Effect of pstC mutation on expression of LEE genes in Escherichia coli O157:H7

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Effect of pstC mutation on expression of LEE genes in Escherichia coli O157:H7

 

Author: Wong, Tsz-yeung
Title: Effect of pstC mutation on expression of LEE genes in Escherichia coli O157:H7
Degree: M.Sc.
Year: 2012
Subject: Escherichia coli O157:H7 -- Genetic aspects.
Hong Kong Polytechnic University -- Dissertations
Department: Dept. of Health Technology and Informatics
Pages: xiv, 109 leaves : ill. (some col.) ; 30 cm.
Language: English
InnoPac Record: http://library.polyu.edu.hk/record=b2475281
URI: http://theses.lib.polyu.edu.hk/handle/200/6445
Abstract: Escherichia coli O157:H7 produces Shiga-like toxin or verotoxin as its virulence factor which can cause diseases such as severe watery diarrhea, bloody diarrhea, fever, abdominal cramps or vomiting. It may lead to serious complications including hemolytic uremic syndrome (HUS), thrombotic thrombocytopenic purpura (TTP) or even death. The pathogen is able to form attaching and effacing (A/E) lesions in infected epithelial cells. Occurrence of A/E lesions involves the localized effacement of the brush microvilli and assembly of complicated actin structures in the epithelial cells beneath the intimately attached bacterium. The locus of enterocyte effacement (LEE) pathogenicity island (PAI) of E. coli O157:H7 encodes both translocators and effector molecules responsible for production of A/E lesions in the infected host cells. Phosphate plays important roles in many biochemical reactions within a bacterial cell. Pst system belonging to the Pho regulon is encoded by the pstSCAB-phoU (pst) operon. Studies have shown that impaired pst operon reduced adhesion of EPEC to host cells. However, the same phenomenon occurs in EHEC is not known. Besides, the underlying mechanism is not well-studied. It was postulated that problematic pstC gene might influence the expression of LEE genes regulator genes, which led to impaired production of A/E lesions. In this study, expression levels of LEE genes and LEE regulators genes were compared between a wild type E. coli serotype O157:H7 and its pstC isogenic mutants. Gene expressions were determined using real-time reverse transcription polymerase chain reaction (RT-PCR). For the LEE genes, it was found that escJ (LEE2), escV (LEE3), espA (LEE4) and tir (LEE5) genes were up-regulated under higher phosphate concentration. This indicated that the mutated pstC gene of E. coli O157:H7 could affect the LEE expression and adhesion of the pathogen, and that phosphate could act as a regulatory factor for LEE genes. For the LEE regulators, this study showed that expression of LEE-encoded regulators ler, grlA and grlR were up-regulated in the pstC mutants under normal and increased phosphate concentration (10mM). grlA and grlR showed similar levels of gene expression regardless of normal or increased phosphate concentrations. This suggested that expression of these three genes, ler, grlA and grlR, could be affected by the impaired pstC in the mutants whilst expression of ler could be dependent on phosphate concentration. pchA in the mutant was up-regulated to similar level under normal and high phosphate concentrations. pstC mutants grown under higher phosphate concentration showed up-regulation of pchB, hns, qseA and sdiA. In summary, mutation of pstC gene of E. coli O157:H7 could affect the expression of LEE genes and LEE regulators whereas phosphate concentration could probably be a factor which could raise effect on gene expression through Pho regulon.

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