Dataset Information

Plasmodium falciparum rosetting protects schizonts against artemisinin.

ABSTRACT:

Background

Artemisinin (ART) resistance in Plasmodium falciparum is thought to occur during the early stage of the parasite's erythrocytic cycle. Here, we identify a novel factor associated with the late stage parasite development that contributes to ART resistance.

Methods

Rosetting rates of clinical isolates pre- and post- brief (one hour) exposure to artesunate (AS, an ART derivative) were evaluated. The effects of AS-mediated rosetting on the post-AS-exposed parasite's replication and survival, as well as the extent of protection by AS-mediated rosetting on different parasite stages were investigated. The rosetting ligands, mechanisms, and gene mutations involved were studied.

Findings

Brief AS exposure stimulated rosetting, with AS-resistant isolates forming more rosettes in a more rapid manner. AS-mediated rosetting enabled infected erythrocytes (IRBC) to withstand AS exposure for several hours and protected the IRBC from phagocytosis. When their rosetting ability was blocked experimentally, the post-AS exposure survival advantage by the AS-resistant parasites was abrogated. Deletions in two genes coding for PfEMP1 exon 2 (PF3D7_0200300 and PF3D7_0223300) were found to be associated with AS-mediated rosetting, and these mutations were significantly selected through time in the parasite population under study, along with the K13 mutations, a molecular marker of ART-resistance.

Interpretation

Rapid ART parasite clearance is driven by the direct oxidative damages on IRBC by ART and the phagocytic destruction of the damaged IRBC. Rosetting serves as a rapid 'buying time' strategy that allows more parasites to complete schizont maturation, reinvasion and subsequent development into the intrinsically less ART-susceptible ring stage.

Funding

A*STAR, NMRC-OF-YIRG, HRC e-ASIA, Wellcome.

SUBMITTER: Lee WC

PROVIDER: S-EPMC8586750 | biostudies-literature | 2021 Nov

REPOSITORIES: biostudies-literature

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Publications

Plasmodium falciparum rosetting protects schizonts against artemisinin.

Lee Wenn-Chyau WC Russell Bruce B Lee Bernett B Chu Cindy S CS Phyo Aung Pyae AP Sriprawat Kanlaya K Lau Yee-Ling YL Nosten François F Rénia Laurent L

EBioMedicine 20211105

<h4>Background</h4>Artemisinin (ART) resistance in Plasmodium falciparum is thought to occur during the early stage of the parasite's erythrocytic cycle. Here, we identify a novel factor associated with the late stage parasite development that contributes to ART resistance.<h4>Methods</h4>Rosetting rates of clinical isolates pre- and post- brief (one hour) exposure to artesunate (AS, an ART derivative) were evaluated. The effects of AS-mediated rosetting on the post-AS-exposed parasite's replica ...[more]

PMID: 34749300

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Project description:Background: The cytoadherence of Plasmodium falciparum is thought to be mediated by variant surface antigens (VSA), encoded by var, rif, stevor and pfmc-2tm genes. The last three families have rarely been studied in the context of cytoadherence. As most VSA genes are unique, the variability among sequences has impeded the functional study of VSA across different P. falciparum strains. However, many P. falciparum genomes have recently been sequenced, allowing the development of specific microarray probes to each VSA gene. Methods: All VSA sequences from the HB3, Dd2 and IT/FCR3 genomes were extracted using HMMer. Oligonucleotide probes were designed with OligoRankPick and added to the 3D7-based microarray chip. As a proof of concept, IT/R29 parasites were selected for and against rosette formation and the transcriptomes of isogenic rosetting and non-rosetting parasites were compared by microarray. Results: From each parasite strain 50-56 var genes, 125-132 rif genes, 26-33 stevor genes and 3-8 pfmc-2tm genes were identified. The ability of the VSA-supplemented microarray chip to detect cytoadherence-related genes was assessed using P. falciparum clone IT/R29, in which rosetting is known to be mediated by PfEMP1 encoded by ITvar9. Whole transcriptome analysis showed that the most highly upregulated gene in rosetting parasites was ITvar9 (19 to 429-fold upregulated over six time points). Only one rif gene (IT4rifA_042) was upregulated by more than 4-fold (5-fold at 12 hours post-invasion), and no stevor or pfmc-2tm genes were upregulated by more than 2-fold. 49 non-VSA genes were upregulated in rosetting parasites by more than 3-fold in at least two time-points, although none as markedly as ITvar9. Conclusions: We demonstrate that the VSA of newly sequenced P. falciparum strains can be added to the 3D7-based microarray chip, allowing the analysis of the entire transcriptome of multiple strains. For the rosetting clone IT/R29, the striking transcriptional upregulation of ITvar9 was confirmed, and the data did not support the involvement of other VSA families in rosette formation.

Project description:BackgroundArtemisinin-based combination therapies are the recommended first-line treatments of falciparum malaria in all countries with endemic disease. There are recent concerns that the efficacy of such therapies has declined on the Thai-Cambodian border, historically a site of emerging antimalarial-drug resistance.MethodsIn two open-label, randomized trials, we compared the efficacies of two treatments for uncomplicated falciparum malaria in Pailin, western Cambodia, and Wang Pha, northwestern Thailand: oral artesunate given at a dose of 2 mg per kilogram of body weight per day, for 7 days, and artesunate given at a dose of 4 mg per kilogram per day, for 3 days, followed by mefloquine at two doses totaling 25 mg per kilogram. We assessed in vitro and in vivo Plasmodium falciparum susceptibility, artesunate pharmacokinetics, and molecular markers of resistance.ResultsWe studied 40 patients in each of the two locations. The overall median parasite clearance times were 84 hours (interquartile range, 60 to 96) in Pailin and 48 hours (interquartile range, 36 to 66) in Wang Pha (P<0.001). Recrudescence confirmed by means of polymerase-chain-reaction assay occurred in 6 of 20 patients (30%) receiving artesunate monotherapy and 1 of 20 (5%) receiving artesunate-mefloquine therapy in Pailin, as compared with 2 of 20 (10%) and 1 of 20 (5%), respectively, in Wang Pha (P=0.31). These markedly different parasitologic responses were not explained by differences in age, artesunate or dihydroartemisinin pharmacokinetics, results of isotopic in vitro sensitivity tests, or putative molecular correlates of P. falciparum drug resistance (mutations or amplifications of the gene encoding a multidrug resistance protein [PfMDR1] or mutations in the gene encoding sarco-endoplasmic reticulum calcium ATPase6 [PfSERCA]). Adverse events were mild and did not differ significantly between the two treatment groups.ConclusionsP. falciparum has reduced in vivo susceptibility to artesunate in western Cambodia as compared with northwestern Thailand. Resistance is characterized by slow parasite clearance in vivo without corresponding reductions on conventional in vitro susceptibility testing. Containment measures are urgently needed. (ClinicalTrials.gov number, NCT00493363, and Current Controlled Trials number, ISRCTN64835265.)

Dataset Information

Plasmodium falciparum rosetting protects schizonts against artemisinin.

Background

Methods

Findings

Interpretation

Funding

Publications

Plasmodium falciparum rosetting protects schizonts against artemisinin.

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