Salmonella typhimurium MMP13 harboring pMMP99, a recombinant plasmid derived by cloning a ghost cassette into
T-vector, were employed for production of ghost cells. Growth of MMP13 (pMMP99) showed an initial increase after a
shift in temperature from 28 to 42°C, and then decreased gradually with ghost formation being complete within 3 h.
Ghost yield of MMP13 (pMMP99) were 99.99% less than 104 CFU/mL. MMP13 (pMMP101-1) showed a ghost yield similar
to MMP13 (pMMP99). Immune responses of BALB/c mice against ghost cells originated from MMP13 (pMMP101-1) were
assessed by measuring total IgG, IgG1, IgG2a, and secretory IgA levels in each sample. Levels of total IgG, IgG1 and
IgG2a and vaginal IgA, increased abruptly after 4 weeks post vaccination, whereas the fecal IgA level did not induce
significant change. Splenocyte proliferation was observed at the cellular level owing to stimulation of ghost cells. Ghost
vaccination protected 25-59% of mice against wild-type S. typhimurium more than those of controls.