Evaluation of the colorimetric malachite green loop-mediated isothermal amplification (MG-LAMP) assay for the detection of malaria species at two different health facilities in a malaria endemic area of western Kenya

Abstract

Background Prompt diagnosis and effective malaria treatment is a key strategy in malaria control. However, the recommended diagnostic methods, microscopy and rapid diagnostic tests (RDTs), are not supported by robust quality assurance systems in endemic areas. This study compared the performance of routine RDTs and smear microscopy with a simple molecular-based colorimetric loop-mediated isothermal amplification (LAMP) at two different levels of the health care system in a malaria-endemic area of western Kenya. Methods Patients presenting with clinical symptoms of malaria at Rota Dispensary (level 2) and Siaya County Referral Hospital (level 4) were enrolled into the study after obtaining written informed consent. Capillary blood was collected to test for malaria by RDT and microscopy at the dispensary and county hospital, and for preparation of blood smears and dried blood spots (DBS) for expert microscopy and real-time polymerase chain reaction (RT-PCR). Results of the routine diagnostic tests were compared with those of malachite green loop-mediated isothermal amplification (MG-LAMP) performed at the two facilities. Results A total of 264 participants were enrolled into the study. At the dispensary level, the positivity rate by RDT, expert microscopy, MG-LAMP and RT-PCR was 37%, 30%, 44% and 42%, respectively, and 42%, 43%, 57% and 43% at the county hospital. Using RT-PCR as the reference test, the sensitivity of RDT and MG-LAMP was 78.1% (CI 67.5–86.4) and 82.9% (CI 73.0–90.3) at Rota dispensary. At Siaya hospital the sensitivity of routine microscopy and MG-LAMP was 83.3% (CI 65.3–94.4) and 93.3% (CI 77.9–99.2), respectively. Compared to MG-LAMP, there were 14 false positives and 29 false negatives by RDT at Rota dispensary and 3 false positives and 13 false negatives by routine microscopy at Siaya Hospital. Conclusion MG-LAMP is more sensitive than RDTs and microscopy in the detection of malaria parasites at public health facilities and might be a useful quality control tool in resource-limited settings.

Naked‐eye detection strategies coupled with isothermal nucleic acid amplification techniques for the detection of human pathogens

“…Focusing in MaG, its low cost, color stability, simplicity of use, and storage are among its advantages over other dyes (Scott et al, 2020). The typical concentration ranges between 0.004% and 0.008% (Nzelu et al, 2014), and this dye seems to have found its niche in LAMP assays targeting different types of parasites (Barazorda et al, 2020;Chahar et al, 2018;Gachugia et al, 2020;Kamber et al, 2020;Lucchi et al, 2016;Nzelu et al, 2016Nzelu et al, , 2019Takagi et al, 2016), even though other pathogens have also been detected, such as bacteria (Cheng et al, 2020;Jothikumar et al, 2014;Li et al, 2019;Li, Liu, et al, 2020). Moving to MeG, it was reported to allow for clear color differentiation when added at a final concentration of 0.004%, and attending to a study from Thapa et al, dealing with the detection of M. tuberculosis, MeG developed the color change in the same time frame as of MaG but with a superior color intensity (Thapa et al, 2019).…”

Impact of Malaria Diagnostic Technologies on the Disease Burden in the Sub-Saharan Africa

“…Despite breakthroughs in malaria control, the illness has long been a major public health burden globally [ 1 ]. It, along with hepatitis, TB, and HIV/AIDS, has claimed many lives globally and is calamitous [ 2 ].…”

Assessment of Commercial Real-Time PCR Assays for Detection of Malaria Infection in a Non-Endemic Setting

“…During amplification reaction, high amounts of DNA and pyrophosphate ion by-product are produced, 24 allowing different ways of detection of amplified products, such as turbidity, fluorescence, or colorimetric dyes. 25,26 This flexibility in the way of detection besides the isothermal nature of the process, which does not require a thermocycler or highly specific devices, have supported LAMP method such as an useful tool to be performed in low resource settings for malaria detection. [26][27][28] In addition, some commercial LAMP methods for malaria diagnosis have been developed.…”

“…25,26 This flexibility in the way of detection besides the isothermal nature of the process, which does not require a thermocycler or highly specific devices, have supported LAMP method such as an useful tool to be performed in low resource settings for malaria detection. [26][27][28] In addition, some commercial LAMP methods for malaria diagnosis have been developed. [29][30][31] Nevertheless, contamination can be also present in LAMP methods 31 and independent amplification processes are needed for species identification.…”