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KMID : 0545120070170020234
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Journal of Microbiology and Biotechnology
2007 Volume.17 No. 2 p.234 ~ p.243
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IVET-based Identification of Virulence Factors in Vibrio vulnificus MO6-24/O
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Lee Ko-Eun
Bang Ji-Sun
Baek Chang-Ho
Park Dae-Kyun
Hwang Won
Choi Sang-Ho
Kim Kun-Soo
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Abstract
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Vibrio vulnificus is an opportunistic pathogen that causes septicemia in humans. To identify the genes associated with its pathogenicity, in vivo expression technology (IVET) was used to select genes specifically expressed in a host, yet not significantly in vitro. Random lacZ-fusions in the genome of V. vulnificus strain MO6-24/O were constructed using an IVET vector, pSG3, which is a suicide vector containing promoterless-aph and -lacZ as reporter genes. A total of ~18,000 resulting library clones were then intraperitoneally injected into BALB/c mice using a colony forming unit (CFU) of 1.6×106. Two hours after infection, kanamycin was administered at 200 μg per gram of mouse weight. After two selection cycles, 11 genes were eventually isolated that were expressed only in the host. Among these genes, VV20781 and VV21007 exhibiting a homology to a hemagglutinin gene and tolC, respectively, were selected based on having the highest frequency. When compared to wild-type cells, mutants with lesions in these genes showed no difference in the rate of growth rate, yet a significant decrease in cytotoxicity and the capability to form a biofilm.
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KEYWORD
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Vibrio vulnificus, IVET, virulence factor, TolC
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