.7, respectively]. The risk of SCH at 2 DIM was associated with the interaction of Ca status at calving and lameness score. Nonlame cows with Ca concentrations ≤2.1 mmol/L (RR = 3.2) and normocalcemic lame cows at parturition (RR = 3.4) were more likely to be SCH at 2 DIM compared with nonlame normocalcemic cows. In conclusion, we identified a prepartum Ca cut-point for identification of cows that are more likely to be classified as SCH at calving. Different risk factors were associated with SCH depending on the timing of diagnosis relative to parturition.

Reproductive management practices and performance of Canadian dairy herds using automated activity-monitoring systems.

The objectives of this study were to describe the characteristics and motivations of producers who had implemented automated activity-monitoring (AAM) systems and to compare herd reproductive performance before and after the implementation of an AAM system and between herds with AAM and herds managing reproduction based on timed artificial insemination (TAI) or based on other programs. Freestall dairy herds located in Ontario and the western provinces of Canada and enrolled in Dairy Herd Improvement were surveyed through a mail questionnaire between April and July 2010. The data describe the characteristics and reproductive management practices of herds using AAM systems. A total of 505 questionnaires (29%) were returned. On average, 21-d pregnancy risk, conception risk, and 21-d insemination risk did not differ between herds managing reproduction based on an AAM system (18, 39, and 50%, respectively) or a TAI-based program (17, 38, and 49%, respectively). Herds that implemented an AAM system had a significant increase in annual pregnancy risk, from 15 to 17%, and insemination risk increased from 42 to 50%, whereas conception risk was unchanged (37 and 35%) following adoption of the system. The majority of respondents with AAM systems first used the system to manage reproduction in lactating cows. Most herds with AAM were performing artificial insemination twice per day, most commonly with an interval from the estrus alarm to artificial insemination of 7 to 12 h. The most commonly reported reason to adopt an AAM system was a desire to improve reproductive performance. These results support the findings from randomized trials that AAM-based programs can yield comparable reproductive performance to TAI-based programs.

Plasma oxylipin profile of postpartum dairy cows categorized into different systemic inflammatory grades in the first week after parturition.

Oxygenated polyunsaturated fatty acids (oxylipins) are important mediators of inflammation ranging from pro- to anti-inflammatory actions. Research investigating differences in the oxylipin profile of dairy cows suffering from different degrees of systemic inflammation in the early postpartum period is lacking and can help advance knowledge on potential mechanisms leading to excessive inflammation. The objective of this preliminary study was to evaluate the plasma oxylipin profile of cows classified in 1 of 4 systemic inflammation categories based on plasma haptoglobin (Hp) concentrations assessed on days in milk (DIM) 1, 2, 3, 4, 5, and 7, in addition to the presence or absence of metritis within 10 DIM, and of cows without any clinical diseases within 21 DIM. Groups were classified as follows: (1) cows with a peak Hp concentration ≤3 DIM (EarlyHp) and diagnosed with metritis; (2) cows with a peak Hp concentration 3 < DIM ≤7 (LateHp) and diagnosed with metritis; (3) cows suffering from persistently elevated Hp concentrations assessed on DIM 4 and 7 while remaining apparently healthy during the first 21 DIM (PersistentHp); and (4) apparently healthy cows not suffering from persistently elevated Hp concentrations (LowHp). Six cows from each category were randomly selected from a plasma bank of a parent cohort study including 380 multiparous cows. Plasma samples on DIM 1 and 2, 3 and 4, and 5 and 7 were proportionally pooled to create 3 samples per cow for lipidomic analysis (i.e., pool 1 = DIM 1 and 2; pool 2 = DIM 3 and 4; pool 3: DIM 5 and 7). Statistical analyses were performed using SAS v9.4 (SAS Institute Inc.) and least squares means adjusted for multiple comparisons using the Tukey-Kramer test. Comparisons for EarlyHp and LateHp were only performed on pooled samples from DIM 1 and 2 (i.e., before metritis diagnosis). EarlyHp cows had decreased concentrations of 9(S)-HOTrE compared with LowHp cows of DIM 1-2 pooled samples. LateHp cows had decreased concentrations of 9(10)-DiHOME compared with LowHp cows. Next, we sought to investigate whether cows classified as PersistentHp had time-dependent differences in oxylipin profile versus LowHp cows. PersistentHp cows had decreased concentrations of 19(R)-HETE compared with LowHp cows in a time-dependent manner (only in pooled samples from DIM 5 and 7). Our results identified oxylipins of interest that warrant further investigation to elucidate their in vitro and in vivo functions in the postpartum inflammatory process of dairy cows.

Comparison of an image cytometry somatic cell count analyzer to a flow cytometry analyzer.

The objective of this study was to evaluate the performance of a small footprint benchtop somatic cell counter based on image cytometry (LactiCyte HD; Page and Pedersen International Ltd., Hopkinton, MA) against a flow cytometer employed at a regional dairy herd improvement (DHI) laboratory. Milk samples collected during monthly DHI testing were split into 2 samples. One sample was evaluated using flow cytometry (Bentley SomaCount FCM; Bentley Instruments, Chaska, MN) at the regional DHI laboratory, whereas the other was evaluated using image cytometry at 2 different image levels (full number of images, 16 pictures per slide; half number of images, 8 pictures per slide). Mean bias of the image cytometer at 16 images was -15,500 cells/mL, whereas at 8 images the bias was 21,800 cells/mL. When considering only cell counts ≤400,000 cells per mL, the bias for both imaging resolutions was positive, meaning the image cytometer read higher than the flow cytometer. Both imaging resolutions (16 and 8) had a concordance correlation coefficient greater than 0.95. Considering ≥200,000 cells/mL to be indicative of subclinical mammary gland infection, the sensitivity and specificity of the image cytometer at 16 images were 92.0% and 91.7%, whereas the sensitivity and specificity of the analyzer at 8 images were 92.0% and 85.7%, respectively. Method precision (repeatability; coefficients of variation) were calculated at 3 different somatic cell counts (100,000, 200,000, and 400,000 cells/mL) where each sample was run repeatedly 12 times. When analyzed at the full number of images the coefficients of variation were 16.9%, 11.7%, and 10.9% for 100,000, 200,000, and 400,000 cells/mL, respectively. Analysis at half the number of images resulted in coefficients of variation of 18.9%, 24.8%, and 8.7% for 100,000, 200,000, and 400,000 cells/mL. We conclude that the image cytometer is an acceptable somatic cell count analyzer for on-farm use for applications such as screening cows for microbiological testing, and that precision is superior when the analysis is performed at the full number of images allowed by the instrument.

Reproductive performance with an automated activity monitoring system versus a synchronized breeding program.

The objective was to compare reproductive performance with management programs based on an automated activity monitoring (AAM) system or a synchronized breeding program under field conditions. In total, 1,429 Holstein cows from 3 commercial herds in Ontario, Canada, were enrolled over 1 yr in a randomized controlled trial. At each farm, primiparous and multiparous animals were housed in separate pens. At the pen level, cows were assigned to reproductive management primarily using an AAM system based on monitoring activity levels (Heatime, SCR Engineers Ltd., Netanya, Israel) or a timed artificial insemination (TAI) program. A crossover occurred after 6 mo of the trial to avoid confounding treatment with parity. Insemination based on additional detection of estrus by visual observation was practiced in all pens. At the individual cow level, time to pregnancy throughout the study (n=1,985 cow-6 mo periods), time to first service, and time to second service were analyzed with a Cox proportional hazards models accounting for herd, and did not differ overall between the AAM and TAI treatment groups. However, we observed an interaction between herd and breeding program, such that association of AAM- or TAI-based program with these outcomes differed between farms. Time to pregnancy was not different in herd A [median=151 and 136 d; hazard ratio (HR) = 0.93] and herd C (median=99 and 124, HR = 1.24), whereas herd B had a median time to pregnancy of 119 d and 146 d (HR = 1.3) in the AAM and TAI groups, respectively. Under conditions in which 19 to 32% of artificial inseminations in both groups were based on visually detected estrus, herd pregnancy rate and cow-level time to pregnancy did not differ overall between TAI- and AAM-based programs, but the effect of reproductive management approach depended on herd. Considering 924 cow-6 mo periods with artificial inseminations only by the assigned program, we also found an interaction between herd and breeding program. Stratified analysis showed no difference in time to pregnancy in herd A (HR = 1.3), whereas in herds B (HR = 1.7) and C (HR = 2.8), cows in the AAM treatment group became pregnant sooner compared with those in the TAI group. Factors that influence the variability in relative performance of these management systems between herds require further investigation. Our study used one commercial activity monitoring system and the results cannot necessarily be generalized to other systems.

Early life indicators of first lactation milk yield and the effect of treatment for bovine respiratory disease on survivability and risk of pregnancy in Holstein dairy cattle.

The primary objective of this study was to evaluate the long-term effects of variables that could be measured early in life on first lactation milk production. The secondary objective was to evaluate the long-term effects of treatment for bovine respiratory disease (BRD), within the first 120 days of life, on survivability to 850 days and reproductive success. Data from Holstein heifer calves was collected from 1 October, 2015-31 January, 2020 and included milk consumption (MC; n = 10,431), bodyweight (BW; n = 9,825), average daily gain (ADG; n = 6,194), heifer conception age (n = 10,112), and 305-day first lactation milk production (305 M; n = 5,823), and treatment records for BRD (n = 12,485). Calves were fed pasteurized whole milk through an automated calf feeding system for 60 days (range, 48 - 126 days), with a 30% crude protein (CP) and 5% crude fat enhancer added at 20 g/L milk. Calves were weighed at birth, weaning, and several other times prior to 2 years of age. Daily BW were predicted for individual animals using a third order orthogonal polynomial to model individual BW regressions. Daily BW predictions were used to calculate ADG and BW predictions on specific days of interest. Season born, ADG (0-400 days), milk production potential, and heifer conception age had a significant impact on 305-day first lactation milk production (all P ≤ 0.05; r2 = 0.31). A retrospective cohort study was conducted utilizing data collected from a commercial farm with cohorts defined by BRD status. Calves treated for BRD from 61 to 120 days old had a significantly lower chance of survival to 850 days of age than animals not treated for BRD. Additionally, calves treated for BRD from 61 to 120 days old had reduced risk of pregnancy. The results show the impact of early life on future milk production, survivability, and pregnancy risk on a commercial dairy herd.