Fluorescence in situ hybridization using peptide nucleic acid probes for rapid detection of Mycobacterium avium subsp. avium and Mycobacterium avium subsp. paratuberculosis in potable-water biofilms

Abstract

Here, we present for the first time a high-affinity peptide nucleic acid (PNA) oligonucleotide sequence for detecting Mycobacterium avium bacteria, including the opportunistically pathogenic subspecies M. avium subsp. avium, M. avium subsp. paratuberculosis, and M. avium subsp. silvaticum, by the fluorescence in situ hybridization (FISH) method. There is evidence that M. avium subsp. avium especially is able to survive and grow in drinking-water biofilms and possibly transmit via drinking water. The designed PNA probe (MAV148) specificity was tested with several bacterial species, including other mycobacteria and mycolic acid-containing bacteria. From the range of bacterial strains tested, only M. avium subsp. avium and M. avium subsp. paratuberculosis strains were hybridized. The PNA FISH method was applied successfully to detect M. avium subsp. avium spiked in water samples and biofilm established within a Propella biofilm reactor fed with potable water from a distribution supply.

FIG. 1.

Epifluorescence photomicrograph of FISH-labeled M. avium subsp. avium bacteria in the outlet water concentrate of the Propella reactor supplied with potable water.