Arrival cortisol measurement in veal calves and its association with body weight, protein fractions, animal health and performance.

Male dairy calves are exposed to an accumulation of transport, social and environmental stressors while transferred to fattening units. As a consequence, calves show high cortisol concentrations upon arrival at the veal facility. Whether cortisol levels as measured on arrival can be associated with animal health, welfare and production results is currently unknown. The first objective of this prospective cohort study was to determine possible associations of arrival serum cortisol concentration with health and production variables of veal calves and other arrival predictors like body weight and γ-globulin concentration. The second aim was to investigate potential clustering of arrival risk factors in veal calves for developing bovine respiratory disease (BRD) based on arrival body weight, serum cortisol concentration, total protein and protein fractions. In total, 105 male Holstein calves from two consecutive production cycles in a single, commercial white veal farm were blood sampled directly at arrival on the farm to determine serum cortisol, total protein and protein fractions. All calves were weighed the day after arrival and clinical signs, average daily weight gain (ADG) and carcass weight were collected. Also, all calves of both production cohorts were repeatedly examined by thoracic ultrasonography at the onset of group respiratory disease symptoms (2-3 weeks after arrival) and four weeks later. Linear and logistic mixed models together with k-means clustering were used for statistical analyses. Calves showed on average high, but individually variable serum cortisol concentrations (mean value = 96.6 ng/mL ± standard deviation (SD) = 48.8; Range (R) = 50.0-317 ng/mL). Arrival cortisol, body weight and γ-globulin content were not significantly associated. Serum cortisol and albumin concentrations at arrival were associated with chronic, unresponsive pneumonia. For each increase of serum cortisol concentration by 10 ng/mL, the odds for lung consolidation of ≥1 cm in depth at the second ultrasonography increased, odds ratio (OR) = 1.03 (95 % confidence interval (CI) = 1.01-1.06; P < 0.050). For every decrease in serum albumin concentration by 1g/L, the OR for developing pneumonia was 1.23 (95 % CI= 1.04-1.46; P < 0.015). Additionally, two clusters of calves were identified based upon arrival status: a low risk cluster with below average stress-induced cortisol values, above average body weight, no acute phase response (APR) and no failure of passive immune transfer (FPIT), and a high risk cluster with above average cortisol values, below average body weight, APR and FPIT. High risk calves had higher odds for developing clinical BRD (OR= 3.88 (95 % CI=1.20-12.53; P < 0.020)) and lung consolidation ≥1 cm in depth at week 6-7 after arrival (OR= 3.93 (95 % CI=1.34-11.53; P < 0.013)). Avoiding high cortisol levels upon arrival of calves is important for animal welfare, but also for reduction of the prevalence of chronic, unresponsive pneumonia and the associated need for (repeated) antimicrobial treatment and production losses.

Associations of barn air quality parameters with ultrasonographic lung lesions, airway inflammation and infection in group-housed calves.

Barn climate is believed to play a major role in the bovine respiratory disease complex. However, the exact air quality parameters associated with (sub)clinical pneumonia or airway inflammation in calves are currently unknown. The objective of this cross-sectional study was to assess associations of air quality parameters with clinical signs, lung consolidation, pulmonary inflammation and infection in group-housed calves. In total, 60 beef and dairy farms were visited from January to April 2017 and 428 calves sampled. Measured air quality parameters included continuous 24-h measurements of ammonia concentration, relative humidity and temperature and punctual measurements of air velocity, ammonia, CO2 and bacterial air load. Calf sampling consisted of clinical examination, thoracic ultrasonography and broncho-alveolar lavage sampling for bacteriological and cytological analysis of broncho-alveolar lavage fluid (BALf). Average air temperature was 14.2 °C (standard deviation (SD) 4.4, range 5.5-23.9) and relative humidity 68.8 % (SD 8.9, range 52.2-91.6). Average ammonia concentration was 1.7 ppm (SD 0.9, range 0-10.0). Lung consolidations of ≥1 cm, ≥3 cm and ≥6 cm in depth were present in 41.1 % (176/428), 27.1 % (116/428) and 16.1 % (69/428) of the calves, respectively. Average pen temperature was positively associated with consolidations of ≥1 cm (P = 0.005), ≥3 cm (P = 0.002) and ≥6 cm (P < 0.01). Ammonia exposure, in hours>4 ppm, was associated with lung consolidation ≥1 cm (odds ratio (OR) = 1.73; confidence interval (CI) = 1.02-3.07; P = 0.04). Ammonia concentration was positively associated with BALf epithelial cell percentage (P = 0.01). Air velocity >0.8 m/s was associated with increased odds of lung consolidation of ≥3 cm (OR = 6.8; CI = 1.2-38.5; P = 0.04) and ≥6 cm (OR = 15.9; CI = 1.2-200.0; P = 0.03). The prevalence of lung consolidations ≥1 cm was higher in the draught (81.8 %; P = 0.0092) and warm, dry and ammonia accumulation clusters (54.2 %; P = 0.02) compared to the presumably normal cluster (31.6 %). In addition, in the warm, dry and ammonia cluster the prevalence of lung consolidations ≥3 cm (38.1 %; P = 0.04) and ≥6 cm (31.4 %; P = 0.01) in depth were higher compared to the presumably normal climate cluster (18.2 % and 9.1 %, respectively). Of all frequently measured indoor air quality parameters, only average temperature, ammonia concentration and air velocity were associated with pneumonia and might therefore be preferable for cost-effective evaluation of calf barn climate.

Effects of omega-3 fatty acids on immune, health and growth variables in veal calves.

Under the present intensive rearing conditions, calves face a series of stressors and multiple pathogens often necessitating antimicrobial use. Multiple feed additives are currently explored for their ability to prevent disease and limit the use of antimicrobials. Supplementation of the polyunsaturated long chain n-3 fatty acids eicosapentaenoic (EPA) and docohexaenoic (DHA) from marine origin has been proposed as a strategy to improve immune function and prevent excessive inflammation reactions. The aim of this randomized clinical trial was to explore the effects of n-3 fatty acids (PUFAs) used as feed supplement on health, production and immune variables in a veal calf setting. One hundred-seventy calves were randomly assigned to 3 treatment groups: microalgae (MA, n = 57, 2.5 g DHA/animal/day), fish oil (FO, n = 57, 2.5 g EPA + DHA/animal/day)] and a control group (CON, n = 56). Average daily gain (ADG), bodyweight at 12 weeks on feed and slaughter weight were determined. Health monitoring consisted of recording of clinical signs and repeated thoracic ultrasonography. After 5, 8 and 11 weeks of supplementation, the function of neutrophils, monocytes and peripheral blood mononuclear cells (PBMCs) was evaluated ex vivo by measuring reactive oxygen species (ROS) production by neutrophils and monocytes and proliferation of and cytokine release by PBMCs. Under the field conditions of this study, dietary supplementation of MA and FO showed very limited immunomodulatory effects. Feeding MA led to increased ROS production by neutrophils, Estimate (E) = 0.38, Standard Error (SE) = 0.14; P < 0.05, compared to the control calves after 5 weeks of in-feed supplementation. FO reduced IL-6 secretion E= -0.29, SE= 0.11; P < 0.05 compared to MA treated animals after 11 weeks on feed. Health and production variables were unaffected by treatments. The doses of EPA and DHA used in this study did not cause immunomodulatory changes in highly stressed calves to such an extent that this led to better health or growth of animals.

Effects of glycerol-esters of saturated short- and medium chain fatty acids on immune, health and growth variables in veal calves.

In veal and dairy beef production systems, Holstein bull calves experience many stressors and excessive pathogen exposure, necessitating the use of antimicrobials for welfare and production reasons. The aim of this randomized clinical trial was to explore the effects of esterified fatty acids used as feed supplement on health, production and immune variables in veal calves. Different glycerol-esters of fatty acids were used: short chain fatty acid (SCFA)-based glycerol-mono- (C4) and tributyrate (C4), and medium chain fatty acid (MCFA)-based glycerol-monocaprylate/monocaprinate (C8/C10) and glycerol-monolaurate (C12) in two different doses. One hundred sixty eight calves (2-to 4-week-old) were randomly assigned to 6 treatment groups; tributyrate (0.5 g/animal/day); monobutyrate (1 g/animal/day); low C8/C10 (7 g/animal/day) and high C8/C10 (10 g/animal/day); low C12 (4 g/animal/day) and high C12 (6 g/animal/day) and a control group (CON). Duration of in-feed supplementation was 14 weeks. Average daily gain, bodyweight at 14 weeks on feed and slaughter weight were determined. Health monitoring consisted of clinical signs and repeated thoracic ultrasonography. After 4, 8 and 12 weeks of supplementation, the function of neutrophils, monocytes and peripheral blood mononuclear cells (PBMCs) was evaluated ex vivo by measuring reactive oxygen species (ROS) production by neutrophils and monocytes, proliferation of and cytokine release by PBMCs. Study power was based upon ROS production by neutrophils and treatment groups were too limited to detect significant differences in growth and health variables. Glycerol-ester supplementation resulted in different effects on immune cell function, depending on the type and dose of the glycerol-ester as well as duration of supplementation. Our main findings were increased secretion of interleukin IL-17A by PBMCs at 4 weeks of feed supplementation in high C8/C10 (P< 0.01), low C12 (P < 0.01) and monobutyrate (P< 0.01) groups, combined with decreased ROS production in neutrophils (P < 0.001) and monocytes (P < 0.05) in the high C8/C10 and monocytes (P < 0.05) in low C12 groups compared to the control animals. After 12 weeks on feed, ROS production by neutrophils (P < 0.001) and monocytes (P < 0.01) of monobutyrate and by monocytes (P < 0.01) of tributyrate groups was decreased compared to control calves. In summary, supplementation of glycerol-esters of MCFAs resulted in immune-modulatory effects, which did not manifest themselves in improved health and growth of calves under the conditions and limitations of this study. Especially doses of high C8/C10 and low C12 show potential to promote an early, robust pro-inflammatory response with diminished ROS production. This might be beneficial for clearance of pathogens in young calves in periods of stress and high pathogen load.

Comparison of bronchoalveolar lavage fluid bacteriology and cytology in calves classified based on combined clinical scoring and lung ultrasonography.

Respiratory tract infections are the leading cause of antimicrobial use in calves. Combining clinical examination and lung ultrasonography allows on-farm classification of calves as healthy or suffering from an upper respiratory tract infection (URTI), subclinical or clinical pneumonia. This might help to improve targeted antimicrobial therapy, restricting treatment to pneumonic cases. However, to what extent these diagnostic categories coincide with expected bacteriological and cytological bronchoalveolar lavage fluid (BALf) characteristics is currently unknown. The objective of this study was therefore to compare BALf bacteriology and cytology between healthy calves and calves with URTI, subclinical and clinical pneumonia. The hypothesis was that calves with subclinical and clinical pneumonia would have higher quantitative bacterial counts, bacterial isolation rates and neutrophil counts than URTIs or healthy animals. A cross-sectional study was performed on 305 indoor group-housed dairy and beef calves, from 62 farms. Calves were classified by combining clinical examination and lung ultrasonography. Clinical respiratory disease was defined using the Wisconsin score card and the Healthy Criterion (HC). The HC classified calves as clinically ill if at least one clinical sign was present. Ultrasonographic lung consolidation with a depth of ≥1 cm was considered indicative for pneumonia. Cytology and bacteriology were performed on BALf sampled by non-endoscopic bronchoalveolar lavage. Calves with clinical pneumonia were further subdivided based on culture result and presence of neutrophils phagocytosing bacteria. Combined lung ultrasonography and clinical examination (HC) classified 25.9 % (79/305) of the calves as healthy, 33.1 % (101/305) as URTI, 10.2 % (31/305) as subclinical and 30.8 % (94/305) as clinical pneumonia. Bacterial isolation rates and quantitative BALf culture results did not differ between groups. Calves with clinical pneumonia and neutrophil phagocytosis showed a significantly higher BALf neutrophil percentage compared to healthy calves (59.0 % vs. 37.7 % in healthy calves, P =.03). Inversely, lymphocyte percentage was lower in these calves (1.8 % vs. 5.3 % in healthy calves, P = .003). Classification of calves using lung ultrasonography and clinical scoring did not correspond with BALf bacteriology and cytology findings, as extrapolated from human and companion animal medicine. Under the current housing conditions of this study high rates of non-infectious airway inflammation or airway colonization by opportunistic pathogens, rather than infection might explain this. Isolation of respiratory pathogens from calves with various signs of respiratory disease or ultrasonographic lesions should be interpreted carefully. Of all cytological features, phagocytosis by neutrophils in BALf might be a useful criterion supporting the diagnosis of bacterial respiratory tract infection.

Factors associated with lung cytology as obtained by non-endoscopic broncho-alveolar lavage in group-housed calves.

BACKGROUND: Respiratory infections are the main indication for antimicrobial use in calves. As in humans and horses, studying inflammation of the deep airways by lung cytology raises the possibility of preventing respiratory disease and targeting its treatment in the future. Whether lung cytology findings coincide with clinical signs and lung ultrasonographic findings is currently unknown. Therefore, the objective of the present study was to determine the association of lung cytology with clinical signs, lung consolidation and broncho-alveolar lavage fluid (BALf) characteristics (including bacteriology). A total of 352 indoor group-housed calves aged between 1 and 6 months from 62 conveniently selected commercial herds were included in this cross-sectional study. Clinical examination, thoracic ultrasound and bacteriology and cytology on non-endoscopic broncho-alveolar lavage (nBAL) samples were performed. RESULTS: Pneumonia, defined as presence of ultrasonographic lung consolidations ≥1 cm in depth, affected 42.4% of the calves. Mean BALf neutrophil percentage was 36.6% (SD 23.8; R 0-97.4) and only a positive induced tracheal cough reflex (P = 0.04), standing posture (P = 0.03) increased breathing rate (P = 0.02) and isolation of Pasteurella multocida (P = 0.005), were associated with increased neutrophil percentage. No significant associations between lung ultrasonographic findings and cytology results were present, except for presence of basophils in BALf and consolidation of > 3 cm in depth (OR = 2.6; CI = 1.2-5.6; P = 0.01). Abnormal lung sounds were associated with detection of eosinophils in BALf (OR = 2.8; CI = 1.0-8.1; P = 0.05). Total nucleated cell count (TNCC) (P <  0.001) was positively and macrophage percentage (P = 0.02) negatively associated with volume of lavage fluid recovered. Macroscopic blood staining of BALf increased TNCC (P = 0.002) and lymphocyte percentage (P = 0.001). CONCLUSIONS: Only a limited number of clinical signs and ultrasonographic findings were associated with nBAL cytology. BALf cytology offers additional and distinct information in calves aiding in detection and prevention of respiratory conditions. In this population, selected from herds not reporting any recent respiratory illness, a high number of calves had ultrasonographic lung consolidation and high neutrophil percentage in BALf, suggesting that subclinical disease presentations frequently occur.

Randomized field trial on the effects of body weight and short transport on stress and immune variables in 2- to 4-week-old dairy calves.

BACKGROUND: Whether underweight calves respond differently to transport stress, enhancing their disease risk, is currently unknown. OBJECTIVE: To determine the effects of low body weight and transport stress on immune variables. ANIMALS: Twenty-one 2- to 4-week-old male Holstein calves, housed on a commercial farm. METHODS: Randomized clinical trial. Full factorial design with 4 treatment groups: low body weight (≤46 kg)/no transport (LOWCON); low body weight/transport (LOWTRANS); normal body weight (>46 kg)/no transport (NORMCON), and normal body weight/transport (NORMTRANS). Transport duration was 2 hours. RESULTS: Transport significantly increased serum cortisol concentration (77.8 µg/mL; 95% confidence interval [CI], 37.8-131.6; P < .001), interleukin (IL)-17A (344.9 pg/mL; 95% CI, 32.2-556.5; P = .04), and tumor necrosis factor-α (TNF-α) (218.2 pg/mL; 95% CI, 32.5-368.3; P = .03) production after lipopolysaccharide (LPS) stimulation. Body weight did not affect any of the studied variables. However, the interaction of transport and body weight was significant. LOWTRANS calves showed increased monocyte count (2.0 × 109 /L; 95% CI, 0.6-4.2; P < .05) and interleukin IL-17A production (106.0 pg/mL; 95% CI, 4.2-306.9; P = .03) compared to normal weight calves and increased TNF-α production (275.6 pg/mL; 95% CI, 2.6-463.0; P = .02) compared to LOWCON calves in unstimulated peripheral blood mononuclear cells (PBMCs) after transport. CONCLUSION AND CLINICAL IMPORTANCE: These findings contribute to our understanding of increased disease susceptibility of underweight calves when transported. Gamma globulin concentration was identified as important interfering factor in studies on immune variables in neonatal calves.