Plasmodium falciparum pfhrp2 and pfhrp3 Gene Deletions from Persons with Symptomatic Malaria Infection in Ethiopia, Kenya, Madagascar, and Rwanda

Abstract

Histidine-rich protein 2 (HRP2)–based rapid diagnostic tests detect Plasmodium falciparum malaria and are used throughout sub-Saharan Africa. However, deletions in the pfhrp2 and related pfhrp3 ( pfhrp2/3 ) genes threaten use of these tests. Therapeutic efficacy studies (TESs) enroll persons with symptomatic P. falciparum infection. We screened TES samples collected during 2016–2018 in Ethiopia, Kenya, Rwanda, and Madagascar for HRP2/3, pan- Plasmodium lactate dehydrogenase, and pan- Plasmodium aldolase antigen levels and selected samples with low levels of HRP2/3 for pfhrp2/3 genotyping. We observed deletion of pfhrp3 in samples from all countries except Kenya. Single-gene deletions in pfhrp2 were observed in 1.4% (95% CI 0.2%–4.8%) of Ethiopia samples and in 0.6% (95% CI 0.2%–1.6%) of Madagascar samples, and dual pfhrp2/3 deletions were noted in 2.0% (95% CI 0.4%–5.9%) of Ethiopia samples. Although this study was not powered for precise prevalence estimates, evaluating TES samples revealed a low prevalence of pfhrp2/3 deletions in most sites.

Portable and cost-effective genetic detection and characterization of Plasmodium falciparum hrp2 using the MinION sequencer

“…Figure 3 Positivity thresholds for previously analyzed TES sample dataset. Based on the positivity threshold established for each sequencing pool, calculated as the rst octile between the median coverage of the Dd2 negative control and 3D7 positive control, largely rei es the ndings from a previous analysis of these samples (McCaffery, et al, 2021;Rogier, et al, 2022). Notably, no samples previously assessed to be pfhrp2were found to be putatively positive using this MinION assay.…”

“…The MR4 set was utilized for nal quality control experiments prior to sequencing eld samples. Remaining clinical isolates from Sub-Saharan Africa therapeutic e cacy studies (TES) that were used for opportunistic pfhrp2 surveillance previously (McCaffery, et al, 2021;Rogier, et al, 2022) were also used. Additionally, numerous domestic and international clinical isolates were utilized.…”

“…Public health decisions regarding the continuation or cessation of HRP2based RDT use must be made at the regional level in response to regional prevalence. Methods used to assess pfhrp2 deletion and diversity have included antigen detection (Rogier, et al, 2022), PCR (Abdallah, et al, 2015;Akinyi, et al, 2013), amplicon sequencing (Nderu, et al, 2019), highthroughput whole genome sequencing, and high-throughput targeted deep sequencing (Feleke, et al, 2021). In many studies two or more of these methods are used in conjunction.…”

Screening strategies and laboratory assays to support Plasmodium falciparum histidine-rich protein deletion surveillance: where we are and what is needed

“…Additionally, testing of blood specimens by RDT can be done at point-of-care, or blood saved under cold storage for later assaying of HRP2 by RDT in another setting. This pragmatic and economical screening of symptomatic persons can provide strong phenotypic evidence for P. falciparum infections not producing HRP2 and HRP3 antigens and requiring molecular interrogation to elucidate pfhrp2/3 genotype [ 18 , 55 , 56 ].…”

Factors Affecting the Performance of HRP2-Based Malaria Rapid Diagnostic Tests

“…The following supporting information can be downloaded at: , Table S1: Reports on pfhrp2/3 deletions between 2018 to 2022 from different countries. References [ 131 , 132 , 133 , 134 , 135 , 136 , 137 , 138 , 139 , 140 , 141 , 142 , 143 , 144 , 145 , 146 , 147 , 148 , 149 , 150 , 151 , 152 , 153 , 154 , 155 , 156 , 157 , 158 , 159 , 160 , 161 ] are cited in the supplementary materials.…”