title: Loop-mediated isothermal amplification with the Procedure for Ultra Rapid Extraction kit for the diagnosis of pneumocystis pneumonia.
abstract: Loop-mediated isothermal amplification (LAMP) is an innovative molecular technique requiring only a heating device and isothermal conditions to amplify a specific target gene. The results of current microscopic diagnostic tools for pneumocystis pneumonia are not sufficiently consistent for detecting infection with a low-density of Pneumocystis jirovecii. Although polymerase chain reaction (PCR) is highly sensitive, it is not suitable for resource-limited facilities. LAMP is a potential diagnostic replacement for PCR in such settings but a critical disadvantage of DNA extraction was still remained. Therefore, we employed the Procedure for Ultra Rapid Extraction (PURE) kit, which uses a porous material, to isolate the DNA from clinical samples in a simple way in combination with previously reported LAMP procedure for diagnosing PCP. The detection limit of the PURE-LAMP method applied to artificial bronchoalveolar lavage fluid samples was 100 copies/tube, even with the use of massive blood-contaminated solutions. In addition, we concluded the diagnostic procedure within 1 h without the need for additional equipment. PURE-LAMP coupled with suitable primers for specific pathogens has good potential for diagnosing various infectious diseases.
authors.affiliation: Division of Infectious Diseases and Pulmonary Medicine, Department of Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Shuichi
authors.initials: S
authors.lastname: Kawano
authors.affiliation: Division of Infectious Diseases and Pulmonary Medicine, Department of Internal Medicine, National Defense Medical College, Saitama, Japan. Electronic address: tmaeda@ndmc.ac.jp.
authors.forename: Takuya
authors.initials: T
authors.lastname: Maeda
authors.affiliation: Department of Pharmacy, National Defense Medical College Hospital, Saitama, Japan.
authors.forename: Takefumi
authors.initials: T
authors.lastname: Suzuki
authors.affiliation: Division of Infectious Diseases and Pulmonary Medicine, Department of Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Tatsuhiro
authors.initials: T
authors.lastname: Abe
authors.affiliation: Division of Infectious Diseases and Pulmonary Medicine, Department of Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Kei
authors.initials: K
authors.lastname: Mikita
authors.affiliation: Division of Infectious Diseases and Pulmonary Medicine, Department of Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Hamakawa
authors.affiliation: Department of Global Infectious Disease and Tropical Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Ono
authors.affiliation: Department of Pharmacy, National Defense Medical College Hospital, Saitama, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Sonehara
authors.affiliation: Department of Global Infectious Disease and Tropical Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Miyahira
authors.affiliation: Division of Infectious Diseases and Pulmonary Medicine, Department of Internal Medicine, National Defense Medical College, Saitama, Japan.
authors.forename: Akihiko
authors.initials: A
authors.lastname: Kawana
mesh_terms: D001992:Bronchoalveolar Lavage Fluid / Q000382:microbiology; D004269:DNA, Bacterial / Q000235:genetics; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D045363:Pneumocystis carinii / Q000235:genetics* / Q000302:isolation & purification; D011020:Pneumonia, Pneumocystis / Q000175:diagnosis* / Q000382:microbiology; D016133:Polymerase Chain Reaction
chemical_list: D004269:DNA, Bacterial
keywords: Loop-mediated isothermal amplification; Pneumocystis jirovecii; Pneumocystis pneumonia; Polymerase chain reaction; Procedure for Ultra Rapid Extraction