pfhrp2 and pfhrp3 Gene Deletions That Affect Malaria Rapid Diagnostic Tests for Plasmodium falciparum: Analysis of Archived Blood Samples From 3 African Countries

Abstract

Background: Malaria rapid diagnostic tests (mRDTs) that target histidine-rich protein 2 (HRP2) are important tools for Plasmodium falciparum diagnosis. Parasites with pfhrp2/3 gene deletions threaten the use of these mRDTs and have been reported in Africa, Asia, and South America. We studied blood samples from 3 African countries to determine if these gene deletions were present.

Methods: We analyzed 911 dried blood spots from Ghana (n = 165), Tanzania (n = 176), and Uganda (n = 570). Plasmodium falciparum infection was confirmed by 18S rDNA polymerase chain reaction (PCR), and pfhrp2/3 genes were genotyped. True pfhrp2/3 gene deletions were confirmed if samples were (1) microscopy positive; (2) 18S rDNA PCR positive; (3) positive for merozoite surface protein genes by PCR or positive by loop-mediated isothermal amplification; or (4) quantitative PCR positive with >5 parasites/µL.

Results: No pfhrp2/3 deletions were detected in samples from Ghana, but deletions were identified in Tanzania (3 pfhrp2; 2 pfhrp3) and Uganda (7 pfhrp2; 2 pfhrp3). Of the 10 samples with pfhrp2 deletions, 9 tested negative by HRP2-based mRDT.

Conclusions: The presence of pfhrp2/3 deletions in Tanzania and Uganda, along with reports of pfhrp2/3-deleted parasites in neighboring countries, reinforces the need for systematic surveillance to monitor the reliability of mRDTs in malaria-endemic countries.

Keywords: pfhrp2; pfhrp3; Ghana; Tanzania; Uganda; deletion; histidine; malaria; mutation; rapid diagnostic test.

Figure 1.

Two-by-two tables showing results of malaria rapid diagnostic tests (mRDTs) based on detection of histidine-rich protein 2 and expert microscopy for human blood samples analyzed for pfhrp2/3 genes. ^aFive samples from Tanzania had no corresponding mRDT result.

Figure 2.

Percentage of samples positive for Plasmodium falciparum in study samples, by detection method. *Denotes κ value of ≥0.6, indicating good agreement between diagnostic methods. Abbreviations: mRDT, malaria rapid diagnostic test; PCR, polymerase chain reaction.

Figure 3.

Flow diagram showing process of determining pfhrp2 and pfhrp3 gene deletions in blood samples from studies in 3 African countries. *The first number in each row denotes the number of samples among pfhrp2-negative samples; the second number denotes the number among pfhrp3-negative samples. Abbreviations: G, Ghana; LAMP, loop-mediated isothermal amplification; msp, merozoite surface protein; PCR, polymerase chain reaction; qPCR, quantitative polymerase chain reaction; T, Tanzania; U, Uganda.