Early Postpartum Dairy Cows Research Articles

The use of ultrasonography in the transition period to estimate adipose tissue depots and their association with risk of early postpartum hyperketonemia in Holstein dairy cattle

An elevated prepartum BCS is a risk factor for postpartum hyperketonemia (HYK) and elevated fatty acid (NEFA) concentrations in dairy cattle. The association between different adipose tissue depots, such as subcutaneous (SCAT) as well as visceral adipose tissue (VAT) depots and HYK and elevated NEFA concentrations remains unknown. The objective of this 2-part study was to describe SCAT and VAT depots using ultrasonography during the transition period and to associate them with metabolic markers of negative energy balance in early postpartum dairy cows. Multiparous Holstein cows were enrolled in a prospective observational cohort study with 2 cohorts in subsequent study years (Cohort 1: n = 31; Cohort 2: n = 28). At −6, −4, −2, and 2 wk relative to calving (Cohort 1) or −2 and 2 wk relative to calving (Cohort 2) BCS was determined and cows were examined via transcutaneous ultrasound at 6 locations for prediction of adipose tissue mass in 5 depots: 1) abdominal adipose tissue (AAT), 2) retroperitoneal adipose tissue (RPAT), 3) omental adipose tissue (OMAT), 4) mesenteric adipose tissue (MAT), and 5) subcutaneous adipose tissue (SCAT). Postpartum serum NEFA and BHB concentrations were determined twice weekly from 1 to 14 DIM. Cows were categorized as having HYK or high NEFA concentrations (NEFAH) if ≥1 sample resulted in a BHB ≥1.2 mmol/L or NEFA ≥720 µEq/L, respectively. Critical thresholds associated with HYK or NEFAH for each depot at −2 wk relative to calving and the time of maximum estimated depot size were evaluated using logistic regression and a receiver operator characteristic analysis. Abdominal AT increased from −6 to −2 wk relative to calving in cohort 1 but did not differ from −2 to 2 wk relative to calving. Subcutaneous AT did not change from −6 to −2 wk but decreased from −2 to 2 wk relative to calving in cohort 1 and 2. Omental AT accurately (AUC = 0.77) predicted HYK in cohort 1 but was non-informative for the prediction of HYK in cohort 2 (AUC = 0.49). Predicted AT depots were not informative for the prediction of NEFAH (AUC ≤0.59) in either cohort. Results from this study suggest that AT can be monitored during the transition period using the described technique; however, estimated prepartum AT depots at −2 wk relative to expected calving were inaccurate in distinguishing between postpartum NEFAH and non-NEFAH or HYK and non-HYK.

Altered bile acid and correlations with gut microbiome in transition dairy cows with different glucose and lipid metabolism status

The transition from pregnancy to lactation is critical in dairy cows. Among others, dairy cows experience a metabolic stress due to a large change in glucose and lipid metabolism. Recent studies revealed that bile acids (BA), other than being involved in both the emulsification and solubilization of fats during intestinal absorption, can also affect the metabolism of glucose and lipids, both directly or indirectly by affecting the gut microbiota. Thus, we used untargeted and targeted metabolomics and 16S rRNA gene sequencing approaches to investigate the concentration of plasma metabolites and BA, the composition of the rectum microbial community, and assess their interaction in transition dairy cows. In Experiment 1, we investigated BA and other blood parameters and gut microbiota in dairy cows without clinical diseases during the transition period, which can be seen as well adapted to the challenge of changed glucose and lipid metabolism. As expected, we detected an increased plasma concentrations of BHB and nonesterified fatty acids (NEFA) but decreased concentrations of glucose, cholesterol, and triglycerides (TG). Untargeted metabolomic analysis of the plasma revealed primary BA biosynthesis was one of the affected pathways, and was consistent with the increased concentration of BA in the plasma. A correlation approach revealed a complex association between BA and microbiota with the host plasma concentration of glucose and lipid metabolites. Among BA, chenodeoxycholic acid derivates such as glycolithocholic acid, taurolithocholic acid, lithocholic acid, taurochenodeoxycholic acid, and taurodeoxycholic acid were the main hub nodes connecting microbe and blood metabolites (such as glucose, TG, and NEFA). In Experiment 2, we investigated early postpartum dairy cows with or without hyperketonemia (HPK). As expected, HPK cows had increased concentration of NEFA and decreased concentrations of glucose and triglycerides. The untargeted metabolomic analysis of the plasma revealed that primary BA biosynthesis was also one of the affected pathways. Even though the BA concentration was similar among the 2 groups, the profiles of taurine-conjugated BA changed significantly. A correlation analysis also revealed an association between BA and microbiota with the concentration in plasma of glucose and lipid metabolites (such as BHB). Among BA, cholic acid and its derivates such as taurocholic acid, tauro α-muricholic acid, and taurodeoxycholic acid were the main hub nodes connecting microbe and blood metabolites. Our results indicated an association between BA, intestinal microbe, and glucose and lipid metabolism in transition dairy cows. These findings provide new insight into the adaptation mechanisms of dairy cows during the transition period.

Customized modified therapies for ovarian cyst and persistent follicle followed with TAI in early postpartum dairy cows.

Postpartum ovarian dysfunction [ovarian cyst (OC) and persistent follicle (PF)] has been an important issue. Finding effective hormonal treatments to improve reproductive performance in dairy cows has become a necessity. Improve reproductive performance and ovarian activity in postpartum cows with specific customized treatment for OC and PFs. The study included 48 cows at 14 days P.P, which received two dosages of 500 μg IM cloprostenol, 14 days apart as presynchronization protocol. Ultrasound ovarian scans 14 days after the last injection for 4 weeks. The cows were divided into three groups according to ovarian status: OC (n = 14), PF (n = 12), and NE (n = 22). In the OC group, received 500 μg IM cloprostenol and 100 μg IM cystoriline, a second dose of cloprostenol 14 days later and a second dose of cystoriline 36 hours later, and AI after 24 hours (GnRH+ PG/PG/GnRH). In the PF group, was fitted with progesterone-releasing intravaginal device (PRID) for 9 days; the same day, they received 100 μg cystoreline then 500 μg cloprostenol 7 days later, after PRID removal AI 56 hours later (PRID + GnRH/PG). In the NE group, artificial insemination was implemented until 28 days depending on estrus detection. The ovarian activity was greatly affected by the customized treatments, leading to enhanced follicular and luteal activity, particularly after the PGF2α injection. The OC and PF groups showed substantial estrus responses of 71.43% and 75.02%, respectively, during AI time. While the NE group had an ovulation rate of 54.5% and a pregnancy rate of 31.8%, the treatment groups showed marked improvements in reproductive performance. The ovulation rates in the OC and PF groups were 71.43% and 75% and the pregnancy rates at the 1st artificial insemination were 64.28% and 66.7%. Improving reproductive performance and minimizing the time to first service are possible advantages of early case-specific treatment for postpartum cows with OC and PFs.

Rumen-protected glucose stimulates the secretion of reproductive hormones and the mTOR/AKT signaling pathway in the ovaries of early postpartum

This study was conducted to determine the response of the reproductive hormones and the mTOR/AKT/PI3K pathway in the ovaries of postpartum dairy cows with dietary rumen-protected glucose (RPG). Twelve Holstein cows were randomly assigned to two groups (n = 6/group): the control group (CT) and the RPG group. Blood samples were collected on d 1, 7, and 14 after calving for the gonadal hormone assay. The expression of the gonadal hormones receptors and PI3K/mTOR/AKT pathways were detected using RT-PCR and Western blot. The RPG addition increased the plasma LH, E2, and P4 concentrations on d 14 after calving and upregulated the mRNA and protein expressions of the ERα, ERβ, 17β-HSD, FSHR, LHR, and CYP17A1 but downregulated StAR expression. Immunohistochemical analysis identified higher expressions of the FSHR and LHR in the ovaries of RPG-fed cows compared to CT cows. Furthermore, the protein expressions of p-AKT/AKT and p-mTOR/mTOR were significantly increased in the ovaries of RPG-fed cows compared to the CT group, but the addition of RPG did not alter the protein expression of p-PI3K/PI3K. In conclusion, the current results indicated that dietary RPG supplementation regulated gonadotropin secretion and stimulated expression of hormone receptors and the mTOR/AKT pathway in the ovaries of early postpartum dairy cows. RPG may be beneficial for the recovery of ovarian activity in post-calving dairy cows.

Effects of a single transdermal administration of flunixin meglumine in early postpartum Holstein Friesian dairy cows: Part 2. Milk yield, culling risk, and reproductive performance.

This study was conducted to assess the effects of a single transdermal administration of flunixin meglumine (FM) in early postpartum Holstein Friesian dairy cows on milk yield, culling risk, and reproductive performance. We hypothesized that FM treatment would reduce systemic inflammation, leading to higher milk yield, reduced culling risk, and better reproductive performance in the subsequent lactation. Holstein Friesian dairy cows [n = 500, 153 primiparous (PRIM), 347 multiparous (MULT)] from 3 farms in northeast Germany were enrolled in a prospective, randomized controlled clinical trial. Farms at risk for cows with excessive postpartum inflammation were identified in a preliminary trial by measuring serum haptoglobin concentrations in their fresh lactating cows. Only cows that had a eutocic birth and delivered a singleton calf alive, with no signs of milk fever or retained fetal membranes and rectal temperature ≤40°C at first clinical examination, were included within 24 to 36 h postpartum. Treatment included a single transdermal administration of either FM (3.33 mg/kg) or a placebo as control (CON). Milk production, milk solids, urea, and somatic cell count were recorded monthly for 8 mo after calving. Culling risk, first-service conception risk, and days open were retrieved from the farms' herd management software. Separate models for PRIM and MULT cows were built for most parameters because of significant effects of parity and parity × treatment interaction. Energy-corrected milk yield from 8 monthly Dairy Herd Improvement-equivalent tests was slightly greater in PRIM cows treated with FM (29.51 and 30.73 ± 1.35 kg, CON vs. FM), whereas it was reduced in treated MULT cows (38.23 and 37.47 ± 1.17 kg, CON vs. FM) compared with CON. Milk fat and protein yields were greater in FM-treated PRIM cows and lower in treated MULT cows compared with CON. Milk urea and somatic cell count were not affected by treatment. No differences in culling risk, first-service conception risk, or days open were observed. We conclude that a single transdermal administration of FM in early postpartum dairy cows on farms at risk for excessive postpartum inflammation slightly increased milk, milk fat, and milk protein yields in PRIM cows and decreased these variables in MULT cows. Neither culling risk nor fertility was affected by treatment in this study.

Reproductive and lactational responses of multiparous dairy cattle to short-term postpartum chromium supplementation during the summer months

The objective of this work was to evaluate the potential benefits of short-duration, high-dose chromium (Cr) supplementation in early postpartum dairy cows during the summer months. Multiparous, early-lactation cows (20.95 ± 0.21 d in milk) were assigned to 1 of 2 treatment groups: (1) control diet (Con; n = 10) or (2) control diet + Cr propionate (CrPro; 12 mg/head per day Cr; n = 12). Measurements of ovarian structures, respiration rates (RR), rectal temperatures (RT), and blood glucose concentrations were recorded every 3 d. Blood was also collected for analysis of plasma progesterone concentrations. Every 6 d, in conjunction with ultrasonography, endometrial cytology samples were collected via cytobrush from each cow to determine the incidences of subclinical endometritis, as determined by polymorphonuclear leukocyte (PMNL)%. No differences were detected in RR, RT, blood glucose, feed intake, milk yield, or change in body weight. The supplementation did, however, improve some reproductive parameters. At cytology sample 6, the PMNL% increased in Con cows, and was greater than the PMNL% in the CrPro group. Chromium consumption did not affect the number or size of most follicles, with the exception being the 6 to 9 mm category where the CrPro group had a greater average diameter and tended to have greater numbers of follicles in this category. While corpus luteum numbers and size did not differ between treatments, the ratio of progesterone to average corpus luteum volume was greater in the CrPro group compared with the Con group. The results from this study indicate that, whereas the short-term, high-dose supplementation strategy did not affect feed intake or milk yield, this Cr supplementation strategy could benefit reproductive performance during periods of stress.

A randomized controlled trial examining the effects of treatment with propylene glycol and injectable cyanocobalamin on naturally occurring disease, milk production, and reproductive outcomes of dairy cows diagnosed with concurrent hyperketonemia and hypoglycemia.

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.

Simultaneous free fatty acid elevations and accelerated desaturation in plasma and oocytes in early postpartum dairy cows under intensive feeding management

A severe negative energy balance and high circulating free fatty acids (FFA) in postpartum cows impair fertility. The lipotoxicity of FFA has been shown to decrease the quality of bovine oocytes in vitro. Therefore, excess FFA in cells is converted to triacylglycerol (TAG), a non-toxic form, to avoid lipotoxicity. We recently reported that the TAG content in oocytes was higher in postpartum lactating cows subjected to grazing management than in heifers (Theriogenology 176: 174–182, 2021). The present study investigated the compositions of the energy metabolism-related lipids, FFA and TAG, in the plasma and oocytes of cows at different lactation stages under indoor intensive feeding management in order to obtain insights into lipotoxicity in oocytes, particularly those in early postpartum cows. Blood and oocytes were collected from 20 lactating cows categorized into the following lactation groups: 20–30 days in milk (DIM) (n = 5), 40–50 DIM (n = 5), 60–80 DIM (n = 5), and 130–160 DIM (n = 5). Daily energy balance data were obtained for 3 weeks prior to oocyte collection using the ovum pick up (OPU) method. The contents and compositions of FFA and TAG in plasma and oocytes were analyzed using liquid chromatography-mass spectrometry. As expected, plasma FFA was high at 20–30 DIM, decreased by 50 DIM, and was maintained at a low level for the remainder of the experimental period. Similar changes were observed in oocyte FFA and TAG with DIM as plasma FFA. Oocyte FFA positively correlated with plasma FFA (P < 0.05), but negatively correlated with the mean energy balance 1 and 21 days before OPU (P < 0.05). Relationships were noted between the composition and content of FFA in plasma and oocytes, with the FFA 16:1/16:0 and 18:1/18:0 ratios positively correlating with the total amount of FFA (P < 0.05). Elevated oocyte FFA in cows in the early postpartum period under intensive feeding management suggested that oocytes were at a high risk of FFA lipotoxicity. Furthermore, the present results implied that the severe negative energy balance in the previous few weeks was closely related to increases in oocyte FFA, which supports the importance of long-term cow feeding management for preserving the quality of oocytes in the early postpartum period. The present results provide insights into the effects of high circulating FFA on the fertility of postpartum cows.

Inhibition of mTOR in bovine monocyte derived macrophages and dendritic cells provides a potential mechanism for postpartum immune dysfunction in dairy cows

Dairy cattle experience a profound nutrient deficit postpartum that is associated with immune dysfunction characterized by heightened inflammation and reduced pathogen clearance. The activation of the central nutrient-sensing mTOR pathway is comparatively reduced in leukocytes of early postpartum dairy cows during this time of most pronounced nutrient deficit. We assessed the effect of pharmacological mTOR inhibition (Torin-1, rapamycin) on differentiation of monocyte derived classically (M1) and alternatively (M2) activated macrophages (MPh) and dendritic cells (moDC) from 12 adult dairy cows. Treatment with mTOR inhibitors generated M1 MPh with increased oxidative burst and expression of IL12 subunits but decreased phagocytosis and expression of IL1B, IL6, and IL10. In M2 MPh, treatment inhibited expression of regulatory features (CD163, ARG2, IL10) skewing the cells toward an M1-like phenotype. In moDC, mTOR inhibition increased expression of pro-inflammatory cytokines (IL12A, IL12B, IL1B, IL6) and surface CD80. In co-culture with mixed lymphocytes, mTOR-inhibited moDC exhibited a cytokine profile favoring a Th1 response with increased TNF and IFNG production and decreased IL10 concentrations. We conclude that mTOR inhibition in vitro promoted differentiation of inflammatory macrophages with reduced regulatory features and generation of Th1-favoring dendritic cells. These mechanisms could contribute to immune dysregulation in postpartum dairy cows.

Concurrent and carryover effects of feeding blends of protein and amino acids in high-protein diets with different concentrations of forage fiber to fresh cows. 2. Protein balance and body composition

Increasing the supply of metabolizable protein (MP) and improving its AA profile may attenuate body protein mobilization in fresh cows and lead to increased milk production. Increasing the concentration of rumen-undegradable protein (RUP) to increase MP supply and replacing RUP sources from forages instead of nonforage fiber sources may further decrease tissue mobilization if it improves dry matter intake (DMI). Our objective was to determine whether increasing MP concentrations and improving the AA profile at the expense of either nonforage or forage fiber (fNDF) would affect MP balance and empty body (EB) composition (measured using the urea dilution method) in early postpartum dairy cows of different parities. In a randomized block design, 40 primigravid [77 ± 1.5 kg of EB crude protein (CP) at 8 ± 0.6 d before calving] and 40 multigravid (92 ± 1.6 kg of EB CP at 5 ± 0.6 d before calving) Holsteins were blocked by calving date and fed a common prepartum diet (11.5% CP). After calving to 25 d in milk (DIM), cows were fed 1 of 4 diets: (1) a diet deficient in MP meeting 87% of MP requirements (DMP, 17% CP, 24% fNDF), (2) 104% of MP requirements using primarily soy protein to make MP adequate (AMP, 20% CP, 24% fNDF), (3) 110% of MP requirements using a blend of proteins and rumen-protected (RP) AA to make MP adequate (Blend, 20% CP, 24% fNDF), or (4) a diet similar to Blend but substituting added RUP for fNDF rather than nonforage NDF (Blend-fNDF, 20% CP, 19% fNDF). Blend was formulated to have a RUP supply with a similar AA profile to that of casein. Cows were fed a common diet (16.3% CP) from 26 to 50 DIM. Calculated MP balance (supply - requirements) was less than zero for DMP and Blend-fNDF from 1 to 4 wk of lactation (WOL), whereas that for AMP was positive from 1 to 4 WOL and that for Blend was close to zero from 3 to 4 WOL. Daily MP balance was greater from 5 to 7 WOL for DMP compared with AMP and Blend (100 vs. 22 g/d). From -7 to 7 d relative to calving, losses of EB CP were greater for DMP than for AMP and Blend (-121 vs. average of 11 g/d). From 7 to 25 DIM, cows fed AMP (-139 g/d) and Blend-fNDF (-147 g/d) lost EB CP but cows fed Blend (-8 g/d) maintained EB CP. Increased DMI for Blend versus AMP led to reduced losses of EB lipid in primiparous cows from 7 to 25 d relative to calving (-1.0 vs. -1.3 kg/d of EB lipid), whereas lipid mobilization was similar in multiparous cows (average -1.1 kg of EB lipid/d). By 50 DIM, EB lipid and CP were similar across treatments and parities (average 60.2 kg of EB lipid and 81.6 kg of EB CP). Overall, feeding fresh cows a high MP diet with a balanced AA profile improved DMI and attenuated EB CP mobilization, which could partly explain positive carryover effects on milk production for multiparous cows and reduced lipid mobilization for primiparous cows.

Evaluation of different acute-phase proteins for herd health diagnostics in early postpartum Holstein Friesian dairy cows.

This research communication describes (1) the comparison of acute-phase protein (APP) concentrations in transition dairy cows on different farms using both pooled and individual blood samples, and (2) the association among different APP and clinical health parameters. The first hypothesis was that early postpartum dairy cows from different farms differ in the level of inflammation, which might be determined using APP assays in both pooled and individual blood samples. The second hypothesis was that the APP haptoglobin (Hp) might be the most sensitive parameter to detect cows at risk of excessive postpartum inflammation concomitant with systemic disease. Serum concentrations of Hp, serum amyloid-A (SAA), total protein (TP), albumin (Alb), coeruloplasmin (Cp) and C-reactive protein (CRP) in 100 fresh lactating cows (within 0-8 d postpartum) from 10 farms were compared and associated to clinical health parameters (rectal body temperature, vaginal discharge (Metricheck™ score), rumen fill, vulvovaginal laceration) using both pooled and individual blood samples. Mean serum concentrations of Hp, SAA and TP revealed significant differences among farms. Pooled serum samples of farms showed high correlations with the mean of individual samples. Only Hp was significantly positively correlated to both body temperature and Metricheck™ score. In conclusion, Hp differentiates dairy farms regarding the inflammatory state of transition cows using individual and pooled serum samples within the first week postpartum. It also mirrors the individual degree of inflammation, thus proving to be a diagnostic parameter of high interest during the periparturient period.

Effects of Intrauterine Infusion of a Chitosan Solution on Recovery and Subsequent Reproductive Performance of Early Postpartum Dairy Cows with Endometritis: A Pilot Field Trial.

Simple SummaryEndometritis is one of the most common disorders during the postpartum period in dairy cows. We investigated the efficacy of intrauterine infusion of a chitosan solution in uterine recovery in early postpartum dairy cows with or without endometritis, and their subsequent reproductive performance. We found that, compared to the absence of treatment, chitosan solution treatment during the early postpartum period (approximately 3 weeks after parturition) accelerated uterine recovery after parturition of dairy cows. These findings suggest that the administration of chitosan solution once in the early postpartum period may have antimicrobial effects on the uterus. We suggest that chitosan solution is a potential therapeutic candidate for endometritis that may replace prostaglandin F2α or antibiotic treatments.This study investigated the efficacy of intrauterine infusion of a chitosan solution (CHT) on uterine recovery in early postpartum dairy cows with or without endometritis, and their subsequent reproductive performance. In Experiment 1, cows with endometritis at 3 weeks postpartum were administered CHT (n = 5) and prostaglandin F2α (PGF2α) (n = 4). Untreated cows (n = 7) served as the control group. In Experiment 2, 18 cows with a normally recovered uterus at the fresh cow check (mean, 35 days postpartum) were assigned to the CHT (n = 10) and control (n = 8) groups, and intrauterine infusion was conducted in the CHT group. Overall, in Experiment 1, the percentage of polymorphonuclear leukocytes significantly declined in the CHT group (32.3 ± 10.2 to 5.5 ± 2.4, p < 0.05) from week 3 to week 5, but no decline occurred in the PGF2α and control groups. In Experiment 2, the CHT and control groups showed no significant differences in reproductive parameters, suggesting the absence of adverse effects of CHT on fertility. These results suggest that intrauterine infusion of CHT in the early postpartum period effectively accelerates uterine recovery from endometritis and might be a suitable replacement for PGF2α administration.

Acute-phase protein α-1-acid glycoprotein is negatively associated with feed intake in postpartum dairy cows

α-1-Acid glycoprotein (AGP) is an acute-phase protein that may suppress dry matter intake (DMI), potentially by acting on the leptin receptor in the hypothalamus. Our objectives were to characterize plasma AGP concentration and associations with DMI during the transition period, and to determine the utility of AGP to identify or predict cows with low DMI. Plasma samples (n = 2,086) from 434 Holstein cows in 6 studies were analyzed on d -21, -13 ± 2, -3, 1, 3, 7 ± 1, 14 ± 1, and 21 ± 1 relative to parturition. A commercially available ELISA kit specific for bovine AGP was validated, and 2 internal controls were analyzed on each plate with interplate variation of 15.0 and 17.3%, respectively. Bivariate analysis was used to assess the relationship between AGP and DMI. For significant associations, treatment(study) was added to the model, and quadratic associations were included in the model if significant. Plasma AGP concentration (±SEM) increased from 213 ± 37.3 μg/mL on d -3 to 445 ± 60.0 μg/mL on d 14. On d 3, AGP was associated negatively with DMI in a quadratic manner for wk 1 and wk 2 and linearly for wk 3. Day 7 AGP was associated negatively with DMI in a quadratic manner for wk 2 and linearly for wk 3. Similarly, d 14 AGP was negatively associated with DMI for wk 3 and wk 4. As d 3 AGP concentration increased over the interquartile range, a calculated 1.4 (8.5%), 0.5 (2.7%), and 0.4 (1.9%) kg/d reduction in predicted DMI was detected during wk 1, 2, and 3, respectively. Using bivariate analysis, d 3 AGP explained 10% of the variation in DMI during wk 1. We explored the clinical utility of d 3 AGP to diagnose low DMI, defined as wk 1 DMI >1 standard deviation below the mean. Receiver operating characteristic analysis identified a threshold of 480.9 μg/mL, providing 76% specificity and 48% sensitivity (area under the curve = 0.60). Limited associations occurred between AGP and blood biomarkers; however, AGP was associated with plasma haptoglobin concentration postpartum and incidence of displaced abomasum, retained placenta, and metritis. These results demonstrate a negative association between plasma AGP concentration and DMI in early-postpartum dairy cows, although its diagnostic performance was marginal. Further investigation into whether AGP directly suppresses DMI in dairy cattle is warranted.

Cytobrush cytology patterns of early postpartum dairy cows

Endometrial cytobrush cytology has been recommended as a reliable method for determining the percentage of polymorphonuclear leukocytes (PMN%) in cattle smears to diagnose cytological endometritis (CE). In this study, the clarity of cytobrush cytological smears and the influence of different sample evaluation methods (number and types of cells counted) on CE diagnosis were evaluated. Samples from 28 lactating Holstein cows were collected weekly between 3 and 7 weeks postpartum. Smear clarity, based on cell density, quality of cell morphology, and red blood cell contamination, was significantly poorer at 3 weeks than between 5 and 7 weeks postpartum. Five different cell counting methods (C100, C200, C300, C400, and C500) were used, where 100–500 nucleated cells (endometrial epithelial cells, PMN consisting of neutrophils, eosinophils and basophils, lymphocytes, and macrophages) were counted. Agreement of diagnostic results for CE between C300 and C500 and between C400 and C500 was excellent at all observation times. In calculations of the PMN% based on whether the number of lymphocytes and macrophages were or were not excluded in the denominator, exclusion of these cells in the calculations did not affect the diagnosis of CE. While reduced clarity in earlier stage samples might interfere with the accuracy of cytobrush cytology, C300 can be recommended to determine the endometrial PMN%.

Hyperketonemia GWAS and parity-dependent SNP associations in Holstein dairy cows intensively sampled for blood β-hydroxybutyrate concentration.

Hyperketonemia (HYK) is a metabolic disorder that affects early postpartum dairy cows; however, there has been limited success in identifying genomic variants contributing to HYK susceptibility. We conducted a genome-wide association study (GWAS) using HYK phenotypes based on an intensive screening protocol, interrogated genotype interactions with parity group (GWIS), and evaluated the enrichment of annotated metabolic pathways. Holstein cows were enrolled into the experiment after parturition, and blood samples were collected at four timepoints between 5 and 18 days postpartum. Concentration of blood β-hydroxybutyrate (BHB) was quantified cow-side via a handheld BHB meter. Cows were labeled as a HYK case when at least one blood sample had BHB ≥ 1.2 mmol/L, and all other cows were considered non-HYK controls. After quality control procedures, 1,710 cows and 58,699 genotypes were available for further analysis. The GWAS and GWIS were performed using the forward feature select linear mixed model method. There was evidence for an association between ARS-BFGL-NGS-91238 and HYK susceptibility, as well as parity-dependent associations to HYK for BovineHD0600024247 and BovineHD1400023753. Candidate genes annotated to these single nuclear polymorphism associations have been previously associated with obesity, diabetes, insulin resistance, and fatty liver in humans and rodent models. Enrichment analysis revealed focal adhesion and axon guidance as metabolic pathways contributing to HYK etiology, while genetic variation in pathways related to insulin secretion and sensitivity may affect HYK susceptibility in a parity-dependent matter. In conclusion, the present work proposes several novel marker associations and metabolic pathways contributing to genetic risk for HYK susceptibility.

Associations between circulating levels of natural antibodies, total serum immunoglobulins, and polymorphonuclear leukocyte function in early postpartum dairy cows

The objective of this study was to investigate the associations between natural antibodies (NAbs) and total serum immunoglobulins (Igs) concentrations (both from IgG and IgM isotypes), with variables of polymorphonuclear leukocytes (PMN) function of dairy cows in the early postpartum period. Seventy-five healthy postpartum dairy cows at 2 ± 1 days in milk were enrolled in a cross-sectional study. Natural and total antibodies in serum samples were measured by ELISA. Flow cytometry was used to determine the phagocytosis and oxidative burst capacities of PMN and the quantification of the adhesion molecule l-selectin. Leukocyte count and differentials, and serum haptoglobin were also measured. A positive correlation between NAbsIgM and total serum IgM and between NAbsIgM and NAbsIgG were found. Before performing the associations between circulating Igs concentrations and PMN function variables, cows were categorized into having low, medium or high circulating antibodies based on their NAbs and total Igs serum concentrations. None of the PMN variables assessed differed between low, medium, and high cows for both NAbsIgM and NAbsIgG. While associations between PMNs function and total IgG were not observed, some associations between total IgM concentrations and PMN activity were found. Cows with high serum IgM had greater phagocytic activity compared to cows in the low IgM group. Finally, the proportion of PMN that performed oxidative burst and PMN surface expression of l-selectin intensity was greater in high total IgM group when compared to medium and low groups. In conclusion, association between NAbs, and PMN activity variables were not observed, but total serum IgM was associated to some PMN function variables in early post-partum dairy cows.

Effects of anionic supplement source in prepartum negative dietary cation-anion difference diets on serum calcium, feed intake, and lactational performance of multiparous dairy cows

Incidence of subclinical hypocalcemia in early postpartum dairy cows continues to be an animal welfare concern and an economic burden for producers. Feeding prepartum negative dietary cation-anion difference (DCAD) diets produces metabolic acidosis, which supports mobilization of bone calcium and reduces the incidence of hypocalcemia. Achieving a sufficient degree of metabolic acidosis without reducing dry matter intake (DMI) can be difficult. This study compared the ability of MegAnion (MA; Origination O2D Inc., Maplewood, MN), a new DCAD supplement designed to be more palatable than typical anionic salt sources, and another palatable commercial DCAD product, SoyChlor (SC; Landus Cooperative, Ralston, IA), to reduce urine pH (a surrogate for metabolic acidosis) without reducing prepartum DMI. A secondary objective was to assess the effect of these anionic supplements on postpartum serum calcium concentrations and DMI. Prepartum multiparous Holstein (HO) and crossbred (XX) cows were blocked by breed and expected calving date and randomly assigned within breed to total mixed rations (TMR) with MA or SC and DCAD values of -215 mEq/kg of DM. Cows (n = 56; 15 MA-HO, 12 SC-HO, 15 MA-XX, 14 SC-XX) consumed the treatment TMR for at least 19 d and completed the 28 d in milk (DIM) phase of the study. Urine and blood samples were collected weekly and at 1, 2, and 3 DIM. Data were analyzed as a randomized block design by repeated measures with week or DIM as the repeated effect. Prepartum urine pH decreased from 8.15 ± 0.27 before treatment to 6.12 ± 0.14 during treatment, was not affected by anionic supplement, and increased immediately after calving when all cows consumed the same early-lactation TMR. Prepartum serum calcium concentrations were not affected (2.34 vs. 2.33 ± 0.02 mmol/L) by treatment, whereas nonesterified fatty acids were lower (86 vs. 120 ± 10 mmol/L) and insulin was greater (215 vs. 174 ± 10 pmol/L) in cows fed MA than in cows fed SC. These differences are supported by the numerically greater prepartum DMI (1.2 kg/d) and energy balance (1.8 Mcal/d) of cows fed MA. However, pre- and postpartum DMI and other production variables, including body weight, body condition score, milk yield, and energy balance, were not affected by treatment. This lack of difference indicates that MA provides another effective source of anionic salts for diets designed to reduce urine pH and induce metabolic acidosis in prepartum dairy cows.

Rumen-Protected Glucose Stimulates the Insulin-Like Growth Factor System and mTOR/AKT Pathway in the Endometrium of Early Postpartum Dairy Cows.

Simple SummaryA rapid recovery of the uterus is important in order to shorten the interval between pregnancies. An insufficient availability of glucose in early lactation can delay the uterine recovery. However, little information is available on dietary rumen-protected glucose addition on the uterine involution and insulin-like growth factor system in the endometrium of dairy cows during the early post-natal period. In this study, dairy cows were fed with 0 or 200 g rumen-protected glucose twice every day from −7 ± 2 to 14 d during the post-natal period. The results indicated that dietary rumen-protected glucose addition promoted the proliferation of endometrial cells in postpartum dairy cows by stimulating the insulin-like growth factor system and rapamycin complex 1 pathway, and might be beneficial for uterine recovery. This study aimed to elucidate the effects of a dietary rumen-protected glucose (RPG) addition on uterine involution through the analysis of an insulin-like growth factor (IGF) system and associated pathways in the post-natal endometrium. Twelve Holstein cows were assigned equally to two groups: a control group (CT) and an RPG group (200 g of RPG per cow per day). The plasma content of insulin-like growth factor 1 (IGF1) was determined by using the ELISA method. Expressions of IGF members, the matrix metalloproteinase, protein kinase B (AKT)/mechanistic target of rapamycin complex1 (mTOR) signaling pathway, and cell proliferation factors (proliferating cell nuclear antigen (PCNA) and Ki67) were detected using real-time polymerase chain reaction, Western blot, immunohistochemistry, and immunofluorescence, respectively. The results showed that the positive cells of PCNA and Ki67 were increased in the endometrium of RPG versus CT cows. The RPG addition significantly increased the plasma IGF1 level 14 d after delivery. The mRNA expressions of the IGF family members (IGF1, IGF2, type 1 IGF receptor (IGF1R) and IGF-binding proteins (IGFBP1, IGFBP2, IGFBP4 and IGFBP5)) were upregulated, and mRNA expressions of matrix metalloproteinase MMP3 and MMP9 were downregulated in cows from the RPG group compared with the CT group. Meanwhile, the protein expressions of IGF1, IGF2, IGF1R, IGFBP1 and IGFBP4 were upregulated in cows from the RPG group compared with the CT group. Immunohistochemical analysis identified a positive response for IGF1R and IGF2R in the endometrium of RPG versus CT cows. Furthermore, the RPG supplementation increased the protein expressions of phosphorylated (p)-AKT to total AKT and p-mTOR to total mTOR ratio in the endometrium. The current results indicated that the RPG supplementation promoted the proliferation of endometrial cells by stimulating the IGFs and mTOR/AKT pathway in the early post-natal endometrium of dairy cows.

Androgen production in response to LH is impaired in theca cells from nonovulatory dominant follicles in early-postpartum dairy cows

Most dairy cows develop a dominant follicle within two weeks postpartum, but 60% of these follicles fail to ovulate. In a previous study, we determined that cows destined to ovulate have higher LH pulse frequency and circulating estradiol. The latter characteristic provided a method for distinguishing ovulatory from nonovulatory follicles during development and we found that nonovulatory follicles have lower estradiol and androstenedione in their follicular fluid. We hypothesized that lower LH pulse frequency impairs androgen production by theca cells of nonovulatory cows, reducing their ability to make estradiol. In the present study, we applied our method for predicting follicle fate to collect dominant follicles from predicted ovulatory (n = 7) and nonovulatory (n = 3) follicles. Theca and granulosa cells were separated and cultured in the absence or presence of LH, FSH, and/or testosterone for three days, with daily collection of culture medium for steroid RIAs. Estradiol and progesterone production by granulosa cells were not different between ovulatory and nonovulatory follicles. By contrast, overall androstenedione production by theca cells from ovulatory follicles was significantly higher compared with nonovulatory follicles on all three days of culture and, as culture progressed, theca from nonovulatory follicles had increasingly poorer responses to LH. In the same cultures, the progesterone production by theca cells was similar in ovulatory and nonovulatory groups. In support of our hypothesis, the results show that estradiol production by granulosa cells from nonovulatory follicles is robust when androgen substrate is present, but that thecal androgen production in response to LH is impaired. This suggests that the initial defect in steroidogenesis in dominant follicles that fail to ovulate postpartum is lower production of androgen by theca cells.

Evaluation of infrared thermography compared to rectal temperature to identify illness in early postpartum dairy cows

This study evaluated and compared infrared thermography (IRT) and rectal temperature (RT) as screening tests to identify sick transition dairy cows. Holstein cows (n = 72; 42 primiparous) had RT and IRT temperatures taken daily from 1 to 12 days in milk (DIM). Health examinations were performed daily to diagnose retained fetal membrane, milk fever and metritis, and blood was analyzed for β-hydroxybutyrate at 6 and 9 DIM to diagnose ketosis. Plasma concentrations of cortisol, interleukin-6, tumor necrosis factor α and serum amyloid A at 3, 6, 9 and 12 DIM were included as additional indicators of illness. Cows were categorized as true sick if clinically diagnosed with an illness, or if at least 2 blood parameters were above the normal range. Diagnostic test performances for RT and IRT variables were determined for each variable at a test referent value that provided the highest Youden's (J) index. The best performing screening test depended on the definition of true sickness. In general, the J index for RT was 0.15–0.17 whereas the highest J index for the IRT variables was 0.22 for the mean eye temperature and 0.19 for the mean cheek temperature. Infrared thermography was at least comparable to RT and some IRT variables performed better as a screening tests than RT. Future studies into the automation of IRT for surveillance of early postpartum diseases is warranted.