Field evaluation of a loop-mediated isothermal amplification test for detection of Plasmodium spp. in the Amazonas region of Peru

Malaria remains a significant public health burden in Peru, particularly in remote endemic areas where access to sensitive diagnostic methods is limited. We evaluated a simplified molecular approach combining Boil and Spin (B&S) DNA extraction with malachite green loop-mediated isothermal amplification (MG-LAMP) for the detection of Plasmodium spp. Under laboratory conditions, 100 Plasmodium spp.-positive and 100 negative samples confirmed by real-time PCR from inhabitants of the Río Santiago District, Amazonas, Peru, were tested with the B&S-MG-LAMP assay. Subsequently, we screened 252 residents from malaria-endemic communities in this area during 2024 as a field evaluation. Standard diagnostic performance measures, including sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and area under the ROC curve (AUC), were calculated using real-time PCR as the reference standard. B&S-MG-LAMP detected one Plasmodium spp. parasite/μl, compared with 2.5 parasites/μl for real-time PCR. In the laboratory evaluation (n = 200), the assay showed 97.0% sensitivity and 100.0% specificity for Plasmodium spp., with 98.5% overall agreement (Cohen's κ = 0.97) with real-time PCR. Sensitivity was 93.6% for P. falciparum and 100% for P. vivax, with 100% specificity for both species. In the field evaluation (n = 252), B&S-MG-LAMP detected 19 positive samples, whereas real-time PCR detected 17, all corresponding to P. vivax. The assay showed 100% sensitivity and 99.1% specificity, with excellent agreement with real-time PCR (Cohen's κ = 0.92; AUC = 0.99). No P. falciparum-positive cases were detected by either method during the field evaluation. The B&S-MG-LAMP assay demonstrated high diagnostic performance for malaria detection in the population studied. Our findings support its potential as a deployable molecular diagnostic approach in areas with limited access to conventional laboratory infrastructure.