Novel diagnostic methods using loop-mediated isothermal amplification (LAMP) for diagnosing malarial infections and preventing clinical relapse

Authors: CHEN, XI - Kunming Medical University; ZHANG, JIAQI - Kunming Medical University; PAN, MAOHUA - Shanglin County People’s Hospital; ZHAO, HUI - Kunming Medical University; QIN, PIEN - Shanglin County People’s Hospital; WANG, SIQI - Kunming Medical University; SI, YU - Kunming Medical University; YANG, QI - Kunming Medical University; LI, XINXIN - Kunming Medical University; ZENG, WEILIN - Kunming Medical University; ZHENG, XIANG - Kunming Medical University; WU, YANRUI - Kunming Medical University; DUAN, MENGXI - Kunming Medical University; LI, XIAOSONG - Kunming Medical University; WANG, XUN - Kunming Medical University; MAXIER, DOMINIQUE - The Sorbonne University; ZHANG, YANMEI - Kunming Medical University; ZHAO, WEI - Kunming Medical University; Rosenthal, Benjamin; HUANG, YAMING - Kunming Medical University; YANG, ZHAOQING - Kunming Medical University

Submitted to: Parasites & Vectors
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/3/2021
Publication Date: 5/24/2021
Citation: Chen, X., Zhang, J., Pan, M., Zhao, H., Qin, P., Wang, S., Si, Y., Yang, Q., Li, X., Zeng, W., Zheng, X., Wu, Y., Duan, M., Li, X., Wang, X., Maxier, D., Zhang, Y., Zhao, W., Rosenthal, B.M., Huang, Y., Yang, Z. 2021. Novel diagnostic methods using loop-mediated isothermal amplification (LAMP) for diagnosing malarial infections and preventing clinical relapse. Parasites & Vectors. https://doi.org/10.1186/s13071-021-04764-9.
DOI: https://doi.org/10.1186/s13071-021-04764-9

Interpretive Summary: Accurate and timely diagnosis aids surveillance and management of parasitic infections, such as those that compromise food safety. Assays based on detecting nucleic acids of parasitic infections contribute such diagnostic information, but can be costly, require specialized equipment, and can be time-consuming. To explore promising alternatives that can deliver a result in just hours using no specialized equipment, USDA researchers partnered with Chinese researchers interested in improving diagnosis of imported malaria, using a technique with potentially wide application. They demonstrated promising sensitivity and specificity, suggesting that this approach may add significantly to the repertoire of diagnostic tools. This work will be of interest to parasitologists, epidemiologists, and others concerned with public health and microbiology.

Technical Abstract: Background Loop-mediated isothermal amplification (LAMP) has been widely used in the diagnosis of various infectious diseases. Malaria is a globally distributed infectious disease attributed to the genus Plasmodium, including P. vivax and P. ovale, prone to relapse of symptomatic blood stage infections. To date, no commercial LAMP or rapid diagnostic tests (RDT) kit of diagnosing infections with P. ovale are available, even no report of P. ovale infection detected by LAMP technique for clinical samples in epidemic studied in large size. Methods An assay was designed to target a portion of mitochondrial DNA (mtDNA) among five species and two other assays were designed to target the nuclear 18S ribosomal RNA gene (18S rDNA) of either P. vivax or P. ovale. The sensitivity of the assays was compared to nested-PCR using defined concentrations of plasmids containing the target sequences, and using limiting dilutions prepared from clinical isolates derived from Chinese workers who became infected in Africa or near the Chinese border with Myanmar. Results 102 copies of the mitochondrial target or 102, 103 copies of 18S rDNA could be detected from genus Plasmodium spp, P. vivax and P. ovale respectively. In 279 clinical samples tests, the malaria Pan mtDNA LAMP test performed well when compared with a nested-PCR assay (95% CI sensitivity= 98.48-100%; specificity 90.75-100%). When diagnosing of clinical samples, P. vivax, the 18S rDNA assay had a great sensitivity (95.85-100%) and specificity (98.1-100%). The same was true for P. ovale (sensitivity 90.76-99.96%; specificity 98.34-100%). By comparing the copies of plasmids/µL, the Limit of Detection (LOD) of PCR was 100 -old higher than that of Malaria Pan LAMP and that of 18S rDNA P. vivax LAMP, and 10-fold higher than that of 18S rDNA P. ovale LAMP. Conclusion The novel LAMP assays can greatly aid the rapid, reliable, and cost-effective diagnosis of infections with genes of human Plasmodium spp, and species of P. vivax and P. ovale respectively.