Construction of a DNA amplification assay for detection of Legionella species in clinical samples

A polymerase chain reaction (PCR) assay for the detection of Legionella spp. in clinical bronchial fluid samples was constructed. The assay could detect a 375 bp fragment of the 16S RNA gene, equivalent to 10 cfu in simulated clinical bronchial fluid and blood samples. Seven clinically relevant Legionella spp. involving 12 serogroups produced a positive signal, whereas three Legionella spp. and none of a panel of 26 bacterial strains produced a signal in the constructed assay. Investigation of bronchial fluid from 51 patients with clinically suspected Legionnaire's disease revealed the presence of Legionella DNA by PCR in seven patients. The results were verified by Southern blot analysis of the amplified DNA fragment. None of 37 control patients produced a positive signal. Only one of the seven bronchial fluid samples positive for Legionella by PCR was positive by conventional culture, thus indicating the increased sensitivity of PCR compared to culture.