Loop-Mediated Isothermal Amplification Assays for the Detection of Antimicrobial Resistance Elements inVibrio cholerae

Daniel Antonio Negrón, Shipra Trivedi, Nicholas Tolli, David Ashford, Gabrielle Melton, Stephanie Guertin, Katharine Jennings, Bryan D. Necciai, Shanmuga Sozhamannan, Bradley W. Abramson
2024 · doi:10.1101/2024.06.29.601342
preprint OA: closed CC-BY-4.0
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Abstract

The bacterium Vibrio cholerae causes diarrheal illness and can acquire genetic material leading to multiple drug resistance (MDR). Rapid detection of resistance-conferring mobile genetic elements helps avoid the prescription of ineffective antibiotics. Colorimetric loop-mediated isothermal amplification (LAMP) assays provide a rapid and cost-effective means for detection at point-of-care, but it can be difficult to design primer sets, determine target specificity, and interpret subjective color changes. We developed an algorithm for the in silico design and evaluation of LAMP assays within the open-source PCR Signature Erosion Tool (PSET) and a computer vision application for the quantitative analysis of colorimetric outputs. As an example, we generated new LAMP assays targeting drug resistance in V. cholerae and evaluated existing ones based on in silico target specificity and in vitro testing. Improvements in the design and testing of LAMP assays, with heightened target specificity and a simple analysis platform, increase utility for in-field applications.
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Abstract The bacterium Vibrio cholerae causes diarrheal illness and can acquire genetic material leading to multiple drug resistance (MDR). Rapid detection of resistance-conferring mobile genetic elements helps avoid the prescription of ineffective antibiotics. Colorimetric loop-mediated isothermal amplification (LAMP) assays provide a rapid and cost-effective means for detection at point-of-care, but it can be difficult to design primer sets, determine target specificity, and interpret subjective color changes. We developed an algorithm for the in silico design and evaluation of LAMP assays within the open-source PCR Signature Erosion Tool (PSET) and a computer vision application for the quantitative analysis of colorimetric outputs. As an example, we generated new LAMP assays targeting drug resistance in V. cholerae and evaluated existing ones based on in silico target specificity and in vitro testing. Improvements in the design and testing of LAMP assays, with heightened target specificity and a simple analysis platform, increase utility for in-field applications. Competing Interest Statement The authors have declared no competing interest.

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License: CC-BY-4.0