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Rapid and Sensitive Detection Method of a Bacterium by Using a GFP Reporter Phage
Author(s) -
Funatsu Takashi,
Taniyama Tadayoshi,
Tajima Takashi,
Tadakuma Hisashi,
Namiki Hideo
Publication year - 2002
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.2002.tb02708.x
Subject(s) - green fluorescent protein , bacteria , escherichia coli , mycobacterium smegmatis , biology , bacteriophage , microbiology and biotechnology , fluorescence , lambda phage , fluorescence microscope , reporter gene , enterobacteriaceae , gene , biochemistry , gene expression , genetics , mycobacterium tuberculosis , medicine , tuberculosis , physics , pathology , quantum mechanics
A rapid, sensitive, and convenient method for detecting a specific bacterium was developed by using a GFP phage. Here we describe a model system that utilizes the temperate Escherichia coli ‐restricted bacteriophage lambda, which was genetically modified to express a reporter gene for GFP to identify the colon bacillus E. coli in the specimen. E. coli infected with GFP phage was detected by GFP fluorescence after 4–6 hr of incubation. The results show that a few bacteria in a specimen can be detected under fluorescence microscopy equipped with a sensitive cooled CCD camera. When E. coli and Mycobacterium smegmatis were mixed in a solution containing GFP phage, only E. coli was infected, indicating the specificity of this method. The method has the following advantages: 1) Bacteria from biological samples need not be purified unless they contain fluorescent impurities; 2) The infection of GFP phage to bacteria is specific; 3) The fluorescence of GFP within infected bacteria enables highly sensitive detection; 4) Exogenous substrates and cofactors are not required for fluorescence. Therefore this method is suitable for any phage‐bacterium system when bacteria‐specific phages are available.