OBJECTIVE: To develop and validate a novel loop-mediated amplification (LAMP) assay for rapid diagnosis (<1 hour) of whooping cough in nasopharyngeal samples versus the gold standard: real-time PCR. METHODS: The study included all nasopharyngeal samples (n = 213) collected from children with clinical suspicion of pertussis admitted to Children's University Hospital Sant Joan de Déu (Barcelona, Spain) during July-December 2014. Fresh samples were routinely analyzed by real-time PCR and stored for retrospective LAMP analysis, following an easy 30 minute DNA extraction step by Chelex-100. RESULTS: Performance results of the LAMP assay were: linearity, 10(5)-10(1) CFU/ml; Limit of Detection, 2 CFU/ml; precision (mean CV), 7.38%; diagnostic sensitivity, 96.55%; diagnostic specificity, 99.46%; time to detection, 12-30 minutes. CONCLUSION: The new test was shown to be 2.5-fold faster than real-time PCR while maintaining similar levels of analytical and clinical performance. Therefore it could become a useful diagnostic tool for molecular point-of-care testing.
OBJECTIVE: To develop and validate a novel loop-mediated amplification (LAMP) assay for rapid diagnosis (<1 hour) of whooping cough in nasopharyngeal samples versus the gold standard: real-time PCR. METHODS: The study included all nasopharyngeal samples (n = 213) collected from children with clinical suspicion of pertussis admitted to Children's University Hospital Sant Joan de Déu (Barcelona, Spain) during July-December 2014. Fresh samples were routinely analyzed by real-time PCR and stored for retrospective LAMP analysis, following an easy 30 minute DNA extraction step by Chelex-100. RESULTS: Performance results of the LAMP assay were: linearity, 10(5)-10(1) CFU/ml; Limit of Detection, 2 CFU/ml; precision (mean CV), 7.38%; diagnostic sensitivity, 96.55%; diagnostic specificity, 99.46%; time to detection, 12-30 minutes. CONCLUSION: The new test was shown to be 2.5-fold faster than real-time PCR while maintaining similar levels of analytical and clinical performance. Therefore it could become a useful diagnostic tool for molecular point-of-care testing.
Entities:
Keywords:
Bordetella pertussis; children; loop-mediated isothermal amplification; point of care; rapid diagnosis; validation
Authors: Maggie R Williams; Robert D Stedtfeld; Tiffany M Stedtfeld; James M Tiedje; Syed A Hashsham Journal: Biomed Microdevices Date: 2017-09 Impact factor: 2.838
Authors: Héctor David de Paz; Pedro Brotons; Cristina Esteva; Carmen Muñoz-Almagro Journal: Front Cell Infect Microbiol Date: 2020-03-24 Impact factor: 5.293