Loop-mediated isothermal amplification (LAMP) assay for rapid detection of viable but non-culturable <i>Vibrio cholerae</i> O1

Chamanrokh, Parastoo; Colwell, Rita R.; Huq, Anwar

doi:10.1139/cjm-2021-0142

Cited by 4 publications

(1 citation statement)

References 41 publications

(51 reference statements)

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“…Several LAMP-based assays were developed previously in the laboratories for the diagnosis of cholera [9,[33][34][35][36][37][38][39][40][41][42][43]. The stool is a complex sample to extract DNA and amplify because of the presence of inhibitors.…”

Section: Discussionmentioning

confidence: 99%

Development of a simple, rapid, and sensitive molecular diagnostic assay for cholera

2023

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Cholera continues to inflict high rates of morbidity and mortality. Prompt identification of cholera cases facilitates rapid outbreak responses in the short term while providing reliable surveillance data to guide long-term policies and interventions. Microbiological stool culture, the current recognized gold standard for diagnosing cholera, has significant limitations. Rapid diagnostic tests (RDTs) represent promising alternatives for diagnosing cholera in areas with limited laboratory infrastructure. However, studies conducted with the current cholera RDTs demonstrated wide variations in sensitivity and specificity. To address this gap in the diagnosis of cholera, we developed a simple, rapid, and sensitive diagnostic assay, "Rapid LAMP based Diagnostic Test (RLDT)." With a novel, simple sample preparation method directly from the fecal samples along with lyophilized reaction strips and using established Loop-mediated Isothermal Amplification (LAMP) platform, cholera toxin gene (ctxA) and O1 (O1rfb) gene could be detected in less than an hour. Cholera RLDT assay is cold chain and electricity-free. To avoid any end-user bias, a battery-operated, handheld reader was used to read the RLDT results. The performance specifications of the cholera RLDT assay, including analytical sensitivity and specificity, were evaluated using direct fecal samples, dried fecal samples on filter paper, and environmental water samples spiked with cholera strain. The limit of detection (LOD) was ~104 CFU/gm of stool for both ctxA and O1 genes, corresponding to about 1 CFU of Vibrio cholerae per reaction within 40 minutes. The LOD was 10 bacteria per ml of environmental water when tested with RLDT directly, without enrichment. Being simple, RLDT has the potential to be applied in resource-poor endemic settings for rapid, sensitive, and reliable diagnosis of cholera.

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Section: Discussionmentioning

confidence: 99%

Development of a simple, rapid, and sensitive molecular diagnostic assay for cholera

2023

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Diagnostic techniques for rapid detection of Vibrio species

et al. 2022

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Molecular Diagnostic Tools Applied for Assessing Microbial Water Quality

Paruch

2022

IJERPH

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Microbial water quality is of vital importance for human, animal, and environmental health. Notably, pathogenically contaminated water can result in serious health problems, such as waterborne outbreaks, which have caused huge economic and social losses. In this context, the prompt detection of microbial contamination becomes essential to enable early warning and timely reaction with proper interventions. Recently, molecular diagnostics have been increasingly employed for the rapid and robust assessment of microbial water quality implicated by various microbial pollutants, e.g., waterborne pathogens and antibiotic-resistance genes (ARGs), imposing the most critical health threats to humans and the environment. Continuous technological advances have led to constant improvements and expansions of molecular methods, such as conventional end-point PCR, DNA microarray, real-time quantitative PCR (qPCR), multiplex qPCR (mqPCR), loop-mediated isothermal amplification (LAMP), digital droplet PCR (ddPCR), and high-throughput next-generation DNA sequencing (HT-NGS). These state-of-the-art molecular approaches largely facilitate the surveillance of microbial water quality in diverse aquatic systems and wastewater. This review provides an up-to-date overview of the advancement of the key molecular tools frequently employed for microbial water quality assessment, with future perspectives on their applications.

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Loop-mediated isothermal amplification (LAMP) assay for rapid detection of viable but non-culturable Vibrio cholerae O1

Cited by 4 publications

References 41 publications

Development of a simple, rapid, and sensitive molecular diagnostic assay for cholera

Development of a simple, rapid, and sensitive molecular diagnostic assay for cholera

Diagnostic techniques for rapid detection of Vibrio species

Molecular Diagnostic Tools Applied for Assessing Microbial Water Quality

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