Polymerase chain reaction to detect the pathogenic Naegleria fowleri: application to water samples

Search of Naegleria fowleri in water samples was performed by using polymerase chain reaction (PCR) and hybridization with an internal digoxigenin labeled probe. First, the specificity of our technique was tested on axenic or monoxenic cultures. Each of the 13 strains Naegleria fowleri of various origins was found PCR positive whereas 28 other strains originated from various organisms were not. The sensitivity with 40 amplification cycles was a single cell (cyst or trophozoite). Secondly, we have several protocols of cell recovery (i.e. sonication, freeze-thaw cycles, formamide, glass beads and chemical lysis) on artificially contaminated tap water samples. We showed that sonication, freeze-thaw cycles and formamide are better than chemical lysis and glass beads with proteinase K. Work is in progress to use this technique on environmental samples.