论文部分内容阅读
Advanced methods are urgently needed to determine the identity and viability of trace amount of pathogenic bacteria in a short time.Existing approaches either fall short in the accurate assessment of microbial viability or lack of specificity in bacterial identification.Bacteriophages are viruses that exclusively infect bacteria with high specificity.Employing a tetracysteine (TC) peptide tag as the phage reporter gene and a biarsenical dye (FlAsH-EDT2) as the fluorescent stain,we developed a sensitive and specific method for bacterial detection in an earlier report.Because phages infect and replicate only in living bacterial hosts,here we exploit the so called "TC-phage-FlAsH" strategy to the discriminative detection of viable target bacteria from dead target cells and other viable but non-targeting bacterial strains.Using recombinant M13KE-TC phage and E.coli ER2738 as a model system,distinct separation between the populations of viable and dead E.coli ER2738 was obtained by flow cytometry and fluorescence microscopy.When the viable and dead E.coli ER2738 were mixed with varying percentages,the measured ratios correlated nicely with the theoretical mixing ratios.With fluorescence microscopy,the proposed method allows specific detection of as few as 1 cfu/mL live target bacteria in the presence of a large excess of dead target cells and other viable but non-targeting bacterial strains in artificially contaminated drinking water samples.The TC-phage-FlAsH method is sensitive,rapid,and simple,and thererefore would be particularly useful for trace detection and identification of viable bacterial pathogens.