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1.
Lipopolysaccharides (LPS) were extracted from a serotype of each of 2 species of Pasteurella isolated from sheep with respiratory tract infections. Lipopolysaccharides from P haemolytica 82-25 (serotype 1A) or P multocida P-1573 (serotype 12) were mixed with sheep lung surfactant and were incubated for 6 hours at 37 C. After incubation, LPS-surfactant mixtures were centrifuged overnight in sucrose density gradients, and fractions were analyzed. Binding occurred between LPS and surfactant vesicles resulting in a stable complex with densities greater than those with the surfactant alone. The surfactant alone had a density of 1.052 to 1.060 g/ml. Diffuse bands of surfactant had a density of 1.075 to 1.092 when incubated with P haemolytica LPS and a density of 1.069 to 1.105 when incubated with P multocida LPS.  相似文献   

2.
A new transtracheal bronchoalveolar lavage technique for the diagnosis of respiratory disease in sheep under field conditions was tested in 76 sheep. The sheep were divided into three groups, normal sheep, sheep with clinical signs of respiratory disease and housed sheep, on the basis of their respiratory disease history and husbandry conditions. The detection of Mannheimia haemolytica and Mycoplasma ovipneumoniae or parainfluenza virus type 3 and bovine respiratory syncytial virus antigen in the lavage samples was closely correlated with clinical disease. The sheep with clinical respiratory disease had a higher mean percentage of neutrophils in the lavage fluid than the sheep in the other two groups.  相似文献   

3.
Three, 5, or 7 days after inoculation with bovine viral diarrhea (BVD) virus (n = 12) or Mycoplasma bovis (n = 12), groups of calves were exposed to aerosols of Pasteurella haemolytica and were euthanatized 4 hours later. Histologic lesions in the lungs and the ratios of neutrophils to alveolar macrophages, collected by bronchoalveolar lavage, were compared with those of clinically healthy calves (n = 8) and calves inoculated with BVD virus only (n = 4), M bovis only (n = 4), or P haemolytica only (n = 2). Inoculation with BVD virus or M bovis did not have a significant (P greater than 0.05) effect on the neutrophil/macrophage ratio in the bronchoalveolar lavage. Aerosol exposure to P haemolytica induced a marked and significant (P less than 0.01) change in the neutrophil/macrophage ratio (from less than 1:9 to greater than 9:1). The reversed neutrophil/macrophage ratio in calves exposed to P haemolytica correlated well with the histologic changes in which small bronchi and bronchioles were plugged with purulent exudate. Inoculation with BVD virus did not induce gross or microscopic lesions in the lungs. Inoculation with M bovis resulted in a severe peribronchial lymphoid hyperplasia with mild exudation of neutrophils and macrophages into the cranioventral parts of the lungs.  相似文献   

4.
Ovine isolates of the 15 known serotypes found within the A and T biotypes of Pasteurella haemolytica were cytotoxic for sheep bronchoalveolar macrophages (BAM). Weaker toxicity for the same target cells was also expressed by non-serotypable ovine isolates of P. haemolytica. The results suggest that cytotoxicity for sheep BAM is a virulence factor common to both A and T biotypes of P. haemolytica.  相似文献   

5.
OBJECTIVE: To evaluate chemotactic, phagocytic, and bactericidal activities of bovine and porcine alveolar macrophages (AM) exposed to tilmicosin. ANIMALS: 12 healthy calves and 12 healthy pigs. PROCEDURES: Lungs were obtained immediately after euthanasia; AM were collected by means of bronchoalveolar lavage and density gradient centrifugation. Chemotactic activity was evaluated by exposing AM to lipopolysaccharide or macrophage inhibitory peptide during incubation with tilmicosin. Phagocytic activity was evaluated by incubating AM with tilmicosin for 24 hours and then with tilmicosin-resistant Salmonella serotype Typhimurium. Bactericidal activity was evaluated by incubating AM with tilmicosin (0, 10, or 20 microg/ml for bovine AM; 0 or 10 microg/ml or 10 microg/ml but washed free of tilmicosin for porcine AM) and then with Mannheimia haemolytica (bovine AM) or with Actinobacillus pleuropneumoniae or Pasteurella multocida (porcine AM). RESULTS: Tilmicosin had no significant effects on chemotactic or phagocytic activities of bovine or porcine AM. The time-course of bactericidal activity was best described by polynomial equations. Time to cessation of bacterial growth and area under the time versus bacterial number curve were significantly affected by incubation of AM with tilmicosin. CONCLUSIONS AND CLINICAL RELEVANCE: Results show that bactericidal activity of bovine and porcine AM was enhanced by tilmicosin, but not in proportion to the reported ability of AM to concentrate tilmicosin intracellularly. With or without exposure to tilmicosin, the time-course of bactericidal activity of bovine AM against M haemolytica and of porcine AM against A pleuropneumoniae or P multocida was too complex to be reduced to a simple linear equation.  相似文献   

6.
Mannheimia haemolytica, one of the agents associated with bovine respiratory disease complex, can cause severe lung pathology including the leakage of vascular products into the airways and alveoli. Previous work by this laboratory has demonstrated that bovine lung endothelial and epithelial cells undergo dramatic permeability increases when exposed to adenosine-5'-triphosphate (ATP). Therefore, we wanted to determine if ATP levels were elevated in bronchoalveolar lavage (BAL) samples from calves experimentally infected with M. haemolytica. In addition, cultured bovine pulmonary epithelial (BPE) cells were stimulated with heat-killed and live M. haemolytica bacteria, lipopolysaccharide (LPS), lipoteichoic acid (LTA), interleukin-1 (IL-1), and zymosan activated plasma (ZAP) to determine whether they might release extracellular ATP during in vitro infection. Calves experimentally exposed to M. haemolytica had an approximately 2-fold higher level of ATP in their BAL samples compared to control. BPE cells exposed to increasing numbers of heat-killed or live M. haemolytica had significantly increased levels of ATP release as compared to time-matched controls. Finally, BPE cells treated with several concentrations of LPS and IL-1 had increases in ATP release as compared to time-matched controls. This increase appeared to be a result of active ATP secretion by the cells, as cell viability was similar between treated and non-treated cells. Neither ZAP nor LTA induced any ATP release by the cells. In conclusion, ATP levels are elevated in lung secretions from calves infected with M. haemolytica. In addition, lung epithelial cells can actively release ATP when exposed to heat-killed or live M. haemolytica, LPS or IL-1.  相似文献   

7.
The distribution of leukocytes in bovine bronchoalveolar lavage fluids was determined in 15 calves at various times after aerosol exposure to Pasteurella haemolytica. For comparison, 10 calves were exposed to aerosols of phosphate-buffered saline solution; 15 calves, to Staphylococcus epidermidis; and 10 calves, to Salmonella typhimurium endotoxin. At 10 minutes after inhalation exposure for each group, the predominant cell type was the macrophage. Macrophages remained the predominant cell type throughout each lavage interval for calves exposed to phosphate-buffered saline solution and Staph epidermidis. For calves exposed to P haemolytica, there was a decrease in the percentage of macrophages detectable by 30 minutes after exposure, with a corresponding increase in the percentage of neutrophils. Sixty minutes after the inhalation exposure to P haemolytica, the percentages of macrophages and neutrophils in the lavage fluid were equal. By 240 minutes after exposure to P haemolytica, greater than 90% of the cells in the lavage fluids was neutrophils. The increase in the percentage of neutrophils in lavage fluids from calves exposed to S typhimurium endotoxin was similar to that seen for the calves exposed to P haemolytica.  相似文献   

8.
Groups of Caesarean-derived, colostrum-deprived lambs were inoculated by the intratracheal route with Pasteurella haemolytica 4 to 6 days after the inoculation of parainfluenza virus type 3 (PI3). Some were killed immediately (0 h) and others 24 h later. Control groups were inoculated with PI3 alone, P. haemolytica alone or media alone. Pulmonary phagocytic cells, P. haemolytica and PI3 were recovered by pulmonary lavage. The phagocytes were separated into alveolar macrophage (AM) and neutrophil fractions by density gradient centrifugation and examined biochemically and microbiologically. Twenty-four hours after the inoculation of P. haemolytica bacterial proliferation to greater than 0 h levels had occurred in four of six animals inoculated with P. haemolytica alone, two of eight inoculated with P. haemolytica 4 days after PI3 and all of eight inoculated with P. haemolytica 6 days after PI3. Mean bacterial numbers in animals inoculated with P. haemolytica 6 days after PI3 and killed at 24 h (10(9.1 +/- 1.9)) were significantly higher than they were in the other two groups killed at this time (PI3 4 days, P. haemolytica 24 h, mean = 10(5.3 +/- 1.7); P. haemolytica alone 24 h, mean = 10(4.5 +/- 2.9)). Pneumonic lesions were also more severe in the first group. This defect in pulmonary clearance and increase in the severity of pneumonia in animals inoculated with P. haemolytica 6 days after PI3 coincided with a 1000-fold decrease in virus titres in the lung between Day 6 and Day 7 after virus inoculation and the first detectable evidence of the host's immune response. The virus infection resulted in a significant increase in the number of AM that could be recovered from the lung and an increase in the number of AM with cytoplasmic vacuolation. However, there was no difference in the total number of AM or the number of vacuolated AM between animals that controlled the P. haemolytica infection and those in which proliferation of P. haemolytica occurred. The inoculation of P. haemolytica resulted in a 100-fold increase in the number of neutrophils in the lavage fluid, but there were no differences between virus-infected and uninfected animals, nor was there a difference between animals that controlled the P. haemolytica infection and those that did not.(ABSTRACT TRUNCATED AT 400 WORDS)  相似文献   

9.
This study was conducted to investigate the in vivo effect of a single intratracheal inoculation of Pasteurella haemolytica cytotoxin on the rat lung. Changes in the biochemical and cytological composition of bronchoalveolar lavage fluid were used to estimate the magnitude of pulmonary cell injury, inflammatory response, vascular permeability and functional status of pulmonary alveolar macrophages. Effect of treatment was compared with rats intratracheally inoculated with supernatants of Pasteurella multocida or with sterile physiological saline solution (vehicle). Results indicated that Pasteurella haemolytica supernatants were not significantly toxic for the lungs of rats.  相似文献   

10.
We have studied the effects of in vivo asbestos exposure on the surface immune-associated (Ia) antigen expression and distribution of alveolar macrophage subpopulations defined by continuous iso-osmotic Percoll gradients (density range: 1.006 to 1.123 g/ml) using a rat model of asbestos inhalation. Two groups of rats were exposed by intermittent inhalation (6 hr/day for 5 days/week over 4 weeks) to either amphibole (crocidolite) or serpentine (chrysotile) asbestos. A group of control rats was sham-exposed to clean air only. Alveolar macrophages from rats of three groups were obtained by bronchoalveolar lavage. During exposure, distinct differences appeared within 7 days of asbestos exposure, and some of these findings persisted in the crocidolite-exposed group for as long as 2 to 5 months after the cessation of exposure. Furthermore, relatively greater proportions of Ia-antigen positive cells were detected in several density fractions obtained from both asbestos-exposed groups (especially the crocidolite-exposed group). Multinucleated alveolar macrophages were seen frequently in all Percoll fractions after both types of asbestos inhalation. A significant proportion of multinucleated alveolar macrophages in these fractions expressed surface Ia-antigen positivity. The finding of enriched numbers of higher-density phagocytes in bronchoalveolar lavage cell subpopulations from asbestos-exposed rats may reflect the presence of newly recruited-immature monocytes and/or macrophages at sites of intrapulmonary asbestos deposition. Also, increased proportions of Ia-antigen positive cells suggest that a part of them were functionally activated.  相似文献   

11.
OBJECTIVE: To determine reference values for cytologic examination results of bronchoalveolar lavage fluid (BALF) and to investigate effects of repeated lavages on pulmonary health and on results of cytologic examination of BALF in dogs. ANIMALS: 16 healthy adult Beagles. PROCEDURE: All dogs underwent pulmonary lavage to obtain BALF. Eleven dogs were repeatedly lavaged 6 times at 5- to 7-week intervals. Analyses for total and differential cell counts and for viability of cells before and after cell processing were performed. Arterial blood gas analysis before and after bronchoalveolar lavage was used to study the safety of the lavage procedure. Histologic and radiologic examinations were used to study effects of repeated lavages on pulmonary health. RESULTS: Mean (+/- SD) cell count was 104 +/- 69 cells/microl, comprising 75 +/- 7% alveolar macrophages, 13 +/- 6% lymphocytes, 5 +/- 4% neutrophils, 4 +/- 5% eosinophils, 2 +/- 2% mast cells, 0.6 +/- 0.7% epithelial cells, and 0.3 +/- 0.4% plasma cells. Centrifugation of samples and washing of cells caused significant cell loss (59 +/- 13%). Repeated lavages did not cause significant variations in cell counts of BALF or results of arterial blood gas analysis, thoracic radiography, or histologic examination of pulmonary specimens. Only a moderate, although significant, decrease in arterial oxygen content was observed after bronchoalveolar lavage. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis indicated that several lavages performed at 5- to 7-week intervals can safely and reliably be used to study the kinetics of pathologic processes in pulmonary tissues or for evaluation of therapeutic efficacy.  相似文献   

12.
Annexins are phospholipid-binding proteins and are abundant in the lung. Annexins I and IV, but not II and VI, have been detected in bronchoalveolar lavage (BAL) fluids from calves inoculated with Pasteurella haemolytica, the pathogen for calf pneumonia. In this study, BAL fluids from calves with experimental pneumonia induced by inoculation to right lung lobes of bovine herpes virus-1 (BHV-1), the major viral pathogen for pneumonia, were examined for detection of annexins I and IV. Of 6 calves inoculated with BHV-1, annexins I and IV were coincidentally detected in BAL fluids from right lung lobes of 4 calves, but not in BAL fluids from left lung lobes of 6 inoculated calves or those from left and right lung lobes of 3 control calves. Annexin II and VI were not found in any BAL fluids examined. These results, together with previous findings on calves inoculated with Pasteurella haemolytica, suggest that the release of annexins I and IV onto the alveolar surface is an essential event occurring in response to pulmonary infections of BHIV-1 and Pasteurella haemolytica.  相似文献   

13.
Pasteurella haemolytica A1 culture supernatant containing leukotoxin, and modifiers of cyclic nucleotide and arachidonate metabolism, were evaluated for their ability to alter oxygen radical production by pulmonary alveolar macrophages obtained from seven Holstein calves. Calves were sedated, and underwent bronchoalveolar lavage to harvest macrophages, which were then incubated with culture supernatant and/or the drugs and toxins under study, and challenged with opsonized zymosan to induce oxygen radical generation. This was measured by a chemiluminescence technique. Pasteurella haemolytica A1 culture supernatant alone delayed the time to maximum oxygen radical production, although total production was increased. The cyclooxygenase inhibitor indomethacin and the phospholipase inhibitor p-bromophenacyl bromide significantly reduced maximum oxygen radical production, but their effects were diminished in the presence of culture supernatant. Although forskolin markedly inhibited oxygen radical generation, this effect was not altered by culture supernatant. Incubation of macrophages with pertussis toxin had no effect on oxygen radical production, while incubation with cholera toxin did inhibit production. This inhibitory effect was significantly lessened by concurrent incubation with P. haemolytica A1 culture supernatant.  相似文献   

14.
A method for bronchoalveolar lavage in live pigs   总被引：3，自引：0，他引：3  
In order to isolate porcine alveolar macrophages and to quantitatively study the components of recovered lung fluid, a bronchoalveolar lavage technique in living pigs was developed. Lung lavage was performed after introducing a catheter through the mouth via the trachea in the diaphragmatical lobe. Thirty ml of phosphate-buffered saline (PBS) was introduced into the lung and the fluid was aspirated after one minute. Following this, another 15 ml of PBS was introduced into the lung and aspirated after one minute. The recovered volume of the second lavage averaged 15 ml (+/- 0.4 S.E.M.). Cells thus obtained from specific-pathogen-free (SPF) pigs were composed of 98% macrophages. Lavage fluids from conventionally bred pigs contained 67% macrophages, 17% neutrophilic granulocytes and about 16% lymphocytes, demonstrated by their morphology and acid phosphatase activity. The viability of the recovered cells was over 98% in both SPF and conventionally bred pigs. The dilution of the aspirated lung liquid was determined by using methylene blue in the introduced fluid. The calculated dilution factor of the recovered lavage fluid was 0.58 (S.E.M. 0.02). No influence was noticed on the number or composition of cells nor on the dilution factor when lung lavage was done in SPF pigs twice a week during a four week period. The protein concentration in lavage fluid from SPF pigs was 142 (SD +/- 26) mcg/ml. In conventionally bred pigs, however, a wide variation (276 +/- 229 mcg/ml) in protein content was noted. Lavage fluid supernatant of some animals had a bactericidal effect on Haemophilus pleuropneumoniae strain 13261, whereas no bactericidal effect was noted in other lavage samples.  相似文献   

15.
The molecular and cellular basis for the enhanced lung pathology and mortality caused by Mannheimia haemolytica in bighorn sheep (BHS, Ovis canadenesis), in comparison to domestic sheep (DS, Ovis aries), is not clear. Polymorphonuclear leukocytes (PMNs) of BHS are four- to eight-fold more susceptible to M. haemolytica leukotoxin-induced cytolysis, which is likely to reduce the number of functional phagocytes in the lung. We hypothesized that enhanced lung pathology is due to defective clearance of M. haemolytica from the lungs of BHS. To test this hypothesis, M. haemolytica (1 × 10(7) colony forming units [cfu]) were inoculated intra-tracheally into three groups each of BHS and DS, which were euthanized and necropsied at 4, 12, and 18 h post-inoculation (hpi). Bacterial and leukocyte counts were performed on broncho-alveolar lavage fluid (BALF) collected at necropsy. BALF from BHS euthanized at 4 and 12 hpi contained a significantly higher number of M. haemolytica than that from DS. More importantly, DS did not have any bacteria in BALF at 18 hpi, while the BHS still had significant numbers. As expected, the BHS did exhibit more extensive lung lesions at 12 and 18 hpi when compared to DS. At 18 hpi, necrotic PMNs were observed in the lesional lung tissues of BHS, but not DS. Furthermore, BALF from BHS had significantly lower titers of antibodies to Lkt and surface antigens of M. haemolytica, than that of DS. These findings suggest that the enhanced pathology in BHS lungs is due to defective clearance of M. haemolytica from the lungs.  相似文献   

16.
The role of recruited neutrophils in Mannheimia haemolytica infection is controversial. We hypothesized that the neutrophilia induced by recombinant bovine granulocyte colony-stimulating factor (GCSF) would lead to rapid bacterial clearance and less severe lesions after infection with M. haemolytica. Two experiments (A and B) were conducted in which four calves per experiment were treated daily with 5 microg/kg GCSF and four calves per experiment were treated with saline. All 16 calves were challenged with 5 x 10(9) colony-forming units (cfu)/ml (experiment A) or 4.5 x 10(8) cfu/ml (experiment B) of M. haemolytica bacteria, into the right bronchus by bronchoscope-placed catheter. The mean maximal blood neutrophil counts in non-GCSF-treated and GCSF-treated calves before bacterial challenge were 5.6 +/- 0.7 x 10(9)/liter and 25.4 +/- 2.7 x 10(9)/liter, respectively. Two untreated calves became neutropenic and were euthanatized 2 days after infection because of severe respiratory distress. GCSF-treated calves had a 37% reduction in lung lesions compared with nontreated calves, and this difference was significant (P=0.04) when the effect of previous antibody titre to leukotoxin was considered. The effect of GCSF treatment on the severity of clinical signs seemed to be influenced by the antibody titre to M. haemolytica leukotoxin, although this effect could not be conclusively addressed. In conclusion, GCSF induced neutrophilia and partially protected calves against experimental infection with M. haemolytica. These results imply that increased numbers of neutrophils may, under some circumstances, protect against severe pneumonia caused by M. haemolytica.  相似文献   

17.
Background: Surfactant alterations are described in horses after exercise, anesthesia, and prolonged transport, in horses with recurrent airway obstruction, and in neonatal foals. The effect of horse age or bronchoalveolar lavage fluid (BALF) sample characteristics on surfactant is unknown.
Objectives: To evaluate surfactant phospholipid composition and function in healthy horses, and to investigate the influence of age and BALF sample characteristics on surfactant.
Animals: Seventeen healthy horses 6–25 years of age maintained on pasture year-round.
Methods: BALF was collected by standard procedures and was assessed for recovery volume, nucleated cell count (NCC), and cytology. Cell-free BALF was separated into crude surfactant pellet (CSP) and surfactant supernatant (Supe) by ultracentrifugation. Phospholipid and protein content were determined from both fractions. CSP phospholipid composition was analyzed by high-performance liquid chromatography with an evaporative light scatter detector. Surface tension of CSP was evaluated with a pulsating bubble surfactometer. Regression analysis was used to evaluate associations between age, BALF sample characteristics, and surfactant variables.
Results: Results and conclusions were derived from 15 horses. Increasing age was associated with decreased phospholipid content in CSP but not Supe. Age did not affect protein content of CSP or Supe, or surfactant phospholipid composition or function. Age-related surfactant changes were unaffected by BALF recovery percentage, NCC, and cytological profile.
Conclusions and Clinical Importance: Older horses have decreased surfactant phospholipid content, which might be because of age-related pulmonary changes. Surfactant composition is unaffected by BALF sample characteristics at a BALF recovery percentage of at least 50%.  相似文献   

18.
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19.
Six Rocky Mountain bighorn sheep were raised in captivity from birth (n = 5) or taken from the wild as a lamb (n = 1). After the bighorn sheep were in captivity for over a year, 6 clinically normal domestic sheep were placed on the 2 ha of pasture on which the bighorn sheep were kept. Nasal swab specimens were obtained from all sheep at the time the domestic sheep were introduced. Pasteurella haemolytica was isolated from swab specimens obtained from 4 of 6 domestic sheep, but not from specimens obtained from the bighorn sheep. All 6 bighorn sheep died of acute hemorrhagic pneumonia after exposure to domestic sheep. Death in the bighorn sheep occurred on days 4, 27, 27, 29, 36, or 71 after initial exposure to domestic sheep. Pasteurella haemolytica was isolated from respiratory tract tissue specimens of all bighorn sheep at the time of death. None of the domestic sheep were clinically ill during the study. At the end of the study, 3 of 6 domestic sheep were euthanatized, and at necropsy, P haemolytica was isolated from 2 of them. The most common serotypes in bighorn and domestic sheep were P haemolytica T-3 and A-2. Other serotypes isolated included P haemolytica A-1, A-9, and A-11 in bighorn sheep and A-1 in domestic sheep. On the basis of results of this study and of other reports, domestic sheep and bighorn sheep should not be managed in proximity to each other because of the potential fatal consequences in bighorn sheep.  相似文献   

20.
The central arterial pharmacokinetics of thiopental were studied in six rabbits, six sheep and six dogs after a short infusion at approximately 10 mg/kg min. Thiopental was infused to a defined electro-encephalographic endpoint (EEG burst suppression). The time to reach early burst suppression was longer in the dog (3.9 +/- 0.5 min) compared with the sheep (3.0 +/- 0.6 min) and the rabbit (2.5 +/- 0.5 min). The total dose required to produce the same level of EEG activity was higher in the dog (35.9 +/- 6.8 mg/kg) compared with the sheep (24.3 +/- 5.3 mg/kg) and the rabbit (21.6 +/- 6.8 mg/kg). The plasma concentration-time data for each animal was fitted using non-linear regression to a bi- or tri-exponential function. In all animals, the plasma-time profile was best described as a tri-exponential decay. The initial volume of distribution was similar in all three species (rabbit, 38.6 +/- 10.0 mg/kg; sheep, 44.5 +/- 9.1 ml/kg; dog, 38.1 +/- 18.4 ml/kg). The maximum arterial plasma thiopental concentration achieved at EEG burst suppression was higher in the sheep (221.8 +/- 27.9 micrograms/ml) than the dog (164.7 +/- 29.9 micrograms/ml) or the rabbit (112.3 +/- 15.1 micrograms/ml). Thiopental distribution clearance was slower in the sheep (43.6 +/- 15.1 ml/min/kg) compared with the rabbit (110.5 +/- 18.7 ml/min kg) and the dog (97.2 +/- 47.2 ml/min kg). Elimination half-life was extended in the sheep (251.9 +/- 107.8 min) and dog (182.4 +/- 57.9 min) relative to the rabbit (43.1 +/- 3.4 min).(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

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