Five strains of Actinobacillus pleuropneumoniae serotype 1 were used to intranasally infect 5 groups of pigs. Using each bacterial strain, infected pigs (termed seeder pigs) were commingled for 48 hours with 5 groups of noninfected test pigs, then were removed. Seeder and test pigs were maintained in isolation and were observed for 14 days. Seeder pigs had mortality that was threefold greater than that of test pigs (24% vs 8%). Rectal temperature in excess of 40.3 C was achieved for 84% of test pigs and 88% of seeder pigs. Neither of these 2 variables was statistically different between the 2 groups of pigs. Clinical impression scores greater than or equal to 2 (on a 0 to 3 scale) were three-fold (64% vs 20%) greater for seeder than for test pigs (P < 0.05). The total number of bacterial isolations or nonrecoverable isolates was tabulated for test and seeder pigs' lungs at necropsy, irrespective of the amount of lesions. The number of A pleuropneumoniae isolations was not statistically different between test and seeder pig populations. Recovery of Pasteurella multocida or other bacteria was greater from the seeder pigs (P < 0.05), whereas the number of non-recoverable isolates was greater from test pigs than from seeder pigs (P < 0.05). Assessment of lung lesions at necropsy by either visual estimation or on a weight basis were in agreement. Fewer test pigs had lung lesions in excess of 5% of total lung volume than did seeder pigs (40% vs 84%) and, according to the odds ratio estimation, seeder pigs were 7 times more likely than test pigs to have such lesions. These results indicate a predictable, moderate intensity, natural exposure model for use in the study of Actinobacillus pleuropneumoniae-induced pneumonia. The seeder pig model appears to mimic field infection in development of clinical illness, febrile response, lung lesions, mortality, and low potential for secondary pneumonic bacterial involvement, thus providing a useful tool for preliminary evaluation of anti-infective modalities.

Alterations in the size and functions of porcine alveolar macrophages exposed to sublytic amounts of heat-labile, hemolytic cytotoxin produced by Actinobacillus pleuropneumoniae (App) serotype 1, strain HS54 into the culture medium were studied in vitro. Alveolar macrophages were sensitive to the cytotoxin; treatment of the macrophages with low concentrations of cytotoxin (0.016 hemolytic unit) resulted in severe, irreversible cell swelling. However, high doses of cytotoxin (2.0 hemolytic units) were required to cause substantial cell death, as indicated by the influx of propidium iodide into and release of lactate dehydrogenase from cells. Macrophages exposed to low, sublytic doses of cytotoxin failed to migrate toward chemoattractant, were unable to attach to glass, and failed to phagocytize optimally opsonized erythrocytes. Macrophages already attached to glass surfaces detached when exposed to sublytic doses of cytotoxin. The swelling and impairment of functions of alveolar macrophages observed in this study could not be attributed to endotoxic effects, because heat treatment of the cytotoxin preparation for 60 minutes at 60 C resulted in complete loss of cytotoxicity. We conclude that sublytic doses of heat-labile, hemolytic cytotoxic substances produced by App depress alveolar macrophage function at concentrations likely to develop in association with acute pulmonary infection with App. The Apx (A pleuropneumoniae Rtx toxins) exotoxins secreted by the bacteria into culture medium were considered responsible for the toxic activity of the cytotoxin preparation. The Apx of the App field strain used in this study were likely to be similar to those of serotype-1 reference strain (S4707). Analysis by use of DNA-DNA hybridization indicated that genomic DNA of the field strain contained sequences similar to those encoding structural protein of ApxI (apxIA) and ApxII (apxIIA) of the serotype-1 reference strain. Therefore, Apx produced by the field strain of App used in this study are likely to be of similar pathogenic importance worldwide.

Clinical trials have shown that currently available commercial vaccines against porcine pleuropneumonia provide inconsistent, serotype-specific protection from the disease. Recovery from naturally acquired infection, however, provides solid, serotype crossprotective immunity. We examined various serum responses of pigs receiving 1 of 4 commercial vaccines or a cell extract, and compared the serologic responses of these pigs after challenge exposure with virulent Actinobacillus pleuropneumoniae serotype 1. Evaluation of serum included complement-mediated killing opsonizing capacity, IgG titers to whole organisms, and cytotoxin neutralization titers. Pigs that received the cell extract had fewer clinical signs of pleuropneumonia than pigs in other vaccinated groups, and also were significantly (P < 0.05) better protected from development of lung lesions and death. Such vaccinates were the only pigs that developed significant (P < 0.05) serum antibody titers (ie, protective immune response) to whole-cell antigens and to cytotoxin.