Characterization of Actinobacillus pleuropneumoniae hemolysin and its role in porcine Actinobacillus pleuropneumonia.

摘要

The present study was conducted to investigate the characteristics of Actinobacillus pleuropneumoniae hemolysin(s) in cell-free culture supernatant fluid (CFCSF) and to further comprehend its role in the disease process. The purpose of the first phase of this study was to determine whether the strains of A. pleuropneumoniae representative of 7 serotypes exhibit any hemolytic activity and to recognize factors that affect the activity of the hemolysin(s). It was found that the hemolysin(s) present in CFCSF of each of 6 of the serotype strains tested exhibited hemolytic activity which was due to the presence of a heat-labile protein in the CFCSF. The hemolysin of the serotype 1 strain was cytotoxic for porcine neutrophils.;The second phase of this study was designed to examine by electron microscopy changes occurring in cultured porcine neutrophils following incubation CFCSF prepared from the 4074 serotype 1 strain of A. pleuropneumoniae or highly purified lipopolysaccharide (LPS) obtained from the same strain. Sodium dodocyl sulphate polyacrylamide electrophoresis (SDS-PAGE) of the CFCSF revealed that the hemolysin (107 kDa protein) was the primary component of the CFCSF, but a small amount of an additional protein (120 kDa) was also present. The hemolysin in the CFCSF was responsible for neutrophil cell swelling, loss of pseudopodia, granule margination, and complete nuclear degeneration after incubation of the porcine neutrophils with the CFCSF from 2 to 15 min. No significant changes were detected in the neutrophils following incubation with LPS for as long as 20 min.;The progressive neutrophil degeneration of the in vitro study provoked the in vivo investigation by inoculating CF