Bovine respiratory disease complex has a major impact on the health and economic performance of cattle. The annual cost of BRDC to the cattle industry in the United States exceeds $3 billion annually, and that disease is believed to be the most frequent cause of death of calves in feedlots.1 Bovine respiratory disease complex is a multifactorial disease involving multiple pathogenic agents.2–6 Mycoplasma bovis is increasingly identified as a contributor to BRDC.7–13 Mycoplasma bovis is recovered from up to 7% of healthy calves that have not recently been transported.12 Results of another study9 indicate the number of calves that shed M bovis at a stocker unit increases after 42 days, and such calves have decreased average daily gain.9
Antemortem diagnoses of BRDC are typically made on the basis of visual observation and rectal temperatures of animals.2 Visual observation has limitations for accurate detection of BRDC in calves, with an estimated sensitivity of 62% and specificity of 63%.14 Determination of CISs can vary substantially among veterinarians evaluating calves for detection of BRDC.15 Rectal temperature, heart rate, and respiratory rate may conflict for determination of a diagnosis of BRDC in calves, and such variables can vary with the time of day.16 Surface temperature, as determined by means of high-resolution thermographic imaging, has been evaluated as a variable for early identification of calves with pneumonia.17 Thermography is a noncontact method that can be applied to groups of calves and may have advantages over rectal temperature assessment. These methods of evaluation of BRDC in cattle have limited reliability and do not yield consistent results, which limits their value as predictors for determination of the onset or severity of disease.18
Few studies have been conducted to evaluate biological variables in calves during the development of BRDC.16 Combinations of changes in results of CBCs or serum biochemical analyses could provide information that would help characterize the inflammatory response associated with pneumonia and aid in the early detection of BRDC and prediction of disease severity. Experimental induction of disease provides temporal control for the study of these variables.
Assays for detection of biomarkers specific for cardiopulmonary diseases in cattle, such as cTnI, are available. Cardiac troponin I is a cardiac muscle protein that is a sensitive and specific biomarker for detection of cardiac muscle injury.19,20 Measurement of cTnI may be useful for evaluation of the severity of pneumonia in humans as an indicator of increased myocardial work.21 Increases in circulating concentrations of cTnI may be detected for patients with respiratory disease because of increased myocardial work associated with pulmonary hypertension and myocardial stress caused by hypoxia.22–26 Circulating cTnI concentrations are significantly increased in cattle with chronic suppurative pneumonia.27 The objectives of the study reported here were to determine associations of circulating cTnI concentrations, hematologic variables, serum biochemical analysis variables, and thermographically determined orbit and nasal planum surface temperatures with the onset and severity of pneumonia associated with M bovis in calves.
Materials and Methods
Animals—Clinically normal Holstein calves (n = 28; 27 bulls and 1 heifer) were used in this study. Study protocols were approved by the Kansas State University Animal Care and Use Committee.
All calves were obtained from a single source when they were approximately 2 months old and group-housed in pens. At the time of arrival at the Kansas State University research facility, the mean ± SD weight of all calves was 77.8 ± 8.0 kg. Criteria for inclusion in the study were negative results of M bovis culture of a nasal swab sample, negative results of a PCR assay for M bovis, low M bovis ELISA serum titer (the highest titer for a calf included in the study had an ELISA value that was 29.6% of the positive control value), and negative results of a bovine viral diarrhea virus real-time PCR assay of an ear notch sample. Of 38 calves tested, 28 met the inclusion criteria for the study. All calves included in the study had negative results for BVDV and M bovis determined by means of culture and PCR assay.
At the time of arrival, calves received 6.6 mg of ceftiofur crystalline free acid/kg at the base of the left ear. The diet for all calves consisted of free choice grass hay and a dairy calf complete starter grain rationa throughout the study period. The ration contained 18.0% crude protein, 3.2% crude fat, 1.0% calcium, and 0.5% phosphorus. During the first week after arrival, calves were fed 0.9 kg of starter ration/animal/day. Then, the ration was increased to 1.4 kg/animal/day for the remainder of the study.
After acclimatization for 14 days, calves were assigned by use of a random number generator to a treatment (M bovis challenge) group (n = 24) or a control (no challenge) group (n = 4). On study day 0, pneumonia was induced with a combination of nebulization and tracheal and bronchial inoculation of 1 × 109 CFUs to 5 × 109 CFUs of M bovis administered in 20 mL of PBS (0.9% NaCl) solution. All M bovis–challenged calves were housed together in a drylot pen (25 × 25 m). All control calves were housed approximately 200 m away from the treatment group calves for the duration of the study in a drylot pen of similar size. All research personnel that interacted with both treatment and control group calves changed protective clothing between pens. All calves were euthanized and necropsies were performed on day 14 of the study period. None of the calves were euthanized or died prior to the conclusion of the study.
CISs—Calves were evaluated twice daily to detect clinical signs of illness and determine a CIS by one of the investigators (BCF). Determination of CISs was started 3 days prior to M bovis challenge and continued until the evening prior to euthanasia. The CIS classifications were 1 (clinically normal), 2 (slight illness [mild lethargy and apparent signs of depression with or without cough]), 3 (moderate illness [moderate lethargy, apparent signs of depression, and labored breathing with or without cough]), and 4 (severe illness [moribund and unresponsive]).
Blood sample collection and analysis—Blood samples (20 mL) were collected by means of jugular venipuncture from each calf on study days 0, 7, and 14. Blood samples were transferred to serum collection tubes,b tubes containing lithium heparin,c and tubes containing EDTAd and placed on ice until processing. Blood samples were submitted to the clinical pathology service within 1 hour after collection for performance of hematologice and serum biochemicalf,g analyses. Hematologic analysis included determination of differential cell countsh by means of direct microscopic examination. Serum total protein was measured with a refractometer, and plasma fibrinogen concentration was determined by use of a heat precipitation method.
Blood samples collected into tubes containing lithium heparinc were used for analysis of cTnI concentrations with an assay that was validated for measurement of bovine cTnI.i Whole blood samples were centrifuged (2,350 × g for 10 minutes at 22°C), and plasma was harvested and stored at −80°C until analysis.
Infrared thermography—Infrared thermographic images were obtained for each calf while they were standing in a restraint chute. Surface thermographic images of the entire left orbit and muzzle were obtained with a high-definition thermography cameraj and the images were analyzed for calculation of maximum surface temperature of each evaluated anatomic site on days 0, 6, and 13. The left orbit was chosen owing to ease of data collection because of equipment position. All thermographic images were obtained at the same time of day in a sheltered area so that environmental conditions were standardized. Each image was captured from a distance of 1 m from the head, and the images were obtained at an angle perpendicular to each anatomic site of interest. Images were imported into an image analysis software program,k the margins of an orbit were traced, and the maximum temperature for that site was determined for each analysis day. Lines were drawn to horizontally divide the muzzle at the level of the ventral aspect of the nares to form a dorsal and ventral area of measurement. The 2 areas of the muzzle were traced separately and considered the dorsal aspect of the nasal planum and the ventral aspect of the nasal planum for analysis. The maximum temperature of each site was used for data analysis.
Necropsy and identification of pneumonia—On day 14, each calf was humanely euthanatized with a captive bolt followed immediately by pithing. Complete gross necropsies were performed, and tissue samples were obtained for microbial culture. Lungs of each calf were assessed by a veterinary pathologist (DAM) by use of a previously validated lung scoring method.28 With this method, the total percentage of lung consolidation was determined with the following formula: (0.053 • percentage consolidation of cranial segment of left cranial lobe) + (0.049 • percentage consolidation of caudal segment of left cranial lobe) + (0.319 • percentage consolidation of left caudal lobe) + (0.043 • percentage consolidation of accessory lobe) + (0.352 • percentage consolidation of right caudal lobe) + (0.061 • percentage consolidation of right middle lobe) + (0.060 percentage consolidation of caudal segment of right cranial lobe) + (0.063 • percentage consolidation of cranial segment of right cranial lobe). Calves were allocated to 2 categories on the basis of lung scores for statistical analysis. Calves with mild lung lesion severity included all calves with ≤ 10% lung consolidation. Calves with moderate lung lesion severity included all calves with > 10% of lung consolidation. A 10% cutoff value was determined on the basis of results of another study.29 Samples of cardiac papillary muscles and lung were obtained and fixed in 10% formalin for histologic examination. Aerobic and M bovis cultures were performed for representative samples of lungs obtained from each calf.
Statistical analysis—Statistical analysis was performed with commercially available software.l Data for treatment and control groups were combined for analyses of associations between respiratory disease severity and other examination and blood sample analysis variables. Variables were organized by sample collection day and pneumonia category (mild or moderate). Generalized linear models were used to determine potential associations between outcomes of interest and study day (0, 7, and 14), pneumonia category (≤ 10% and > 10% lung consolidation), and the potential interactions between these variables. A random effect was also included in each model to account for repeated collection of samples and measurements of calves. Values of P ≤ 0.05 were considered significant.
Results
CIS—The frequency of detection of CISs ≥ 2 (indicating detectable illness) increased during the study for the 24 calves in the M bovis challenge group (Figure 1). On day 0, all calves had a CIS of 1 (clinically normal), but by day 14, all 24 M bovis challenge group calves had at least 1 CIS ≥ 2. The CIS was 1 on all days for the 4 calves in the control group.
Results for 24 Holstein calves challenged with Mycoplasma bovis for experimental induction of pneumonia indicating the number of calves with CISs ≥ 2 on various study days. Data for control calves (n = 4) in the study are not included, and all calves in that group had CIS values of 1 (clinically normal) during the study. Calves were inoculated with M bovis on day 0 and euthanized on day 14. The CISs were determined twice daily (in the morning [AM] and evening [PM]).
Citation: American Journal of Veterinary Research 75, 2; 10.2460/ajvr.75.2.200
Plasma cTnI concentrations—Results of analysis of data for all 28 calves in the study indicated the plasma concentration of bovine cTnI had a significant (P = 0.04) positive association with blood sample collection day. Plasma cTnI concentrations were significantly higher on day 14 (least squares mean, 0.02 ng/mL) than they were on days 0 (least squares mean, 0 ng/mL) and 7 (least squares mean, 0 ng/mL). There was no significant (P < 0.05) association between severity of pneumonia and plasma cTnI concentration. At all times, plasma cTnI concentrations were within reported30–33 reference ranges (< 0.05 ng/mL to < 0.08 ng/mL).
Serum biochemical analyses, hematologic analyses, and CBCs—Multiple serum biochemical analysis variables were significantly associated with blood sample collection day and pneumonia severity category, and some significant interactions were found (Table 1). Variables significantly associated with blood sample collection day included anion gap and albumin, BUN, chloride, creatinine, globulin, glucose, bicarbonate, potassium, and phosphorus concentrations. Variables significantly associated with pneumonia severity category included anion gap and albumin, chloride, globulin, bicarbonate, phosphorus, and total protein concentrations. Serum biochemical analysis variables that had interactions between blood sample collection day and pneumonia severity category included albumin-to-globulin concentration ratio and ALP, calcium, and sodium concentrations. The only values with least square means outside of reference ranges included serum calcium and glucose concentrations and ALP activity. Serum calcium concentration for calves with moderately severe pneumonia on day 14 was within the reference range.34,35 At all other times, serum calcium concentrations were higher than the reference range for all calves. Serum glucose concentration and ALP activity were higher than reference ranges for all calves at all times.
Least squares mean ± SEM values of various serum biochemical analysis variables controlled for blood sample collection day, percentage of lung consolidation, or the interaction between sample collection day and percentage of lung consolidation for 2.5-month-old Holstein calves that were (n = 24) or were not (4) challenged with Mycoplasma bovis for induction of pneumonia.
| Variable | Effect | ||
|---|---|---|---|
| Albumin (g/L) | Sample collection day | — | < 0.001 |
| 0 | 3.52 (0.04)a | — | |
| 7 | 3.35 (0.04)b | — | |
| 14 | 3.18 (0.06)c | — | |
| Lung consolidation | — | < 0.01 | |
| < 10% | 3.44 (0.05)a | — | |
| ≥ 10% | 3.26 (0.03)b | — | |
| Anion gap (mmol/L) | Sample collection day | — | < 0.01 |
| 0 | 19.80 (0.44)a | — | |
| 7 | 17.70 (0.31)b | — | |
| 14 | 19.66 (0.40)a | ||
| Lung consolidation | — | < 0.01 | |
| < 10% | 19.70 (0.38)a | — | |
| ≥ 10% | 18.40 (0.26)b | — | |
| BUN (mg/dL) | Sample collection day | — | 0.04 |
| 0 | 12.60 (0.59)a | — | |
| 7 | 12.25 (0.39)a,b | — | |
| 14 | 14.00 (0.48)c | — | |
| Chloride (mmol/L) | Sample collection day | — | < 0.01 |
| 0 | 97.26 (0.30)a | — | |
| 7 | 97.18 (0.33)a | — | |
| 14 | 95.65 (0.39)b | — | |
| Lung consolidation | — | 0.05 | |
| < 10% | 97.11 (0.35)a | — | |
| ≥ 10% | 96.28 (0.24)b | — | |
| Creatinine (mg/dL) | Sample collection day | — | 0.02 |
| 0 | 0.70 (0.02)a | — | |
| 7 | 0.64 (0.02)b | — | |
| 14 | 0.64 (0.02)b | — | |
| Globulin (g/dL) | Sample collection day | — | < 0.01 |
| 0 | 2.97 (0.09)a | — | |
| 7 | 3.10 (0.09)a | — | |
| 14 | 3.46 (0.08)b | — | |
| Lung consolidation | — | 0.02 | |
| < 10% | 3.31 (0.09)a | — | |
| ≥ 10% | 3.04 (0.06)b | — | |
| Glucose (mg/dL) | Sample collection day | — | < 0.001 |
| 0 | 88.89 (1.88)a | — | |
| 7 | 76.64 (1.68)b | — | |
| 14 | 82.10 (1.59)c | — | |
| Bicarbonate (mmol/L) | Sample collection day | — | 0.01 |
| 0 | 28.22 (0.30)a | — | |
| 7 | 28.90 (0.34)a | — | |
| 14 | 27.11 (0.48)b | — | |
| Lung consolidation | — | 0.04 | |
| < 10% | 27.59 (0.39)a | — | |
| ≥ 10% | 28.56 (0.27)b | — | |
| Potassium (mmol/L) | Sample collection day | — | < 0.01 |
| 0 | 5.03 (0.09)a | — | |
| 7 | 4.77 (0.05)b | — | |
| 14 | 4.73 (0.05)b | — | |
| Phosphorus (mg/dL) | Sample collection day | — | < 0.01 |
| 0 | 7.71 (0.19)a | — | |
| 7 | 7.21 (0.17)b | — | |
| 14 | 6.89 (0.18)b | — | |
| Lung consolidation | — | < 0.01 | |
| < 10% | 7.61 (0.17)a | — | |
| ≥ 10% | 6.92 (0.12)b | — | |
| Protein (g/dL) | Lung consolidation | — | < 0.01 |
| < 10% | 6.75 (0.11)a | — | |
| ≥ 10% | 6.30 (0.07)b | — | |
| Albumin-to-globulin ratio | Interaction of sample collection day and lung consolidation | — | 0.03 |
| Day 0 and < 10% lung consolidation | 1.11 (0.06)a | — | |
| Day 0 and ≥ 10% lung consolidation | 1.28 (0.05)b | — | |
| Day 7 and < 10% lung consolidation | 1.08 (0.06)a | — | |
| Day 7 and ≥ 10% lung consolidation | 1.13 (0.04)a | — | |
| Day 14 and < 10% lung consolidation | 1.00 (0.05)a | — | |
| Day 14 and ≥ 10% lung consolidation | 0.90 (0.03)a | — | |
| ALP (U/L) | Interaction of sample collection day and lung consolidation | — | 0.05 |
| Day 0 and < 10% lung consolidation | 207.89 (7.55)a | — | |
| Day 0 and ≥ 10% lung consolidation | 196.11 (13.03)a | — | |
| Day 7 and < 10% lung consolidation | 195.56 (10.86)a | — | |
| Day 7 and ≥ 10% lung consolidation | 149.05 (10.03)b | — | |
| Day 14 and < 10% lung consolidation | 183.56 (8.99)a | — | |
| Day 14 and ≥ 10% lung consolidation | 105.16 (9.01)b | — | |
| Calcium (mg/dL) | Interaction of sample collection day and lung consolidation | — | 0.02 |
| Day 0 and < 10% lung consolidation | 10.64 (0.11)a | — | |
| Day 0 and ≥ 10% lung consolidation | 10.72 (0.08)a | — | |
| Day 7 and < 10% lung consolidation | 10.74 (0.12)a | — | |
| Day 7 and ≥ 10% lung consolidation | 10.55 (0.11)a | — | |
| Day 14 and < 10% lung consolidation | 10.57 (0.12)a | — | |
| Day 14 and ≥ 10% lung consolidation | 9.83 (0.10)b | — | |
| Sodium (mmol/L) | Interaction of sample collection day and lung consolidation | — | 0.03 |
| Day 0 and < 10% lung consolidation | 139.22 (0.36)a | — | |
| Day 0 and ≥ 10% lung consolidation | 138.79 (0.28)a | — | |
| Day 7 and < 10% lung consolidation | 139.33 (0.52)a | — | |
| Day 7 and ≥ 10% lung consolidation | 136.79 (0.29)b | — | |
| Day 14 and < 10% lung consolidation | 136.67 (0.31)a | — | |
| Day 14 and ≥ 10% lung consolidation | 136.47 (0.46)b | — |
— = Not determined.
Within a variable and effect, values with different superscript letters are significantly (P < 0.05) different (determined with a Student t test).
Multiple CBC variables were associated with blood sample collection day and pneumonia severity category or had a significant interaction (Table 2). Variables associated with sample collection day included band (nonsegmented) neutrophil concentration, basophil concentration, Hct, hemoglobin concentration, mean corpuscular hemoglobin concentration, and RBC concentration. Variables associated with pneumonia severity category included lymphocyte concentration, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, and RBC concentration. The only values with least square means outside of reference ranges included RBC and plasma fibrinogen concentrations. At all times, RBC concentrations were higher than the reference range.34,35 The only hematologic variable that had an interaction between study day and pneumonia severity category was plasma fibrinogen concentration. The plasma fibrinogen concentration was higher than the reference range on day 14 for calves with moderate severity pneumonia. Plasma fibrinogen concentrations of calves with mild and moderate severity of pneumonia were summarized (Figure 2). The mean plasma fibrinogen concentration for M bovis challenge group calves on day 14 was 0.7 g/dL (range, 0.3 to 1.0 g/dL; reference range, 0.1 to 0.6 g/dL). The mean fibrinogen concentration for the 4 control calves on day 14 was 0.3 g/dL (range, 0.3 to 0.5 g/dL).
Least squares mean ± SEM plasma fibrinogen concentrations for 28 calves (24 calves challenged with M bovis for experimental induction of pneumonia and 4 control calves) with a mild severity of pneumonia (< 10% lung consolidation; solid line; n = 9 calves) or moderate severity of pneumonia (≥ 10% lung consolidation; dashed line; 19) on study days 0, 7, and 14 (M bovis inoculation performed on day 0). *On day 14, the calves with a moderate severity of pneumonia had significantly (P ≤ 0.05) higher plasma fibrinogen concentration versus calves with a mild severity of pneumonia.
Citation: American Journal of Veterinary Research 75, 2; 10.2460/ajvr.75.2.200
Least squares mean ± SEM values of various CBC variables controlled for blood sample collection day, percentage of lung consolidation, or the interaction between sample collection day and percentage of lung consolidation for the calves in Table 1.
| Variable | Effect | Mean (SEM) | P value |
|---|---|---|---|
| Band neutrophils (X 103/μL) | Sample collection day | — | 0.02 |
| 0 | 0.01 (0.01)a | — | |
| 7 | 0.00 (0.01)a | — | |
| 14 | 0.09 (0.04)b | — | |
| Basophils (X 103/μL) | Sample collection day | — | 0.01 |
| 0 | 0.15 (0.02)a | — | |
| 7 | 0.12 (0.02)a,b | — | |
| 14 | 0.07 (0.01)b | — | |
| Lymphocytes (X 103/μL) | Lung consolidation | — | 0.02 |
| < 10% | 5.57 (0.27)a | — | |
| ≥ 10% | 4.80 (0.18)b | — | |
| Hct (%) | Sample collection day | — | < 0.01 |
| 0 | 34.21 (0.58)a | — | |
| 7 | 31.68 (0.56)b | — | |
| 14 | 31.47 (0.62)b | — | |
| Hemoglobin (g/dL) | Sample collection day | — | 0.01 |
| 0 | 12.26 (0.23)a | — | |
| 7 | 11.50 (0.22)b | — | |
| 14 | 11.32 (0.24)b | — | |
| MCHC (g/dL) | Sample collection day | — | 0.04 |
| 0 | 35.94 (0.15)a | — | |
| 7 | 36.44 (0.16)b | — | |
| 14 | 36.05 (0.14)a,b | — | |
| Lung consolidation | — | 0.00 | |
| < 10% | 36.48 (0.15)a | — | |
| ≥ 10% | 35.81 (0.10)b | — | |
| RBCs (X 106/μL) | Sample collection day | — | 0.02 |
| 0 | 10.37 (0.24)a | — | |
| 7 | 9.45 (0.24)b | — | |
| 14 | 9.60 (0.28)b | — | |
| Lung consolidation | — | 0.01 | |
| < 10% | 10.23 (0.33)a | — | |
| ≥ 10% | 9.38 (0.23)b | — | |
| MCH (pg) | Lung consolidation | — | 0.02 |
| < 10% | 11.85 (0.15)a | — | |
| ≥ 10% | 12.28 (0.10)b | — | |
| MCV (fL) | Lung consolidation | — | < 0.01 |
| < 10% | 32.41 (0.46)a | — | |
| ≥ 10% | 34.40 (0.32)b | — | |
| Fibrinogen (g/dL) | Interaction of sample collection day and lung consolidation | — | < 0.01 |
| Day 0 and < 10% lung consolidation | 0.42 (0.02)a | ||
| Day 0 and ≥ 10% lung consolidation | 0.44 (0.03)a | — | |
| Day 7 and < 10% lung consolidation | 0.43 (0.03)a | — | |
| Day 7 and ≥ 10% lung consolidation | 0.47 (0.03)a | — | |
| Day 14 and < 10% lung consolidation | 0.44 (0.04)a | — | |
| Day 14 and ≥ 10% lung consolidation | 0.74 (0.04)b | — |
MCH = Mean corpuscular hemoglobin. MCHC = Mean corpuscular hemoglobin concentration. MCV = Mean corpuscular volume.
See Table 1 for remainder of key.
Infrared thermography—No significant associations between study day or pneumonia severity category and orbit temperature were detected. Although no significant effect of study day was found for temperatures of dorsal and ventral aspects of the nasal planum, there was a significant association between pneumonia severity category and temperatures of dorsal and ventral aspects of the nasal planum. Summary least squares mean values for all days on which such data were obtained were used. Calves with ≥ 10% of lung consolidation had significantly lower surface temperature on the dorsal aspect of the nasal planum (18.7°C; P < 0.01) and ventral aspect of the nasal planum (23.1°C; P = 0.01) compared with values for calves with < 10% lung consolidation (dorsal aspect of the nasal planum temperature = 22.9°C; ventral aspect of the nasal planum temperature = 25.8°C).
Necropsy and histologic examination of tissue samples—The gross morphological features of pneumonia in M bovis challenge group calves were variable and depended on the stage of disease progression (including various severities of lung consolidation, parenchymal and airway exudate, abscessation, and pleural and interlobular septal fibrin). The total percentage of lung consolidation ranged from 0% to 54%. Of the 28 calves in the study, 9 (32%) had mild (< 10% lung consolidation) pneumonia (range, 0% to 8% lung consolidation; median, 4% lung consolidation; mean, 3.1% lung consolidation) and 19 (68%) had moderate (≥ 10% lung consolidation) pneumonia (range, 12% to 54% lung consolidation; median, 28% lung consolidation; mean, 27.7% lung consolidation). Most calves in the M bovis challenge group had a histologic diagnosis of lobular bronchopneumonia (n = 23) and bronchitis-bronchiolitis (21). No substantial gross or histologic abnormalities were detected in the 4 control group calves. No gross or histologic abnormalities in the cardiac muscles of any calves were detected.
Microbial culture—Mycoplasma bovis was isolated from lung samples obtained from 26 of the 28 calves in the study (24 M bovis–challenged and 2 control calves). Results of aerobic microbial cultures of the lung samples obtained from the 28 calves included no growth (n = 10), Pasteurella multocida (18), and beta-hemolytic Streptococcus sp (4). Lung samples obtained from 1 calf had negative M bovis culture results but positive P multocida and beta-hemolytic Streptococcus sp culture results. Pasteurella multocida was isolated without isolation of Streptococcus sp for 13 M bovis–challenged calves with ≥ 10% lung consolidation and 1 M bovis–challenged calf with < 10% lung consolidation. Pasteurella multocida was isolated in conjunction with Streptococcus sp for 3 M bovis–challenged calves with ≥10% lung consolidation and 1 control calf with < 10% lung consolidation.
Discussion
Although results of this study indicated various variables for calves changed, values of most variables remained within reference ranges and were not determined to be associated with the presence or severity of pneumonia in calves. Although plasma cTnI concentrations were higher on study day 14 than they were on days 0 and 7, this increase was detected for calves in both the M bovis challenge and control groups, and none of the plasma cTnI concentrations were higher than reported30–33 reference ranges. There was no histologic evidence of cardiac injury in any calf. Results of a retrospective study27 indicate circulating cTnI concentrations are significantly higher in cattle with chronic suppurative pneumonia than they are in healthy cattle.27 Cattle with chronic respiratory disease may have increased myocardial work as a result of pulmonary hypertension.25,26 However, the increase in plasma cTnI concentration detected in the present study seemed to be incidental and was not a unique finding for calves with pneumonia. It was unlikely that the acute pneumonia induced in calves in this study was of adequate duration and severity to induce cardiac alterations and high circulating cTnI concentrations that can accompany chronic pulmonary injury. Any prognostic value of circulating cTnI concentrations for identification of cattle with pneumonia may be restricted to animals with more severe or chronic pneumonia.
The only hematologic variable that had a significant interaction between study day and pneumonia severity category was plasma fibrinogen concentration. The circulating fibrinogen concentration was higher for calves with moderate severity pneumonia than it was for calves with mild pneumonia on day 14, and that value was higher than the reference range.35 Fibrinogen is an acute-phase protein that is synthesized in and released by the liver in response to immunologic and inflammatory stimuli, such as infection.36 Plasma fibrinogen concentrations in calves are not specific indicators of respiratory disease and can increase during stressful events such as abrupt weaning, transportation, and comingling.37–41 This may be a result of stimulation of the reticuloendothelial system or cytokine production. In this study, the high circulating fibrinogen concentration in calves with a moderate severity of pneumonia most likely indicated the higher severity of inflammation in these calves, compared with that in calves with mild pneumonia.
Although several serum biochemical analysis variables were affected by study day, none of these were predictive of the presence or severity of BRDC in calves. The only values that were higher than reference ranges were serum calcium and glucose concentrations and ALP activity, but values of these variables are typically high in young animals.19,42,43 The albumin-to-globulin concentration ratio was of limited value for prediction of BRDC because it was not outside of the reported34,35 reference ranges. Circulating glucose concentration is expected to be higher in calves than the reference range for adult cattle.19 The magnitude of changes in blood glucose concentrations was small and may have been caused by responses of individual animals to stress. The low serum ALP activity found for calves with the highest percentages of lung consolidation may have been attributable to decreased physical activity and an anabolic state associated with pneumonia.42 The serum ALP activity differences between groups were small, and values were likely clinically normal for animals of this age. Similarly, serum calcium concentration is expected to be higher in calves than it is in mature cows.43 Several other variables had significant associations with pneumonia severity category or study day, but the clinical importance of these findings may have been small because the values were within reference ranges during the study.
Various CBC variables had a significant association with study day or pneumonia severity category in this study. The only hematologic analysis variable that was higher than the reference range was RBC concentration. The differences between groups for this variable were small and likely clinically normal for animals of this age.44 These significant associations may have been attributable to physiologic differences among animals or changes related to inflammation or stress. However, these variables were of minimal value for prediction of BRDC because they were not specific for the disease and values were similar to reference ranges.34,35
High-resolution digital infrared thermography is useful for evaluation of animals with naturally occurring BRDC.17,45–47 In the present study, no significant changes in orbit temperatures were identified during progression of respiratory disease in calves; however, significant changes in nasal planum temperatures were detected. The low temperatures of the nasal planum for calves with moderate severity pneumonia may have been attributable to hypotension, hypovolemia, or a catabolic state.
Mycoplasma bovis was isolated from 2 of the control calves. There may not have been sufficient distance or biosecurity between the groups of calves. Those calves had no substantial gross or histologic changes in their lungs; therefore, disease attributable to M bovis infection was not identified. None of the gross or histologic lesions were specific for M bovis or P multocida infection. This may have been attributable to the early stage of disease in the calves. The calves may have been subclinically infected during transport and comingling.
Bovine respiratory disease complex is an important disease of cattle, and further research is warranted to determine the responses of individual animals to this disease. Circulating cTnI concentrations in calves increased during the study, although that variable was not predictive for early detection of BRDC. Infection with M bovis may not have resulted in a pneumonia as severe as that caused by other pathogens, including Mannheimia haemolytica, which may cause greater changes in serum cTnI concentrations. No substantial cardiac histologic lesions were identified in calves. None of the variables evaluated in this study were specific or accurate early indicators of BRDC in calves. Although circulating fibrinogen concentration is a nonspecific indicator of inflammation, results of this study suggested that plasma fibrinogen concentrations for calves with M bovis pneumonia could be useful in determination of the severity of pneumonia in individual animals.
ABBREVIATIONS
| ALP | Alkaline phosphatase |
| BRDC | Bovine respiratory disease complex |
| CIS | Clinical illness score |
| cTnI | Cardiac troponin I |
Land O'Lakes, Herd Maker Supreme B90, Shoreview, Minn.
10-mL serum collection tube, Becton, Dickinson & Co, Franklin Lakes, NJ.
1.3-mL tube containing lithium heparin, Sarstedt, Numbrecht, Germany.
2-mL tube containing EDTA, Becton, Dickinson & Co, Franklin Lakes, NJ.
Cell-Dyn 3700 hematology analyzer, Abbott Diagnostics, Abbott Park, Ill.
Cobas 6000, Roche Diagnostics, West Sussex, England.
Clinical Pathology Laboratory, College of Veterinary Medicine, Kansas State University, Manhattan, Kan.
Geisma-Wright quick stain, Wescor, Logan, Utah.
ADVIA Centaur TnI-Ultra, Siemens Medical Solutions Diagnostics, New York, NY.
ThermaCAM S65, FLIR Systems, Wilsonville, Ore.
ThermaCAM Researcher 2.8 Professional, FLIR Systems, Wilsonville, Ore.
JMP 7.0, SAS Institute Inc, Cary, NC.
References
1. Griffin D. Economic impact associated with respiratory disease in beef cattle. Vet Clin North Am Food Anim Pract 1997; 13: 367–377.
2. Apley M. Bovine respiratory disease: pathogenesis, clinical signs, and treatment in lightweight calves. Vet Clin North Am Food Anim Pract 2006; 22: 399–411.
3. Allen JW, Viel L, Bateman KG, et al. Serological titers to bovine herpesvirus 1, bovine viral diarrhea virus, parainfluenza 3 virus, bovine respiratory syncytial virus and Pasteurella haemolytica in feedlot calves with respiratory disease: associations with bacteriological and pulmonary cytological variables. Can J Vet Res 1992; 56: 281–288.
4. Confer AW. Update on bacterial pathogenesis in BRD. Anim Health Res Rev 2009; 10: 145–148.
5. Poulsen KP, McGuirk SM. Respiratory disease of the bovine neonate. Vet Clin North Am Food Anim Pract 2009; 25: 121–137.
6. Wiggins MC, Woolums AR, Hurley DJ, et al. The effect of various Mycoplasma bovis isolates on bovine leukocyte responses. Comp Immunol Microbiol Infect Dis 2011; 34: 49–54.
7. Maunsell FP, Donovan GA. Mycoplasma bovis infections in young calves. Vet Clin North Am Food Anim Pract 2009; 25: 139–177.
8. Caswell JL, Bateman KG, Cai HY, et al. Mycoplasma bovis in respiratory disease of feedlot cattle. Vet Clin North Am Food Anim Pract 2010; 26: 365–379.
9. Hanzlicek GA, White BJ, Renter DG, et al. Associations between the prevalence of Mollicutes and Mycoplasma bovis and health and performance in stocker calves. Vet Rec 2011; 168: 21.
10. Caswell JL, Archambault M. Mycoplasma bovis pneumonia in cattle. Anim Health Res Rev 2007; 8: 161–186.
11. Fulton RW, Blood KS, Panciera RJ, et al. Lung pathology and infectious agents in fatal feedlot pneumonias and relationship with mortality, disease onset, and treatments. J Vet Diagn Invest 2009; 21: 464–477.
12. Francoz D, Fortin M, Fecteau G, et al. Determination of Mycoplasma bovis susceptibilities against six antimicrobial agents using the E test method. Vet Microbiol 2005; 105: 57–64.
13. Maunsell FP, Woolums AR, Francoz D, et al. Mycoplasma bovis infections in cattle. J Vet Intern Med 2011; 25: 772–783.
14. White BJ, Renter DG. Bayesian estimation of the performance of using clinical observations and harvest lung lesions for diagnosing bovine respiratory disease in post-weaned beef calves. J Vet Diagn Invest 2009; 21: 446–453.
15. Amrine DE, White BJ, Larson R, et al. Precision and accuracy of clinical illness scores, compared with pulmonary consolidation scores, in Holstein calves with experimentally induced Mycoplasma bovis pneumonia. Am J Vet Res 2013; 74: 310–315.
16. Hanzlicek GA, White BJ, Mosier D, et al. Serial evaluation of physiologic, pathological, and behavioral changes related to disease progression of experimentally induced Mannheimia haemolytica pneumonia in postweaned calves. Am J Vet Res 2010; 71: 359–369.
17. Schaefer AL, Cook NJ, Church JS, et al. The use of infrared thermography as an early indicator of bovine respiratory disease complex in calves. Res Vet Sci 2007; 83: 376–384.
18. Griffin D, Chengappa MM, Kuszak J, et al. Bacterial pathogens of the bovine respiratory disease complex. Vet Clin North Am Food Anim Pract 2010; 26: 381–394.
19. O'Brien PJ. Cardiac troponin is the most effective translational safety biomarker for myocardial injury in cardiotoxicity. Toxicology 2008; 245: 206–218.
20. Hasić S, Kiseljaković E, Jadrić R, et al. Cardiac troponin I: the gold standard in acute myocardial infarction diagnosis. Bosn J Basic Med Sci 2003; 3: 41–44.
21. Moammar MQ, Ali MI, Mahmood NA, et al. Cardiac troponin I levels and alveolar-arterial oxygen gradient in patients with community-acquired pneumonia. Heart Lung Circ 2010; 19: 90–92.
22. Hamilton MA, Toner A, Cecconi M. Troponin in critically ill patients. Minerva Anestesiol 2012; 78: 1039–1045.
23. Heresi GA, Tang WH, Aytekin M, et al. Sensitive cardiac troponin I predicts outcomes in pulmonary arterial hypertesion. Eur Respir J 2012; 39: 939–944.
24. McLaughlin V. Managing pulmonary arterial hypertension and optimizing treatment options: prognosis of pulmonary artery hypertension. Am J Cardiol 2013; 111 (suppl 8): 10C–15C.
25. Angel KL, Tyler JW. Pulmonary hypertension and cardiac insufficiency in three cows with primary lung disease. J Vet Intern Med 1992; 6: 214–219.
26. Jubb TF, Malmo J. Cor pulmonale in an Angus cow. Aust Vet J 1989; 66: 257–259.
27. Mellanby RJ, Henry JP, Cash R, et al. Serum cardiac troponin I concentrations in cattle with cardiac and noncardiac disorders. J Vet Intern Med 2009; 23: 926–930.
28. Fajt VR, Apley MD, Roth JA, et al. The effects of danofloxacin and tilmicosin on neutrophil function and lung consolidation in beef heifer calves with induced Pasteurella (Mannheimia) haemolytica pneumonia. J Vet Pharmacol Ther 2003; 26: 173–179.
29. White BJ, Anderson DE, Renter DG, et al. Clinical, behavioral, and pulmonary changes following Mycoplasma bovis challenge in calves. Am J Vet Res 2012; 73: 490–497.
30. Fraser BC, Anderson DE, White BJ, et al. Assessment of a commercially Available point-of-care assay for the measurement of bovine cardiac troponin I concentration. Am J Vet Res 2013; 74: 870–873.
31. Varga A, Schober KE, Walker WL, et al. Validation of a commercially Available immunoassay for the measurement of bovine cardiac troponin I. J Vet Intern Med 2009; 23: 359–365.
32. Varga A, Schober KE, Holloman CH, et al. Correlation of serum cardiac troponin I and myocardial damage in cattle with monensin toxicosis. J Vet Intern Med 2009; 23: 1108–1116.
33. Peek SF, Apple FS, Murakami MA, et al. Cardiac isoenzymes in healthy Holstein calves and calves with experimentally induced endotoxemia. Can J Vet Res 2008; 72: 356–361.
34. Mohri M, Sharifi K, Eidi S. Hematology and serum biochemistry of Holstein dairy calves: age related changes and comparison with blood composition in adults. Res Vet Sci 2007; 83: 30–39.
35. Knowles TG, Edwards JE, Bazeley KJ, et al. Changes in the blood biochemical and haematological profile of neonatal calves with age. Vet Rec 2000; 147: 593–598.
36. Baumann H, Gauldie J. The acute phase response. Immunol Today 1994; 15: 74–80.
37. Holland BP, Step DL, Burciaga-Robles LO, et al. Effectiveness of sorting calves with high risk of developing bovine respiratory disease on the basis of serum haptoglobin concentration at the time of arrival at a feedlot. Am J Vet Res 2011; 72: 1349–1360.
38. Berry BA, Confer AW, Krehbiel CR, et al. Effects of dietary energy and starch concentration for newly received feedlot calves: II. Acute-phase protein response. J Anim Sci 2004; 82: 845–850.
39. Hickey MC, Drennan M, Earley B. The effect of abrupt weaning of suckler calves on the plasma concentrations of cortisol, catecholamines, leukocytes, acute phase proteins and in vitro interferon-gamma production. J Anim Sci 2003; 81: 2847–2855.
40. Arthington JD, Eicher SD, Kunkle WE, et al. Effect of transportation and commingling on the acute-phase protein response, growth, and feed intake of newly weaned beef calves. J Anim Sci 2003; 81: 1120–1125.
41. Carter JN, Meredith GL, Montelongo M, et al. Relationship of vitamin E supplementation and antimicrobial treatment with acute-phase protein responses in cattle affected by naturally acquired respiratory tract disease. Am J Vet Res 2002; 63: 1111–1117.
42. Thompson JC, Pauli JV. Colostral transfer of gamma glutamyl transpeptidase in calves. N Z Vet J 1981; 29: 223–226.
43. Szenci O, Chew BP, Bajcsy AC, et al. Total and ionized calcium in parturient dairy cows and their calves. J Dairy Sci 1994; 77: 1100–1105.
44. Brun-Hansen HC, Kampen AH, Lund A. Hematologic values in calves during the first 6 months of life. Vet Clin Pathol 2006; 35: 182–187.
45. Stewart M, Webster JR, Stafford KJ, et al. Effects of an epinephrine infusion on eye temperature and heart rate variability in bull calves. J Dairy Sci 2010; 93: 5252–5257.
46. Stewart M, Verkerk GA, Stafford KJ, et al. Noninvasive assessment of autonomic activity for evaluation of pain in calves, using surgical castration as a model. J Dairy Sci 2010; 93: 3602–3609.
47. Stewart M, Webster JR, Verkerk GA, et al. Non-invasive measurement of stress in dairy cows using infrared thermography. Physiol Behav 2007; 92: 520–525.
