title: PCR methods for the rapid detection and identification of four pathogenic Legionella spp. and two Legionella pneumophila subspecies based on the gene amplification of gyrB.
abstract: A total of 25 gyrB gene sequences from 20 Legionella pneumophila subsp. pneumophila strains and five L. pneumophila subsp. fraseri strains were obtained and analyzed, and a multiplex PCR for the simultaneous detection of Legionella bozemanae, Legionella longbeachae, Legionella micdadei and Legioenella pneumophila, and two single PCRs for the differentiation of L. pneumophila subsp. pneumophila and L. pneumophila subsp. fraseri were established. The multiplex PCR method was shown to be highly specific and reproducible when tested against 41 target strains and 17 strains of other bacteria species. The sensitivity of the multiplex PCR was also analyzed and was shown to detect levels as low as 1 ng of genomic DNA or 10 colony-forming units (CFUs) per milliliter in mock water samples. Sixty-three air conditioner condensed water samples from Shanghai City were examined, and the result was validated using 16S rRNA sequencing. The data reported here demonstrate that the multiplex PCR method described is efficient and convenient for the detection of Legionella species in water samples. Twenty L. pneumophila subsp. pneumophila strains and five L. pneumophila subsp. fraseri strains were used for the validation of the two L. pneumophila subspecies-specific PCR methods, and the results indicated that the two PCR methods were both highly specific and convenient for the identification of L. pneumophila at the subspecies level.
authors.affiliation: Key Laboratory of Molecular Microbiology and Technology of the Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China.
authors.forename: Guangpeng
authors.initials: G
authors.lastname: Zhou
authors.forename: Boyang
authors.initials: B
authors.lastname: Cao
authors.forename: Yan
authors.initials: Y
authors.lastname: Dou
authors.forename: Yanwei
authors.initials: Y
authors.lastname: Liu
authors.forename: Lu
authors.initials: L
authors.lastname: Feng
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
mesh_terms: D015373:Bacterial Typing Techniques / Q000379:methods; D001483:Base Sequence; D027081:DNA Gyrase / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D005784:Gene Amplification; D007875:Legionella / Q000235:genetics* / Q000302:isolation & purification*; D007876:Legionellosis / Q000175:diagnosis* / Q000235:genetics / Q000382:microbiology; D007877:Legionnaires' Disease / Q000175:diagnosis / Q000235:genetics / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013997:Time Factors; D014871:Water Microbiology
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D027081:DNA Gyrase