Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of Mycoplasma pneumoniae
1 Department of Infection Control and Prevention, University of Tokyo Hospital, Bunkyo-ku, Tokyo 113-8655, Japan 2 Department of Microbiology and Immunology, Graduate School of Allied Health Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan 3 Kitasato Institute for Life...
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| Veröffentlicht in: | Journal of medical microbiology 2005-11, Vol.54 (11), p.1037-1041 |
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| container_title | Journal of medical microbiology |
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| creator | Saito, Ryoichi Misawa, Yoshiki Moriya, Kyoji Koike, Kazuhiko Ubukata, Kimiko Okamura, Noboru |
| description | 1 Department of Infection Control and Prevention, University of Tokyo Hospital, Bunkyo-ku, Tokyo 113-8655, Japan 2 Department of Microbiology and Immunology, Graduate School of Allied Health Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan 3 Kitasato Institute for Life Sciences and Graduate School of Infection Control Sciences, Kitasato University, Minato-ku, Tokyo 108-8641, Japan
Correspondence Ryoichi Saito saito-lab{at}h.u-tokyo.ac.jp
Received March 3, 2005
Accepted July 29, 2005
A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Mycoplasma pneumoniae was developed and evaluated. The assay specifically amplified only M. pneumoniae sequences, and no cross-reactivity was observed for other Mycoplasma species or respiratory bacterial species. The detection limit for this assay was found to be 2 x 10 2 copies, corresponding to 220 colour changing units of M. pneumoniae in 1 h, as observed in a real-time turbidimeter and electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease analysis as well as direct sequencing of the amplified product. The assay was applied to 95 nasopharyngeal swab samples collected from patients or from healthy individuals, and compared to a real-time PCR assay in-house. A concordance of 100 % was observed between the two assays. The LAMP assay is easy to perform, shows a rapid reaction and is inexpensive. It may therefore be applied in the routine diagnosis of M. pneumoniae infection in the clinical laboratory.
Abbreviation: LAMP, loop-mediated isothermal amplification. |
| doi_str_mv | 10.1099/jmm.0.46071-0 |
| format | Article |
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Correspondence Ryoichi Saito saito-lab{at}h.u-tokyo.ac.jp
Received March 3, 2005
Accepted July 29, 2005
A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Mycoplasma pneumoniae was developed and evaluated. The assay specifically amplified only M. pneumoniae sequences, and no cross-reactivity was observed for other Mycoplasma species or respiratory bacterial species. The detection limit for this assay was found to be 2 x 10 2 copies, corresponding to 220 colour changing units of M. pneumoniae in 1 h, as observed in a real-time turbidimeter and electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease analysis as well as direct sequencing of the amplified product. The assay was applied to 95 nasopharyngeal swab samples collected from patients or from healthy individuals, and compared to a real-time PCR assay in-house. A concordance of 100 % was observed between the two assays. The LAMP assay is easy to perform, shows a rapid reaction and is inexpensive. It may therefore be applied in the routine diagnosis of M. pneumoniae infection in the clinical laboratory.
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Correspondence Ryoichi Saito saito-lab{at}h.u-tokyo.ac.jp
Received March 3, 2005
Accepted July 29, 2005
A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Mycoplasma pneumoniae was developed and evaluated. The assay specifically amplified only M. pneumoniae sequences, and no cross-reactivity was observed for other Mycoplasma species or respiratory bacterial species. The detection limit for this assay was found to be 2 x 10 2 copies, corresponding to 220 colour changing units of M. pneumoniae in 1 h, as observed in a real-time turbidimeter and electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease analysis as well as direct sequencing of the amplified product. The assay was applied to 95 nasopharyngeal swab samples collected from patients or from healthy individuals, and compared to a real-time PCR assay in-house. A concordance of 100 % was observed between the two assays. The LAMP assay is easy to perform, shows a rapid reaction and is inexpensive. It may therefore be applied in the routine diagnosis of M. pneumoniae infection in the clinical laboratory.
Abbreviation: LAMP, loop-mediated isothermal amplification.</description><subject>Adolescent</subject><subject>Adult</subject><subject>Aged</subject><subject>Bacteriology</subject><subject>Biological and medical sciences</subject><subject>Child</subject><subject>Child, Preschool</subject><subject>DNA Restriction Enzymes - metabolism</subject><subject>DNA, Bacterial - analysis</subject><subject>DNA, Bacterial - chemistry</subject><subject>Electrophoresis, Agar Gel</subject><subject>Female</subject><subject>Fundamental and applied biological sciences. 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Psychology</topic><topic>Humans</topic><topic>Infant</topic><topic>Infectious diseases</topic><topic>Male</topic><topic>Medical sciences</topic><topic>Microbiology</topic><topic>Middle Aged</topic><topic>Miscellaneous</topic><topic>Molecular Diagnostic Techniques</topic><topic>Mycoplasma pneumoniae - genetics</topic><topic>Mycoplasma pneumoniae - isolation & purification</topic><topic>Nasopharynx - microbiology</topic><topic>Nephelometry and Turbidimetry</topic><topic>Nucleic Acid Amplification Techniques</topic><topic>Pneumonia, Mycoplasma - diagnosis</topic><topic>Polymerase Chain Reaction</topic><topic>Sensitivity and Specificity</topic><topic>Sequence Analysis, DNA</topic><topic>Temperature</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Saito, Ryoichi</creatorcontrib><creatorcontrib>Misawa, Yoshiki</creatorcontrib><creatorcontrib>Moriya, Kyoji</creatorcontrib><creatorcontrib>Koike, Kazuhiko</creatorcontrib><creatorcontrib>Ubukata, Kimiko</creatorcontrib><creatorcontrib>Okamura, Noboru</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of medical microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Saito, Ryoichi</au><au>Misawa, Yoshiki</au><au>Moriya, Kyoji</au><au>Koike, Kazuhiko</au><au>Ubukata, Kimiko</au><au>Okamura, Noboru</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of Mycoplasma pneumoniae</atitle><jtitle>Journal of medical microbiology</jtitle><addtitle>J Med Microbiol</addtitle><date>2005-11-01</date><risdate>2005</risdate><volume>54</volume><issue>11</issue><spage>1037</spage><epage>1041</epage><pages>1037-1041</pages><issn>0022-2615</issn><eissn>1473-5644</eissn><coden>JMMIAV</coden><abstract>1 Department of Infection Control and Prevention, University of Tokyo Hospital, Bunkyo-ku, Tokyo 113-8655, Japan 2 Department of Microbiology and Immunology, Graduate School of Allied Health Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan 3 Kitasato Institute for Life Sciences and Graduate School of Infection Control Sciences, Kitasato University, Minato-ku, Tokyo 108-8641, Japan
Correspondence Ryoichi Saito saito-lab{at}h.u-tokyo.ac.jp
Received March 3, 2005
Accepted July 29, 2005
A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Mycoplasma pneumoniae was developed and evaluated. The assay specifically amplified only M. pneumoniae sequences, and no cross-reactivity was observed for other Mycoplasma species or respiratory bacterial species. The detection limit for this assay was found to be 2 x 10 2 copies, corresponding to 220 colour changing units of M. pneumoniae in 1 h, as observed in a real-time turbidimeter and electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease analysis as well as direct sequencing of the amplified product. The assay was applied to 95 nasopharyngeal swab samples collected from patients or from healthy individuals, and compared to a real-time PCR assay in-house. A concordance of 100 % was observed between the two assays. The LAMP assay is easy to perform, shows a rapid reaction and is inexpensive. It may therefore be applied in the routine diagnosis of M. pneumoniae infection in the clinical laboratory.
Abbreviation: LAMP, loop-mediated isothermal amplification.</abstract><cop>Reading</cop><pub>Soc General Microbiol</pub><pmid>16192434</pmid><doi>10.1099/jmm.0.46071-0</doi><tpages>5</tpages><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; Microbiology Society; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Alma/SFX Local Collection |
| subjects | Adolescent Adult Aged Bacteriology Biological and medical sciences Child Child, Preschool DNA Restriction Enzymes - metabolism DNA, Bacterial - analysis DNA, Bacterial - chemistry Electrophoresis, Agar Gel Female Fundamental and applied biological sciences. Psychology Humans Infant Infectious diseases Male Medical sciences Microbiology Middle Aged Miscellaneous Molecular Diagnostic Techniques Mycoplasma pneumoniae - genetics Mycoplasma pneumoniae - isolation & purification Nasopharynx - microbiology Nephelometry and Turbidimetry Nucleic Acid Amplification Techniques Pneumonia, Mycoplasma - diagnosis Polymerase Chain Reaction Sensitivity and Specificity Sequence Analysis, DNA Temperature |
| title | Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of Mycoplasma pneumoniae |
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