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Identification of outer membrane proteins as target antigens of Pseudomonas aeruginosa Homma serotype M
1Sumitomo Pharmaceuticals Research Center, Osaka, Japan.
Abstract:
Pseudomonas aeruginosa is routinely serotyped in Japan by using the Homma scheme. The serotypes (O serotypes) are based on the chemical structure of the O-polysaccharide portion of the lipopolysaccharide (LPS). However, the nature of the Homma serotype M antigen has remained obscure because strains classified as serotype M usually have the rough phenotype. I characterized the target antigen of serotype M. The results of Western blotting (immunoblotting) showed that commercially available typing monoclonal antibody (MAb) against serotype M specifically bound to outer membrane protein (Opr) G and that typing rabbit antiserum specific for serotype M mainly contained antibodies against Oprs F and H2. These Oprs were distributed among all P. aeruginosa strains tested, including the serotype standard, serotype M and nontypeable strains, and a series of LPS-core-defective mutants derived from strain PAC1. However, the rough mutants derived from strain PAC1 agglutinated with the anti-serotype M antibodies, whereas the smooth strains did not. LPS preparations from serotype M strains possessed few or no polysaccharide chains. These strains had higher levels of binding activity with anti-serotype M MAb, as well as with anti-lipid A MAb, which specifically bound to the cell surface of the rough-natured gram-negative bacterial strains with high activity. The anti-serotype M antiserum also contained rough-LPS-specific antibodies, but the epitope was distributed among only a few strains. The results suggested that the Oprs acted as the serotype M antigen and that LPS did not. In conclusion, the rough strains agglutinated with anti-Opr antibodies and were distinguished as serotype M from the smooth strains of other serotypes, because the antibodies were accessible to the cell surface lacking O polysaccharides. I supposed that Homma serotype M is an index of the rough nature of P. aeruginosa strains rather than one of the O serotypes.
Insights
Homma serotype M in Pseudomonas aeruginosa is not determined by lipopolysaccharide (LPS) but by outer membrane proteins (Oprs). This study reveals serotype M indicates a rough bacterial phenotype, not a specific O antigen.
Area of Science:
- Microbiology
- Immunology
- Bacterial Pathogenesis
Background:
- Pseudomonas aeruginosa serotyping in Japan uses the Homma scheme, based on O-polysaccharide structure of lipopolysaccharide (LPS).
- The antigen for Homma serotype M remains unclear, as M strains typically exhibit a rough phenotype.
Purpose of the Study:
- To characterize the target antigen of Homma serotype M in Pseudomonas aeruginosa.
- To investigate the relationship between serotype M and bacterial phenotype (rough vs. smooth).
Main Methods:
- Western blotting (immunoblotting) using monoclonal antibodies (MAbs) and rabbit antiserum against serotype M.
- Analysis of outer membrane proteins (Oprs) and lipopolysaccharide (LPS) in various P. aeruginosa strains and mutants.
- Agglutination assays with anti-serotype M antibodies.
Main Results:
- Anti-serotype M MAb bound to outer membrane protein G (OprG).
- Anti-serotype M antiserum contained antibodies against Oprs F and H2.
- Serotype M strains showed high binding to anti-serotype M MAb and anti-Lipid A MAb, characteristic of rough strains.
- Rough mutants agglutinated with anti-serotype M antibodies, while smooth strains did not.
Conclusions:
- Outer membrane proteins (Oprs), not LPS, likely function as the serotype M antigen.
- Homma serotype M appears to be an indicator of the rough phenotype in P. aeruginosa, rather than a distinct O serotype.