Examination of surface polysaccharides of Actinobacillus pleuropneumoniae serotype. 1 Grown under iron-restricted conditions

Abstract

We investigated the expression of important Actinobacillus pleuropneumoniae surface polysaccharides, namely, capsular polysaccharides (CPS) and lipopolysaccharides (LPS), after growth under iron-restricted conditions. Iron restriction did not seem to affect the production of CPS, as determined by labelling with a monoclonal antibody (mAb) against the serotype 1 K-antigen and flow cytometry analysis, and also as determined by electron microscopy. SDS-PAGE revealed that the LPS profiles of these cells were also unaffected by iron restriction. Using flow cytometry analysis, however, we observed that binding of mAb against serotype 1 O-antigen was altered in cells of A. pleuropneumoniae serotype 1 reference strain (4074) grown under iron-restricted conditions. This strain exhibited two subpopulations with distinct patterns of reactivity with the mAb against the O-antigen. When strain 4074 was grown under iron-restricted conditions, a shift from one cell subpopulation (moderately fluorescent) to another cell subpopulation (highly fluorescent, thus binding more antibodies) was observed. Our results indicate that growth of A. pleuropneumoniae serotype 1 under iron-restricted conditions did not seem to affect CPS production, but might alter, at least for the reference strain, the expression of LPS.

Production and characterization of murine monoclonal antibodies against Haemophilus parasuis and study of their protective role in mice

“…LPS is an important virulence factor of A. pleuropneumoniae (Haesebrouck et al, 1997;Tascon et al, 1996) and a major adhesin involved in adherence to porcine respiratory tract cells (Jacques et al, 1991;Bélanger et al, 1990), mucus (Bélanger et al, 1994(Bélanger et al, , 1992 and to host glycosphingolipids (Abul-Milh et al, 1999). Paradis et al (1996) demonstrated that LPS can traverse the thick capsular material and reach the outer-most region of the cell. This may explain the positive reaction obtained in the colony-blotting assay and electron microscopy with MAb 4G9 using whole cells in our studies (Fig.…”

Actinobacillus pleuropneumoniaesurface polysaccharides: their role in diagnosis and immunogenicity

“…On the other hand, elucidation of LPS O-chain structures (Perry et al, 1990) revealed that some serotype specificity is associated with this important antigen, which can be purified and obtained in large quantities in a few steps. In addition, the surface exposure of A. pleuropneumoniae LPS was demonstrated by Paradis et al (1996), who showed that outer membrane blebs crossed the capsular layer. These antigens can trigger the immune response and thus establish humoral immunity (Munford, 1991).…”

“…Even though a capsule is present at the surface of this microorganism, recent studies have revealed that LPS can traverse the thick capsular material and reach the outmost region of the cell (Paradis et al, 1996). This observation is of prime importance if we consider that the development of a diagnostic tool or a vaccine should be based on molecules that are easily accessible to the host's immunological responsive cells and antibodies during the infection process.…”

“…Recently, microparticlebased agglutination tests have been used for the detection or identification of a wide variety of antigens and antibodies. In an attempt to develop a simple, convenient and more reliable serotyping method, a serotype 5 murine monoclonal antibody directed against the LPS O-chain was used to sensitize protein A-coated polystyrene microparticles (Dubreuil et al, 1996). The advantage of such a test resides in the use of monoclonal instead of polyclonal antibodies.…”