Although the invasion and inflammatory phenotypes are the best studied pathogenic mechanisms of Shigella infection, clinical data show that a considerable number of patients develop a self-limiting watery diarrhea (Keusch et al., 1986; Vargas et al., 1999). These clinical observations led to the description of two candidate enterotoxins

in Shigella flexneri, called ShET-1 and ShET-2, encoded on the chromosome and the Inv virulence plasmid, respectively (Fasano et al., 1995; Nataro et al., 1995). ShET-2 was initially described in enteroinvasive Escherichia coli strain EI-34, but was also found in most isolates of the Doxorubicin research buy genus Shigella (Nataro et al., 1995; Vargas et al., 1999). The protein was purified after recombinant gene expression and was found to induce rises in short-circuit current in rabbit intestinal tissue mounted in the Ussing chamber (Nataro et al., 1995). Recently, vaccine trials using live attenuated Shigella strains with deletions in the genes encoding ShET-1 and ShET-2 suggested that one or both of these toxins contribute to virulence in humans (Kotloff et al.,

2000, 2004, 2007). More thorough characterization of these two factors is therefore warranted. Multiple virulence factors of Shigella spp. are secreted by type III secretion systems (T3SS) or by the autotransporter (type V) mechanisms. However, no experimental data have been published implicating Galunisertib clinical trial either of these mechanisms for ShET-1 or ShET-2 secretion. over Notably, neither putative toxin exhibits a typical Gram-negative signal sequence (Nataro et al., 1995) and no signature suggesting T3SS-dependent translocation has been reported. The Shigella T3SS, encoded on the 31-kb Inv plasmid-encoded entry region, comprises a multiprotein bacterial complex that forms a needle-like structure, termed the injectosome; this nanomachine mediates the translocation

of bacterial effector proteins directly to the eukaryotic cytoplasm (Mota & Cornelis, 2005). In Shigella, the T3SS is induced upon contact of the bacteria with epithelial cells (Watarai et al., 1995) or by adding Congo red (CR) dye to the growth medium (Bahrani et al., 1997). Constitutive secretion of T3SS effectors is observed after inactivation of the ipaB or the ipaD genes (Menard et al., 1994). In an S. flexneriΔipaBCDA mutant, 14 other type III effectors encoded on the Inv virulence plasmid were identified and designated as outer Shigella proteins (Osp proteins). These proteins were organized in groups OspB to OspG according to similarities in their amino-acid sequence (Buchrieser et al., 2000). The OspD group includes three members: OspD1 (a proven type III effector) (Parsot et al., 2005), OspD2 (of unknown function) and OspD3 (also known as ShET-2). Notably, this first report did not directly document dependence of OspD3 secretion on the T3SS.