Effect of oxytocin use during colostrum harvest and association of cow characteristics with colostrum yield and IgG concentration in Holstein dairy cows.

Our objectives were to determine the effect of oxytocin use during colostrum harvest on colostrum yield (CY) and IgG concentration in Holstein dairy cows on a commercial dairy in New York and to describe associations of cow characteristics with these outcomes. Animals were enrolled between July and October 2023 using a randomized block design, with day of enrollment as the unit of randomization. A median (range) of 10 (3 to 19) cows were enrolled/d. Treatments were 1) 40 IU Oxytocin (OXY40), 2) 20 IU Oxytocin (OXY20), and 3) an untreated control group (CNTR). Oxytocin was administered intramuscularly (IM) approximately 45 s before unit attachment in a rotary parlor. Colostrum weight was measured using the colostrum bucket-embedded scale or a platform scale when the yield was less than the smallest bucket scale. The concentration of IgG ([IgG]) in colostrum was determined using radial immunodiffusion (RID) and used to calculate total IgG, and dry matter (%) was determined by oven drying. Individual cow characteristics such as parity, calf sex, weight, dystocia score, stillbirth, milk production in wk 4 of lactation, and for multiparous cows, dry period length, previous lactation dry off linear score (LS) were collected. Data were analyzed separately for primiparous and multiparous cows using backward stepwise elimination to produce final mixed effects ANOVA models Primiparous cows (n = 201) were randomized to 35.8% (n = 72) OXY40, 32.8% (n = 66) OXY20, and 31.3% (n = 63) CNTR. Multiparous groups (n = 435) were randomized to 34.7% (n = 151) OXY40, 29.7% (n = 129) OXY20, and 35.6% (n = 155) CNTR. The median (range) CY was 6.0 (0 to 20.6) kg and [IgG] was 98.5 (0.1 to 293.6) g/L in the study population. In primiparous cows, OXY40 had a higher colostrum yield (LSM [95% CI]) of 5.4 (4.9 to 5.9) kg compared with both OXY20 (4.1 [3.5 to 4.7] kg) and CNTR (3.8 [3.3 to 4.3] kg) (P < 0.001). In multiparous cows, OXY40, OXY20, and CTNR did not differ in CY (5.9 [5.3 to 6.5], 5.7 [6.3 to 5.1], and 5.4 [6.0 to 4.8] kg, respectively, P = 0.43), but colostrum yield was greater in parity 2 compared with all other parities, cows giving birth to male calves, cows with the highest milk production at wk 4 of lactation, and with a dry period of >65 d. Oxytocin use did not affect [IgG] in either primiparous or multiparous cows (P > 0.56), but [IgG] was highest in cows in parity ≥4 and lowest in cows dry >65 d. In summary, oxytocin use at 40 IU IM in primiparous was associated with a higher CY but not [IgG]. Oxytocin use likely addressed disturbed milk ejection and therefore increased CY in heifers milked for the first time in a rotary parlor. This study confirms cow characteristics associated with colostrum production within a single herd.

Development of a bovine continuing education program for early-career veterinarians to address clinical service shortage.

There is currently a shortage of production animal veterinarians worldwide. Access to adequate continuing education (CE) increases retention of health-care workers in rural areas. However, little information is available about the preferences of bovine veterinarians in their first years after graduation. Our objective was to design a readily available CE program tailored for recent veterinary graduates working in bovine medicine. For this, we conducted a cross-sectional survey to identify early-career bovine veterinarian needs and their preferences to access CE conducted while practicing full time. Subsequently, we conducted a second survey to triangulate their responses with the opinions of experienced veterinarians, where the experienced veterinarians ranked the relevance of the CE topics that resulted from the early-career veterinarian survey. We received a total of 132 valid responses from US veterinarians in their first 5 years after graduation, and 32 responses associated with bovine veterinarians with 10 or more years of experience. Our results showed that a combination of distance education and workshops to practice hands-on skills was the preferred method for accessing CE among early-career veterinarians. Furthermore, recent graduates were willing to commit 1 to 2 h/wk to distance education. From the list of 20 topics identified by early-career veterinarians, the experienced veterinarians ranked "calf/heifer management" and "immunology and vaccinology" as the most relevant ones for practice. With the information gathered from these surveys, we designed, implemented, and piloted a 180-h CE program on bovine health management that is delivered over 2 yr through asynchronous distance education and annual hands-on workshops. Participant feedback has been very positive. Overall, the survey results will serve to develop CE programs targeted to bovine veterinarians in their first years of practice. Future research is needed to evaluate this program's success in retaining the bovine veterinary workforce within the United States, particularly in rural underserved areas.

The effect of heat treatment on colostral and newborn calf redox status and oxylipid biomarkers.

Newborn calves experience altered redox balance upon transition to extrauterine life. In addition to its nutritional value, colostrum is rich in bioactive factors, including pro- and antioxidants. The objective was to investigate differences in pro- and antioxidants as well as oxidative markers in raw and heat-treated (HT) colostrum and in the blood of calves fed either raw or HT colostrum. Eleven colostrum samples (≥8 L) of Holstein cows were each divided into a raw or HT (60°C, 60 min) portion. Both treatments were stored for <24 h at 4°C and tube-fed in a randomized-paired design at 8.5% of body weight to 22 newborn female Holstein calves within 1 h after birth. Colostrum samples were obtained before feeding, and calf blood samples were taken immediately before feeding (0 h) and at 4, 8, and 24 h after feeding. All samples were analyzed for reactive oxygen and nitrogen species (RONS) and antioxidant potential (AOP), from which the oxidant status index (OSi) was calculated. In 0-, 4-, and 8-h plasma samples, targeted fatty acids (FA) were analyzed using liquid chromatography-mass spectrometry, and oxylipids and isoprostanes (IsoP) using liquid chromatography-tandem mass spectrometry. Results for RONS, AOP, and OSi were analyzed by mixed-effects ANOVA or mixed-effects repeated-measures ANOVA, for colostrum and calf blood samples, respectively, whereas FA, oxylipid, and IsoP were analyzed using false discovery rate-adjusted analysis of paired data. Compared with control, HT colostrum showed lower RONS [least squares means (LSM) 189, 95% confidence interval (95% CI): 159-219 vs. 262, 95% CI: 232-292) relative fluorescence units] and OSi (7.2, 95% CI: 6.0-8.3 vs. 10.0, 95% CI: 8.9-11.1), but AOP remained unchanged (26.7, 95% CI: 24.4-29.0 vs. 26.4, 95% CI: 24.1-28.7 Trolox equivalents/µL). Changes in colostrum oxidative markers due to heat treatment were minor. No changes in RONS, AOP, OSi, or oxidative markers were detected in calf plasma. In both groups of calves, plasma RONS activity declined considerably at all postfeeding time points compared with precolostral values, and AOP reached its maximum 8 to 24 h after feeding. Generally, oxylipid and IsoP plasma abundance reached nadirs at 8 h post-colostrum in both groups. Overall, effects due to heat treatment on redox balance of colostrum and newborn calves and on oxidative biomarkers were minimal. In this study, heat treatment of colostrum reduced RONS activity but did not lead to detectable changes in calf oxidative status overall. This indicates that there were only minor changes in colostral bioactive components that could alter newborn redox balance and markers of oxidative damage.

Effects of dietary branched-chain amino acid supplementation on serum and milk metabolome profiles in dairy cows during early lactation.

The 3 branched-chain AA (BCAA), Val, Leu, and Ile, are essential AA used by tissues as substrates for protein synthesis and energy generation. In addition, BCAA are also involved in modulating cell signaling pathways, such as nutrient sensing and insulin signaling. In our previous study, dietary BCAA supplementation was shown to improve protein synthesis and glucose homeostasis in transition cows. However, a more detailed understanding of the changes in metabolic pathways associated with an increased BCAA availability is desired to fine-tune nutritional supplementation strategies. Multiparous Holstein cows (n = 20) were enrolled 28 d before expected calving and assigned to either the BCAA treatment (n = 10) or the control group (n = 10). Cows assigned to BCAA were fed 550 g/d of rumen-protected BCAA mixed with 200 g/d of dry molasses from calving until 35 DIM, whereas the cows assigned to the control were fed only 200 g/d of dry molasses. Serum samples were collected on d 10 before expected calving, as well as on d 4 and d 21 postpartum. Milk samples were collected on d 14 postpartum. From a larger cohort, we selected 20 BCAA-supplemented cows with the greatest plasma urea nitrogen concentration, as an indicator for greater BCAA availability, for the metabolomics analysis herein. Serum and milk samples were subjected to a liquid chromatography-mass spectrometry-based assay, detecting and measuring the abundance of 241 serum and 211 milk metabolic features, respectively. Multivariable statistical analyses revealed that BCAA supplementation altered the metabolome profiles of both serum and milk samples. Increased abundance of serum phosphocholine and glutathione and of milk Val, Ile, and Leu, and decreased abundance of milk acyl-carnitines were associated with BCAA supplementation. Altered phosphocholine and glutathione abundances point to altered hepatic choline metabolism and antioxidant balance, respectively. Altered milk acyl-carnitine abundances suggest changes in mammary fatty acid metabolism. Dietary BCAA supplementation was associated with a range of alterations in serum and milk metabolome profiles, adding to our understanding of the role of BCAA availability in modulating dairy cow protein, lipid, and energy metabolism on a whole-body level and how it affects milk composition.

Ambidirectional cohort study on the agreement of ultrasonography and surgery in the identification of parathyroid pathology, and predictors of postoperative hypocalcemia in 47 dogs undergoing parathyroidectomy due to primary hyperparathyroidism.

OBJECTIVE: To investigate (1a) agreement of ultrasonographic, surgical, and histopathologic findings in the diagnosis of a neoplastic etiology underlying primary hyperparathyroidism (PHPT), (1b) the ability of ultrasonographically determined parathyroid gland size to distinguish between malignant (carcinoma) and non-malignant (hyperplasia, adenoma) pathology, and (2) variables associated with postoperative hypocalcemia in dogs undergoing surgical treatment of PHPT. STUDY DESIGN: Ambidirectional cohort study. ANIMALS: Forty-seven client owned dogs with PHPT (34 retrospective; 13 prospective). METHODS: Data were extracted from medical records. Method agreements were explored using Cohen's Kappa statistic. A receiver operating characteristic curve (ROC) was used to determine a cut-off separating parathyroid pathologies. Univariable and multivariable models assessed associations between postoperative hypocalcemia and potential risk factors. RESULTS: Agreement of ultrasound and surgery for number and side of affected glands was 31/47 (65.9%) and 34/47 (72.3%), respectively. In 37/47 (78.7%) cases, parathyroid tissue was correctly assessed as pathologic by the surgeon. An ultrasonographic cut-off of ≥8.0 mm (ROC AUC = 0.82) best distinguished malignant from benign pathologies. Dogs with a preoperative serum ionized calcium (iCa) concentration ≥1.75 mEq/L had 7.5 times greater odds of becoming hypocalcemic postoperatively. CONCLUSION: A fair agreement existed between ultrasonographic and surgical findings in dogs with PHPT. A parathyroid mass ≥8.0 mm on ultrasonographic examination was suggestive of malignancy, while dogs with a preoperative serum iCa concentration ≥1.75 mEq/L were at increased risk for postoperative hypocalcemia in this study. CLINICAL SIGNIFICANCE: This study supports the use of bilateral cervical surgical exploration to identify abnormal parathyroid glands for the treatment of PHPT.

The effect of ex vivo lipopolysaccharide stimulation and nutrient availability on transition cow innate immune cell AKT/mTOR pathway responsiveness.

Postpartum dairy cows experience a heightened inflammatory state coinciding with the time of greatest nutrient deficit. Nutrient availability is sensed on the cellular level by nutrient sensing kinases, such as the PI3K/AKT/mTOR (mTOR) pathway, a key orchestrator of immune cell activation and inflammatory balance. Our objective was to determine the responsiveness of this pathway to inflammatory stimulation with and without nutrient supplementation ex vivo. Blood samples were collected from Holstein cows (n = 14) at -42, -14, 7, 21, and 42 d relative to calving. Control samples and samples pretreated with a mixture of amino acids, glucose, and insulin (AAM) were stimulated with 100 ng/mL E. coli lipopolysaccharide (LPS; LPS, AAMLPS) or left unstimulated (control, AAM). After 1 h, ratios of mean fluorescence intensity for phosphorylated to total protein of AKT and mTORC1 substrates S6RP and 4EBP1 were analyzed in polymorphonuclear cells (PMN), and monocytes by flow cytometry. A separate aliquot was stimulated with LPS for 2 h and relative mRNA abundance of IL10, IL12A, IL12B, and TNFA in whole blood leukocytes from 10 cows was measured by reverse-transcription quantitative PCR. Repeated measures ANOVA was performed with fixed effects of time, treatment, and their interaction. Cells had different ratios of pathway proteins with PMN having the highest phosphorylation of AKT, S6RP, and 4EBP1. Stimulation with LPS consistently activated mTOR signaling in PMN regardless of nutrient supplementation except for postpartum 4EBP1, which increased in response to nutrients alone. In monocytes, AKT baseline phosphorylation was lower and activation could not be induced by either treatment, whereas activation of 4EBP1 responded to nutrient supplementation. Treatment with LPS increased phosphorylation of S6RP in both innate immune cell types. Nutrient supplementation increased baseline IL10 expression and decreased baseline as well as LPS-induced IL12B and TNFA expression. We conclude that the mTOR pathway in bovine innate immune cells can be differentially activated in response to inflammatory stimulation and nutrient supplementation in monocytes versus PMN. Effects of nutrient supplementation on cytokine mRNA abundance are likely specific to immune cell type.

Alopecia in Belgian Blue crossbred calves: a case series.

BACKGROUND: Alopecia is defined as the partial or complete absence of hair from areas of the body where it normally grows. Alopecia secondary to an infectious disease or parasitic infestation is commonly seen in cattle. It can also have metabolic causes, for example in newborn calves after a disease event such as diarrhoea. In the article, the investigation of a herd problem of acquired alopecia in Belgian Blue (BB) crossbred calves is described. CASE PRESENTATION: Several BB crossbred calves had presented with moderate to severe non-pruritic alopecia in a single small herd located in Southern Germany. The referring veterinarian had ruled out infectious causes, including parasitic infection and had supplemented calves with vitamins (vitamins A, B1, B2, B3, B5, B6, B7, B9, B12, C, and K3) orally. Results of the diagnostic workup at the Clinic for Ruminants are presented for three affected calves and findings from a farm visit are discussed. Because of these investigations, an additional four calves were brought to the referral clinic within the first week of life, and before onset of alopecia, in order to study the course of the condition; however, these calves never developed any signs of alopecia during their clinic stay. CONCLUSIONS: Because all other plausible differential diagnoses were ruled out during our investigation, we concluded that the documented alopecia was due to malabsorption of dietary fat and consecutive disruption of lipid metabolism leading to telogen or anagen effluvium. In this particular case, this was caused by a mixing error of milk replacer in conjunction with insufficiently tempered water. We conclude that nutritional, management or environmental factors alone can lead to moderate to severe alopecia in calves in the absence of a prior or concurrent disease event or infectious cause.

The effect of the transition period and postpartum body weight loss on macrophage infiltrates in bovine subcutaneous adipose tissue.

The transition period in dairy cattle is characterized by a rapid increase in metabolic demands due to the onset of lactation in parallel with a voluntary drop in dry matter intake, which slowly increases compared with the rapidly increasing milk production. The resulting deficit in energy intake is largely met by lipolysis of adipose tissue stores liberating nonesterified fatty acids (NEFA) as energy substrates to support milk production and for the synthesis of milk fat. Previous work in cattle and other species has documented an increase in adipose tissue macrophages (ATM) during periods of feed restriction and lipolysis; however, alterations in ATM during the transition period have only recently received interest because of the role that adipose tissue remodeling and inflammation may play during this time. The primary objective of this study was to evaluate the number of macrophages in subcutaneous adipose tissue from dairy cattle at multiple time points throughout the transition period. Secondary objectives included the evaluation for potential associations of ATM with changes in body condition score (BCS) and concentrations of circulating NEFA and ß-hydroxybutyrate. Holstein cows (n = 25) were enrolled in the study 28 d before expected parturition and fed the same prepartum and postpartum rations for ad libitum consumption. On d 10 before expected parturition, and on d 4 and 21 after calving, subcutaneous adipose tissue biopsies were taken from the paralumbar fossa and preserved in formalin. Paraffin-embedded sections of tissue from each of the 25 cows at the 3 time points (total of 75 samples) were labeled with an immunohistochemical marker of macrophages, ionized calcium-binding adapter molecule 1 (Iba1, also known as allograft inflammatory factor 1). The number of Iba1-positive cells, the number of crown-like structures, and the number of immunopositive aggregates were enumerated in each sample. The number of Iba1-positive macrophages increased by 60 and 93% on d 4 and 21, respectively, when compared with the average prepartum number. Additionally, cows that lost ≥0.50 points of BCS had a 57 and 52% higher number of ATM on d 4 and 21 postpartum, respectively, when compared with cows that lost ≤0.25 points of BCS. No association was found between ATM numbers and NEFA and ß-hydroxybutyrate concentrations or milk production in early lactation. Cows experiencing a more pronounced decrease in BCS postpartum had the largest increase in adipose tissue macrophage counts, suggesting that a greater degree of adipose tissue remodeling during the period of greatest nutrient deficit may be associated with higher numbers of ATM.

Comparison of three clinical scoring systems for Culicoides hypersensitivity in a herd of Icelandic horses.

BACKGROUND: Culicoides hypersensitivity (CH), an intensely pruritic and seasonal allergic dermatitis, is a common allergic disease affecting horses worldwide. Currently, there is no validated clinical scoring system for the quantification of clinical signs associated with CH. OBJECTIVES: To (i) determine the best cut-off point of three scoring systems, (ii) test the accuracy of each system when compared to the clinical diagnosis of an experienced veterinarian and (iii) assess agreement between systems. ANIMALS: Icelandic horses (n = 20); eight with CH and 12 unaffected, from a research herd receiving no treatments for allergic dermatitis. METHODS AND MATERIALS: Lesion scores were recorded biweekly from April until September with three clinical scoring systems (A, B and C) by a single observer initially blinded to CH status. Separate logistic regression analyses for each time point were used to determine appropriate cut-offs for CH classification. Spearman's rho and Cohen's kappa were calculated to analyze correlation of scores and agreement of CH categorization between systems, respectively. RESULTS: The best allergic cut-off scores for system A, B and C were determined to be three, eight and 12, respectively. For each system median areas under the curve (>0.85) were excellent and discriminatory ability for correctly classifying CH status was strong. Excellent correlation between scores for each system (Spearman's rho > 0.96) and excellent intersystem agreement for CH categorization (kappa ≥ 0.73) were found across scoring time points. CONCLUSION AND CLINICAL IMPORTANCE: Results support the use of these scoring systems as templates for the future standardization of a CH clinical scoring system.

A randomized blind placebo-controlled trial investigating the effects of photobiomodulation therapy (PBMT) on canine elbow osteoarthritis.

The effect of photobiomodulation therapy (PBMT) or sham light therapy on pain, nonsteroidal anti-inflammatory drug (NSAID) requirement, and lameness was studied in 20 dogs with naturally occurring elbow osteoarthritis. Dogs (n = 20) were randomly assigned to receive either PBMT (group PBMT; n = 11) 10 to 20 J/cm2 or a placebo treatment (sham light group S; n = 9) treatment 0 J/cm2, to both elbows for 6 weeks. Lameness score, pain score, and NSAID dose were recorded by blinded study personnel before and 7 to 10 days after last treatment. Reduction in NSAID dose occurred in 9/11 dogs in the PBMT group, and in 0/9 of group S dogs (P = 0.0003). There was greater improvement in lameness score post PMBT versus S therapy (P = 0.001). A greater reduction in pain score was detected in 9/11 parameters in group PBMT (P < 0.05). Regularly scheduled PBMT at 10 to 20 J/cm2 per joint for 6 weeks was successful in improving lameness and pain scores, and in lowering NSAID requirement in canine elbow osteoarthritis patients.

Essai clinique randomisé à double insu examinant les effets de la thérapie par photobiomodulation (PBMT) en comparaison à un placebo pour le traitement de l'ostéoarthrite du coude canin. Les effets de la thérapie par photobiomodulation, PBMT (anciennement thérapie au laser froid, thérapie au laser de basse énergie, ou LLLT, abréviation anglaise) ont été évalués et comparés à un placebo lumineux chez 20 chiens souffrant d'ostéoarthrite bilatérale du coude. Les chiens (n = 20) ont été assignés aléatoirement au groupe recevant le traitement au laser (PBMT; n = 11), ou à celui recevant le traitement placebo (S; n = 9). Les deux groupes ont été traités à double insu pendant 6 semaines, recevant soit 10 à 20 J/cm2 (groupe PBMT) ou 0J/cm2 (S) pendant 3 à 5 minutes sur chaque coude. Avant (pré) et 7 à 10 jours après chaque traitement (post), la fréquence d'administration et le dosage d'anti-inflammatoire non stéroïdien (AINS), le degré de boiterie, évalué par un clinicien, ainsi que le degré de confort selon l'index d'Helsinki pour la douleur chronique, évalué par le propriétaire, ont été notés. Une réduction du besoin en AINS a été possible chez 9 des 11 chiens du groupe PBMT, tandis qu'aucun chien du groupe S n'a pu réduire sa consommation de médicament (P = 0,0003). Les grades de boiterie se sont améliorés de façon plus marquée chez le groupe PBMT que chez le groupe S (P = 0,001). Il en va de même pour l'index d'Helsinki chez le groupe PBMT pour lequel les propriétaires ont remarqué une amélioration du confort de leur animal pour 9 des 11 paramètres évalués (P < 0,05). Ces données suggèrent qu'un traitement au laser pour 6 semaines à un dosage de 10 à 20 J/cm2 a un effet bénéfique pour les chiens atteints d'ostéoarthrite bilatérale du coude en améliorant leur niveau de boiterie et de confort en plus de diminuer leur besoin en AINS.(Traduit par Dre Lauri Jo Gamble).

Evaluation of plasma inflammatory cytokine concentrations in racing sled dogs.

In human athletes significant changes in cytokine concentrations secondary to exercise have been observed. This prospective study evaluated the effect of a multi-day stage sled dog race on plasma concentrations of monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-alpha (TNF-α), interleukin-2 (IL-2), interleukin-6 (IL-6), interleukin-8 (IL-8), and interleukin-10 (IL-10). Samples from 20 dogs were harvested prior to and on days 2 and 8 of an 8-day race. Exercise resulted in significantly decreased TNF-α and IL-8 as well as increases of MCP-1, IL-6, and IL-10 concentrations (P-value between 0.01 and < 0.0001 for all parameters). The proportion of values for IL-2 that were below the detection limit increased from 40% on day 0 to 75% on day 2 and decreased on day 8 to 40% (P = 0.04). Racing sled dogs show cytokine-concentration changes that are different from those in humans.

Évaluation des concentrations plasmatiques de cytokines inflammatoires chez des chiens de traîneau de course. Chez les athlètes humains, des changements importants des concentrations de cytokines secondaires à l'exercice ont été observés. Cette étude prospective a évalué l'effet d'une course de chiens de traîneau par étapes de plusieurs jours sur les concentrations plasmatiques des protéines-1 chimio-attractives des monocytes (MCP-1), du facteur-alpha nécrosant des tumeurs (TNF-α), d'interleukine-2 (IL-2), d'interleukine-6 (IL-6), d'interleukine-8 (IL-8) et d'interleukine-10 (IL-10). Des échantillons ont été prélevés sur 20 chiens avant la course et aux jours 2 et 8 d'une course de 8 jours. L'exercice a produit des valeurs significativement réduites de TNF-α et d'IL-8 ainsi qu'une hausse des concentrations de MCP-1, d'IL-6 et d'IL-10 (la valeur-P entre 0,01 et < 0,0001 pour tous les paramètres). La proportion des valeurs pour IL-2 qui étaient inférieures au seuil de détection a augmenté de 40 % le jour 0 à 75 % le jour 2 et a baissé le jour 8 à 40 % (P = 0,04). Les chiens de traîneau de course montrent des changements de la concentration des cytokines qui sont différents de ceux observés chez les humains.(Traduit par Isabelle Vallières).

[Outbreak of Staphylococcus aureus mastitis in a dairy herd: A case report]. / Gehäuftes Auftreten von Staphylococcus-aureus-Mastitiden in einer Milchviehherde: Ein Fallbericht.

The report describes an outbreak of Staphylococcus aureus mastitis in a dairy herd. The risk assessment included the analysis of data from milk recordings, bacteriological milk cultures, clinical mastitis records and influence of infected status on the culling risk of affected animals, as well as an evaluation of the milking routine. The milking routine and the treatment protocol of animals with Staphylococcus aureus infection were identified as possible risk factors. The implemented measures included changes to the milking routine, a different treatment protocol for infected animals, and culling and segregation of infected animals to reduce overall prevalence.

Hyperketonemia: A Marker of Disease, a Sign of a High-Producing Dairy Cow, or Both?

This review covers the history and nomenclature of ketosis, the source and use of ketones in transition cows, and the controversial role of hyperketonemia's association with health and production outcomes in dairy cows. With the goal of assisting veterinarians with on-farm diagnostic and treatment methods, the authors present current and evolving means of direct and indirect hyperketonemia detection as well as a summary of treatment modalities and their efficacy. They encourage veterinarians to include hyperketonemia testing as part of their routine physical examinations and contemplate day in milk at hyperketonemia diagnosis when designing treatment and management strategies.

Metabolic Factors at the Crossroads of Periparturient Immunity and Inflammation.

Periparturient cows have the highest risk for disease and culling in the adult dairy herd. This risk is compounded by the multiple physiological changes of metabolism and immune function occurring around calving that alter the cow's inflammatory response. In this article, the authors summarize the current knowledge on immunometabolism in the periparturient cow, discussing major changes in immune and metabolic function around parturition that will facilitate the assessment of periparturient cow management programs.

Case-control exercise challenge study on the pathogenesis of high serum gamma-glutamyl transferase activity in racehorses.

BACKGROUND: High serum γ-glutamyl-transferase (GGT) activity syndrome in racehorses has been associated with maladaption to exercise. Investigation of affected horses before and immediately after standard exercise may provide critical insight into the syndrome's pathophysiology. OBJECTIVES: To investigate blood biomarker changes in actively competing racehorses with high GGT activity associated with an exercise challenge. STUDY DESIGN: Case-control study. METHODS: High GGT case (age: 2-3 years) and normal GGT control (age: 2-7 years) pairs (3 Thoroughbred, 4 Standardbred pairs) at least 3 months into their training/racing season were included. Horses with a recent history of high GGT activity (≥50 IU/L) without additional biochemical evidence of liver disease were identified by veterinarians. Horses were tested again in the week prior to a planned exercise challenge to confirm persistent increases in GGT activity. Controls from the same stable with similar training/racing intensity and serum GGT activity ≤36 IU/L were matched with each case. Blood samples were obtained immediately before, 15 and 120 min after exercise. Pre-exercise serum samples were analysed for baseline select serum chemistries, selenium and vitamin E concentrations. Cortisol concentration and markers of oxidative status were measured in serum or plasma for all time points. Individual serum bile acid and coenzyme Q10 concentrations, plasma lipid mediator (fatty acids, oxylipids, isoprostanes) concentrations and targeted metabolomics analyses were performed using liquid chromatography-mass spectrometry. Serum viral PCR for equine hepaci- and parvovirus was performed in each animal. RESULTS: Cases had higher baseline concentrations of total glutathione, taurocholic acid, cortisol and cholesterol concentrations and higher or lower concentrations of specific oxylipid and isoprostane mediators, but there were no case-dependent changes after exercise. MAIN LIMITATIONS: Small sample size. CONCLUSIONS: Results indicated that glutathione metabolism was altered in high GGT horses. Enhanced glutathione recycling and mild cholestasis are possible explanations for the observed differences.

Evaluation of Fecal Egg Count Tests for Effective Control of Equine Intestinal Strongyles.

The American Association of Equine Practitioners strongly advocates evidence-based intestinal strongyle control in horses. It recommends targeted treatment of all heavy egg shedders (>500 eggs per gram (EPG) of feces), while the low shedders (0-200 EPG) are left untreated. As 50-75% of adult horses in a herd are low shedders, preventing them from unnecessary anthelmintic exposure is critical for tackling resistance. There are various fecal egg count (FEC) techniques with many modifications and variations in use, but none is identified as a gold standard. The hypothesis of the study was that the diagnostic performance of 12 commonly used quantitation methodologies (three techniques with four variants) differs. In this regard, method comparison studies were performed using polystyrene beads as proxy for intestinal strongyle eggs. Mini-FLOTAC-based variants had the lowest coefficient of variation (CV%) in bead recovery, whereas McMaster variants had the highest. All four variants of Mini-FLOTAC and the NaNO3 1.33 specific gravity variant of modified Wisconsin followed a linear fit with R2 > 0.95. In contrast, the bead standard replicates for modified McMaster variants dispersed from the regression curve, causing a lower R2. The Mini-FLOTAC method seems less influenced by the choice of floatation solution and has better repeatability parameters and linearity for bead standard recovery. For FEC tests with high R2 (>0.95) but that underestimated the true bead count, a correction factor (CF) was determined to estimate the true count. Finally, the validity of CF was analyzed for 5 tests with R2 > 0.95 to accurately quantify intestinal strongyle eggs from 40 different horses. Overall, this study identified FEC methodologies with the highest diagnostic performance. The limitations in standardizing routine FEC tests are highlighted, and the importance of equalization of FEC results is emphasized for promoting uniformity in the implementation of parasite control guidelines.

Pharmacological inhibition of the mTOR pathway alters phenotype and cytokine expression in bovine monocyte-derived dendritic cells.

Epidemiological studies have long demonstrated the association of nutrient status and immune dysfunction in dairy cows. Postpartum dairy cows experiencing a nutrient deficit show a propensity for increased inflammatory response, decreased pathogen clearance, and increased incidence of infectious disease. Studies in cows and other species show that the nutrient sensing mechanistic target of rapamycin (mTOR) signaling pathway could be one potential causal pathway connecting the deficit in nutrient availability and the heightened inflammatory response. Our objective was to investigate the effects of pharmacological mTOR pathway inhibition on phenotype and cytokine expression of bovine monocyte derived dendritic cells (moDC). We differentiated CD14+ monocytes from dairy cows (n = 14) into moDC in the presence or absence of first- or second-generation mTOR inhibitor rapamycin and PP242 (both 100 nM), respectively. On day seven cells were matured with E. coli lipopolysaccharide (LPS, 100 ng/mL) or left unstimulated to represent naïve moDC. Surface expression of CD14, CD40, CD80, and MHCII was measured via flow cytometry. We measured mRNA expression of IL10, IL12A, IL12B, and TNFα by rt-qPCR, and protein concentrations of IL-10 and TFN-α in cell culture supernatants with a bead-based multiplex assay. Cultures from ten cows successfully developed the moDC phenotype in culture without inhibitors, defined as increased surface expression of CD40, CD80, and MHCII compared with naïve moDC. Only data from these cows were considered for the results on effects of mTOR inhibitors. In naïve and mature moDC mTOR inhibition increased MHCII expression compared to controls. In mature moDC, in addition to MHCII, CD80 expression was increased compared with untreated LPS-stimulated controls. Expression of IL12A mRNA was upregulated in mature, mTOR inhibited moDC compared with untreated controls. In cell culture supernatants mTOR inhibition reduced IL-10 and increased TNF-α concentrations in naïve and mature moDCs compared with untreated controls. Overall rapamycin had a more consistent effect on altering phenotype and cytokine expression of moDC than PP242. In summary we observed an increased expression of co-stimulatory molecules and antigen presentation potential in mature moDC differentiated under mTOR inhibition, and a cytokine pattern that would potentially favor a Th1 type response. This study provides novel data indicating a role for mTOR signaling in bovine moDC phenotype and mediator profile. This proof-of-concept study demonstrates the role of the mTOR pathway in shaping the bovine immune response and may help to provide mechanistic insight and opportunities for modulation of the immune response during the nutrient deficit of early lactation.

Development of a bead-based multiplex assay to quantify bovine interleukin-10, tumor necrosis factor-α, and interferon-γ concentrations in plasma and cell culture supernatant.

The quantification of cytokines can improve our understanding of immune response and inflammation dynamics in dairy cows. Bead-based assays provide a sensitive, high-throughput platform, allowing for simultaneous quantification of multiple cytokines within a wide linear detection range. Our objective was to develop a multiplex bead-based assay using monoclonal antibodies for simultaneous quantification of bovine tumor necrosis factor (TNF)-α, IL-10, and IFN-γ in plasma and peripheral blood mononuclear cell (PBMC) culture supernatants. Recombinant cytokine standards produced in mammalian cells were used to determine the lower limit of detection and the linear detection range for each cytokine. The lower limit of detection was 110 pg/mL for IL-10, 95 pg/mL for TNF-α, and 20 pg/mL for IFN-γ. The linear quantification range was 110 to 241,000 pg/mL for IL-10, 95 to 620,000 pg/mL for TNF-α, and 20 to 130,000 pg/mL for IFN-γ. All 3 monoclonal capture and detection antibodies were specific for their respective cytokine analyte when using the recombinant IL-10, TNF-α, and IFN-γ standards. Intraassay and interassay coefficients of variation (CV) were <10% and <12%, respectively, for all analytes and samples matrices. Next, concentrations of native cytokines were determined in PBMC culture supernatants (n = 4) and in plasma from whole-blood samples (n = 6) with or without stimulation with Escherichia coli lipopolysaccharide or a mix of phorbol myristate acetate (PMA) and ionomycin. Peak concentrations of all 3 cytokines were secreted from PBMC after PMA/ionomycin stimulation (TNF-α, 8 h, range: 39,266-506,422 pg/mL; IL-10, 18 h, range: 15,770-63,415 pg/mL; IFN-γ 18 h, range: 189,977-492,659 pg/mL). In contrast, the highest concentrations in plasma from whole-blood stimulation were observed for IL-10 and TNF-α after LPS stimulation (TNF-α, 4 h, range: 1,764-13,460 pg/mL; IL-10, 24 h, range: 2,401-6,371 pg/mL), whereas PMA and ionomycin induced the highest secretion of IFN-γ (18 h, range: 53-20,215 pg/mL). In conclusion, the multiplex assay can quantify native IL-10, TNF-α, and IFN-γ across a broad concentration range in bovine plasma and cell culture supernatant, thereby providing a novel tool to evaluate inflammatory profiles in cattle and especially in dairy cows with inflammatory conditions. The existing multiplex assay can be expanded in the future by adding bead assays for additional bovine cytokines.

Investigating the pathogenesis of high-serum gamma-glutamyl transferase activity in Thoroughbred racehorses: A series of case-control studies.

BACKGROUND: High-serum γ-Glutamyl Transferase (GGT) activity has been associated with and thought to be a marker of maladaptation to training and possibly poor performance in racehorses, but the cause is unknown. OBJECTIVES: To investigate possible metabolic and infectious causes for the high GGT syndrome. STUDY DESIGN: Pilot case-control study and nested case-control study. METHODS: The case-control study in 2017 included 16 horses (8 cases and 8 controls with median [range] serum GGT 82 [74-148] and 22 [19-28] IU/L, respectively) from the same stable. In 2018, similar testing was performed in a nested case-control study that identified 27 case (serum GGT 50 ≥ IU/L)-control pairs from three stables for further testing. Serum liver chemistries, selenium measurements, viral PCR and metabolomics were performed. RESULTS: No differences were found in frequency of detection of viral RNA/DNA or copy numbers for equine hepacivirus (EqHV) and parvovirus-hepatitis (EqPV-H) between cases and controls. Mild increases in hepatocellular injury and cholestatic markers in case vs control horses suggested a degree of liver disease in a subset of cases. Metabolomic and individual bile acid testing showed differences in cases compared with controls, including increased abundance of pyroglutamic acid and taurine-conjugated bile acids, and reduced abundance of Vitamin B6. Selenium concentrations, although within or above the reference intervals, were also lower in case horses in both studies. MAIN LIMITATIONS: Observational study design did not allow us to make causal inferences. CONCLUSIONS: We conclude that high GGT syndrome is likely a complex metabolic disorder and that viral hepatitis was not identified as a cause for this syndrome in this cohort of racehorses. Our results support a contribution of oxidative stress and cholestasis in its pathophysiology.

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.