Specific detection of Stachybotrys chartarum in pure culture using quantitative polymerase chain reaction

Research was conducted with laboratory cultures to establish a protocol for the rapid detection and quantitation of the toxigenic fungus Stachybotrys chartarum by means of polymerase chain reaction (PCR). Sequences for the 18 S rRNA gene of S. chartarum were obtained from GenBank and compared against all other available sequences on-line with the Basic Local Alignment Search Tool (BLAST). Two sets of TaqManTMprimers and one fluorescently labelled probe were designed and tested for selectivity, specificity and sensitivity of detection. A fluorogenic nuclease assay in conjunction with a sequence detector were used for the amplification and quantitation of S. chartarum. The primers designed amplified allS. chartarum isolates tested and did not amplify DNA extracted from other Stachybotrys species or 15 other fungal genera. The primer set selected had a sensitivity of