Immunoglobulin containing cells in the bronchopulmonary system of non-pneumonic and pneumonic calves

In an examination of the immunoglobulin containing cells in the respiratory mucosa of three-month-old non-pneumonic calves, IgA was found to be the predominant plasma cell type. In calves with cuffing pneumonia there was a marked increase in the plasma cell population in the bronchial wall and IgG1 cells became the major cell type. Similarly in calves with an exudative type of pneumonia IgG1 plasma cells were the significant cell type in the respiratory mucosa but the total number of plasma cells was similar to that found in the non-pneumonic animals. The immunohistochemical results suggest that IgA and, to a lesser extent, IgM are both transported across the mucous membrane via bronchial epithelial secretory cells and submucosal glands.     

Changes in blood and bronchoalveolar lavage fluid components in calves with experimentally induced pneumonic pasteurellosis

Pneumonic pasteurellosis was experimentally induced in calves by inoculation of 5 x 10(8) Pasteurella haemolytica organisms into the right diaphragmatic lung lobe. Blood and bronchoalveolar lavage fluid samples were obtained prior to inoculation and at postinoculation hour (PIH) 2, 4, and 6. Calves developed acute lung injury, characteristic of pneumonic pasteurellosis. Lesions were found only in the right diaphragmatic lobe. By PIH 4, significant (P less than 0.01) increases were detected in lavage fluid total cell count, neutrophil count, total protein and albumin concentrations, and alkaline phosphatase (ALP) and lactic dehydrogenase (LD) activities. Myeloperoxidase and elastase activities did not increase. Neutrophil depletion ameliorated the lung lesions and prevented the increase in lavage fluid cell count, total protein, and albumin concentrations and ALP and LD activities. Treatment with the iron chelator, deferoxamine mesylatehydroxyethyl starch, attenuated the increase in total protein and albumin concentrations and ALP and LD activities at PID 4, but not PIH 6. Treatment with a neutrophil function inhibitor, pentoxifylline, prevented the increase in lavage fluid neutrophil numbers, but accentuated the increase in total protein and albumin concentrations, and ALP, LD, myeloperoxidase, and elastase activities.     

Pathological and microbiological studies on pneumonic lungs from Danish calves

During 1 year, the association between microbiological and pathological findings in 72 lungs from calves submitted to the Danish Veterinary Laboratory for diagnostic purposes was studied. All cases were evaluated pathologically and bacteriologically, whereas only 68 cases were examined for the presence of bovine respiratory syncytial virus (BRSV), parainfluenza-3 virus (PI-3 virus) and bovine coronavirus, 62 cases for bovine viral diarrhoea virus (BVD), 45 cases for bovine adenovirus and 51 cases for mycoplasmas. Based on histopathological examination, the cases were diagnosed as fibrinous and/or necrotizing bronchopneumonia, suppurative bronchopneumonia, embolic pneumonia and others. The diagnoses were based on the dominating and most severe lesions in each lung. Haemophilus somnus, Pasteurella multocida, Actinomyces pyogenes, P. haemolytica and BRSV were the most commonly found bacterial and viral lung pathogens, respectively. Pasteurella spp. and H. somnus were often associated with the more severe fibrinonecrotizing type of bronchopneumonia, whereas BRSV was primarily detected in cases of suppurative bronchopneumonia. Mycoplasma bovis was isolated from one case only, whereas M. dispar, M. bovirhinis and Ureaplasma diversum were present, often concomitantly, in the majority of cases. Aspergillus fumigatus was isolated from one case.     

Serum and tissue concentrations of erythromycin in calves with induced pneumonic pasteurellosis

The effects of pneumonia on the pharmacokinetics of erythromycin administered IM and the tissue concentration changes with time were evaluated in 2-month-old calves. Pneumonia was induced by injection of Pasteurella haemolytica cultures through the thoracic wall into each lung. Six days prior to induction of pneumonia, erythromycin (15 mg/kg) was administered in a single IM dose. Erythromycin was administered again 48, 72, and 96 hours after injection of P haemolytica. On the third day of erythromycin administration (96 hours), the calves were serially euthanatized in groups of 4 calves each at 2, 5, 8, 12, 18, and 24 hours after the final dose was given. Tissue concentrations of erythromycin in kidney, liver, lung, muscle, CSF, and serum were determined. Neither the serum concentrations nor the overall pharmacokinetic values were significantly (P less than or equal to 0.05) changed by pneumonia. The concentrations of erythromycin were maximal at 5 hours for liver, muscle, and serum and at 8 hours for CSF, kidney, and lung. Serum and muscle concentrations were similar, whereas concentrations in CSF were lower than in serum and higher in kidney, liver, and lung. The lung/serum ratios were approximately 2.5 to 3 at 8 through 24 hours after IM administration. The peak concentration in lung was approximately 6 micrograms/g at 8 hours.     

Immunoperoxidase evaluation of pneumonic lesions induced by Pasteurella haemolytica in calves

Calves inoculated with Pasteurella haemolytica serovar 1 developed lesions of coagulation necrosis in the lungs that were sharply demarcated by leukocytes. The P haemolytica antigen was detected in the area of coagulation necrosis in histologic sections, using an immunoperoxidase technique. In the central area of the necrotic tissue, the bacterial antigen was diffusely presented in the necrotic alveolar wall, fibrin, serous exudate, and degenerated leukocyte. The bacterial antigen also was found in some groups of degenerating leukocytes around the necrotic tissue. The bacterial colonies among these leukocytes had strong specific reactions against P haemolytica. The bacterial antigen was observed in the cytoplasm of macrophages in alveoli around the necrotic lesion. These findings confirmed that coagulation necrosis is an important lesion in calves with pneumonia caused by P haemolytica.     

Prevalence of mycoplasmas in the respiratory tracts of pneumonic calves.

The prevalence of mycoplasmas in the respiratory tracts of 148 pneumonic calves originating from 25 herds in the Netherlands is reported. Four types of culture media were used to isolate mycoplasmas: solid modified Edward medium, 2 types of Friis media, and A7B differential agar medium. Mycoplasmas were isolated both from nasal swab specimens and lung lavage fluids collected from live calves and from nasal mucosa and lung tissue specimens collected post mortem. All of the mycoplasma strains isolated could be identified as either Ureaplasma diversum (isolated from 80% of 25 herds), Mycoplasma dispar (92%), M. bovirhinis (88%), M. bovis (20%), M. bovigenitalium (4%), M. arginini (16%), or M. canis (12%). Isolation rates of M. dispar and U. diversum were considerably higher from lung lavage fluids than from nasal swab specimens. M. bovis was detected only in fattening herds and not in dairy herds. The respiratory tracts of 75% of the calves examined contained at least 2 mycoplasma species. In total, 25 different combinations of mycoplasma species were detected in specimens collected from noses and lungs. The pathogenic species U. diversum and M. dispar had not been isolated before in the Netherlands.     

Efficacy evaluation of the use of oral tilmicosin in pneumonic calves

A trial was carried out on a beef cattle farm to obtain efficacy data for the treatment of naturally occurring bronchopneumonia in calves under commercial production conditions. The treatment was administered by medicating milk with three different tilmicosin doses mixed in the first 80% of the daily milk ration. According to the clinical results, all three treatment regimens proved to be effective. The efficacies of the reduced dose of 12.5 mg tilmicosin/kg body weight daily for 5 days and of the reduced treatment duration with 25 mg/kg b.w. daily for 3 days were equivalent to a previously proven dose of 25 mg/kg b.w. daily for 5 days.     

Efficacy evaluations of the use of oral tilmicosin in pneumonic calves

The therapeutic effect of oral tilmicosin was compared with enrofloxacin, and the efficacy of three doses of the drug was examined in two fully randomized blinded field trials. Pneumonic milk-fed calves between 3 days and 2.5 months of age were allocated into two groups in trial 1 (50 animals) and into three groups in trial 2 (69 calves). In the first trial, the animals were treated with 25 mg/kg/day tilmicosin or 2.5 mg/kg/day enrofloxacin in milk for 5 days. In the second trial, the calves received either 25 mg/kg/day tilmicosin for 5 days or 3 days, or else 12.5 mg/kg tilmicosin for 5 days. All calves were clinically examined for 10 days. In the first trial, oral tilmicosin at a dose of 25 mg/kg/day for 5 days proved to be effective for the treatment of endemic pasteurellosis of milk-fed calves. The efficacy was the same as that of enrofloxacin. All three doses in the second trial were effective and were statistically equivalent to the original dose tested.     

Immunoperoxidase evaluation of pneumonic lesions induced by Pasteurella multocida in calves

To evaluate the relationship between pneumonic lesions and distribution of bacteria, lungs from calves inoculated with Pasteurella multocida were examined histologically by use of immunoperoxidase technique. Pneumonic lesions fundamentally consisted of broncho-pneumonia with fibrinopurulent pleuritis. The lesions were confirmed to be associated with inoculated P multocida, using the immunoperoxidase technique. The P multocida antigen was detected not only in the bacterial clusters in the lesions, but also in the cytoplasm of infiltrating neutrophils and macrophages. Further, immunoelectron microscopy indicated that the inoculated bacteria generally were phagocytosed and digested by neutrophils.     

Bovine pneumonic pasteurellosis: experimental induction in vaccinated and nonvaccinated calves.

A study was undertaken to investigate the effects of the immune response induced by combined aerosol and parenteral vaccination on the lung lesions induced in calves by Pasteurella haemolytica AI. Twenty-four calves, twelve of which had been vaccinated with killed P. haemolytica by aerosol and subcutaneous injection in Freund's complete and incomplete adjuvant were challenged by intratracheal inoculation of live P. haemolytica. Serological response to vaccination was not marked but was best measured by the whole cell agglutination test or by indirect bacterial agglutination rather than by the passive haemagglutination test. Titres of vaccinates were positively correlated with the degree of pneumonic change following challenge while in nonvaccinated controls, titres were negatively correlated with lung lesions. These findings suggest the occurrence of an immunologically mediated hypersensitivity pneumonitis in the lungs of vaccinates and point to the potential efficacy of live bacterial aerosols for stimulation of protective immunity in pneumonic pasteurellosis.     

Chloramphenicol, lincomycin and oxytetracycline disposition in calves with experimental pneumonic pasteurellosis

The effects of pneumonia on the pharmacokinetics of chloramphenicol, lincomycin, and oxytetracycline were evaluated in two-month-old calves. Pneumonia was induced by injection of Pasteurella haemolytica cultures directly through the thoracic wall into each lung. Six days prior to induction of pneumonia, the antibiotics were administered in a single i.v. dose. The antibiotics were administered again 48 (i.v.), 60 and 72 h (i.m.) following injection of P. haemolytica. The pharmacokinetics of chloramphenicol (25 mg/kg) and lincomycin (10 mg/kg) were not significantly different in calves with pneumonia. The hybrid rate constant beta for oxytetracycline was increased in calves with pneumonia from 0.0034 +/- 0.0003/min to 0.0048 +/- 0.0007/min between 2 h and 8 h. Thus the elimination half-life in serum was shortened from 212.4 +/- 20.3 min to 149.3 +/- 19.5 min. In addition, there was an apparent but not statistically significant decrease in K12 with pneumonia. These findings accentuate the need for observance of 12-h dose intervals with oxytetracycline.     

Importance of neutrophils in the pathogenesis of acute pneumonic pasteurellosis in calves

Acute lung injury was induced in 24 calves by intratracheal inoculation with Pasteurella haemolytica. Calves in groups 1 and 2 were neutrophil depleted, using hydroxyurea given IV. Group 1 calves (n = 7) were inoculated intratracheally with saline solution, and group 2 calves (n = 7) were inoculated with P haemolytica. Group 3 calves (n = 7) had normal numbers of neutrophils and were inoculated with P haemolytica. Group 4 calves (n = 3) were treated acutely with hydroxyurea IV, had normal numbers of neutrophils, and were inoculated with P haemolytica. After inoculation, calves with normal numbers of neutrophils (groups 3 and 4) became hypoxemic 2 hours after inoculation, and hypoxemia persisted until necropsy (6 hours after inoculation). These calves also developed tachypnea, bradycardia, neutropenia, and lymphopenia. Lung lesions consisted of necrosis of the alveolar walls, intra-alveolar hemorrhage, and a severe exudative and necrotizing bronchopneumonia, with accumulation of proteinaceous fluid in alveoli and lymphatics. In neutrophil-depleted calves (groups 1 and 2), blood gas values, heart and respiratory rates, and numbers of circulating leukocytes did not change after inoculation with saline solution or with P haemolytica. At necropsy, the lungs of neutrophil-depleted calves were grossly normal. Therefore, neutrophils were required for the acute lung injury induced by P haemolytica. The protective effect of neutrophil depletion was a specific effect of hydroxyurea because calves with high circulating concentrations of hydroxyurea and calves with normal numbers of neutrophils (group 4) developed lung injury.     

Treatment of experimentally induced pneumonic pasteurellosis of young calves with tilmicosin.

Twenty four (24) healthy male Holstein calves (< 70 kg) were each experimentally infected by intrabronchial inoculation of 4.0 x 10(9) viable cells of Pasteurella haemolytica-AI (B122) at Time = 0 h. At 1 h following inoculation animals received either: 1) Sham treatment with sterile 0.85% saline SC (n = 12); or 2) a single injection of 10 mg tilmicosin per kg body weight (n = 12). Calves that were non-infected and tilmicosin-treated were also included for determining tilmicosin concentrations in serum and lung tissue at 1, 2, 4, 6, 8, 24, 48, and 72 h (n = 3-per time). In the infected calves, response to therapy was monitored clinically. Serum samples were collected for determination of tilmicosin concentrations using HPLC. Any animal becoming seriously ill was humanely killed. Complete necropsy examinations were performed on all animals and included gross pathologic changes, bacteriologic analysis, histopathology, and determination of pulmonary concentrations of tilmicosin. Tilmicosin treated animals responded significantly better to therapy than saline-treated control calves. Clinical assessment of calves during the study indicated that tilmicosin-treated calves had significantly improved by T = 8 h compared to satine-treated animals (P < 0.05). At necropsy tilmicosin-treated calves had significantly less severe gross and histological lesions (P < 0.05) of the pulmonary tissue. Of the 12 saline-treated calves, 92% (11/12) had Pasteurella haemolytica-A1 in lung tissue, while of the tilmicosin-treated calves 0% (0/12) cultured positive for P. haemolytica. Mean (+/- standard error) serum tilmicosin concentrations in infected calves peaked at 1 h post-injection (1.10 +/- 0.06 micrograms/mL) and rapidly decreased to 0.20 +/- 0.03 microgram/mL, well below the MIC of 0.50 microgram/mL for P. haemolytica-A1 (B122), by 12 h. These serum concentrations were very similar to serum concentrations of tilmicosin in non-infected tilmicosin-treated calves. Lung tissue concentrations of the antibiotic were comparatively high, even at 72 h post-infection (6.50 +/- 0.75 ppm). Lung tissue concentrations at 72 h were significantly higher in experimentally infected calves than in non-infected tilmicosin-treated animals (P < 0.05). These data demonstrate that tilmicosin was effective in treating experimentally-induced pneumonic pasteurellosis as determined by alleviation of clinical signs, pathological findings at post mortem, and presence of viable bacteria from the lung. Concentrations substantially above MIC for P. haemolytica were present in lung tissue even at 72 h following a single subcutaneous injection of 10 mg tilmicosin per kg body weight.     

Induction of immunity against pneumonic pasteurellosis following experimental infection in calves.

Immunity against pneumonic pasteurellosis was studied in calves after recovery from experimental respiratory disease with Pasteurella haemolytica. Nine calves were exposed to aerosols of parainfluenza-3 virus and Pasteurella haemolytica A1 six days apart to produce respiratory disease. After recovery from the disease, these nine principal and four control calves were challenged with aerosols of bovine herpesvirus 1 and P. haemolytica A1 four days apart. With this viral-bacterial challenge, the nine principal animals failed to develop clinical responses to this bacterial challenge and their lungs did not show the growth of P. haemolytica on cultures, whereas two of four control calves had elevated temperatures and developed necropurulent pneumonia with the isolation of P. haemolytica from the lungs. The principal calves had developed high levels of cytotoxin neutralizing antibodies in their sera following parainfluenza-3 virus-P. haemolytica infection. This demonstrated that immunity against pneumonic pasteurellosis can be achieved, with a suggestion that further search for an effective vaccine for P. haemolytica is warranted.     

Immunoperoxidase evaluation of pneumonic lesions in calves naturally infected with Pasteurella haemolytica

Immunoperoxidase technique was applied for pathological study on naturally occurring pneumonic tissues of calves from which Pasteurella haemolytica was isolated. Multifocal necrosis occurred in the lungs of 25 out of 42 calves (59.5%) and P. haemolytica antigen was detected in 22 out of the 25 calves (88.0%). The calves were divided into 3 groups according to the number of P. haemolytica isolated. The positive rate of the bacterial antigen detected by the technique was 66.6% (28/42) on the average, reaching up to 85.7% (18/21) in the group from which the largest number of P. haemolytica was isolated.     

Effects of dexamethasone on distribution and function of peripheral mononuclear blood cells in pneumonic calves

Experimental pneumonic pasteurellosis was used in 30 Holstein calves to compare the effects on the immune response of different treatments: antibiotics alone or antibiotics in combination with either steroidal (SAID) or non-steroidal anti-inflammatory drugs (NSAID). Clinical parameters and the effect of treatments on the mitogen-induced proliferation of peripheral mononuclear blood cells (PMBCs) and on their distribution were evaluated.Calves in all three treated groups showed rapid improvement, but clinical signs were less marked in both groups receiving anti-inflammatory drugs. Limited difference in the effect on the immune system of treatments was observed. No inhibition of lymphocytes proliferation was detected in these experimental conditions in the dexamethasone (DM)-treated group. There was no variation among groups for the CD4+, CD5+, CD8+ and CD21+ populations. The only noteworthy change was a transient increase in the percentage of the monocyte population (CD14+) in the DM-treated group compared to the group treated with NSAID.     

Experimentally induced pneumonic pasteurellosis: dose-response relationships and protection against natural reinfection in calves

Calves were inoculated intranasally with 2 X 10(6.2) tissue culture infective doses of infectious bovine rhinotracheitis virus, followed in 7 days by intratracheal inoculations with 1 of 4 challenge doses of pathogenic Pasteurella haemolytica. Severity and duration of the ensuing clinical signs of respiratory tract disease were correlated with the challenge dose of bacteria. Calves given 1 X 10(6) colony-forming units (CFU) of bacteria did not develop reliable clinical evidence of disease, whereas those given 1 X 10(8) CFU or 1 X 10(10) CFU of bacteria developed clinical signs of pneumonic pasteurellosis within 12 to 24 hours of bacterial challenge. Severity of clinical signs was equal at the 10(8) and 10(10) doses of bacteria, but duration of clinical signs was greater in calves given the 10(10) dose. Calves given 1 X 10(12) CFU of bacteria developed relatively severe respiratory tract disease in excess of what was necessary for positive clinical detection. Positive correlations were found between the bacterial challenge dose and the height and duration of increased rectal temperature, amount and duration of increases in ocular and nasal discharges, and the subjective evaluation of depressed attitude and appetite. Correlations were not found between challenge dose and respiratory rate or character, or between challenge dose and complete blood cell count. Convalescent calves were resistant to naturally occurring pneumonic pasteurellosis, which caused severe disease in nontreated calves. Adverse effects of P haemolytica were not observed after the first 4 to 15 days after bacterial administration; however, the bacteria were isolated from nasal secretions of convalescent calves 89 to 116 days after bacterial inoculation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Selective medium for the isolation of Mycoplasma bovis from nasal discharges of pneumonic calves

Strains of bovine Mycoplasmatales exhibited two growth forms on a plate medium containing 0.1 per cent Tween 80. Colonies of Mycoplasma bovis could easily be differentiated from those of concomitant mycoplasmatales in that the former grew rapidly with film and spots, while colonies of M arginini and Acholeplasma species did not produce film and spots; the growth of M bovirhinis was completely inhibited.     

Effect of deferoxamine pretreatment on acute pneumonic pasteurellosis and neutrophil oxidative metabolism in calves.

Iron plays a central role in bacterial infections, influencing both bacterial virulence and host cellular defense mechanisms. We investigated whether iron chelation might be of benefit in the treatment of pneumonic pasteurellosis of calves. Neutrophils obtained from calves previously treated with the iron chelator, deferoxamine, were studied for their responses to latex and opsonized zymosan by luminol-enhanced chemiluminescence and to phorbol myristate acetate and opsonized zymosan by superoxide generation. Treatment with deferoxamine in vivo failed to influence these in vitro measures of neutrophil oxidative metabolism. Furthermore, iron depletion with deferoxamine failed to modify the pathophysiological derangements that occurred in calves following experimental induction of pneumonia by intratracheal inoculation with Pasteurella haemolytica. These data indicate that iron chelation using deferoxamine cannot be recommended as an adjunct to conventional therapy in the treatment of pneumonic pasteurellosis of cattle.