Enterohemorrhagic Escherichia coli O157∶H7 Gene Expression Profiling in Response to Growth in the Presence of Host Epithelia

BACKGROUND: The pathogenesis of enterohemorrhagic Escherichia coli (EHEC) O157:H7 infection is attributed to virulence factors encoded on multiple pathogenicity islands. Previous studies have shown that EHEC O157:H7 modulates host cell signal transduction cascades, independent of toxins and rearrangement of the cytoskeleton. However, the virulence factors and mechanisms responsible for EHEC-mediated subversion of signal transduction remain to be determined. Therefore, the purpose of this study was to first identify differentially regulated genes in response to EHEC O157:H7 grown in the presence of epithelial cells, compared to growth in the absence of epithelial cells (that is, growth in minimal essential tissue culture medium alone, minimal essential tissue culture medium in the presence of 5% CO(2), and Penassay broth alone) and, second, to identify EHEC virulence factors responsible for pathogen modulation of host cell signal transduction. METHODOLOGY/PRINCIPAL FINDINGS: Overnight cultures of EHEC O157:H7 were incubated for 6 hr at 37 degrees C in the presence or absence of confluent epithelial (HEp-2) cells. Total RNA was then extracted and used for microarray analyses (Affymetrix E. coli Genome 2.0 gene chips). Relative to bacteria grown in each of the other conditions, EHEC O157:H7 cultured in the presence of cultured epithelial cells displayed a distinct gene-expression profile. A 2.0-fold increase in the expression of 71 genes and a 2.0-fold decrease in expression of 60 other genes were identified in EHEC O157:H7 grown in the presence of epithelial cells, compared to bacteria grown in media alone. CONCLUSION/SIGNIFICANCE: Microarray analyses and gene deletion identified a protease on O-island 50, gene Z1787, as a potential virulence factor responsible for mediating EHEC inhibition of the interferon (IFN)-gamma-Jak1,2-STAT-1 signal transduction cascade. Up-regulated genes provide novel targets for use in developing strategies to interrupt the infectious process

Getting to Know Your Microbiota in Health and Disease

<p><strong>Review of:</strong><em> </em><em>The Human Microbiota: How Microbial Communities Affect Health and Disease</em>; David N. Fredricks (ed.); (2013). John Wiley and Sons, Hoboken, NJ. 362 pages.</p

Clicker Student Response Systems: Dedicated Physical Devices or Web-Enabled Systems that Allow Student to Bring Their Own Device (BYOD)

The implementation of clickers and clicker questions into classroom teaching is a well-established active-learning technique that provides more interactive classrooms and engages students with course content.  Throughout the years, however, the actual clickers themselves have undergone an evolution from physical hand-held devices to web-based applications.  This review explores the general advantages and disadvantages of two-main clicker styles, dedicated physical devises and web enabled clickers

Introductory Immunology: Begin the Journey

<p><strong>Review of</strong><em>: </em><em>Primer</em><em> to the Immune Response</em>, 2^nd ed.; Tak Mak, Mary Saunders, and Bradley Jett; (2014). Elsevier Inc., London, UK. 702 pages.</p

An Incredibly Fun and Easy Way to Learn Clinical Microbiology

<em></em><strong>Review of</strong>: <em>Clinical Microbiology Made Ridiculously Simple</em>, 5th ed.; Mark Gladwin and William Trattler; (2011). MedMaster Inc., Miami, FL. 400 pages

Conditioned Medium from Enterohemorrhagic Escherichia coli-Infected T84 Cells Inhibits Signal Transducer and Activator of Transcription 1 Activation by Gamma Interferon

Gamma interferon (IFN-γ) is a cytokine important to host defense which can signal through signal transducer and activator of transcription 1 (Stat1). Enterohemorrhagic Escherichia coli (EHEC) modulates host cell signal transduction to establish infection, and EHEC serotypes O113:H21 and O157:H7 both inhibit IFN-γ-induced Stat1 tyrosine phosphorylation in vitro. The aim of this study was to delineate both bacterial and host cell factors involved in the inhibition of Stat1 tyrosine phosphorylation. Human T84 colonic epithelial cells were challenged with direct infection, viable EHEC separated from T84 cells by a filter, sodium orthovanadate, isolated flagellin, bacterial culture supernatants, and conditioned medium treated with proteinase K, trypsin, or heat inactivation. Epithelial cells were then stimulated with IFN-γ and protein extracts were analyzed by immunoblotting. The data showed that IFN-γ-inducible Stat1 tyrosine phosphorylation was inhibited when EHEC adhered to T84 cells, but not by bacterial culture supernatants or bacteria separated from the epithelial monolayer. Conditioned medium from T84 cells infected with EHEC O157:H7 suppressed Stat1 activation, and this was not reversed by treatment with proteinases or heat inactivation. Use of pharmacological inhibitors showed that time-dependent bacterial, but not epithelial, protein synthesis was involved. Stat1 inhibition was also independent of bacterial flagellin, host proteasome activity, and protein tyrosine phosphatases. Infection led to altered IFN-γ receptor domain 1 subcellular distribution and decreased expression in cholesterol-enriched membrane microdomains. Thus, suppression of host cell IFN-γ signaling by production of a contact-dependent, soluble EHEC factor may represent a novel mechanism for this pathogen to evade the host immune system