Development of an improved PCR-ICT hybrid assay for direct detection of Legionellae and Legionella pneumophila from cooling tower water specimens.
A novelly improved polymerase chian reaction and immunochromatography test (PCR-ICT) hybrid assay comprising traditional multiplex-nested PCR and ICT, (a lateral-flow device) was developed for direct detection of Legionella bacteria from environmental cooling tower samples. The partial 16S rDNA (specific for Legionella spp.) and dnaJ (specific for Legionella pneumophila) genes from Legionella chromosome were first specifically amplified by multiplex-nested PCR, respectively, followed by detection using ICT strip. Reading of results was based on presence or absence of the two test lines on the strips. Presence of test line 1 indicated existence of Legionella spp. specific 16S rDNA and identified Legionella spp. Presence of test line 2 further indicated existence of dnaJ and thus specifically identified L. pneumophila. In contrast, for non-Legionellae bacteria no test line formation was observed. Results of direct detection of Legionella bacteria and L. pneumophila from water tower specimens by this assay showed 100% sensitivity, and 96.6% and 100% specificity, respectively compared with traditional culture, biochemical and serological identification methods. The PCR-ICT hybrid assay does not require sophisticated equipment and was proved to be practically useful in rapid and direct Legionellae detection from environmental water samples.
Horng
Yu-Tze
YT
Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan University College of Medicine, No. 1. Chan-Dar Street 100, Taipei, Taiwan, ROC.
Soo
Po-Chi
PC
Shen
Bin-Jon
BJ
Hung
Yu-Li
YL
Lo
Kai-Yin
KY
Su
Hsun-Pi
HP
Wei
Jun-Rong
JR
Hsieh
Shang-Chen
SC
Hsueh
Po-Ren
PR
Lai
Hsin-Chih
HC
D001483:Base Sequence; D015169:Colony Count, Microbial; D007159:Immunologic Tests / Q000379:methods*; D007875:Legionella / Q000235:genetics / Q000302:isolation & purification*; D016952:Legionella pneumophila / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014871:Water Microbiology; D014881:Water Supply / Q000032:analysis*