The objective of this study was to evaluate the effect of administering 200 µg versus 100 µg of GnRH as gonadorelin hydrochloride (Factrel; Zoetis, Parsippany, NJ) at the first GnRH injection (G1) of the breeding Ovsynch in a Double-Ovsynch (DO) protocol on pregnancy per artificial insemination (P/AI) at first service in lactating dairy cows. The study was conducted across 4 commercial dairies located in California (n = 2) and Wisconsin (n = 2). We also assessed the effect of GnRH dose on ovulatory response and LH release. Each week, cows were blocked in a 2 × 2 factorial design by parity (primiparous or multiparous) and GnRH dose (100 µg or 200 µg) within each herd. The primary outcome was P/AI following estrus synchronization and timed AI. Cows were excluded if they were inseminated outside the synchronization window, culled before pregnancy diagnosis, or missed scheduled GnRH or PGF2α treatments. Dinoprost tromethamine (12.5 mg/mL; Lutalyse HighCon, Zoetis) served as the PGF2α source. A total of 6,621 cows met the inclusion criteria (2,525 primiparous; 4,096 multiparous). Of these, 3,310 received 100 µg GnRH (2 mL), and 3,311 received 200 µg GnRH (4 mL). A subset of 505 cows from herd A underwent transrectal ultrasonography to assess ovulatory response at G1 and 7 d later (first PGF2α of DO), with concurrent serum progesterone (P4) sampling. An additional subset of 18 cows had blood samples collected at 0 (pretreatment) and at 30, 60, 90, 120, 150, 180, 210, 240, 300, and 360 min post-treatment to measure LH concentrations. The final statistical model for P/AI included fixed effects of treatment, parity, and their interaction, with random effects of herd, service sire, herd-by-treatment-by-parity, and month fresh nested within herd-by-parity. Treatment and parity significantly affected P/AI, although their interaction was not significant. Least squares means for P/AI were higher with 200 µg than 100 µg GnRH (51.8% vs. 48.6%). When stratified, the effect in primiparous cows was minimal (52.8% vs. 51.8%), but substantial in multiparous cows (50.7% vs. 45.4%). Cows treated with 200 µg GnRH had a greater ovulatory response than those receiving 100 µg (83.0% vs. 70.8%). Parity also influenced ovulation, though no treatment-by-parity interaction was observed. When P4 was categorized (Low <3.0 ng/mL, High ≥3.0 ng/mL), a 3-way interaction emerged: in primiparous cows with Low-P4, ovulation was greater with 200 µg; in multiparous cows with High-P4, ovulation was greater with 200 µg. No treatment effect was observed on P4 concentrations 7 d post-treatment. Cows receiving 200 µg GnRH had higher peak LH and area under the curve than those receiving 100 µg, though peak timing did not differ. In summary, increasing the GnRH dose to 200 µg at G1 of the breeding Ovsynch in a DO program improved LH release, ovulatory response, and first-service P/AI, with the greatest benefit in multiparous cows.

Heat stress profoundly compromises reproductive efficiency in dairy cattle by disrupting follicular and oocyte development, altering steroid hormone profiles, impairing estrous expression and ovulation, reducing fertilization success, hindering embryonic development, and increasing early embryonic loss. Although the molecular mechanisms underlying these disruptions remain incompletely understood, significant advances in reproductive technologies have been developed to mitigate the negative impact of heat stress on reproductive performance. Estrous cycle synchronization programs, enabling timed artificial insemination (AI), emerged in the mid-1990s and reduced heat stress-related reproductive inefficiencies by eliminating the need for estrus detection, improving submission rates, and optimizing the ovulation-to-insemination interval, thereby enhancing pregnancy rates per AI. Embryo transfer (ET) has been adopted as a complementary strategy to bypass fertilization failure during hot periods, whereas genetic selection, most notably the introgression of the SLICK-1 allele of the prolactin receptor gene (PRLR), confers thermotolerance via a sleek hair coat and improved heat dissipation. Nevertheless, heat stress also exacerbates common postpartum disorders such as metritis, which negatively affect fertility regardless of AI, ET, or thermotolerant genotypes, underscoring the importance of preventive herd-health management. Furthermore, exposure to heat stress during fetal development imposes transgenerational consequences, diminishing the health, productivity, longevity, and fertility of subsequent generations. This review evaluates current and emerging technologies, including timed AI, ET, genetic selection for thermotolerance and reproductive resilience, and preventive approaches to postpartum disease, while also addressing strategies to mitigate the transgenerational impairments induced by heat stress in dairy cattle.

The goal of this study was to calculate the cost of purulent vaginal discharge (PVD) in dairy cows. The dataset included 11,051 cows from 16 dairy herds located in 4 regions of the United States. Purulent vaginal discharge was characterized as a mucopurulent, purulent, or reddish-brownish vaginal discharge collected at 28 f 7 DIM. Gross profit was calculated as the difference between incomes and expenses, and the cost of PVD was calculated by subtracting the gross profit of cows with PVD from the gross profit of cows without PVD. Continuous outcomes such as milk production (kg/cow), milk sales ($/cow), cow sales ($/cow), feed costs ($/cow), reproductive management costs ($/cow), replacement costs ($/ cow), and gross profit ($/cow) were analyzed using linear mixed effects models. Pregnancy and culling by 305 DIM were analyzed by generalized linear mixed effects models using logistic regression. Models included the fixed effects of PVD, metritis, parity, region, season of calving, and morbidity in the first 60 DIM, as well as the interactions between PVD and metritis, PVD and parity group, and PVD and morbidity. Farm and the interaction between PVD and farm were considered random effects in all the statistical models. A stochastic analysis was conducted using 10,000 iterations with varying relevant inputs. Cows with PVD produced less milk (9,753.2 f 333.6 vs. 9,994.6 f 330.9 kg/cow), were less likely to be pregnant (70.7 f 1.7% vs. 78.9 f 1.2%), and were more likely to be culled by 305 DIM (34.6 f 1.7% vs. 27.2 f 1.3%) compared with cows without PVD. Consequently, milk sales (4,744.7 f 162.3 vs. 4,862.1 f 161.0 $/cow) and residual cow value (1,079.6 f 23.0 vs. 1,179.3 f 20.3 $/cow) were lesser for cows with PVD. Replacement (639.4 f 26.4 vs. 526.0 f 23.4 $/cow) and reproductive management costs (76.3 f 2.5 vs. 69.0 f 2.4 $/cow) were greater for cows with PVD. The mean cost of PVD was $202. The stochastic analysis also showed a mean cost of $202, ranging from $152 to $265. The robust dataset and the stochastic analysis strengthen both the external and internal validity of our findings, offering a deeper understanding of the economic consequences of PVD. In conclusion, PVD resulted in large economic losses to dairy herds by being associated with decreased milk yield, impaired reproduction, and greater culling.

The objective of this study was to evaluate the differences in milk production, reproductive performance, and culling associated with antimicrobial therapy and clinical cure of metritis in dairy cows. In this study, data from 2 randomized controlled trials consisting of 900 Holstein cows from 5 dairy farms in California, Florida, and Texas were used. Cows were examined for metritis (fetid, watery, reddish-brownish vaginal discharge; VD) at 4, 7, and 10 DIM (study d 0). Cows diagnosed with metritis (n = 900) were randomly assigned (regardless of fever) to receive ceftiofur (CEF = 457) or to remain untreated (NT = 443). Clinical cure of metritis was assessed at study d 13 ± 1 based on the visual evaluation of the VD; cows that underwent clinical cure displayed clear, mucopurulent, or purulent discharge (cured = 710), whereas cows with clinical cure failure had persistent fetid, watery, reddish-brownish VD (not cured = 190). The data were analyzed considering antimicrobial treatment, cure status on d 13 ± 1, and their interaction. Risk of pregnancy and culling at 300 DIM were analyzed using logistic regression, time to pregnancy and culling by 300 DIM were analyzed using Cox's proportional hazard, and milk production within 300 DIM was analyzed using ANOVA with repeated measures. Antimicrobial treatment and cure status were included as fixed effects and farm as a random effect in all models. Both randomized trials that generated data used in this study reported a greater risk of clinical cure associated with CEF treatment. Differences in risk of pregnancy at 300 DIM were associated with cure status (cured = 70.9% vs. not cured = 61.7%), but not with antimicrobial treatment or antimicrobial treatment by cure interaction. The risk of culling at 300 DIM was not associated with cure status, antimicrobial treatment, or antimicrobial treatment by cure interaction. Reduced median time to pregnancy was associated with cure status (cured = 134 vs. not cured = 154 d) but not with antimicrobial treatment or antimicrobial treatment by cure interaction. No interaction of antimicrobial treatment by cure status was observed on milk production; however, interactions between antimicrobial treatment, parity, and month of lactation and cure, parity, and month of lactation were observed. Differences in milk production between multiparous CEF and NT cows were observed in the third month of lactation. Conversely, differences in milk production associated with cure status were observed for primiparous and multiparous cows in the first and second months of lactation, whereas only multiparous cows in the eighth, ninth, and tenth months of lactation presented differences in milk production. Clinical cure of metritis was positively associated with milk production and reproduction, regardless of antimicrobial therapy, warranting further investigation regarding selective therapy of metritis.

Genomic prediction of daughter pregnancy rate (GDPR) and cow conception rate (GCCR) are fertility traits developed to help improve selection for reproductive performance. Although these traits overlap, their denominator can vary, and programs with different strategies combining estrus detection (ED) and timed AI might experience different associations with these traits (e.g., shorter interval for first service for high GDPR quartiles, but not for GCCR). The objectives were to assess days from calving to first service (TP1), AI at ED (AIE), pregnancy at the first service (P1), pregnancy loss for the first service (PL), and number of services to conception (NSFC) in 2 reproductive programs (RepP) combining variable ED length and timed AI (TAI) and their relationship with GDPR and GCCR. Holstein cows from a single farm were randomly allocated to 2 different programs, the short RepP (n = 982) or long RepP (n = 942). In the short RepP, cows were enrolled in a Presynch-Ovsynch (PGF2 alpha: 36 +/- 3 and 50 +/- 3 DIM) followed by ED and AI from 50 +/- 3 to 62 +/- 3 DIM. Cows not detected in estrus underwent in the Ovsynch (GnRH: 62 +/- 3, PGF2 alpha: 69 +/- 3, GnRH: 71 +/- 3 and TAI :72 +/- 3 DIM). In the long RepP, cows received a PGF2 alpha at 50 +/- 3 DIM followed by AI at ED up to 81 +/- 3 DIM. Cows with no ED were enrolled in an Ovsynch (GnRH:82 +/- 3, PGF2 alpha:89 +/- 3, GnRH:91 +/- 3 and TAI:92 +/- 3 DIM). Cows were categorized into quartiles (Q1 to Q4) considering their GDPR (qGDPR) and GCCR (qGCCR). Statistical analyses included logistic regression used for AIE, P1, and PL; Poisson regression for the NSFC; and linear regression for TP1. Models included AIE, P1, PL, NSFC, and TP1 as dependent variables, with RepP and qGDPR for GDPR and RepP and qGCCR for GCCR as independent variables. Time to pregnancy was analyzed using Cox's proportional hazard model. The short RepP had a shorter TP1 (short = 64.3 vs. long = 72.1) and fewer NSFC than the long RepP (short = 2.9 vs. long = 3.1). The long RepP had a higher AIE (short = 45.2% vs. long = 73.2%). The P1 tended to be greater in the short than the long RepP (short = 33.7% vs. long = 30.0%). Cows in Q4 (GDPR and GCCR) had lower TP1 and NSFC, greater AIE and P1 compared with the lowest quartiles (Q1). Interactions between RepP and GDPR were present for TP1, AIE, and P1, but no interactions were observed between RepP and GCCR. The short program had a shorter interval from calving to pregnancy than the long program. Cows in the highest quartiles for GDPR and GCCR had shorter intervals from calving to pregnancy and higher pregnancy hazards than the lowest quartiles. The current study revealed that cows enrolled in long RepP that relied on longer ED intervals had lower reproductive outcomes than short RepP cows, and cows ranked in the highest GDPR and GCCR quartile for most responses had better outcomes independent of the RepP used.

Effective isolation of bacterial pathogens for bovine respiratory disease (BRD) is a critical step for accurate diagnosis of the agent associated with this disease on the dairy. Limited information is available on factors associated with herd-level bacterial pathogen recovery for BRD clinical cases, which are important data to help identify strategies to support the successful collection of a minimum number of each organism over time to generate cumulative antibiogram susceptibility testing reports. Our objective was to evaluate factors associated with the recovery of common pathobionts (Pasteurella multocida and Mannheimia haemolytica) in BRD clinical cases from preweaning calves, heifers, and cows at 3 California dairy farms over 2 yr. A second objective was to test the predictability of isolating these organisms in BRD clinical cases using the factors evaluated in the first objective utilizing machine learning (ML). During monthly herd visits, deep nasopharyngeal samples were collected from calves, heifers, and cows diagnosed with BRD over 2 yr. Samples were cultured in aerobic conditions, and pathogens were isolated through colony morphology and validated with MALDI-TOF MS. Evaluation of factors influencing bacterial recovery was initially tested for independence, followed by a logistic regression model and a stepwise logistic feature selection in SAS, and ML classifiers with leave-one-out cross-validation in Python packages. For our study, samples were collected from a total of 301 BRD clinical cases: 152 samples with a culture-positive for P. multocida, 63 samples with a culture-positive for M. haemolytica, and 3 samples with a culture-positive for Histophilus somni. For factors associated with the culture-positive of P. multocida in BRD clinical cases, an interaction between age and season was identified, where cows had overall lower odds of being culture-positive independently of the season when compared with calves in the spring and summer and heifers in the fall and winter. For factors associated with the culture-positive of M. haemolytica in BRD clinical cases, an interaction was also observed between age and season, but the farm further played a role in the odds of being culture-positive, with one farm having considerably greater odds than the remaining ones. Machine learning models with cross-validation showed a weak ability to distinguish positive from negative cases when using age, season, and farm for all scenarios according to F1-scores and receiver operating characteristic analysis. Differences in predictive abilities, factor importance, and the still limited number of predictors in these ML analyses further indicate a potential for building more robust future models once datasets are expanded and more robust algorithms are considered. Overall, correctly identifying factors that may be associated with the prevalence of BRD pathogens and, therefore, recovery of these pathogens is critical for the development of antibiogram programs for evaluation of the antimicrobial susceptibility of BRD pathogens.

The objectives of this study were to assess differences in lactational performance (reproduction, milk production, and removal from the herd) associated with time to clinical cure of metritis. This retrospective cohort study was conducted using data from 3 experiments conducted at 5 dairy herds in Florida. Cows with metritis were randomly assigned to receive either ampicillin or ceftiofur in 2 experiments (1 experiment only implemented ceftiofur). Vaginal discharge assessment using the Metricheck device within 12 DIM was used to diagnose metritis, characterized by presence of fetid, watery, reddish-brown vaginal discharge (VD; n = 1,410), and the day of diagnosis was defined as d 0. A total of 556 cows were treated with ampicillin, and 854 cows were treated with ceftiofur. Clinical cure was visually assessed on d 5 and 12 and defined as clear, mucopurulent, or purulent VD. Cows were grouped as follows: CureD5 (cured by d 5; n = 705); CureD12 (cured between d 6 and 12; n = 406); NoCure (not cured by d 12; n = 299); NoMet (not diagnosed with metritis; n = 1,194). Data regarding risk of receiving a first service, risk and hazard of pregnancy by 300 DIM, risk and hazard of removal from the herd by 300 DIM, and total milk production within 10 mo were collected from the herd management software. Statistical analyses were performed using multivariable logistic regression, Cox proportional hazards, and ANOVA models. Orthogonal contrasts were set to assess the effects of metritis, clinical cure, and time to clinical cure. The antimicrobial treatment formulation was not associated with the odds of cure by d 5. A greater proportion of CureD5 cows received a first service (91.6% vs. 87.1%) and were pregnant by 300 DIM (69.6% vs. 62.9%) compared with CureD12 cows. However, time to cure was not associated with reduced hazard of pregnancy within 300 DIM. A smaller proportion of CureD5 cows were removed from the herd by 300 DIM compared with CureD12 cows (15.5% vs. 23.7%). Additionally, a smaller hazard of removal from the herd within 300 DIM was observed for CureD5 compared with CureD12 cows (0.61). No differences in milk production within 10 mo were associated with time to clinical cure. Our findings highlight that expedited clinical cure of metritis was associated with positive effects on reproductive performance and removal from the herd.

Feed efficiency is critical in dairy farming, affecting production costs and environmental sustainability. The development of the trait residual feed intake (RFI) has provided an opportunity to select dairy cows that are more efficient in converting nutrients into milk. Note that RFI requires individual daily intake records, which are typically collected on a limited number of research farms. In this context, the identification of biomarkers that can be used to identify and select more feed-efficient cows is of great interest. As such, this study aimed to identify ruminal and serum biomarkers associated with RFI in mid-lactation Holstein cows. A selected subset of 24 out of 454 Holstein cows was used in this study. This subset was strategically selected to represent extremes of least feed-efficient (LFE; n = 12, RFI = 2.44) and most feed-efficient (MFE; n = 12, RFI = -2.69) cows with no difference in the 3 energy sinks, namely BW change, metabolic BW, and energy secreted in milk. Rumen fluid and serum samples were collected between 60 and 90 DIM. Rumen fluid samples were collected using an oroesophageal tubing procedure. Serum samples were used to measure fatty acids using a 2-step assay. The fatty acid methyl ester was assessed using solid-phase extraction and quantified using the chromatographic peak area and internal standard-based calculations. Ruminal ammonia nitrogen was measured using a phenol-hypochlorite assay, and serum urea was measured using a commercial ELISA kit. Cows in the MFE group had higher ruminal ammonia nitrogen concentrations than cows in the LFE group. There were no differences in serum urea concentration between MFE and LFE cows. Serum fatty acid concentrations differed between groups, with myristic acid (C14:0), palmitic acid (C16:0), cis-heptadecenoic acid (cis-9-17:1), stearic acid (C18:0), and total SFA having greater concentrations in the MFE group than in the LFE group. The total PUFA concentration was lower in the MFE group than in the LFE group. A model incorporating C14:0, C16:0, palmitoleic acid (trans-9-C16:1), anteiso-heptadecanoic acid plus palmitoleic acid (C17: 0+trans-13-C16:1), oleic acid (cis-9-C18:1), cis-vacce-nic acid (cis-11-C18:1), petroselinic acid (cis-12-C18:1), C18:0, linoleic acid (C18:2n-6), dihomo-gamma-linolenic acid (C20:3n-6), cis-MUFA, n-6 PUFA, total PUFA, total SFA, and other or unknown fatty acids was used to assess goodness-of-fit for RFI and showed an adjusted R2 of 0.74. When ruminal ammonia nitrogen was added to the previous model, the adjusted R2 improved to 0.84. Our findings provide evidence that ruminal ammonia nitrogen and serum fatty acids are associated with RFI, thus suggesting that these metabolites might be helpful in identifying more feed-efficient dairy cows.

The objective was to evaluate the performance of exploratory models containing routinely available on-farm data, behavior data, and the combination of both to predict metritis self-cure (SC) and treatment failure (TF). Holstein cows (n = 1,061) were fitted with a collar-mounted automated- health monitoring device (AHMD) from −21 ± 3 to 60 ± 3 d relative to calving to monitor rumination and activity. Cows were examined for diagnosis of metritis at 4 ± 1, 7 ± 1, and 9 ± 1 DIM. Cows diagnosed with metritis (n = 132), characterized by watery, fetid, reddish/brownish vaginal discharge (VD) were randomly allocated to one of 2 treatments: Control (CON; n = 62) - no treatment at the time of metritis diagnosis (d 0); Ceftiofur (CEF; n = 70) - subcutaneous injection of 6.6 mg/kg of ceftiofur crystalline-free acid on d 0 and 3 relative to diagnosis. Cure was determined 12 d after diagnosis and was considered when VD became mucoid and not fetid. Cows in CON were used to determine SC and cows in CEF were used to determine TF. Univariable analyses were performed using farm-collected data (parity, calving season, calving-related disorders, body condition score, rectal temperature, and days in milk at metritis diagnosis) and behavior data (i.e., daily averages of rumination, activity generated by AHMD, and derived variables) to assess their association with metritis SC or TF. Variables with a P ≤ 0.20 were included in the multivariable logistic regression exploratory models. To predict SC, the area under the curve (AUC) for the exploratory model containing only data routinely available on-farm was 0.75. The final exploratory model to predict SC combining routinely available on-farm data and behavior data increased the AUC to 0.87, sensitivity (Se) 87% and specificity (Sp) 71%. To predict TF, the AUC for the exploratory model containing only data routinely available on-farm was 0.90. The final exploratory model combining routinely available on-farm data and behavior data increased the AUC to 0.93, Se of 93% and Sp of 82%. Cross-validation analysis revealed that generalizability of the exploratory models was poor, which indicates that the findings are applicable to the conditions of the present exploratory study. In summary, the addition of behavior data contributed to increasing the prediction of SC and TF. Developing and validating accurate prediction models for SC could lead to a reduction in antimicrobial use, whereas accurate prediction of cows that would have TF may allow for better management decisions.

The initial ovulatory response during synchronization programs is often low in dairy heifers, largely due to follicular dynamics and hormonal dynamics. Specifically, the progesterone concentration (P4) at the time of the first GnRH treatment in a breeding program can influence the LH response, often resulting in a suboptimal ovulatory response. The objective of this study was to determine the effect of the highest label dose 200 μg (100 μg vs. 200 μg) of GnRH (50 μg gonadorelin hydrochloride per mL; Factrel®; Zoetis Inc. Madison, NJ) at the first GnRH of a 6-d CoSynch plus P4 device program on ovulatory response and pregnancy per AI (P/AI) in first service in Holstein heifers. A total of 1308 Holstein heifers were randomly allocated at the beginning of a 6-d CIDR-Synch program, Day 0, to receive either i.m. treatment of 100 μg (2CC, n = 655) or 200 μg (4CC, n = 653) of GnRH. Also, at Day 0, heifers received an intravaginal insert with 1.38 g of P4 (Eazi-Breed CIDR® Cattle Insert; Zoetis Inc., Madison, NJ). On Day 6, the insert was removed, and i.m. treatment of 25 mg of PGF2α (12.5 mg dinoprost tromethamine/mL; Lutalyse® HighCon Injection Zoetis) was administered. On Day 7, a second i.m. treatment of 25 mg of PGF2α was given, followed on Day 9 by concurrent i.m. treatment of 100 μg of GnRH and timed AI (TAI). A subset of 396 heifers had their ovaries scanned to evaluate ovulatory response, and blood samples were collected to measure the serum concentration of P4 at Day 0 and Day 6 of the study. The P4 concentrations at Day 0 were categorized as Low (≤3ng/mL) or High (>3ng/mL). The ovulatory response was greater for heifers receiving 4CC than 2CC at Day 0 (54.7% vs. 42.8%). The ovulatory response was greater for Low P4 than High P4 at Day 0 (54.3% vs. 37.8%). However, there was not an interaction between treatment and P4 concentrations (Low P4 2CC = 48.6% vs. High P4 2CC = 30.0%; Low P4 4CC = 60.0% vs. High P4 4CC = 45.5%). The ROC curve analysis indicates that P4 concentrations at Day 0 treatment could predict the ovulatory response, although the area under the curve was only 0.6. As expected, heifers that ovulated had increased P/AI (No = 55.6% vs. Yes = 67.7%); however, there was no effect of treatment on P/AI (2CC = 63.3% vs. 4CC = 59.6%), nor interactions between treatment and ovulation and treatment and P4 (HIGH vs LOW) for pregnancy outcomes. In summary, P4 concentration and increasing the dose of GnRH at Day 0 positively impacted ovulatory response in Holstein heifers. However, there was no interaction between treatment and P4 on ovulation and no subsequent impact of GnRH dose on P/AI.

Rumen-protected choline (RPC) promotes benefits in milk production, immunity, and health in dairy cows by optimizing lipid metabolism during transition period management and early lactation. However, the RPC success in dairy cows depends on choline bioavailability which is affected by the type of protection used in rumen-protected choline. Therefore, our objectives were to determine the effects of a novel RPC on DMI, markers of metabolism and immunity, and lactation performance. Dry Holstein (n = 48) cows at 245 ± 3 d of gestation were blocked by parity and assigned to Control or RPC treatment within each block. Cows enrolled in the RPC treatment received 15 g/d of CholiGEMTM (Kemin Industries, Cavriago RE, Italy) from 21 d prepartum and 30 g/d of CholiGEMTM from calving to 21 d postpartum. During the transition period, DMI was measured daily, and blood was sampled weekly for energy-related metabolites [e.g., β-hydroxybutyrate (BHB), glucose, and nonesterified fatty acids (NEFA)] and immune function markers [e.g., haptoglobin (Hp) and lipopolysaccharide-binding protein (LPB)]. Vaginal discharge samples were collected at the calving and 7 d postpartum (DPP) and stored in microcentrifuge tubes at −80°C until 16S rRNA sequencing. The main responses of body condition score, body weight, DMI, milk yield, milk components, and immune function markers were analyzed using the GLIMMIX procedure of SAS with the effect of treatment, time, parity, and relevant covariates added to the models. The relative abundance of microbiome α-diversity was evaluated by 3 indexes (Chao1, Shannon, and Simpson) and β-diversity by principal coordinate analysis and PERMANOVA. There were no differences in DMI in the pre- and postpartum periods. Cows fed RPC increased the yields of energy and 3.5% fat-corrected milk and fat yield in primiparous and multiparous cows, with an interaction between treatment and parity for these lactation variables. However, there were no differences in milk protein and lactose up to 150 DIM between treatments. Glucose, NEFA, and BHB had no differences between the treatments. However, RPC decreased numerically BHB (Control = 1.07 ± 0.13 vs. RPC = 0.63 ± 0.13) in multiparous on the third week postpartum and tended to reduce the incidence of subclinical ketosis (12.7% vs. 4.2%). No effects for Hp and LPB were found in cows fed RPC. Chao1, Shannon, and Simpson indices were lower at calving in the RPC treatment than in the Control. However, no differences were found 7 d later for Chao1, Shannon, and Simpson indices. The vaginal discharge microbiome (VDM) was altered in cows fed RPC at 7 DPP. Fusobacterium, a common pathogen associated with metritis, was reduced in cows fed RPC. Rumen-protected choline enhanced lactation performance and health and altered the vaginal discharge microbiome, a potential proxy for uterine healthy in dairy cows. The current study's findings corroborate that RPC is a tool to support adaptation to lactation and shed light on opportunities for further research in reproductive health.

Automatic milking system (AMS) adoption in the United States is trending upward, with issues such as lower availability and increased cost of labor being factors frequently listed as motives for AMS implementation. In addition, more interest in precision dairy farming by the new generation of farmers may also help increase AMS adoption. The objective of this scoping review was to characterize the nature of the literature investigating non-pasture-based AMS and the opportunities and challenges for future research. The eligibility criteria included studies published in or after the year 2000, with full text in English, of at least 500 words, examining various outcomes related to AMS in non-pasture-based dairy farms. Six electronic databases were searched: Biosis (Web of Science), CAB Abstracts (CAB Direct), Medline (PubMed), PubAg, AGRIS (FAO), and Scopus (Elsevier). The review focused on studies with objectives, characteristics, farms, and AMS information. A total of 4,292 titles and abstracts were screened, and 536 studies were finally included. Most of the studies were conducted in Europe (73.5%), among commercial herds (67.9%), comprising Holstein cows (57.7%), using Lely and DeLaval brands (45.4% vs. 39.7%), with free-flow traffic (52.7%). The main research topics investigated were milk production, milk composition, and AMS efficiency, followed by behavior and welfare, health disorders (especially mastitis), and nutrition in Europe and other regions. At the same time, in the United States, trends were similar, except for nutrition. Since 2016, there has been an increased interest in studies on energy and water consumption, technological development, environment (enteric emissions), reproduction, genetics, and longevity or culling. However, the small number of studies and unclear characterization of what is optimum for reproductive management, other health disorders, economics, and water and energy consumption suggest a need for future research.

The most frequently reported definition of cystic ovarian disease in cattle is an abnormally persistent follicle (>7 to 10 d) with a diameter >25 mm. Dis-crimination between luteal and follicular ovarian cystic structures has traditionally been conducted by mea-suring the rim width of luteal tissue. The most com-mon practice used in the field for diagnosis of cystic ovarian disease is examination by rectal palpation with or without the use of a B-mode ultrasound. Color Dop-pler ultrasound technology allows assessment of blood flow area measurements in the ovary, which has been proposed as a potential indirect measure for plasma progesterone (P4) concentrations. The objective of this study was to compare the diagnostic accuracy of differentiating luteal structures from follicular ovarian cysts using measures collected with B-mode and color Doppler transrectal ultrasonography. The definition of an ovarian cyst was a follicle greater than 20 mm in diameter in the absence of a corpus luteum that per -sisted for at least 10 d. A 3-mm luteal rim width was used to differentiate follicular and luteal cysts. A total of 36 cows were enrolled in the study during routine herd reproductive examination visits, with 26 and 10 having follicular and luteal cysts, respectively. Cows en -rolled in the study were examined using a Mini-ExaPad mini ultrasound with color Doppler capabilities (IMV Imaging Ltd.). Blood samples were collected from each cow to measure P4 serum concentrations. History and signalment of each cow, including days in milk, lacta-tion, times bred, days since last heat, milk composi-tion, and somatic cell counts, were retrieved from an online database (DairyComp 305, Valley Agricultural Software). The accuracy of diagnosing follicular from luteal cysts based on luteal rim thickness was analyzed by receiver operating characteristic (ROC) curve using P4 as the gold standard, where P4 concentrations exceeding 1 ng/mL was defined as luteal, and all other structures with less P4 were considered follicular. Lu-teal rim and blood flow area were selected for further analysis because they presented the best ROC curves for differentiating cystic ovarian structures, with areas under the curve of 0.80 and 0.76, respectively. Luteal rim width of 3 mm was used as the cutoff standard in the study, resulting in sensitivity and specificity of 50% and 86%, respectively. Blood flow area of 0.19 cm2 was used as the cutoff standard in the study, resulting in sensitivity and specificity of 79% and 86%, respectively. When combining the use of luteal rim width and blood flow area to differentiate cystic ovarian structures, a parallel approach resulted in sensitivity and specific-ity of 73% and 93%, respectively, whereas an in-series approach resulted in sensitivity and specificity of 35% and 100%, respectively. In conclusion, the use of color Doppler ultrasonography when discriminating between luteal and follicular ovarian cysts in dairy cattle re-sulted in higher diagnostic accuracy compared with using B-mode ultrasonography alone.

The objective of this study was to assess the effects of treatment with propylene glycol (PG) and cyanocobalamin (B12) on health, milk production, and reproductive outcomes of cows diagnosed with hyperketonemia (HK), hypoglycemia (HG), or concurrent HKHG. Glucose and β-hydroxybutyric acid (BHBA) concentrations were assessed in whole blood using a handheld device in lactating dairy cows (n = 2,418) between 3 and 9 d postpartum. Cows categorized as HK (n = 232, BHBA ≥1.2 mmol/L), HG (n = 161, glucose ≤2.2 mmol/L), and concurrent HKHG (n = 204, BHBA ≥1.2 mmol/L, and glucose ≤2.2 mmol/L) were randomized to receive treatment or to remain untreated (control). Treatment consisted of a single dose of B12 (10 mg, intramuscularly) and 300 mL of PG orally for 5 d, starting on the day of cow-side testing. Milk production, health, and reproductive outcomes were analyzed according to groups. Statistical analysis was carried out using SAS version 9.4 (SAS/STAT, SAS Institute Inc.). Treatment in HG cows decreased clinical ketosis, increased milk production in the fifth week of lactation for multiparous cows, and tended to increase 305-d mature-equivalent milk yield (305ME) for primiparous cows compared with untreated cows with the same metabolic profile. For cows with HKHG, treatment increased 305ME in multiparous cows and tended to increase 305ME in primiparous cows. No differences were found for treatment among any of the metabolic groups regarding reproductive outcomes, nor were any treatment effects found among HK cows. Glycemic status may help identify metabolically challenged early postpartum dairy cows, which may have differential response to PG and B12 treatment.

The objectives of this study were to determine the effects of nerve growth factor-β (NGF), purified from bulls' seminal plasma and administered at the time of artificial insemination (AI), on progesterone post-AI, interferon-stimulated genes (ISG), and pregnancy per AI (P/AI) for lactating Holstein dairy cows enrolled in a timed-AI protocol. We hypothesized that administration of NGF at the time of AI would increase plasma progesterone post-AI, upregulate relative abundance of ISG, and improve P/AI in lactating dairy cows. Holstein cows (n = 557) from a single commercial dairy farm were blocked by parity and randomly assigned to receive an intramuscular injection containing 296 µg of bovine purified NGF at the time of AI, diluted in 2 mL of phosphate-buffered saline (NGF: n = 275), or receive only the 2 mL of phosphate-buffered saline (control: n = 282). Plasma progesterone and corpus luteum size were assessed in a subset of cows (NGF: n = 32; control: n = 36) at d 7, 14, and 19 post-AI. Relative mRNA abundance of ISG (ISG15, MX1, MX2, and RTP4) was assessed in peripheral blood leukocytes on d 19 post-AI. Pregnancy diagnosis was performed at 37 and 65 d post-AI. There was an interaction effect between treatment and parity for plasma progesterone; however, plasma progesterone and ISG did not differ between treatments. There were no effects of NGF for P/AI at 37 d post-AI (NGF = 40.0% vs. control = 41.6%), 65 d post-AI (NGF = 36.0% vs. control = 38.1%), and for pregnancy loss (NGF = 8.4% vs. control = 7.7%). The current study revealed that effects to NGF in lactating Holstein cows were minor and contingent with parity for progesterone, and no improvement in ISG relative abundance and P/AI were observed.

The objective of this prospective cohort study was to characterize the metabolic profile, health, milk production, and reproductive outcomes of cows diagnosed with hyperketonemia (HK; β-hydroxybutyrate ≥1.2 mmol/L), hypoglycemia (HG; glucose ≤2.2 mmol/L), or concurrent HK and HG (HKHG). Glucose and β-hydroxybutyrate concentrations in whole blood were assessed using a handheld device (Precision Xtra, Abbott Laboratories) in lactating dairy cows (n = 2,418) between 3 and 9 d postpartum. Cows were categorized into 4 groups: no HK or HG (healthy; Norm = 1,821), HK only (HK = 232), HG only (HG = 161), and concurrent HK and HG (HKHG = 204). Subsequent milk production, along with health and reproductive outcomes, as recorded by farm personnel, were analyzed according to metabolic category. Serum collected on the day of cow-side diagnosis of hyperketonemia and hypoglycemia was evaluated for total calcium (tCa), magnesium (Mg), nonesterified fatty acids (NEFA), triglycerides (TG), and urea using an automated chemistry analyzer (Randox Daytona; Randox Laboratories Ltd.). Statistical analysis was carried out using SAS version 9.4 (SAS Institute Inc.). Hyperketonemia in multiparous cows was associated with greater incidence of metabolic abnormalities (hypomagnesemia, hypocalcemia, and elevated NEFA and urea). Hyperketonemia in primiparous and multiparous cows led to increased adverse health events (culling rate, retained fetal membranes, puerperal metritis, clinical ketosis, left displaced abomasum) relative to Norm cows. Multiparous cows with HKHG had fewer metabolic disturbances (hypomagnesemia, hypocalcemia, elevated NEFA) than HK cows. Cows with HKHG had an increased incidence of clinical ketosis and left displaced abomasum relative to Norm cows. Cows with HG had similar metabolic profiles to Norm cows and had lower incidence of retained fetal membranes and puerperal metritis than cows with HK. Multiparous cows with HG produced more milk than Norm cows from wk 10 to 20, whereas multiparous cows with HK produced less milk than Norm cows. For primiparous cows, HK did not have a negative effect on milk production compared with Norm cows.