Evaluation of the hemoglobin-binding activity of Actinobacillus pleuropneumoniae using fluorescein-labeled pig hemoglobin and flow cytometry.
In the present study, the hemoglobin (Hb)-binding activity of Actinobacillus pleuropneumoniae was examined using fluorescein-labeled pig Hb and flow cytometry. Comparison of the Hb-binding activity of A. pleuropneumoniae serotype 1 strain 4074 grown under iron-restricted conditions with cells grown under iron-sufficient conditions indicated that iron-restriction in A. pleuropneumoniae promotes the expression of Hb receptors, and that Hb-binding activity is, at least in part, iron-repressible. Hb-binding activity was also observed in representative strains of A. pleuropneumoniae belonging to serotypes 1 and 2. In addition, A. pleuropneumoniae serotype 1 LPS or capsule isogenic mutants were tested in flow cytometry in order to understand the influence of surface polysaccharides on Hb-binding activity. Experiments with an acapsulated mutant indicated that surface molecules with Hb-binding activity are more exposed at the cell surface in the absence of capsular polysaccharides. However, the Hb-binding activity of LPS mutants analyzed in this study was unchanged compared to the parent strain. The outer membrane proteins profile of A. pleuropneumoniae serotype 1 grown under iron-restricted or iron-sufficient conditions was also evaluated by polyacrylamide gel electrophoresis. Iron-regulated outer membrane proteins were observed under iron-restricted growth conditions which suggests that one or more of these outer membrane proteins may play a role in the Hb-binding activity detected by flow cytometry.
Archambault
M
M
Groupe de Recherche sur les Maladies Infectieuses du Porc, Faculté de Médecine Vétérinaire, Université de Montréal, Qué, Canada.
Rioux
S
S
Jacques
M
M
D000189:Actinobacillus Infections / Q000382:microbiology / Q000662:veterinary; D016977:Actinobacillus pleuropneumoniae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry / Q000379:methods; D019793:Fluorescein / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism*; D007501:Iron / Q000378:metabolism; D012703:Serotyping; D013552:Swine
D003470:Culture Media; D006454:Hemoglobins; D007501:Iron; D019793:Fluorescein