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Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test

OBJECTIVES: Rickettsia rickettsii is the causative agent of Rocky Mountain spotted fever, which is the most severe spotted fever group (SFG) rickettsiosis. Developing a simple and reliable detection method is required. METHODS: A detection method for R. rickettsii was established based on a recombin...

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Autores principales: Qi, Yong, Shao, Yinxiu, Rao, Jixian, Shen, Wanpeng, Yin, Qiong, Li, Xiaoling, Chen, Hongxia, Li, Jiameng, Zeng, Wenwen, Zheng, Shulong, Liu, Suyun, Li, Yuexi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6257923/
https://www.ncbi.nlm.nih.gov/pubmed/30475889
http://dx.doi.org/10.1371/journal.pone.0207811
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author Qi, Yong
Shao, Yinxiu
Rao, Jixian
Shen, Wanpeng
Yin, Qiong
Li, Xiaoling
Chen, Hongxia
Li, Jiameng
Zeng, Wenwen
Zheng, Shulong
Liu, Suyun
Li, Yuexi
author_facet Qi, Yong
Shao, Yinxiu
Rao, Jixian
Shen, Wanpeng
Yin, Qiong
Li, Xiaoling
Chen, Hongxia
Li, Jiameng
Zeng, Wenwen
Zheng, Shulong
Liu, Suyun
Li, Yuexi
author_sort Qi, Yong
collection PubMed
description OBJECTIVES: Rickettsia rickettsii is the causative agent of Rocky Mountain spotted fever, which is the most severe spotted fever group (SFG) rickettsiosis. Developing a simple and reliable detection method is required. METHODS: A detection method for R. rickettsii was established based on a recombinase polymerase amplification (RPA) assay and the lateral flow (LF) test. A specific target sequence was screened, and corresponding primers and probes were designed, synthesized, and screened for establishing an RPA assay with high amplification efficiency. Reagent concentrations, amplification time, and loading volume for strip development were optimized. The detection limit, analytic sensitivity and specificity were evaluated. RESULTS: A rapid, visual, sensitive and specific method for the detection of R. rickettsii based on RPA and the LF test was successfully established. The novel method had a limit of detection of 10 to 50 copies/reaction without recognizing other organisms. Analytical sensitivity and specificity were ≥90% and 100%, respectively, as evaluated by animal and simulative human samples. CONCLUSIONS: Using the established method, detection could be completed in 30 min with visually detectable results by the naked eye, without requirement of any instrument except a constant temperature equipment. The technique shows superior detection performance and is promising for wide use in the field as well as resource-limited areas for R. rickettsii detection.
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spelling pubmed-62579232018-12-06 Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test Qi, Yong Shao, Yinxiu Rao, Jixian Shen, Wanpeng Yin, Qiong Li, Xiaoling Chen, Hongxia Li, Jiameng Zeng, Wenwen Zheng, Shulong Liu, Suyun Li, Yuexi PLoS One Research Article OBJECTIVES: Rickettsia rickettsii is the causative agent of Rocky Mountain spotted fever, which is the most severe spotted fever group (SFG) rickettsiosis. Developing a simple and reliable detection method is required. METHODS: A detection method for R. rickettsii was established based on a recombinase polymerase amplification (RPA) assay and the lateral flow (LF) test. A specific target sequence was screened, and corresponding primers and probes were designed, synthesized, and screened for establishing an RPA assay with high amplification efficiency. Reagent concentrations, amplification time, and loading volume for strip development were optimized. The detection limit, analytic sensitivity and specificity were evaluated. RESULTS: A rapid, visual, sensitive and specific method for the detection of R. rickettsii based on RPA and the LF test was successfully established. The novel method had a limit of detection of 10 to 50 copies/reaction without recognizing other organisms. Analytical sensitivity and specificity were ≥90% and 100%, respectively, as evaluated by animal and simulative human samples. CONCLUSIONS: Using the established method, detection could be completed in 30 min with visually detectable results by the naked eye, without requirement of any instrument except a constant temperature equipment. The technique shows superior detection performance and is promising for wide use in the field as well as resource-limited areas for R. rickettsii detection. Public Library of Science 2018-11-26 /pmc/articles/PMC6257923/ /pubmed/30475889 http://dx.doi.org/10.1371/journal.pone.0207811 Text en © 2018 Qi 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 (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Qi, Yong
Shao, Yinxiu
Rao, Jixian
Shen, Wanpeng
Yin, Qiong
Li, Xiaoling
Chen, Hongxia
Li, Jiameng
Zeng, Wenwen
Zheng, Shulong
Liu, Suyun
Li, Yuexi
Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
title Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
title_full Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
title_fullStr Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
title_full_unstemmed Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
title_short Development of a rapid and visual detection method for Rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
title_sort development of a rapid and visual detection method for rickettsia rickettsii combining recombinase polymerase assay with lateral flow test
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6257923/
https://www.ncbi.nlm.nih.gov/pubmed/30475889
http://dx.doi.org/10.1371/journal.pone.0207811
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