Type III Secretion-Dependent Sensitivity of Escherichia coli O157 to Specific Ketolides

Abstract

A subset of Gram-negative bacterial pathogens uses a type III secretion system (T3SS) to open up a conduit into eukaryotic cells in order to inject effector proteins. These modulate pathways to enhance bacterial colonization. In this study, we screened established bioactive compounds for any that could repress T3SS expression in enterohemorrhagic Escherichia coli (EHEC) O157. The ketolides telithromycin and, subsequently, solithromycin both demonstrated repressive effects on expression of the bacterial T3SS at sub-MICs, leading to significant reductions in bacterial binding and actin-rich pedestal formation on epithelial cells. Preincubation of epithelial cells with solithromycin resulted in significantly less attachment of E. coli O157. Moreover, bacteria expressing the T3SS were more susceptible to solithromycin, and there was significant preferential killing of E. coli O157 bacteria when they were added to epithelial cells that had been preexposed to the ketolide. This killing was dependent on expression of the T3SS. Taken together, this research indicates that the ketolide that has accumulated in epithelial cells may traffic back into the bacteria via the T3SS. Considering that neither ketolide induces the SOS response, nontoxic members of this class of antibiotics, such as solithromycin, should be considered for future testing and trials evaluating their use for treatment of EHEC infections. These antibiotics may also have broader significance for treating infections caused by other pathogenic bacteria, including intracellular bacteria, that express a T3SS.

Ribosome maturation by the endoribonuclease YbeY stabilizes a type 3 secretion system transcript required for virulence of enterohemorrhagic Escherichia coli

“…Recent work has demonstrated that the ketolide antibiotics solithromycin and telithromycin inhibit ribosome maturation ( 51 ) and are potent inhibitors of T3S in EHEC ( 52 ). Our results suggest that these antibiotics block T3S by depleting mature ribosomes in an analogous manner to the ybeY deletion.…”

“…Type 3 secretion profiles and Western blotting for RecA and EspD were performed as described previously ( 52 ) using antibodies described previously ( 7 ). Where indicated, 50 μg·ml −1 kasugamycin was added to the supplemented MEM-HEPES before inoculation.…”

A New, Potent, and Placenta-Permeable Macrolide Antibiotic, Solithromycin, for the Prevention and Treatment of Bacterial Infections in Pregnancy

“…Solithromycin has been shown to exhibit excellent activity against many of the microbial species known to be associated with infection-associated preterm delivery (Table 1 ), including Ureaplasma spp., Mycoplasma spp., Group B streptococci, staphylococci, and Chlamydia trachomatis ( 59 , 61 , 65 – 67 , 73 , 74 ). Indeed, although solithromycin has not yet been tested on all relevant microorganisms, from the established efficacy profile of its parent drug clarithromycin, it is likely that solithromycin will be effective against all bacteria known to be associated with intra-amniotic infection.…”

An RNA-dependent mechanism for transient expression of bacterial translocation filaments

“…For bacterial lysates, bacterial pellets were suspended directly in SDS PAGE loading buffer. Proteins were separated by SDS-PAGE using standard methods and Western blotting performed as described previously for EspD, RecA and EscJ ( 33 , 34 ). Bacteria were stained for EspA filaments following fixation with 4% paraformaldehyde for 5 min.…”