Membrane vesicles derived from Pseudomonas aeruginosa and Shigella flexneri can be integrated into the surfaces of other Gram-negative bacteria

Incubation of intact Salmonella typhi Ty21a, Salmonella enterica serovar Ty phimurium (Salmonella typhimurium) aroA or Escherichia coli DH5 alpha with membrane vesicles (MVs) derived from either Shigella flexneri M90T or Pseud omonas aeruginosa dsp89 resulted in a significant incorporation of vesicle antigens into the outer membrane of the bacteria; each recipient strain pos sessed a surface mosaic of new Shigella and Pseudomonas antigens intermixed with the native antigens of the Salmonella or Escherichia strains. Electro n microscopy of preparations during the integration of vesicle antigens rev ealed that the MVs rapidly fused with the outer membrane of the host strain s. Western blot analysis of host bacteria confirmed the integration of fore ign antigens, Quantitative analysis far binding and fusion of antigens usin g an ELISA showed that approximately 78.7 +/- 12.8 ng of the Pseudomonas an d 67.5 +/- 13.8 ng of the Shigella LPSs (mu g host protein)(-1) were integr ated into the Sal, typhimurium strain, Similar integrations of the Shigella or Pseudomonas vesicles were found with the E. coli or Sal. typhi strains. There was no loss of viability in the recipient bacteria after incorporati on of the MVs, although vesicle antigens became diluted during continued gr owth as daughter cells shared the vesicle antigens, The new antigens were h ighly stable after being incorporated into recipient strains, being able to withstand storage of several months at 4 degrees C as well as several cycl es of freezing and thawing. Since the recipient bacteria are common vaccine strains, the procedure described here offers a simple efficient means of i ntroducing exogenous surface antigens, in their native form, into the outer membranes of Cram-negative bacteria for possible vaccine use.