Open AccessDevelopment and Clinical Evaluation of an Internally Controlled, Single-Tube Multiplex Real-Time PCR Assay for Detection of Legionella pneumophila and Other Legionella Species
Author(s) -
Kate Templeton,
Sitha A. Scheltinga,
Peter Sillekens,
Jantine W. Crielaard,
Alje P. van Dam,
Herman Goossens,
Eric C. J. Claas
Publication year - 2003
Publication title -
journal of clinical microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.349
H-Index - 255
eISSN - 1070-633X
pISSN - 0095-1137
DOI - 10.1128/jcm.41.9.4016-4021.2003
Subject(s) - legionella pneumophila , legionella , multiplex , microbiology and biotechnology , legionnaires' disease , biology , false positive paradox , multiplex polymerase chain reaction , real time polymerase chain reaction , virology , bacteria , polymerase chain reaction , gene , genetics , machine learning , computer science
A multiplex real-time PCR assay for detection of Legionella pneumophila and Legionella spp. and including an internal control was designed. Legionella species, L. pneumophila, and the internal control were detected simultaneously by probes labeled with 6-carboxy-fluorescein, hexachlorofluorescein, and indodicarbocyanine, respectively. Therefore, no postamplification analysis was required in order to distinguish the targets. The sensitivity of both assays was 2.5 CFU/ml, and from analysis of 10 culture-positive and 74 culture-negative samples from patients investigated for legionellosis, 100% agreement was observed by both assays in comparison to culture. Four additional positives were found by the multiplex real-time PCR assay in the Legionella culture-negative samples.