First report of pfhrp2 and pfhrp3 gene deletions compromising HRP2-based malaria rapid diagnostic tests in Malawi
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Background
Histidine Rich Protein 2-based rapid diagnostic tests (HRP2-based RDTs) are widely used for malaria diagnosis in Malawi, but their accuracy may be compromised by Plasmodium falciparum parasites lacking the P. falciparum histidine rich protein 2 ( pfhrp2 ) and P. falciparum histidine rich protein 3 ( pfhrp3 ) genes. While such deletions have been reported in other malaria-endemic countries, their presence and diagnostic impact in Malawi remain unknown. This study aimed to determine the prevalence of pfhrp2/pfhrp3 gene deletions in Malawi and their effect on the diagnostic accuracy of HRP2-based RDTs relative to light microscopy and qPCR.
Methods
A cross-sectional study was conducted between December 2020 and June 2021, enrolling 1582 participants from referral hospitals in Mzuzu ( n = 1186) and Lilongwe ( n = 396). Malaria diagnosis was performed using RDTs, microscopy, and qPCR. A total of 391 P. falciparum positive samples were analyzed for pfhrp2/pfhrp3 gene deletions using multiplex qPCR. Diagnostic accuracy metrics, such as sensitivity and specificity, were calculated with 95% confidence intervals. Spearman correlation was applied to assess associations involving log-transformed parasitemia, unpaired t -tests were used to compare diagnostic methods, and Mann–Whitney tests were used to compare symptomatic and asymptomatic groups.
Results
Malaria prevalence was higher in Lilongwe (45.2%) than in Mzuzu (22.9%). Infections in Lilongwe were predominantly asymptomatic (94.2%), whereas Mzuzu had mostly symptomatic cases (97.1%) ( P < 0.0002). RDTs demonstrated higher sensitivity of 78.5% (95% CI : 74.6–82.1%) than microscopy 64.8% (95% CI : 60.3–69.1), but slightly lower specificity, with 93.6% (95% CI : 92.0–95.0%) for RDT compared to 95.4% (95% CI : 94.0–96.6%) for microscopy. Dual pfhrp2/3 gene deletions were found in 24 (15.0%) isolates from Lilongwe and 24 (10.4%) from Mzuzu. All dual-deleted samples were false negative by RDT but were positive by microscopy and qPCR.
Conclusions
This study is the first to report pfhrp2/3 gene deletions in Malawi. The presence of these deletions may compromise the performance of HRP2-based RDTs, indicating the need to reassess diagnostic strategies in affected regions.
