Knockout of the mltA gene gives bacteria that spontaneously release vesicles that are rich in immunogenic outer membrane proteins and that can elicit cross-protective antibody responses with higher bactericidal titres than OMVs prepared by normal production processes. Thus the invention provides a bacterium, such as an E.coli, having a knockout mutation of its mltA gene. The invention also provides a Escherichia bacterium, wherein the bacterium: (i) has a cell wall that includes peptidoglycan； and (ii) does not express a protein having the lytic transglycosylase activity of MItA protein. Tol-Pal knockouts of pathogenic E.coli are also disclosed. The invention also provides compositions comprising vesicles that, during culture of bacteria of the invention, are released into the culture medium.
