Effects of low molecular weight compounds on growth, virulence gene expression and cytotoxic activity of Shiga toxin-producing Escherichia coli O157:H7
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[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT REQUEST OF AUTHOR.] Shiga toxin-producing Escherichia coli (STEC) infections are clinically associated with hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS), and are the leading cause of acute renal failure in children. A series of novel low molecular weight chemical compound (LMWC) capable of inhibiting various virulence gene expressions in E. coli O157:H7 has been identified, which could help mitigate its pathogenicity at certain time points in its infection. Our own previous studies showing significant down-regulation of virulence gene expression in group A Streptococcus and Staphylococcus aureus by this novel class of low molecular weight compounds, prompted the theory that their analogs may cause similar effects with STEC virulence genes. Recent preliminary experiments have generated quite promising evidence that this is indeed true. The aim of this study was to systematically determine the influence of three LMWC on the growth and expression of the Shiga toxin genes, stx1 and stx2, and the intimin gene, eaeA, by STEC serotypes (O157:H7 and non-O157), and to investigate the effect of compound NV002 on human kidney (HK-2) cells based on the influence of the compound on the expression of major virulence genes in E. coli O157:H7 strains. Five STEC O157:H7 strains (C7927, 43894, 93-111, OK-1, G5101) growth was monitored by measuring optical density (OD600 nm) and viable cell count over 10 h. qPCR was performed on cDNA generated at mid-logarithmic (MLP) (3.5 h), late logarithmic (LLP) (6.5 h) and stationary (SP) (20 h) growth phases to monitor gene expression. HK-2 cells were exposed to different concentrations of cell-free extracts or live cells following treatment of the cells with NV002 or DMSO for 3.5 h, 6.5 h, or 20 h and cell viability was measured by the MTT assay. Results showed that these compounds function by inhibiting virulence gene expression in STEC, but not affecting the viability of the bacteria, while there was no significant difference in HK-2 cell viability for either bacterium treated with NV002 and DMSO.
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