IPAB MEDIATES MACROPHAGE APOPTOSIS INDUCED BY SHIGELLA-FLEXNERI

Shigella flexneri kills macrophages through apoptosis, involving the induction of host cell DNA fragmentation and characteristic morphological changes. Shigella can only cause damage it it escapes from the phagolysosome into the cytoplasm. The S. flexneri cytotoxic genes have been localized to the ipa operon of shigella's virulence plasmid. ipaB, C and D deletion mutants are not invasive and therefore not cytotoxic. In order to distinguish genes involved in the escape from the phagolysosome as distinct from cytotoxicity, we constructed Shigella strains that secrete law amounts of Escherichia call haemolysin (hly(low)). These strains can escape into the cytoplasm of the macrophage even in the absence of the invasion plasmid as verified by electron microscopy and resistance to chloroquine. Macrophages were infected with different ipa mutants expressing hly(low). Both Delta ipaC hly(low) and Delta ipaD hly(low) were cytotoxic whilst Delta ipa hly(low) and a hly(low) strain cured of shigella's pathogenicity plasmid were not. Furthermore, both Delta ipaC hly(low) and Delta ipaD hly(low) killed through apoptosis as shown by both changes in ultrastructural morphology and fragmentation of the host cell DNA. These results demonstrate that ipaB is essential for S. flexneri to induce apoptosis in macrophages.