Optimization of a Loop-Mediated Isothermal Amplification Assay as a Point-of-Care Tool for the Detection of Wuchereria bancrofti in Human Blood in Tana River Delta, Kenya

Abstract

Introduction. Accurate detection of filarial parasites in humans and vectors is essential for the implementation and evaluation of Global and National Programs to eliminate lymphatic filariasis. Immunological methods to detect infection are available; however, cross-reactivity issues have been reported in most of them. Nucleic acid-based molecular assays offer high levels of specificity and sensitivity and can be used to detect the infections. Methods. In this study, we evaluated loop-mediated isothermal amplification (LAMP) tests to amplify Wuchereria bancrofti DNA in patients’ blood. The amplicons were tested by both pH-sensitive dyes for enhanced visual detection and agarose gel electrophoresis. A closed-tube LAMP assay was also evaluated. Cohen’s Kappa statistics was used for statistical analysis of the assays. 125 patients consented for blood sampling which were used for clinical analysis of LAMP assays with the PCR method used as the “gold standard.” Results. The sensitivity of the evaluated Wuchereria bancrofti LAMP was 92.3%, with a specificity of 97.3% and kappa statistics value of 0.84, which is in a strong agreement. Conclusion. In this study, LAMP assays coupled with fluorescence dye detection have been found to be suitable for diagnosis and monitoring of Wuchereria bancrofti infections in the Kenyan population.

LAMP assays for the simple and rapid detection of clinically important urinary pathogens including the detection of resistance to 3rd generation cephalosporins

“…These can be avoided by adding dimethylsulfoxide (DMSO) to the reaction. De-Guo Wang et al (2015) and Kinyatta Nanc et al (2021) demonstrated that adding 7.5% of DMSO in the reaction mix improved the LAMP specificity [ 34 , 35 ].The potential benefit of the LAMP assays developed in this study is their ability to detect not only bacteria responsible for UTI but also the most common resistance to C3G antibiotics, the CTX-M group 1. The early diagnosis of this gene could lead to improving the antibiotic therapy and antimicrobial stewardship (Additional file 1 , Additional file 2 , Additional file 3 , Additional file 4 ).…”

The Revolution of Lateral Flow Assay in the Field of AMR Detection

“…Indeed, the amplicons can be visualized by electrophoresis [ 137 ] and by monitoring the turbidity or the fluorescence. It is possible to look for color changes induced by chemical reaction [ 137 , 138 , 139 ]. The amplicons can be also detected using labelled primers with fluorescent dyes [ 140 ] or fluorescent intercalating dyes [ 137 ].…”

“… Detection methods for amplicons; ( A ) Gel electrophoresis [ 137 ]; ( B ) Chemical reaction, visual inspection with fluorescence [ 137 ]; ( C ) Chemical reaction, visual inspection with color change [ 139 ]; ( D ) Fluorescent primers, fluorescence monitoring [ 140 ]; ( E ) Chemical reaction, in real-time, turbidity monitoring [ 138 ]. …”