Evaluation of Histidine-Rich Proteins 2 and 3 Gene Deletions in Plasmodium falciparum in Endemic Areas of the Brazilian Amazon

Abstract

Histidine-rich proteins 2 and 3 gene (pfhrp2 and pfhrp3) deletions affect the efficacy of rapid diagnostic tests (RDTs) based on the histidine-rich protein 2 (HRP2), compromising the correct identification of the Plasmodium falciparum species. Therefore, molecular surveillance is necessary for the investigation of the actual prevalence of this phenomenon and the extent of the disappearance of these genes in these areas and other South American countries, thus guiding national malaria control programs on the appropriate use of RDTs. This study aimed to evaluate the pfhrp2 and pfhrp3 gene deletion in P. falciparum in endemic areas of the Brazilian Amazon. Aliquots of DNA from the biorepository of the Laboratory of Basic Research in Malaria, Evandro Chagas Institute, with a positive diagnosis for P. falciparum infection as determined by microscopy and molecular assays, were included. Monoinfection was confirmed by nested-polymerase chain reaction assay, and DNA quality was assessed by amplification of the merozoite surface protein-2 gene (msp2). The pfhrp2 and pfhrp3 genes were amplified using primers for the region between exons 1 and 2 and for all extension of exon 2. Aliquots of DNA from 192 P. falciparum isolates were included in the study, with 68.7% (132/192) from the municipality of Cruzeiro do Sul (Acre) and 31.3% (60/192) from Manaus (Amazonas). Of this total, 82.8% (159/192) of the samples were considered of good quality. In the state of Acre, 71.7% (71/99) showed pfhrp2 gene deletion and 94.9% (94/99) showed pfhrp3 gene deletion, while in the state of Amazonas, 100.0% (60/60) of the samples showed pfhrp2 gene deletion and 98.3% (59/60) showed pfhrp3 gene deletion. Moreover, 79.8% (127/159) of isolates displayed gene deletion. Our findings confirm the presence of a parasite population with high frequencies of pfhrp2 and pfhrp3 gene deletions in the Brazilian Amazon region. This suggests reconsidering the use of HRP2-based RDTs in the Acre and Amazonas states and calls attention to the importance of molecular surveillance and mapping of pfhrp2/pfhrp3 deletions in this area and in other locations in the Amazon region to guarantee appropriate patient care, control and ultimately contribute to achieving P. falciparum malaria elimination.

A Comprehensive Analysis of the Genetic Diversity of Plasmodium falciparum Histidine-Rich Protein 2 (PfHRP2) in the Brazilian Amazon

“…Isolates in samples collected from Roraima were found to have the highest proportion (15%) of pfhrp2 gene deletion, followed by the Mato Grosso (10%), and Rondônia (6%) isolates. These findings contrast with previous observations in other areas of the Brazilian Amazon, with rates of 32% to 72% and 54% to 100% being reported in the states of Acre and Amazonas, respectively ( Rachid Viana et al, 2017 ; Góes et al, 2021 ). More consistent with our present findings, Rachid Viana et al (2017) reported a low proportion (3%) of pfhrp2 deletions in Rondônia state for isolates sampled in the Monte Negro municipality.…”

“…Isolates in samples collected from Roraima were found to have the highest proportion (15%) of pfhrp2 gene deletion, followed by the Mato Grosso (10%), and Rondonia (6%) isolates. These findings contrast with previous observations in other areas of the Brazilian Amazon, with rates of 32% to 72% and 54% to 100% being reported in the states of Acre and Amazonas, respectively (Rachid Viana et al, 2017;Goés et al, 2021).…”

“…However, there is evidence to indicate that some monoclonal antibodies used in PfHRP2-based RDTs show cross-reactivity to PfHRP2 and PfHRP3 ( Lee et al, 2006 ). In Brazil, similar to the findings reported from other South American countries (40%–70% of pfhrp3 deletion), the proportion of pfhrp3 deletions is typically very high (35%–98%) ( Gamboa et al, 2010 ; Murillo Solano et al, 2015 ; Dorado et al, 2016 ; Rachid Viana et al, 2017 ; Góes et al, 2021 ). One of the limitations of the present study is that we were unable to assess PfHRP2 levels or the performance of a second brand of RDT to confirm initial results indicating a lack of the protein.…”

“…The sensitivity of RDTs is dependent on detection of the target protein; however, since the 2000s, isolates with deletion of the gene pfhrp2 gene have been reported in Africa and South America ( Baker et al, 2005 ; Baker et al, 2010 ; Gamboa et al, 2010 ; Houzé et al, 2011 ). In Brazil, pfhrp2 deletions have been reported in some endemic states, such as Acre, Rondônia, and Amazonas ( Rachid Viana et al, 2017 ; Góes et al, 2021 ).…”

High-throughput Plasmodium falciparum hrp2 and hrp3 gene deletion typing by digital PCR to monitor malaria rapid diagnostic test efficacy

“…Brazil and Ecuador share no borders, and the amount of human migration is limited. The results add to the heterogeneous pattern of hrp2 / hrp3 deletion in South America, with high frequency of hrp2 deletion in the Amazon ( Gamboa et al, 2010 ; Murillo Solano et al, 2015 ; Góes et al, 2020 ), but low levels among the Pacific coast ( Sáenz et al, 2015 ; Murillo Solano et al, 2015 ). An outbreak of parasites with hrp2 deletion at the Peruvian Pacific coast was caused by infections imported from the Amazon ( Baldeviano et al, 2015 ).…”