The Immunochemistry of Serratia Marcescens

Abstract

Protein profiles of the outer membranes (OMs) of Serratia marcescens New CDC 014:H12, isolated from cells grown in various iron-rich and iron-restricted media, were investigated using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Three major proteins were separated: 43.5 and 42 kDal (the porins), 38 kDal (the OmpA protein), and a group of iron-regulated proteins (IRMPs) of 63–70 kDal. Immunoblotting of whole cells or OMs using antiserum raised against whole cells revealed similar complex patterns of antigens. The OmpA protein was the major immunogen, although other OM proteins were also detected; the porins and IRMPs reacted only weakly with antibodies in this system. Immunoabsorption of antiserum with whole cells showed that only the O-antigenic chains of lipopolysaccharide (LPS) and the H (flagella) antigens were accessible to antibodies on the cell surface. Failure to detect the OmpA and other envelope proteins suggests that their antigenic sites are not able to react with antibodies and are possibly masked by the O antigen. The sensitivities of two 014:H12 strains, New CDC and 4444-60, to a range of antimicrobial agents were very similar. The third 014:H12 strain, S1220, was particularly resistant, suggesting the presence of a plasmid. The response of the cells to killing by normal pooled human serum varied significantly. NewCDC was either sensitive or resistant, depending on the media it was grown in, whereas 4444-60 was resistant, and S1220 sensitive, independent of growth media. Complement binding, studied by measuring hydrophobicity and using rocket immunoelectrophoresis with antihuman C3, showed that the sensitive cells (S1220) rapidly bound complement, whereas the resistant cells (4444-60) bound less C3b. Crossed immunoelectrophoresis suggested that in S1220 cells, the polysaccharide material, including LPS, was less antigenic and present in smaller amounts than in 4444-60. Examining extracted polysaccharide material chemically and by SDS-PAGE showed that the resistant strain had 33% more material than the sensitive strain, comprising LPS with longer O-antigen chain lengths and/or a microcapsule of O-antigen polysaccharide. The extra polysaccharide material on the surface of 4444-60 cells likely prevents complement components from binding and reaching the hydrophobic membrane, where lytic lesions occur.