Rapid and sensitive detection method of a bacterium by using a GFP reporter phage

Microbiol Immunol. 2002;46(6):365-9. doi: 10.1111/j.1348-0421.2002.tb02708.x.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacteriophage lambda / genetics*
  • Bacteriophage lambda / metabolism
  • Escherichia coli / isolation & purification*
  • Escherichia coli Infections / microbiology*
  • Green Fluorescent Proteins
  • Humans
  • Indicators and Reagents / metabolism
  • Luminescent Proteins / genetics*
  • Microscopy, Fluorescence
  • Microscopy, Phase-Contrast
  • Sensitivity and Specificity

Substances

  • Indicators and Reagents
  • Luminescent Proteins
  • Green Fluorescent Proteins