The pulmonary clearance of Pasteurella hemolytica in calves infected with bovine parainfluenza-3 virus.

The purpose of this study was to determine if parainfluenza-3 virus in calves interfered with normal pulmonary bacterial clearance. Three groups (A, B and C) of four calves each were exposed to an aerosol of parainfluenza-3 virus. Three days later the four hour pulmonary clearance of Pasteurella hemolytica was determined on the first group (A), seven days later on the second (B) and 11 days later on the third (C). Group A had a mean pulmonary bacterial retention of 3.6 +/- 3.5%. Group B was 83.1 +/- 35.9% and Group C was 41.2 +/- 30.9%. The results demonstrate that parainfluenza-3 virus interfered with the pulmonary bacterial clearance of Pasteurella hemolytica particularly on day 7 and also on day 11 but not on day 3. This inhibition of pulmonary clearance caused by the virus may be a key factor in the pathogenesis of pneumonic pasteurellosis. Histological examination of the lungs did not demonstrate a correlation between pulmonary retention of bacteria and the development of pathological changes.     

A Three Year Comparison of Acute Respiratory Disease, Shrink and Weight Gain in Preconditioned and Non-preconditioned Illinois Beef Calves Sold at the Same Auction and Mixed in a Feedlot

During 1969 to 1971, 78 preconditioned (PC) and 79 non-preconditioned (NPC) beef calves were purchased at the same auction and mixed in a feedlot. Preconditioned calves were weaned 30 days before the sale, used to drinking from a tank, and vaccinated against blackleg, malignant edema, infectious bovine rhinotracheitis (IBR), parainfluenza-3 (PI3) and bovine virus diarrhea (BVD) in 1970 and 1971, and Pasteurella hemolytica and multocida in 1971. All vaccinations were completed two to three weeks before the sale. PC calves were given thiabenzadole. PC calves had significantly less shrink after shipment and in 1971 significantly more rapid daily gain during the first weeks of the feeding period. In 1969 more PC calves were treated for acute respiratory disease than NPC calves during an outbreak of PI3 and BVD infection. In 1970 and 1971 fewer PC than NPC calves were treated for acute respiratory tract disease during outbreaks of PI3 infection. The differences in clinical respiratory disease were significant in 1971. Inclusion of two doses of P. hemolytica and P. multocida bacterin before the sale in 1971 and use of an intranasal PI3 vaccine was considered to improve the PC program. Fecal egg counts for gastrointestinal nematodes were much lower in PC calves treated with thiabenzadole than untreated NPC calves.     

The Pulmonary Clearance of Bacteria by Calves and Mice

Using a modified aerosol generator, white mice and calves were exposed to aerosols of viable Staphylococcus aureus and Pasteurella haemolytica and the clearance of the inhaled organisms by the lungs of the experimental animals was measured. Fifty-seven percent of inhaled S. aureus were cleared in two hours by the mouse lungs, 79% were cleared in four hours and 93% were cleared in eight hours. Fifty-six percent of inhaled P. haemolytica were cleared in two hours by the mouse lungs, 76% were cleared in four hours and 93% were cleared in eight hours. Seventy percent of inhaled S. aureus were cleared in two hours by the calf lungs, 90% were cleared in four hours and 95% were cleared in eight hours. Seventy-five percent of inhaled P.haemolytica were cleared in two hours by the calf lungs, 90% were cleared in four hours and 92% were cleared in eight hours.     

The pulmonary clearance of Pasteurella haemolytica in calves given Corynebacterium parvum and infected with parainfluenza-3 virus.

Four control calves were aerosolized with parainfluenza-3 and one week later with Pasteurella haemolytica. Three calves were given Corynebacterium parvum at a dose of 15 mg/m2 body surface area, infected with parainfluenza-3 virus one week later, and aerosolized with P. haemolytica two weeks after C. parvum injection. All calves were killed four hours after P. haemolytica exposure and the bacterial retention in the lung was determined. Parainfluenza-3 viral infection did not exert any suppressive effect on pulmonary clearance of P. haemolytica in six out of seven calves used. However, the bacterial colony counts in the lungs of control calves were higher (P less than 0.05) than those in calves given C. parvum. Hence, C. parvum appeared to enhance bacterial clearance. Despite the marked influx of neutrophils into the lungs after the bacterial inoculation, the neutrophil:macrophage ratio in lavage samples was less in calves given C. parvum than in the control calves. The alveolar macrophages in C. parvum treated calves were generally larger but did not differ significantly (P less than 0.05) from those in the controls. There was no significant (P less than 0.05) correlation between the percentages of alveolar macrophages and the bacterial clearance. In calves given C. parvum, bacterial clearance was enhanced in those calves which had larger macrophages.     

Experimental Bovine Pneumonic Pasteurellosis II. Genesis and Prevention

Two experiments were conducted. In the first, 16 crossbred Hereford calves were divided into two equal groups. The first group was vaccinated intranasally with a commercial vaccine against bovid herpesvirus 1 and the second group was unvaccinated. The calves were later exposed to an aerosol of bovid herpesvirus 1 (strain 108) for five minutes. Four calves from each group were subjected to transportation and four calves from each group were kept in an environmental chamber for four days. Four days after viral aerosol all calves were exposed to an aerosol of Pasteurella haemolytica and the same subgroups were again transported or held in the chamber for a further four days.

The calves that did not die from pneumonia were necropsied ten days after the final day of transport. Pulmonary lesions were present in both vaccinated and control animals but were less extensive in the vaccinated calves. Six of eight vaccinated but none of the eight control calves survived.

In the second experiment, eight crossbred Hereford calves were divided into two equal groups. One group was vaccinated with bovid herpesvirus 1 (strain 108) and the other acted as controls. Four weeks later all calves were sequentially exposed to aerosols of bovid herpesvirus 1 (strain 108) and P. haemolytica four days apart. Three of the four controls but none of the vaccinates died from pneumonia. Every lobe of the lungs in all the controls was affected by pneumonia while no pulmonary lesions were found in the vaccinated calves. The differences in efficacy of the modes of vaccination and the possible role of transport stress are discussed.

     

Influence of Immunization Procedures on Upper Respiratory Tract Immunity in Cattle

Aspects of respiratory tract immunity have been investigated in the bovine species. Using Past. hemolytica type I as the antigen for this model the relationship of nasal and serum antibody production to the route of vaccination and type of vaccine was investigated in a series of 15 dairy calves from two to four months of age. Experimental results indicated that an aerosol vaccination with live Past. hemolytica resulted in a significant nasal antibody response while parenterally vaccinated gave calves with equivalent serum titers had no significant nasal antibody response.     

Exposure of calves to aerosols of parainfluenza-3 virus and Pasteurella haemolytica.

The present study was undertaken to investigate whether sequential exposure to aerosols of parainfluenza-3 virus followed by Pasteurella haemolytica, or P. haemolytica followed by parainfluenza-3 virus, could lead to the production of pulmonary lesions in conventionally-raised calves. Twenty male calves with low serum antibody titres to both organisms were placed in five equal groups. Synergism of parainfluenza-3 virus and P. haemolytica was not demonstrated in any of the sequentially infected groups and pulmonary lesions were mild in all challenged calves. Clinical signs of disease were not present after exposure to parainfluenza-3 virus although the virus was repeatedly isolated from nasal secretions of all inoculated calves. Exposure to P. haemolytica produced a transient response which consisted of increased rectal temperatures and respiratory rates, with a mild neutrophilic leukocytosis and a mild left shift present six hours postinoculation and returning to normal within 24 hours. Results from this study suggest, although do not confirm, that reduced pulmonary clearance of inhaled P. haemolytica in parainfluenza-3 virus infected calves does not necessarily lead to production of severe pulmonary lesions and that previous exposure to aerosols of P. haemolytica may not enhance secondary parainfluenza-3 virus infection.     

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.     

Preliminary Observations on the Effect of Specific Immunity on Nasal Bacterial Flora

This investigation has shown that Pasteurella hemolytica could actively colonize the nasal cavity of calves in the presence of indirect hemagglutinating IHA antibody against that particular strain. It was not determined if the level of nasal IHA antibody to Past. hemolytica had an effect on colonization of the nasal passage by Past. hemolytica. Specific immunity to Past. hemolytica would appear to be important in respiratory disease induced by this organism.     

Studies with Radioactive Endotoxin II. Clearance of 3H-Labelled Endotoxin from the Blood of Calves

The clearance of ³H-labelled Pseudomonas endotoxin from the blood was studied in a nontolerant and in an endotoxin tolerant state. Calves were rendered tolerant to the toxic effects of the endotoxin by four daily intravenous injections of endotoxin at the dose rate of 5 µg/kg body weight.

Clearance of ³H-endotoxin from the blood of nontolerant calves occurred more slowly than did clearance of ⁵¹Cr-endotoxin and was not significantly (P<0.05) affected by the development of tolerance. The lungs and liver were the major organs involved in the clearance of ³H-endotoxin from the blood of calves. Leukocytes and erythrocytes, but not platelets, were shown to participate in endotoxin clearance in calves. ³H₂O, the control substance used in calves, was not concentrated within any particular organ but rapidly equilibrated with total body water and was slowly excreted.

     

Studies with Radioactive Endotoxin I. Clearance of 51Cr-Labelled Endotoxin from the Blood of Calves

The clearance of ⁵¹Cr-labelled Pseudomonas endotoxin from the blood was studied in calves in a nontolerant and in an endotoxin-tolerant state. Calves were rendered tolerant to the toxic effects of the endotoxin by four daily intravenous injections of endotoxin at the dose rate of 5 µg/kg body weight.

Clearance of a small amount of ⁵¹Cr-endotoxin from the blood of nontolerant calves was almost complete within three minutes of injection and was not significantly faster in tolerant calves. The lungs and liver were the major organs involved in clearance of endotoxin from the blood. The ⁵¹Cr label was slowly excreted by the kidneys. Neither platelets nor leukocytes were demonstrated to participate in endotoxin clearance in calves. ⁵¹CrCl₃ was injected into control calves. Relative to the distribution and loss of labelled endotoxin, the ⁵¹CrCl₃ was cleared slowly from the blood, was distributed uniformly throughout the body and was excreted rapidly.

     

Colonization of the nasal passages of calves with Pasteurella haemolytica serotype 1 and regeneration of colonization after experimentally induced viral infection of the respiratory tract

Healthy nonstressed calves were inoculated intranasally with or subjected to aerosol exposure to Pasteurella haemolytica serotype 1. Only 4 of 28 calves harbored the bacterium in enough numbers to be isolated from the nasal passages for more than 7 days. After apparent clearing from the nasal passages, 8 calves were inoculated intranasally with infectious bovine rhinotracheitis virus; 2 of these calves shed the P haemolytica during clinical illness due to the virus. The remaining 20 calves were aerosol-exposed to parainfluenza-3 virus; 6 of these calves shed P haemolytica during clinical illness due to the parainfluenza-3 virus.     

Deposition in the Respiratory Tract of Cattle of Spores of Bacillus subtilis var niger by Inhalation and by Nasal Instillation

Spores of Bacillus subtilis var niger were deposited in the lungs, tracheae and nasal cavities of four calves by aerosol inhalation and in three calves by intranasal instillation. From each calf 20 specimens of lung tissue, each weighing one gm, three of trachea and three of nasal mucosa were examined for spore content. The average numbers of spores per gm of lung tissue from animals exposed to aerosols were 3.05 and 4.84, 2.35 and 2.02 x 10⁴. Lungs from animals exposed intranasally contained only 747, 62 and 1424 spores per gm of tissue respectively. Animals exposed intranasally had a hundred to a thousand fold more spores on nasal mucosa than animals exposed by aerosol and the latter had a thousand fold more spores on tracheal mucosa than calves exposed intranasally. Aerosol inhalation exposed the lung and trachea more densely and uniformly than did intranasal instillation.     

Ascaris suum Infection in Calves III. Pathology

Gross changes in the lungs of Ascaris suum- infected calves consisted of atelectasis and hemorrhagic foci, edema and emphysema, frequently with bullae. Prominent microscopic lung lesions were edema and emphysema of the interlobular septa with large numbers of eosinophils within and around lymphatics, peribronchiolar lymphoid nodules and parasitic granulomas. Many of the microscopic features were consistent with those found in atypical interstitial pneumonia. Changes in the alveoli were atelectasis, the exudation of plasma proteins, mononuclear cells and eosinophils, and alveolar wall thickening. Lesions found later included fibrosis and fetalization of the alveolar walls. Plasma cells and neutrophils were not common. Challenge with Toxocara canis after sensitization with A. suum resulted in the lungs developing a few areas of atelectasis. Migration of T. canis to lungs of calves is slower than A. suum. A. suum larvae were always found in bronchi, bronchioles and alveoli of calves that died. Lesions were observed in the liver but not the kidney of A. suum infected calves; both lung and liver lesions tended to resolve with time.     

Pneumonia in Calves Produced with Aerosols of Bovine Herpesvirus 1 and Pasteurella haemolytica

In each of 11 experiments, four calves were exposed first to an aerosol of bovine herpesvirus 1 (BHV1, virus of infectious bovine rhinotracheitis) and second to an aerosol of Pasteurella haemolytica. The interval between aerosols was three to five days. In two other experiments, calves were exposed only to a bacterial aerosol. Climate was controlled for all experiments from the day of viral exposure and for eight of the experiments it was also controlled for four to six days before the first aerosol. The concentration of infectious doses of virus in the aerosols and the number of bacteria in the aerosols of each calf were determined. Macroscopically recognizable rhinitis, tonsillitis, laryngitis, tracheitis and pneumonia of lobar distribution in 42 lobes from 11 calves were seen in five experiments in which bacterial aerosol followed the viral aerosol by at least four days. One calf died with marked respiratory disease in each of four experiments within four days of exposure to the bacterial aerosol. Production of pneumonia was dependent on an interval between aerosols of at least four days but not on the condition of controlled climate on the environmental chamber either before or after the viral aerosol nor on the period of habituation allowed calves of some experiments.     

A Three Year Field Study of Preconditioning Native Illinois Beef Calves Sold Through a Cooperative Marketing Association — 1969 to 1971

During the fall of 1969, 1970 and 1971, Central Illinois practitioners preconditioned (PC) 1,576 beef calves at a cost range of $3.02 to $4.72. The PC program included weaning calves 30 days before sale, having calves eating grain from a bunk and drinking from a tank, vaccinated against blackleg, malignant edema, infectious bovine rhinotracheitis (IBR) and bovine parainfluenza virus (PI3). Calves were tagged in the right ear with the green certified preconditioned for health (CPH) tag of the American Association of Bovine Practitioners. Bovine virus diarrhea (BVD) vaccine was added to the program in 1970 and 1971 since clinical cases of the disease occurred in PC calves not receiving the vaccine in 1969. Intranasal PI3 vaccine and Pasteurella hemolytica and multocida bacterin was added to the 1971 program to attempt better immunization. In 1969, more PC than non-preconditioned (NPC) calves were treated for acute respiratory disease after sale. The 1970 study showed that fewer PC than NPC calves were treated for acute respiratory tract disease. The 1971 finding showed the greatest differences. A comparison of 389 PC and 227 NPC calves sold at auction in 1971 and moved into the same feedlots showed a statistically significant reduction in number of cases of acute respiratory tract disease treated in PC calves. Generally, PC calves sold at a higher price than NPC calves.     

Pulmonary bacterial deposition and clearance during ascarid larval migration in weanling pigs.

Pulmonary deposition and clearance of bacteria were measured in weanling pigs, half of which had been inoculated at age 31 days with larvated Ascaris suum ova. Seven days later, when breathing signs of larval migration were pronounced, all pigs were exposed to aerosolized Escherichia coli (strain B). Then, either immediately after aerosol exposure (for deposition assessment) or immediately after a 120 minute period in filtered air (for clearance), bacteria in the pigs' lungs were counted. Ascarid ova-inoculated pigs did not differ significantly from control pigs for number of bacteria in the lungs after aerosol exposure, but after the 120 minute clearance period they had 7.2 times more than did the control pigs. Thus, in weanling pigs, the breathing-pattern changes that were evident during ascarid-larval migration did not affect pulmonary deposition of inhaled bacteria significantly, but the presence of ascarid larvae in the lungs was associated with impaired pulmonary bacterial clearance.     

Bovine Viral Diarrhea Virus and Escherichia Coli in Neonatal Calf Enteritis

This study was initiated to determine the etiologic and pathogenic significance of an American strain of bovine viral diarrhea (BVD) virus (strain NADL-MD) in enteritis of neonatal calves (calf scours).

Three colostrum-fed calves from dams exposed intravenously to BVD virus at 6, 16 and 25 days prepartum, respectively, had moderate diarrhea persisting until the eighth day of life. The BVD virus was isolated from all 3 calves and persisted up to 93 days in 1 calf, indicating either that BVD was transmitted in utero or via the dam's milk.

Three specific pathogen free (SPF) calves permitted dams' colostrum for the first 4 feedings and then given milk replacer were exposed orally on the day of birth to BVD virus. One calf died of neonatal enteritis 28 hours post-exposure and at necropsy the BVD virus was isolated from several of its organs. The remaining 2 calves had a mild diarrhea persisting to the eighth day of age.

Two calves permitted dams' colostrum ad lib. for 72 hours, and then weaned, were exposed orally to BVD virus. Both calves had a mild persistent diarrhea and BVD virus was isolated from their blood for 56 days post-exposure.

Of 13 SPF, colostrum-deprived calves exposed orally or intranasally at birth to the BVD virus, 4 had severe diarrhea and died of neonatal enteritis from 38 hours to 13 days postexposure. Isolations of BVD virus were made from several of the organs of the calves at necropsy. All of the 9 surviving calves had a moderate to severe diarrhea frequently persisting for 7 to 10 days, and BVD virus was isolated from the survivors up to 103 days postexposure.

Several strains of Escherichia coli were isolated from calves after the second day of life, but were neither pathogenic for mice, nor serologically related to strains of E. coli usually associated with outbreaks of calf scours. Four colostrum-deprived SPF calves were exposed orally at birth to a strain of E. coli isolated from the intestine of the calf with the most acute symptoms and fatal neonatal enteritis. None of the four calves receiving the E. coli had diarrhea. One calf, however, had respiratory distress and died on day 5.

Two SPF colostrum-deprived control calves had neither diarrhea nor respiratory distress.

The above findings support the conclusion that BVD virus should not be overlooked as a primary cause of the neonatal calf enteritis complex.

     

Respiratory disease in calves produced with aerosols of parainfluenza-3 virus and Pasteurella haemolytica.

In four experiments, 22 calves were exposed to aerosols of parainfluenza-3 virus, followed by Pasteurella haemolytica at intervals of three to 13 days. The purpose of each experiment was to study viral-bacterial interactions in the respiratory tracts. Two experiments, in which the viral aerosols were diluted by the addition of air, produced sporadic temperature elevations while two experiments with undiluted viral aerosols produced consistent temperature elevations. Diluted viral aerosols produced lobular sized lesions in the lungs and hemagglutinating inhibition antibodies in sera, whilst undiluted aerosols produced a synergistic effect in the form of purulent pneumonia in ten of 14 calves when the interval between viral and bacterial aerosols was from three to ten days. Histopathological changes attributable to the virus only were seen in all experiments, and the histopathological changes due to mixed infection of parainfluenza-3 virus and P. haemolytica are described in detail. This is the first report of extensive purulent pneumonia in calves after parainfluenza-3 virus and P. haemolytica exposure. This was achieved using much smaller inocula than in experiments previously reported.     

Proliferation of Pasteurella haemolytica in the calf respiratory tract after an abrupt change in climate

Two groups of four calves were exposed to a poly-disperse aerosol of 1.3 X 10(4) to 16 X 10(4) colony forming units (CFU) of nalidixic acid resistant Pasteurella haemolytica type A1 litre-1 of air with a mass median aerodynamic diameter of 2.6 +/- 0.8 microns. One group of calves was kept at 5 degrees C, 72 per cent relative humidity (RH) and the second was subjected to an abrupt change in climate directly after aerosol exposure from 5 degrees C, 75 per cent RH to 13 degrees C, 84 per cent RH. Clearance of the organism from the respiratory tract of the calves was monitored over the subsequent 23 hours by a method of tracheal and nasopharyngeal swabbing. Clearance measured at the trachea in all calves in both groups was not a continuous, uninterrupted process but exhibited a temporary decline between eight and 14 hours. Calves subjected to an abrupt change in climate after aerosol challenge had raised respiratory rates eight to 14 hours later and P haemolytica proliferated in the nasopharynx over the entire 23 hours. There was no apparent effect of climate on P haemolytica in the trachea. It is suggested that rapid growth of P haemolytica accompanying a change in climate may be an important aetiological factor in pneumonic pasteurellosis of calves.