Rapid and accurate detection of Pseudomonas aeruginosa by real-time polymerase chain reaction with melting curve analysis targeting gyrB gene.
Laboratory detection of Pseudomonas spp., particularly Pseudomonas aeruginosa, is an important assay in the nosocomial control. The study was designed firstly to establish a new assay-applied LightCycler polymerase chain reaction (PCR) technology with melting curve analysis (MCA). A total of 224 Gram-negative isolates were used to verify the assay system. The PCR with MCA method using the P. aeruginosa-specific gyrase B gene primers was rapid and accurate; the total run is approximately 3 h, and the sensitivity and specificity relative to the Vitek (bioMerieux, Hazelwood, MO) results were 98.1% and 100%, respectively. Vitek identification system was not able to identify the isolates from the new Pseudomonas otitidis spp. opposite to the real-time PCR. This assay was validated to be accurate with an overall sensitivity and specificity of 98.7% and 98.9%, respectively. Conclusively, this rapid and accurate PCR assay with MCA will help to manage and control infections with P. aeruginosa.
Motoshima
Maiko
M
Department of Laboratory Medicine, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki City, Nagasaki 852-8501, Japan.
Yanagihara
Katsunori
K
Fukushima
Kazuko
K
Matsuda
Junichi
J
Sugahara
Kazuyuki
K
Hirakata
Yochi
Y
Yamada
Yasuaki
Y
Kohno
Shigeru
S
Kamihira
Shimeru
S
D015373:Bacterial Typing Techniques; D001483:Base Sequence; D027081:DNA Gyrase / Q000235:genetics*; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D011552:Pseudomonas Infections / Q000175:diagnosis* / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D013997:Time Factors; D044366:Transition Temperature
D017931:DNA Primers; D004269:DNA, Bacterial; D027081:DNA Gyrase