A randomized controlled trial evaluating the efficacy of systemic ceftiofur administration for metritis therapy in dairy cows and the effect of metritis cure on economically important outcomes.

The main objective of this study was to evaluate the effect of ceftiofur on metritis cure, milk yield, reproductive performance, and culling up to 300 DIM. The secondary objective was to evaluate the effect of metritis cure at 5 (ECURE) and 14 (LCURE) d after diagnosis on milk production, reproduction, and culling. A total of 422 Holstein cows diagnosed with metritis from 4 herds located in TX, CA, and FL were enrolled in a randomized clinical trial. Cows diagnosed with metritis (fetid, watery, reddish/brownish uterine discharge) were blocked by herd and parity and were randomly allocated to receive systemic administration of ceftiofur (CEF) or to remain untreated (CON). In addition, 399 non-metritic cows (NMET) were included for comparison purposes. Metritis cure was evaluated at 5 and 14 d after diagnosis and was defined as the absence of metritis clinical signs. Logistic regression models were fitted to the data to assess the effect of treatment on metritis cure. Milk yield was analyzed using a mixed linear model, while logistic regression, Cox proportional hazard and Kaplan-Meier survival analysis models were fitted to culling and reproduction data. Cows treated with CEF had 1.86 (95% CI: 1.22 - 2.81) and 1.68 (95% CI: 1.02 - 2.75) greater odds of being cured than CON cows at 5 and 14 d after diagnosis, respectively. No effect of CEF was observed for milk yield; however, NMET cows had greater milk yield compared with metritic cows (CEF = 36.0, 95% CI = 33.8 - 38.1; CON = 36.1, 95% CI = 33.9 - 38.2; NMET = 36.9 kg/d, 95% CI = 34.8 - 39.4). Likewise, no effect of CEF was observed on reproductive performance and culling. Nonetheless, the likelihood of conceiving for NMET cows was 1.72 (95% CI = 1.41 - 2.12) and 1.64 (95% CI = 1.33 - 2.00) times greater than for CEF and CON cows, respectively. Ceftiofur-treated and CON cows had 2.93 (95% CI = 1.90 - 4.51) and 2.37 (95% CI = 1.51 - 3.71) greater hazard of culling compared with NMET, respectively. Regardless of treatment, no differences between ECURE and LCURE were observed on milk yield, reproduction, and culling throughout the entire lactation, but cows that cured at 5 or 14 d after diagnosis had greater milk production in the first 60 DIM compared with cows that did not cure (NCURE). Cows in ECURE and LCURE also had a 1.59 (95% CI = 1.16 - 2.16) and 1.49 (95% CI = 1.08 - 2.05) greater hazard of pregnancy and 0.43 (95% CI = 0.26-0.71) and 0.56 (95% CI = 0.34-0.92) hazard of culling compared with NCURE. Ceftiofur therapy increased metritis cure, but benefits to productivity and longevity were not observed. Also, cows that fail to cure have impaired lactation performance, but no differences regarding timing of cure were observed.

Evaluating differences in milk production, reproductive performance, and survival associated with vaginal discharge characteristics and fever in postpartum dairy cows.

The objective was to assess differences in productive and reproductive performance, and survival associated with vaginal discharge characteristics and fever in postpartum dairy cows located in Western and Southern states of the U.S.A. This retrospective cohort study included data from 3 experiments conducted in 9 dairies. Vaginal discharge was evaluated twice within 12 DIM and scored on a 5-point scale. The highest score observed for each cow was used for group assignment (VD group) as follows: VD 1 and 2 (VD 1/2; n = 1,174) = clear mucus/lochia with or without flecks of pus; VD 3 (n = 1,802) = mucopurulent with < 50% pus; VD 4 (n = 1,643) = mucopurulent with ≥50% of pus or non-fetid reddish/brownish mucous, n = 1,643; VD 5 = fetid, watery, and reddish/brownish, n = 1,800. All VD 5 cows received treatment according to each herd's protocol. Rectal temperature was assessed in a subset of VD 5 cows, and subsequently divided into Fever (rectal temperature ≥39.5°C; n = 334) and NoFever (n = 558) groups. A smaller proportion of cows with VD 5 (67.6%) resumed ovarian cyclicity compared with VD 1/2 (76.2%) and VD 4 (72.9%) cows; however, a similar proportion of VD5 and VD 3 (72.6%) cows resumed ovarian cyclicity. A smaller proportion of VD 5 (85.8%) cows received at least one artificial insemination (AI) compared with VD 1/2 (91.5%), VD 3 (91.0%), or VD 4 (91.6%) cows. Although we did not detect differences in pregnancy at first AI according to VD, fewer cows with VD 5 (64.4%) were pregnant at 300 DIM than cows with VD 1/2 (76.5%), VD 3 (76.2%), or VD 4 (74.7%). Hazard of pregnancy by 300 DIM was smaller for VD 5 compared with VD 1/2, VD 3, or VD 4 cows. A greater proportion of VD 5 cows were removed from the herd within 300 DIM compared with other VD groups. There was 760 kg lesser milk production within 300 DIM for VD 5 compared with VD 2, VD 3, and VD 4, whereas VD 2, VD 3, and VD 4 had similar milk production. We did not detect an association between fever at diagnosis of VD 5 and reproductive performance or milk production. A greater proportion of VD 5 cows without fever were removed from the herd by 300 DIM compared with VD 5 cows with fever. Differences in productive and reproductive performance, and removal of the herd were restricted to fetid, watery, and reddish/brownish vaginal discharge, which was independent of fever.

Predictive models for metritis cure using farm-collected data, metabolic and inflammation biomarkers, and hemogram variables measured at diagnosis.

Our objective was to evaluate the accuracy of predictive models for metritis spontaneous cure (SC) and cure among ceftiofur-treated cows using farm-collected data only, and with the addition of hemogram variables and circulating concentration of metabolites, minerals, and biomarkers (BM) of inflammation measured at time of diagnosis. Data related to parity, calving-related issues, BCS, rectal temperature, and DIM at metritis diagnosis were collected from a randomized clinical trial that included 422 metritic cows from 4 herds in Texas, California, and Florida. Metritis was defined as the presence of red-brownish, watery, and fetid vaginal discharge, and cure was defined as the absence of metritis 14 d after initial diagnosis. Cows were randomly allocated to receive systemic ceftiofur therapy (2 subcutaneous doses of 6.6 mg/kg of ceftiofur crystalline-free acid on the day of diagnosis and 3 d later; CEF) or to remain untreated (control). At enrollment (day of metritis diagnosis), blood samples were collected and submitted to complete blood count (CBC) and processed for the measurement of 13 minerals and BM of metabolism and inflammation. Univariable analysis to evaluate the association of farm-collected data and blood-assessed variables with metritis cure were performed, and variables with P ≤ 0.20 were offered to multivariable logistic regression models and retained if P ≤ 0.15. The areas under the curve for models predicting SC using farm data only and farm + BM were 0.70 and 0.76, respectively. Complete blood count variables were not retained in the models for SC. For models predicting cure among CEF cows, the area under the curve was 0.75, 0.77, 0.80, and 0.80 for models using farm data only, farm + CBC, farm + BM, and farm + CBC + BM, respectively. Predictive models of metritis cure had fair accuracy, with SC models being less accurate than models predictive of cure among CEF cows. Additionally, adding BM variables marginally improved the accuracy of models using farm collected data, and CBC data did not improve the accuracy of predictive models.

Association between prepartum body condition score and prepartum and postpartum dry matter intake and energy balance in multiparous Holstein cows.

The objectives of this retrospective observational study were to investigate the association between BCS at 21 d before calving with prepartum and postpartum DMI, energy balance (EB), and milk yield. Data from 427 multigravid cows from 11 different experiments conducted at the University of Florida (Gainesville, FL) were used. Cows were classified according to their BCS at 21 d before calving as fat (BCS ≥ 4.00; n = 83), moderate (BCS = 3.25-3.75; n = 287), and thin (BCS ≤ 3.00; n = 57). Daily DMI from -21 to -1 and from +1 to +28 DIM was individually recorded. Energy balance was calculated as the difference between net energy for lactation consumed and required. Dry matter intake in fat cows was lower than that in moderate and thin cows both prepartum (fat = 9.97 ± 0.21, moderate = 11.15 ± 0.14, and thin = 11.92 ± 0.22 kg/d) and postpartum (fat = 14.35 ± 0.49, moderate = 15.47 ± 0.38, and thin = 16.09 ± 0.47 kg/d). Dry matter intake was also lower for moderate cows compared with thin cows prepartum, but not postpartum. Energy balance in fat cows was lower than in moderate and thin cows both prepartum (fat = -4.16 ± 0.61, moderate = -1.20 ± 0.56, and thin = 0.88 ± 0.62 Mcal/d) and postpartum (fat = -12.77 ± 0.50, moderate = -10.13 ± 0.29, and thin = -6.14 ± 0.51 Mcal/d). Energy balance was also lower for moderate cows compared with thin cows both prepartum and postpartum. There was a quadratic association between BCS at 21 d before calving and milk yield. Increasing BCS from 2.5 to 3.5 was associated with an increase in daily milk yield of 6.0 kg and 28 d cumulative milk of 147 kg. Increasing BCS from 3.5 to 4.5 was associated with a decrease in daily milk yield of 4.4 kg and 28 d cumulative milk of 116 kg. In summary, a moderate BCS at 21 d before calving was associated with intermediate DMI and EB pre- and postpartum but greater milk yield compared with thinner and fatter cows. Our findings indicate that a moderate BCS is ideal for ensuring a successful lactation.

Blood metabolomics and impacted cellular mechanisms during transition into lactation in dairy cows that develop metritis.

The objective of this study was to identify metabolites associated with metritis and use them for identification of cellular mechanisms affected during transition into lactation. Holstein cows (n = 104) had blood collected in the prepartum period (d -14 ± 6 relative to calving), at calving (d 0), and at the day of metritis diagnosis (d 7 ± 2 after calving). Cows with reddish or brownish, watery, and fetid discharge were diagnosed with metritis (n = 52). Cows with metritis were paired with herdmates without metritis (n = 52) based on days in milk. The metabolome of plasma samples was evaluated using untargeted gas chromatography time-of-flight mass spectrometry. Univariate analyses included t-tests and fold change analyses. Metabolites with false discovery rate adjusted P ≤ 0.10 on t-tests were used for partial least squares discriminant analysis coupled with permutational analysis using 2,000 permutations. Metabolites with false discovery rate adjusted P ≤ 0.10 on t-tests were also used for enriched pathway analyses and identification of cellular processes. Cows that developed metritis had affected cellular processes associated with lower amino acid metabolism in the prepartum period, greater lipolysis, cell death, and oxidative stress at calving and at metritis diagnosis, and greater leukocyte activation at calving, but lower immune cell activation at metritis diagnosis. In summary, cows that developed metritis had plasma metabolomic changes associated with greater lipolysis, oxidative stress, and a dysregulated immune response which may predispose cows to metritis development.

Unraveling the immune and metabolic changes associated with metritis in dairy cows.

The objective was to unravel the peripartum immune and metabolic changes associated with metritis in Holstein cows. Holstein cows (n = 128) had blood collected at -14, 0, 3, and 7 d relative to parturition (DRP). Flow cytometry was used to evaluate blood leukocyte counts, proportions, and activation. Total cells, live cells, single cells, monocytes (CD172α+/CD14+), polymorphonuclears (CD172α+/CD14-/SSChigh), B-cells (CD21+/MHCII+), CD4+ T-cells (CD4+), CD8+ T-cells (CD8+), and γδ T-cells (γδTCR+) were evaluated. Both CD62L and CD11b were used as markers of cell activation. Major histocompatibility complex class II was used as a marker of antigen presentation in monocytes. A Milliplex Bovine Cytokine/Chemokine 08-plex kit was used to evaluate plasma concentrations of IFN-γ, IL-1α, IL-1ß, IL-4, IL-6, IL-8, IL-10, and tumor necrosis factor-α. The body weight (BW) change prepartum was calculated as the difference between calving BW and prepartum BW divided by the number of days between measurements. Plasma fatty acids (FA) were measured at -14 and 0 DRP using untargeted gas chromatography with time-of-flight mass spectrometry. Data were analyzed by ANOVA for repeated measures. Cows that developed metritis (n = 57) had greater prepartum BW, prepartum BW loss, and greater FA concentrations at calving. Plasma FA at calving was positively correlated with IL-1ß. Cows that developed metritis had persistent systemic inflammation, which was demonstrated by greater B-cell activation, greater pro-inflammatory cytokine concentrations, and greater cell damage pre- and postpartum. Postpartum, we observed greater polymorphonuclear cell activation and extravasation but lesser monocytes and CD4+ T-cells activation and extravasation, which suggests postpartum immune tolerance. Greater prepartum adiposity in cows that developed metritis may lead to systemic inflammation pre- and postpartum and immune tolerance postpartum, which may lead to failure to prevent bacterial infection, and development of puerperal metritis.

Combined effect of purulent vaginal discharge and anovulation on pregnancy status in a large multi-state population of Holstein cows.

The objective of this observational prospective cohort study was to evaluate the combined effect of purulent vaginal discharge (PVD) and anovulation (ANOV) on the reproductive performance of a large multi-state population of Holstein cows. Data were prospectively collected from 11,729 cows in 16 herds located in 4 regions in the United States [Northeast (4 herds), Midwest (6), Southeast (1), and Southwest (5)]. Cows were enrolled at calving and monitored weekly for disease occurrence, reproductive events, and survival. Prevalence of PVD was evaluated at 28 ± 3 d in milk and defined by the presence of mucopurulent to fetid vaginal discharge. Resumption of ovarian cyclicity was determined via transrectal ultrasonography at 40 ± 3 and 54 ± 3 d postpartum. Pregnancy diagnosis was performed by ultrasonography on d 32 ± 3 after artificial insemination (AI) and reconfirmed at d 60 ± 3 of gestation. Pregnancy loss (PL) was defined as a cow diagnosed pregnant at 32 ± 3 but nonpregnant at 60 ± 3 d after AI. The association of PVD and ANOV with pregnancy traits was analyzed using 4 PVD-cyclicity categories that considered the following combinations: NPVD-CYC = absence of PVD and cycling; PVD-CYC = presence of PVD and cycling; NPVD-ANOV = absence of PVD and anovular; and PVD-ANOV = presence of PVD and anovular. Multiple logistic regression and Cox proportional regression were used for the analysis of potential associations between PVD and cyclicity categories and pregnancy at first AI (PAI1), days from calving to pregnancy, and PL at first AI. The odds (95% confidence intervals) of pregnancy increased from cows in the PVD-ANOV category (reference category) to cows in NPVD-ANOV [2.09 (1.62-2.50)], PVD-CYC [2.52 (2.02-3.14)], and NPVD-CYC [3.46 (2.84-4.23)]. Similarly, days from calving to pregnancy were less for NPVD-CYC, followed by PVD-CYC, NPVD-ANOV, and PVD-ANOV (121.4, 137.2, 137.3, and 157.4 d, respectively). On the contrary, no clear association was identified between groups and PL. The results indicate that both PVD and ANOV had a negative impact on PAI1 and days from calving to pregnancy. The results indicated a variable magnitude in the negative impact on the reproductive traits analyzed when both conditions were combined.

Differences in uterine and serum metabolome associated with metritis in dairy cows.

Objectives were to evaluate differences in the uterine and serum metabolomes associated with metritis in dairy cows. Vaginal discharge was evaluated using a Metricheck device (Simcro) at 5, 7, and 11 d in milk (DIM; herd 1) or 4, 6, 8, 10, and 12 DIM (herd 2). Cows with reddish or brownish, watery, and fetid discharge were diagnosed with metritis (n = 24). Cows with metritis were paired with herdmates without metritis (i.e., clear mucous vaginal discharge or clear lochia with ≤50% of pus) based on DIM and parity (n = 24). Day of metritis diagnosis was considered study d 0. All cows diagnosed with metritis received antimicrobial therapy. The metabolome of uterine lavage collected on d 0 and 5, and serum samples collected on d 0 were evaluated using untargeted gas chromatography time-of-flight mass spectrometry. Normalized data were subjected to multivariate canonical analysis of population using the MultBiplotR and MixOmics packages in R Studio. Univariate analyses including t-test, principal component analyses, partial least squares discriminant analyses, and pathway analyses were conducted using Metaboanalyst. The uterine metabolome differed between cows with and without metritis on d 0. Differences in the uterine metabolome associated with metritis on d 0 were related to the metabolism of butanoate, amino acids (i.e., glycine, serine, threonine, alanine, aspartate, and glutamate), glycolysis and gluconeogenesis, and the tricarboxylic acid cycle. No differences in the serum metabolome were observed between cows diagnosed with metritis and counterparts without metritis on d 0. Similarly, no differences in uterine metabolome were observed between cows with metritis and counterparts not diagnosed with metritis on d 5. These results indicate that the establishment of metritis in dairy cows is associated with local disturbances in amino acid, lipid, and carbohydrate metabolism in the uterus. The lack of differences in the uterine metabolome on d 5 indicates that processes implicated with the disease are reestablished by d 5 after diagnosis and treatment.

Flow cytometry panels for immunophenotyping dairy cattle peripheral blood leukocytes.

Many aspects of the bovine immune system remain poorly characterized, which poses an obstacle to improving dairy cow health. Herein, we describe two flow cytometry panels that included antibodies against CD8α, CD4, TCR-δ, CD172α, CD14, MHCII, CD21, CD62L, and CD11b. These panels were used to characterize the phenotype of leukocyte subpopulations from the peripheral blood of 30-day old Holstein calves and Holstein cows at 260 d of gestation and calving. No leukocyte subset differences were found between the pre- and post-partum cows. However, calf leukocytes presented a higher proportion of CD3+ lymphocytes, γδ T-cells, CD8+ γδ T-cells, and monocytes when compared with mature cows. Conversely, cow lymphocytes had a higher proportion of CD4+ and CD8+ T-cells, and B-cells than calf lymphocytes. The proportion of CD4+ T-cells and B-cells expressing CD62L was greater in calves than in cows, while cow B-cells expressed greater levels of CD11b than calf B-cells. In contrast, calf polymorphonuclear cells (PMN) and monocytes expressed greater levels of CD11b compared to cows. Moreover, calf monocytes expressed higher levels of MHCII compared with those of cows. Collectively, our data provides a resource to better understand the bovine immune system as well as immune-related diseases that affect dairy cattle.

Effect of body condition change and health status during early lactation on performance and survival of Holstein cows.

Body condition score (BCS) and disease records are commonly available in dairy operations. However, the effect of BCS changes (ΔBCS) considering specific health profiles has not been investigated extensively. The objective of this study was to assess the effects of different levels of ΔBCS on fertility, milk yield, and survival of Holstein cows diagnosed with reproductive disorders (REP; dystocia, twins, retained fetal membranes, metritis, and clinical endometritis), other health disorders (OTH; subclinical ketosis, left displaced abomasum, lameness, clinical mastitis, and respiratory disease), or with no disease events (HLT) within 40 days in milk (DIM). Data included lactation information from 11,733 cows calving between November 2012 and October 2014 in 16 herds across 4 geographical regions in the United States (Northeast, Midwest, Southwest, Southeast). Cows were evaluated for BCS at 5 ± 3 DIM (BCS5) and at 40 ± 3 DIM (BCS40) and the difference between BCS40 and BCS5 was classified as excessive loss of BCS (EL; ΔBCS ≤-0.75), moderate loss (ML; ΔBCS = -0.5 to -0.25), no change (NC; ΔBCS = 0), or gain of BCS (GN; ΔBCS ≥0.25). Multivariable logistic regression was used for assessing potential associations between the outcomes of interest and ΔBCS and health. The effect of the interaction term ΔBCS by health group was not statistically significant for any of the study outcomes. The odds of resumption of ovarian cyclicity (ROC), in GN, NC, and ML cows were 1.94 (95% CI: 1.57-2.40), 1.59 (1.28-1.97), and 1.27 (1.10-1.47) times greater than the odds of ROC in EL cows, respectively. The odds of pregnancy at 150 DIM (P150) in GN cows were 1.61 (1.20-2.17) times greater than the odds of P150 in EL cows. Cows with REP or OTH disorders had smaller odds of ROC compared with HLT cows [REP: OR = 0.65 (0.56-0.76) and OTH: OR = 0.79 (0.68-0.92)]. For pregnancy outcomes, REP cows had smaller odds of pregnancy at the first artificial insemination compared with HLT cows [0.70 (0.58-0.84)]. Similarly, REP cows had smaller odds of being diagnosed pregnant by 150 and 305 DIM compared with HLT cows [P150: 0.73 (0.59-0.87), P305: 0.58 (0.49-0.69)]. Overall, average daily milk within the first 90 DIM was greater in EL (39.5 ± 1.13 kg/d) and ML (38.9 ± 1.11 kg/d) cows than in NC (37.8 ± 1.12 kg/d) and GN (36.2 ± 1.12 kg/d) cows. On the other hand, average daily milk within the first 90 DIM was lower in REP (37.0 ± 1.11 kg/d) cows compared with OTH (38.7 ± 1.12 kg/d) and HLT cows (38.6 ± 1.11 kg/d). The magnitude of ΔBCS and the health status of early lactation cows should be considered when assessing subsequent cow performance and survival.

Integration of statistical inferences and machine learning algorithms for prediction of metritis cure in dairy cows.

The study's objectives were to identify cow-level and environmental factors associated with metritis cure to predict metritis cure using traditional statistics and machine learning algorithms. The data set used was from a previous study comparing the efficacy of different therapies and self-cure for metritis. Metritis was defined as fetid, watery, reddish-brownish discharge, with or without fever. Cure was defined as an absence of metritis signs 12 d after diagnosis. Cows were randomly allocated to receive a subcutaneous injection of 6.6 mg/kg of ceftiofur crystalline-free acid (Excede, Zoetis) at the day of diagnosis and 3 d later (n = 275); and no treatment at the time of metritis diagnosis (n = 275). The variables days in milk (DIM) at metritis diagnosis, treatment, season of the metritis diagnosis, month of metritis diagnostic, number of lactation, parity, calving score, dystocia, retained fetal membranes, body condition score at d 5 postpartum, vulvovaginal laceration score, the rectal temperature at the metritis diagnosis, fever at diagnosis, milk production from the day before to metritis diagnosis, and milk production slope up to 5, 7, and 9 DIM were offered to univariate logistic regression. Variables included in the multivariable logistic regression model were selected from the univariate analysis according to P-value. Variables were offered to the model to assess the association between these factors and metritis cure. Additionally, the univariate logistic regression variables were offered to a recursive feature elimination to find the optimal subset of features for a machine learning algorithms analysis. Cows without vulvovaginal laceration had 1.91 higher odds of curing of metritis than cows with vulvovaginal laceration. Cows that developed metritis at >7 DIM had 2.09 higher odds of being cured than cows that developed metritis at ≤7 DIM. For rectal temperature, each degree Celsius above 39.4°C led to lower odds to be cured than cows with rectal temperature ≤39.4°C. Furthermore, milk production slope and milk production difference from the day before to the metritis diagnosis were essential variables to predict metritis cure. Cows that had reduced milk production from the day before to the metritis diagnosis had lower odds to be cured than cows with moderate milk production increase. The results from the multivariable logistic regression and receiver operating characteristic analysis indicated that cows developing metritis at >7 DIM, with increase in milk production, and with a rectal temperature ≤39.40°C had increased likelihood of cure of metritis with an accuracy of 75%. The machine learning analysis showed that in addition to these variables, calving-related disorders, season, and month of metritis event were needed to predict whether the cow will cure or not from metritis with an accuracy ≥70% and F1 score (harmonic mean between precision and recall) ≥0.78. Although machine learning algorithms are acknowledged as powerful tools for predictive classification, the current study was unable to replicate its potential benefits. More research is needed to optimize predictive models of metritis cure.

Associations between dry period length and time to culling and pregnancy in the subsequent lactation.

The association between dry period length (DPL) and time to culling and pregnancy in the subsequent lactation may be important for the economically optimal length of the dry period. Therefore, this study aimed to (1) quantify the association between DPL and hazard of culling and pregnancy in the subsequent lactation; (2) develop continuous functions of DPL for the hazard ratios of culling and pregnancy; and (3) investigate the effect of a cause-specific hazards model and a subdistribution model to analyze competing events. The data used in this observational cohort study were from dairy herd improvement milk test lactation records from 40 states in the United States. After edits, there remained 1,108,515 records from 6,730 herds with the last days dry in 2014 or 2015. The records from 2 adjacent lactations (current, subsequent) were concatenated with the DPL of interest, 21 to 100 d, in between both lactations. We defined 8 DPL categories of 10 d each. Kaplan-Meier survival curves were used to show associations between DPL and time to culling or pregnancy for 3 lactation groups: lactation 1 and 2, lactation 2 and 3, and lactation 3 and greater. To control for confounding factors in Cox proportional models, we included 6 current lactation covariates and 3 time-dependent variables in the survival models. Hazard ratios of culling were estimated for 4 days in milk (DIM) categories from 1 to 450 DIM. Hazard ratios of pregnancy were estimated for 3 DIM categories from 61 to 300 DIM. Competing risk analysis of 8 disposal codes (i.e., farmer reported reasons) for culling and the culling event for pregnancy were conducted by a cause-specific hazards model and a subdistribution model. Hazard ratios were also estimated as quadratic polynomials of DPL. Compared with the reference DPL category of 51 to 60 d, hazard ratios of culling and pregnancy of the other 7 DPL categories ranged between 0.70 and 1.49, and 0.93 and 1.15, respectively. Short DPL were associated with lower risk of culling in the early lactation but not over the entire lactation. Short DPL were associated with greater hazard of pregnancy. Trends in hazard ratios over the ranges of the 8 DPL categories were not always consistent. Competing risk analysis with both models provided little differences in hazard ratios of culling and pregnancy. In conclusion, variations in DPL were associated with meaningful differences in the hazard ratios for culling and pregnancy and minor differences in the relative frequency of disposal codes. Subdistribution hazards models produced hazard ratios similar to cause-specific hazard models. The quadratic polynomials may be useful for decision support on customization of DPL for individual cows.

Economic comparison between ceftiofur-treated and nontreated dairy cows with metritis.

The objective was to investigate the economic effect of treating dairy cows with metritis using ceftiofur-free acid or leaving them untreated at the time of diagnosis. Cows with a fetid, watery, red-brownish vaginal discharge were diagnosed with metritis (d 0). Data from 875 dairy cows (506 primiparous and 369 multiparous) from 1 herd in northern Florida that had been part of a larger study evaluating different treatments for metritis were used for the economic analysis. Holstein cows with metritis had been randomly assigned to: Ceftiofur (CEF, n = 239) = subcutaneous injection of 6.6 mg/kg of ceftiofur crystalline-free acid in the base of the ear at d 0 and d 3; Untreated (UNT, n = 233) = no treatment applied at metritis diagnosis. Both groups could receive escape therapy if condition worsened. A group of nonmetritic healthy cows (NMET; n = 403) from the same cohort was randomly selected for comparison. Continuous outcomes such as 300-d milk production (kg/cow), milk sales ($/cow), cow sales ($/cow), treatment cost by 60 days in milk ($/cow), reproduction cost ($/cow), replacement cost ($/cow), feeding cost ($/cow), and gross profit per cow ($/cow) were analyzed using the ANOVA (MIXED procedure of SAS version 9.4). Dichotomous outcomes such as pregnancy and culling by 300 d were analyzed using logistic regression (GLIMMIX procedure of SAS). Models included the fixed effects of treatment, parity, and the interaction between treatment and parity. A stochastic analysis was performed with 10,000 iterations using the observed results from each group. The CEF treatment resulted in greater treatment cost by 60 DIM than UNT ($112 vs. $37), but resulted in a greater proportion of pregnant cows (71 vs. 61%) and decreased culling by 300 DIM (29 vs. 39%) compared with UNT. Gross profit was lesser for UNT than NMET ($2,969 vs. $3,426), and CEF was intermediate ($3,219). The stochastic analysis showed that the mean difference in gross profit between UNT and NMET was -$457; saleable milk (49%) and replacement cost (24%) accounted for most of the variation. The mean difference in gross profit between CEF and NMET group was -$207; saleable milk (82%) and initial metritis treatment cost (9%) accounted for most of the variation. The mean difference in gross profit between the UNT and the CEF group was -$250; replacement cost (41%) and cow sales (31%) accounted for most of the variation. In summary, metritis caused large economic losses when left untreated, and CEF reduced those losses by improving fertility, reducing culling and replacement cost, and reducing milk yield losses.

Failure of clinical cure in dairy cows treated for metritis is associated with reduced productive and reproductive performance.

Objectives were to assess reproductive and productive outcomes associated with failure of clinical cure in dairy cows diagnosed with metritis following antimicrobial therapy. This retrospective cohort study included data from 3 experiments performed in 5 dairies. Metritis was characterized by the presence of watery, fetid, reddish-brownish vaginal discharge within 21 DIM (study d 0). Cows not diagnosed with metritis (i.e., cows may have had other diseases postpartum; NoMT; n = 1,194) were paired based on lactation number and calving date. All cows with metritis received antimicrobial therapy (ampicillin or ceftiofur). Clinical cure was evaluated on d 10 based on vaginal discharge score, and cows were categorized as cured (MTC; n = 1,111) or not cured (MTnoC; n = 299). Purulent vaginal discharge (28 ± 3 or 32 ± 3 DIM), cytological endometritis (35 ± 3 or 39 ± 3 DIM), and estrous cyclicity (50 ± 3 and 64 ± 3, 36 ± 3 and 50 ± 3, or 37 ± 5 and 51 ± 5 DIM) were evaluated in subgroups of cows. Proportions of cows with purulent vaginal discharge and cytological endometritis were greatest for MTnoC (91.7 and 91.4%), intermediate for MTC (74.0 and 73.3%), and smallest for NoMT (38.1 and 36.4%). Proportion of cyclic cows was smaller for MTnoC compared with MTC and NoMT (62.0, 71.0, and 71.0%). Pregnancy per artificial insemination following first service was smaller for cows with metritis compared with their counterparts with no metritis (NoMT = 28.1, MTC = 26.1, MTnoC = 22.0%). Pregnancy loss tended to be greater for MTnoC compared with MTC (NoMT = 11.5, MTC = 11.1, MTnoC = 18.4%). Hazard of pregnancy by 300 DIM was smallest for MTnoC, intermediate for MTC, and greatest for NoMT. Death by 60 DIM (3.9, 1.1, and 0.6%) and removal from herd by 300 DIM (26.3, 17.4, and 15.4%) were greatest for MTnoC compared with MTC and NoMT, respectively. Milk production among multiparous cows was smaller for MTnoC compared with MTC and NoMT in the first 10 mo postpartum, whereas MTC produced less milk compared with NoMT only during the first 2 mo postpartum (NoMT = 42.0 ± 0.22, MTC = 40.6 ± 0.28, MTnoC = 37.7 ± 0.54 kg/d). Failure of clinical cure was not associated with milk yield in primiparous cows (NoMT = 35.2 ± 0.31, MTC = 33.9 ± 0.31, MTnoC = 35.0 ± 0.52 kg/d). Cows diagnosed with metritis that do not undergo clinical cure by 10 d of onset of antimicrobial therapy have impaired reproductive performance, reduced milk production, and increased risk of leaving the herd.

The economic cost of metritis in dairy herds.

The objective of this study was to estimate the cost of metritis in dairy herds. Data from 11,733 dairy cows from 16 different farms located in 4 different regions of the United States were compiled for up to 305 d in milk, and 11,581 cows (2,907 with and 8,674 without metritis) were used for this study. Metritis was defined as fetid, watery, red-brownish vaginal discharge that occurs ≤21 d in milk. Continuous outcomes such as 305-d milk production, milk sales ($/cow), cow sales ($/cow), metritis treatment costs ($/cow), replacement costs ($/cow), reproduction costs ($/cow), feeding costs ($/cow), and gross profit per cow ($/cow) were analyzed using mixed effect models using the MIXED procedure of SAS (SAS Institute Inc., Cary, NC). Gross profit was also compared using the Kruskal-Wallis test. Dichotomous outcomes such as pregnant and culling by 305 d in milk were analyzed using the GLIMMIX procedure of SAS. Time to pregnancy and culling were analyzed using the PHREG procedure of SAS. Models included the fixed effects of metritis, parity, and the interaction between metritis and parity, and farm as the random effect. Variables were considered significant when P ≤ 0.05. Metritis cost was calculated by subtracting the gross profit of cows with metritis from the gross profit of cows without metritis. A stochastic analysis was performed with 10,000 iterations using the observed results from each group. Milk yield and proportion of cows pregnant were lesser for cows with metritis than for cows without metritis, whereas the proportion of cows leaving the herd was greater for cows with metritis than for cows without metritis. Milk sales, feeding costs, residual cow value, and gross profit were lesser for cows with metritis than for cows without metritis. Cow sales and replacement costs were greater for cows with metritis than for cows without metritis. The mean cost of metritis from the study herds was $511 and the median was $398. The stochastic analysis showed that the mean cost of a case of metritis was $513, with 95% of the scenarios ranging from $240 to $884, and that milk price, treatment cost, replacement cost, and feed cost explained 59%, 19%, 12%, and 7%, respectively, of the total variation in cash flow differences. In conclusion, metritis caused large economic losses to dairy herds by decreasing milk production, reproduction, and survival in the herd.

Early-lactation diseases and fertility in 2 seasons of calving across US dairy herds.

The objective of this study was to characterize incidences of health disorders during early lactation in a large population of Holstein cows calving in 2 seasons across multiple US dairy herds. In addition, cumulative effects of combinations of health-related events on fertility and survival by season of calving and parity number were tested. Data were prospectively collected from a total of 11,729 cows in 16 herds located in 2 regions in the United States [north (7,820 cows in 10 herds) and south (3,909 cows in 6 herds)]. Cows were enrolled at parturition and monitored weekly for disease occurrence, reproductive events, and survival. Health-related events were grouped into reproductive disorders (REP; dystocia, twins, retained fetal membranes, metritis, and clinical endometritis) and other disorders (OTH; subclinical ketosis, mastitis, displaced abomasum, and pneumonia). Counts of health events within 50 d postpartum were added into each of the groups and categorized as 0, 1, 2, 3, and ≥4 for REP and 0, 1, 2, and ≥3 for OTH. Multivariable logistic regression was used for testing potential associations between categories of disease occurrence and outcome variables, including resumption of ovarian cyclicity, pregnancy per artificial insemination (AI), pregnancy loss, and survival up to and after 50 DIM. The incidence of disease varied with season of calving and parity, and these 2 variables were associated with the reproductive and survival outcomes. The size of the detrimental effect of disease incidence on reproduction and survival depended on disease group and varied for each specific outcome. Resumption of ovarian cyclicity decreased as incidences of disorders increased in both REP and OTH categories. Pregnancy at first AI also was smaller in greater number of REP categories, but the effect of number of OTH categories on pregnancy at first AI was not consistent. Similarly, pregnancy loss at first AI was not affected consistently by REP or OTH. Survival was reduced by REP and OTH. The magnitude of these negative effects was variable, depending on season of calving and parity, but consistently increased with the number of health events during early lactation.

The association of cow-related factors assessed at metritis diagnosis with metritis cure risk, reproductive performance, milk yield, and culling for untreated and ceftiofur-treated dairy cows.

Our objective was to assess the association of cow-related factors with metritis cure risk and economically important outcomes. In this prospective cohort study nested inside a randomized clinical trial, cows enrolled in a clinical trial that aimed to evaluate an alternative metritis therapy that had available plasma samples collected at metritis diagnosis were included. Metritis was defined as fetid, watery, reddish-brownish discharge with or without fever, and cure was defined as the absence of metritis signs 12 d after diagnosis. Cows were randomly allocated to remain untreated (CON; n = 147) or receive subcutaneous injections of 6.6 mg/kg of ceftiofur crystalline-free acid at enrollment and 72 h later (CEF, n = 168). Additionally, a random subset of 150 nonmetritic cows (NMET) was also included to compare milk production, reproductive performance, and culling responses. Cow-related factors evaluated include plasma concentrations of nonesterified fatty acids, ß-hydroxybutyrate, and haptoglobin (Hp), parity, rectal temperature, and days in milk (DIM) at metritis diagnosis, vulvovaginal laceration (VL), BCS, dystocia, twins, and retained placenta. Among CON cows, DIM at metritis diagnosis was positively associated with metritis cure [threshold = 8, area under the curve (AUC) = 0.67], whereas plasma Hp concentration tended to be negatively associated with cure of metritis (threshold = 0.54 mg/mL, AUC = 0.64). Among CEF cows, DIM at metritis diagnosis (threshold = 5, AUC = 0.67) and dystocia were positively associated with metritis cure, whereas VL and Hp (threshold = 0.78 mg/mL, AUC = 0.76) were negatively associated with cure. For CON cows that were diagnosed with metritis after 8 DIM or had plasma Hp concentration ≤0.54 mg/mL, milk production, pregnancy, and culling risk were comparable to NMET cows. However, performance was impaired when cows that developed metritis at ≤8 DIM or had Hp >0.54 mg/mL were left untreated. Among CEF cows, Hp, DIM at metritis diagnosis, dystocia, and VL were associated with metritis cure. Milk yield, reproductive performance, and culling losses are more pronounced among CEF cows when metritis was diagnosed at ≤5 DIM, Hp >0.78 mg/mL, or if they had VL or dystocia. In conclusion, these data indicate that timing of the onset of metritis and inflammatory biomarkers could be used for the development of a selective therapy strategy for metritis, but more research is needed to identify more accurate predictors of metritis spontaneous cure and treatment failure.

Using chitosan microparticles to treat metritis in lactating dairy cows.

The main objective of this study was to evaluate the efficacy of intrauterine administration of chitosan microparticles (CM) in curing metritis in dairy cows. A secondary objective was to evaluate the effects of metritis treatments on milk yield, survival, and reproductive performance. Cows with a fetid, watery, red-brownish vaginal discharge were diagnosed with metritis. Holstein cows (n = 826) with metritis from 3 dairies located in northern Florida were blocked by parity (primiparous or multiparous) and, within each block, randomly assigned to one of 3 treatments: CM (n = 276) = intrauterine infusion of 24 g of CM dissolved in 40 mL of sterile distilled water at the time of metritis diagnosis (d 0), 2 (d 2), and 4 (d 4) d later; ceftiofur (CEF; n = 275) = subcutaneous injection of 6.6 mg/kg ceftiofur crystalline-free acid in the base of the ear at d 0 and d 3; Control (CON; n = 275) = no treatment applied at metritis diagnosis. All groups could receive escape therapy if condition worsened. Cure was considered when vaginal discharge became mucoid and not fetid. A group of nonmetritic (NMET; n = 2,436) cows was used for comparison. Data were analyzed by generalized linear mixed and Cox's proportional hazard models. Cows in CM and CON had lesser risk of metritis cure on d 12 than cows in CEF (58.6 ± 5.0 vs. 61.9 ± 4.9% vs. 77.9 ± 3.9, respectively). The proportion of cows culled within 60 days in milk (DIM) was greater for cows in CM than for cows in CEF and CON (21.5 ± 2.7 vs. 9.7 ± 1.9 vs. 11.3 ± 2.0%, respectively). Treatment did not affect rectal temperature or plasma nonesterified fatty acids, ß-hydroxybutyrate, and haptoglobin concentrations. Milk yield in the first 60 DIM differed for all treatments, and it was lowest for CM (35.8 ± 0.3 kg/d), followed by CON (36.8 ± 0.3 kg/d) and CEF (37.9 ± 0.3 kg/d). The hazard of pregnancy up to 300 DIM was lesser for CM than CEF (hazard ratio = 0.62; 95% CI: 0.50-0.76), for CM than CON (hazard ratio = 0.77; 95% CI: 0.62-0.95) and for CON than CEF (hazard ratio = 0.80; 95% CI: 0.65-0.99). Culling was greater, and milk yield and fertility were lesser for CEF than NMET. In summary, CM did not improve the cure of metritis, and was detrimental to milk yield, survival, and fertility compared with CON. In contrast, CEF increased the cure of metritis, milk yield, and fertility compared with CM and CON. Finally, the negative effects of metritis on milk yield culling and fertility could not be completely reversed by CEF.

Associations of reproductive indices with fertility outcomes, milk yield, and survival in Holstein cows.

The study is part of a research effort investigating potential associations between genomic variation and fertility of Holstein cows. The objective was to compare the reproductive performance of Holstein cows in 3 categories of 2 reproductive indices (RI) that were developed for the allocation of cows in a ranking for potential fertility, based on the predicted probability of pregnancy. The associations between categories of the developed indices and multiple fertility variables in a large multistate population of Holstein cows were tested. In addition, we analyzed associations among the RI categories with milk yield and survival. Based on phenotypic information from individual cows, 2 reproductive indices (RI1 and RI2) were developed, representing a predicted probability that a cow will become pregnant at first artificial insemination postpartum, as a function of explanatory variables used in a logistic model. Data from a total of 11,733 cows calving in 16 farms located in 4 regions of the United States (Northeast, Midwest, Southeast, and Southwest) were available. Cows were enrolled at parturition and monitored weekly for reproductive events, health status, milk yield, and survival. To develop the indices, potential significant effects were initially tested by univariate analyses. Effects with P ≤ 0.05 were offered to the multivariate analysis, and the final models were determined through backward elimination, considering potentially significant interactions. The final model for RI1 included the random effect of farm and a complement of significant fixed effects as explanatory variables influencing a pregnancy outcome: (1) incidence of retained fetal membranes; (2) metritis; (3) clinical endometritis; (4) lameness at 35 days in milk (DIM); (5) resumption of postpartum ovulation by 50 DIM; (6) season of calving; and (7) parity number. The model for RI2 included (1) parity number; (2) body condition score at 40 DIM; (3) incidence of retained fetal membranes; (4) metritis; (5) resumption of postpartum ovulation by 50 DIM; (6) region; (7) subclinical ketosis; (8) mastitis; (9) clinical endometritis; and (10) milk yield at the first milk test after calving; as well as the interaction effects of postpartum resumption of ovulation by 50 DIM × region; mastitis × region; and milk yield at the first milk test after calving × parity number. Multivariate logistic regression, ANOVA, and survival analysis were used to test the correspondence between the resulting RI and individual fertility, milk yield, and survival from the population. To facilitate the analyses, the resulting RI values were categorized as low for cows in the lowest quartile, medium for cows within the interquartile range, or high for cows in the top quartile. We found consistent agreement between categories of the predicted RI and the measures of fertility and survival collected from individual cows. We conclude that the proposed RI represent a viable approach to refine the allocation of cows into potential low- and high-fertility populations.

Genome-enable prediction for health traits using high-density SNP panel in US Holstein cattle.

The objective of this study was to compare accuracies of different Bayesian regression models in predicting molecular breeding values for health traits in Holstein cattle. The dataset was composed of 2505 records reporting the occurrence of retained fetal membranes (RFM), metritis (MET), mastitis (MAST), displaced abomasum (DA), lameness (LS), clinical endometritis (CE), respiratory disease (RD), dystocia (DYST) and subclinical ketosis (SCK) in Holstein cows, collected between 2012 and 2014 in 16 dairies located across the US. Cows were genotyped with the Illumina BovineHD (HD, 777K). The quality controls for SNP genotypes were HWE P-value of at least 1 × 10-10 ; MAF greater than 0.01 and call rate greater than 0.95. The FImpute program was used for imputation of missing SNP markers. The effect of each SNP was estimated using the Bayesian Ridge Regression (BRR), Bayes A, Bayes B and Bayes Cπ methods. The prediction quality was assessed by the area under the curve, the prediction mean square error and the correlation between genomic breeding value and the observed phenotype, using a leave-one-out cross-validation technique that avoids iterative cross-validation. The highest accuracies of predictions achieved were: RFM [Bayes B (0.34)], MET [BRR (0.36)], MAST [Bayes B (0.55), DA [Bayes Cπ (0.26)], LS [Bayes A (0.12)], CE [Bayes A (0.32)], RD [Bayes Cπ (0.23)], DYST [Bayes A (0.35)] and SCK [Bayes Cπ (0.38)] models. Except for DA, LS and RD, the predictive abilities were similar between the methods. A strong relationship between the predictive ability and the heritability of the trait was observed, where traits with higher heritability achieved higher accuracy and lower bias when compared with those with low heritability. Overall, it has been shown that a high-density SNP panel can be used successfully to predict genomic breeding values of health traits in Holstein cattle and that the model of choice will depend mostly on the genetic architecture of the trait.