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Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)

Naegleria fowleri is the causative agent of the fatal disease primary amebic meningoencephalitis. Detection of N. fowleri using conventional culture and biochemical-based assays is time-consuming and laborious, while molecular techniques, such as PCR, require laboratory skills and expensive equipmen...

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Autores principales: Mahittikorn, Aongart, Mori, Hirotake, Popruk, Supaluk, Roobthaisong, Amonrattana, Sutthikornchai, Chantira, Koompapong, Khuanchai, Siri, Sukhontha, Sukthana, Yaowalark, Nacapunchai, Duangporn
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4379150/
https://www.ncbi.nlm.nih.gov/pubmed/25822175
http://dx.doi.org/10.1371/journal.pone.0120997
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author Mahittikorn, Aongart
Mori, Hirotake
Popruk, Supaluk
Roobthaisong, Amonrattana
Sutthikornchai, Chantira
Koompapong, Khuanchai
Siri, Sukhontha
Sukthana, Yaowalark
Nacapunchai, Duangporn
author_facet Mahittikorn, Aongart
Mori, Hirotake
Popruk, Supaluk
Roobthaisong, Amonrattana
Sutthikornchai, Chantira
Koompapong, Khuanchai
Siri, Sukhontha
Sukthana, Yaowalark
Nacapunchai, Duangporn
author_sort Mahittikorn, Aongart
collection PubMed
description Naegleria fowleri is the causative agent of the fatal disease primary amebic meningoencephalitis. Detection of N. fowleri using conventional culture and biochemical-based assays is time-consuming and laborious, while molecular techniques, such as PCR, require laboratory skills and expensive equipment. We developed and evaluated a novel loop-mediated isothermal amplification (LAMP) assay targeting the virulence-related gene for N. fowleri. Time to results is about 90 min and amplification products were easily detected visually using hydroxy naphthol blue. The LAMP was highly specific after testing against related microorganisms and able to detect one trophozoite, as determined with spiked water and cerebrospinal fluid samples. The assay was then evaluated with a set of 80 water samples collected during the flooding crisis in Thailand in 2011, and 30 natural water samples from border areas of northern, eastern, western, and southern Thailand. N. fowleri was detected in 13 and 10 samples using LAMP and PCR, respectively, with a Kappa coefficient of 0.855. To the best of our knowledge, this is the first report of a LAMP assay for N. fowleri. Due to its simplicity, speed, and high sensitivity, the LAMP method described here might be useful for quickly detecting and diagnosing N. fowleri in water and clinical samples, particularly in resource-poor settings.
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spelling pubmed-43791502015-04-09 Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP) Mahittikorn, Aongart Mori, Hirotake Popruk, Supaluk Roobthaisong, Amonrattana Sutthikornchai, Chantira Koompapong, Khuanchai Siri, Sukhontha Sukthana, Yaowalark Nacapunchai, Duangporn PLoS One Research Article Naegleria fowleri is the causative agent of the fatal disease primary amebic meningoencephalitis. Detection of N. fowleri using conventional culture and biochemical-based assays is time-consuming and laborious, while molecular techniques, such as PCR, require laboratory skills and expensive equipment. We developed and evaluated a novel loop-mediated isothermal amplification (LAMP) assay targeting the virulence-related gene for N. fowleri. Time to results is about 90 min and amplification products were easily detected visually using hydroxy naphthol blue. The LAMP was highly specific after testing against related microorganisms and able to detect one trophozoite, as determined with spiked water and cerebrospinal fluid samples. The assay was then evaluated with a set of 80 water samples collected during the flooding crisis in Thailand in 2011, and 30 natural water samples from border areas of northern, eastern, western, and southern Thailand. N. fowleri was detected in 13 and 10 samples using LAMP and PCR, respectively, with a Kappa coefficient of 0.855. To the best of our knowledge, this is the first report of a LAMP assay for N. fowleri. Due to its simplicity, speed, and high sensitivity, the LAMP method described here might be useful for quickly detecting and diagnosing N. fowleri in water and clinical samples, particularly in resource-poor settings. Public Library of Science 2015-03-30 /pmc/articles/PMC4379150/ /pubmed/25822175 http://dx.doi.org/10.1371/journal.pone.0120997 Text en © 2015 Mahittikorn et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Mahittikorn, Aongart
Mori, Hirotake
Popruk, Supaluk
Roobthaisong, Amonrattana
Sutthikornchai, Chantira
Koompapong, Khuanchai
Siri, Sukhontha
Sukthana, Yaowalark
Nacapunchai, Duangporn
Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)
title Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)
title_full Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)
title_fullStr Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)
title_full_unstemmed Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)
title_short Development of a Rapid, Simple Method for Detecting Naegleria fowleri Visually in Water Samples by Loop-Mediated Isothermal Amplification (LAMP)
title_sort development of a rapid, simple method for detecting naegleria fowleri visually in water samples by loop-mediated isothermal amplification (lamp)
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4379150/
https://www.ncbi.nlm.nih.gov/pubmed/25822175
http://dx.doi.org/10.1371/journal.pone.0120997
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