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  • Title: Filter-based PNA in situ hybridization for rapid detection, identification and enumeration of specific micro-organisms.
    Author: Perry-O'Keefe H, Stender H, Broomer A, Oliveira K, Coull J, Hyldig-Nielsen JJ.
    Journal: J Appl Microbiol; 2001 Feb; 90(2):180-9. PubMed ID: 11168720.
    Abstract:
    AIMS: A method for rapid and simultaneous detection, identification and enumeration of specific micro-organisms using Peptide Nucleic Acid (PNA) probes is presented. METHODS AND RESULTS: The method is based on a membrane filtration technique. The membrane filter was incubated for a short period of time. The microcolonies were analysed by in situ hybridization, using peroxidase-labelled PNA probes targeting a species-specific rRNA sequence, and visualized by a chemiluminescent reaction. Microcolonies were observed as small spots of light on film, thereby providing simultaneous detection, identification and enumeration. The method showed 95-100% correlation to standard plate counts along with definitive identification due to the specificity of the probe. CONCLUSION: Using the same protocol, results were generated approximately three times faster than culture methods for Gram-positive and -negative bacterial species and yeast species. SIGNIFICANCE AND IMPACT OF THE STUDY: The method is an improvement on the current membrane filtration technique, providing rapid determination of the level of specific pathogens, spoilage or indicator micro-organisms.
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