Smooth lipopolysaccharide is the major protective antigen for mice in the surface extract from IATS serotype 6 contributing to the polyvalent Pseudomonas aeruginosa vaccine PEV

Abstract

The nature of the protective antigen in one of the sixteen monovalent extracts (viz., extract-6) contributing to the pseudomonas polyvalent extract vaccine (PEV) was studied in a mouse challenge assay. Selective removal, by filtration through Sep-Pak C18 cartridges, of two major protein antigens with molecular weights of 16,200 and 21,000 had no effect on the protection afforded by extract-6. When analyzed on the basis of 2-keto-3-deoxyoctonate, lipopolysaccharide (LPS) purified by hot phenol extraction (LPS-A) from Pseudomonas aeruginosa (International Antigenic Typing System serotype 6) could account in full for the protective capacity of extract-6. Comparative analysis of LPS heterogeneity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by silver staining indicated that both extract-6 and LPS-A possessed similar spectra of smooth LPS molecules, containing between 10 and approximately equal to 50 O-antigen repeating units. Differences in the profiles of heterogeneity displayed by LPS in LPS-A and extract-6 were restricted to molecular species with short O-antigen chains. Subfractionation of LPS molecules on the basis of number of O-antigen repeating units was achieved by gel filtration in the presence of deoxycholate. Protection experiments performed on the subfractionated species of LPS-A revealed a relationship between O-antigen chain length and protective capacity; molecules with over 18 O-antigen repeating units being 50 to 100 times more protective than those with zero-two repeating units. The results indicate that most of the protection afforded by LPS-A and extract-6 can be accounted for by LPS molecules possessing extended (10 or more) O-antigen repeating units.