[Biological threats to global malaria elimination I Antimalarial drug resistance].

Malaria is an infectious disease that seriously threatens human health. Currently, malaria control mainly depends on antimalarial chemotherapy. However, antimalarial drug resistance is becoming increasingly severe, which poses a great challenge to malaria control, notably treatment of Plasmodium falciparum malaria. To address this challenge, there is a need to facilitate development of novel antimalarial drugs and innovation of treatment strategies, as well as reinforce surveillance and research on antimalarial drug resistance. This article reviews the main categories and use guidelines of current antimalarial agents, summarizes the current status and monitoring methods of antimalarial drug resistance, and proposes the response to antimalarial drug resistance, so as to provide insights into the use of antimalarial drugs and response to antimalarial drug resistance, and contribute to global malaria elimination.

3-Position Biaryl Endochin-like Quinolones with Enhanced Antimalarial Performance.

ELQ-300 is a potent antimalarial drug with activity against blood, liver, and vector stages of the disease. A prodrug, ELQ-331, exhibits reduced crystallinity and improved in vivo efficacy in preclinical testing, and currently, it is in the developmental pipeline for once-a-week dosing for oral prophylaxis against malaria. Because of the high cost of developing a new drug for human use and the high risk of drug failure, it is prudent to have a back-up plan in place. Here we describe ELQ-596, a member of a new subseries of 3-biaryl-ELQs, with enhanced potency in vitro against multidrug-resistant Plasmodium falciparum parasites. ELQ-598, a prodrug of ELQ-596 with diminished crystallinity, is more effective vs murine malaria than its progenitor ELQ-331 by 4- to 10-fold, suggesting that correspondingly lower doses could be used to protect and cure humans of malaria. With a longer bloodstream half-life in mice compared to its progenitor, ELQ-596 highlights a novel series of next-generation ELQs with the potential for once-monthly dosing for protection against malaria infection. Advances in the preparation of 3-biaryl-ELQs are presented along with preliminary results from experiments to explore key structure-activity relationships for drug potency, selectivity, pharmacokinetics, and safety.

Establishment and evaluation of a qPCR method for the detection of pfmdr1 mutations in Plasmodium falciparum, the causal agent of fatal malaria.

Drug resistance surveillance is a major integral part of malaria control programs. Molecular methods play a pivotal role in drug resistance detection and related molecular research. This study aimed to develop a rapid and accurate detection method for drug resistance of Plasmodium falciparum (P. falciparum). A quantitative real-time PCR (qPCR) assay has been developed that identifies the mutation at locus A256T in the P.falciparum multi-drug resistance(pfmdr1) gene producing amino acid change at position 86. The results of 198 samples detected by qPCR were consistent with nested PCR and sequencing, giving an accuracy of 94.3%. The sensitivity, specificity, positive and negative predictive value of qPCR were 85.7%, 97.6%, 90.0% and 96.4%, respectively. The results of qPCR are basically consistent with the nested PCR, which is expected to replace the nested PCR as a new molecular biological method for drug resistance detection, providing reliable technical support for global malaria prevention and control.

Efficacy of artemether-lumefantrine and dihydroartemisinin-piperaquine and prevalence of molecular markers of anti-malarial drug resistance in children in Togo in 2021.

BACKGROUND: Artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) are the currently recommended first- and second-line therapies for uncomplicated Plasmodium falciparum infections in Togo. This study assessed the efficacy of these combinations, the proportion of Day3-positive patients (D3 +), the proportion of molecular markers associated with P. falciparum resistance to anti-malarial drugs, and the variable performance of HRP2-based malaria rapid diagnostic tests (RDTs). METHODS: A single arm prospective study evaluating the efficacy of AL and DP was conducted at two sites (Kouvé and Anié) from September 2021 to January 2022. Eligible children were enrolled, randomly assigned to treatment at each site and followed up for 42 days after treatment initiation. The primary endpoint was polymerase chain reaction (PCR) adjusted adequate clinical and parasitological response (ACPR). At day 0, samples were analysed for mutations in the Pfkelch13, Pfcrt, Pfmdr-1, dhfr, dhps, and deletions in the hrp2/hrp3 genes. RESULTS: A total of 179 and 178 children were included in the AL and DP groups, respectively. After PCR correction, cure rates of patients treated with AL were 97.5% (91.4-99.7) at day 28 in Kouvé and 98.6% (92.4-100) in Anié, whereas 96.4% (CI 95%: 89.1-98.8) and 97.3% (CI 95%: 89.5-99.3) were observed at day 42 in Kouvé and Anié, respectively. The cure rates of patients treated with DP at day 42 were 98.9% (CI 95%: 92.1-99.8) in Kouvé and 100% in Anié. The proportion of patients with parasites on day 3 (D3 +) was 8.5% in AL and 2.6% in DP groups in Anié and 4.3% in AL and 2.1% DP groups in Kouvé. Of the 357 day 0 samples, 99.2% carried the Pfkelch13 wild-type allele. Two isolates carried nonsynonymous mutations not known to be associated with artemisinin partial resistance (ART-R) (A578S and A557S). Most samples carried the Pfcrt wild-type allele (97.2%). The most common Pfmdr-1 allele was the single mutant 184F (75.6%). Among dhfr/dhps mutations, the quintuple mutant haplotype N51I/C59R/S108N + 437G/540E, which is responsible for SP treatment failure in adults and children, was not detected. Single deletions in hrp2 and hrp3 genes were detected in 1/357 (0.3%) and 1/357 (0.3%), respectively. Dual hrp2/hrp3 deletions, which could affect the performances of HRP2-based RDTs, were not observed. CONCLUSION: The results of this study confirm that the AL and DP treatments are highly effective. The absence of the validated Pfkelch13 mutants in the study areas suggests the absence of ART -R, although a significant proportion of D3 + cases were found. The absence of dhfr/dhps quintuple or sextuple mutants (quintuple + 581G) supports the continued use of SP for IPTp during pregnancy and in combination with amodiaquine for seasonal malaria chemoprevention. TRIAL REGISTRATION: ACTRN12623000344695.

Molecular mechanisms of mitochondria-mediated ferroptosis: a potential target for antimalarial interventions.

Ferroptosis is an iron-dependent form of regulated cell death characterized by glutathione (GSH) depletion, glutathione peroxidase 4 (GPX4) inactivation, and the build-up of lipotoxic reactive species. Ferroptosis-targeted induction is a promising therapeutic approach for addressing antimalarial drug resistance. In addition to being the primary source of intracellular energy supply and reactive oxygen species (ROS) generation, mitochondria actively participate in diverse forms of regulated cell death, including ferroptosis. Altered mitochondrial morphology and functionality are attributed to ferroptosis. Diverse mitochondria-related proteins and metabolic activities have been implicated in fine-tuning the action of ferroptosis inducers. Herein, we review recent progress in this evolving field, elucidating the numerous mechanisms by which mitochondria regulate ferroptosis and giving an insight into the role of the organelle in ferroptosis. Additionally, we present an overview of how mitochondria contribute to ferroptosis in malaria. Furthermore, we attempt to shed light on an inclusive perspective on how targeting malaria parasites' mitochondrion and attacking redox homeostasis is anticipated to induce ferroptosis-mediated antiparasitic effects.

Putative molecular markers of Plasmodium falciparum resistance to antimalarial drugs in malaria parasites from Ghana.

Introduction: Antimalarial drugs including artemisinin-based combination therapy (ACT) regimens and sulphadoxine-pyrimethamine (SP) are used in Ghana for malaria therapeutics and prophylaxis respectively. The genetic basis of Plasmodium falciparum development of drug resistance involves single nucleotide polymorphisms in genes encoding proteins for multiple cellular and metabolic processes. The prevalence of single nucleotide polymorphisms in nine P. falciparum genes linked to ACT and SP resistance in the malaria parasite population was determined. Methods: Archived filter paper blood blot samples from patients aged 9 years and below with uncomplicated malaria reporting at 10 sentinel sites located in three ecological zones for the Malaria Therapeutic Efficacy Studies were used. The samples used were collected from 2007-2018 malaria transmission seasons and mutations in the genes were detected using PCR and Sanger sequencing. Results: In all 1,142 samples were used for the study. For falcipain-2 gene (pffp2), Sanger sequencing was successful for 872 samples and were further analysed. The prevalence of the mutants was 45% (392/872) with pffp2 markers V51I and S59F occurring in 15.0% (128/872) and 3.0% (26/872) of the samples respectively. Prevalence of other P. falciparum gene mutations: coronin (pfcoronin) was 44.8% (37/90); cysteine desulfurase (pfnfs) was 73.9% (68/92); apicoplast ribosomal protein S10 (pfarps10) was 36.8% (35/95); ferredoxin (pffd) was 8.8% (8/91); multidrug resistance protein-1 (pfmrp1) was 95.2.0% (80/84); multidrug resistance protein-2 (pfmrp2) was 91.4% (32/35); dihydrofolate reductase (pfdhfr) was 99.0% (84/85); dihydropteroate synthase (pfdhps) was 72% (68/95). Discussion: The observation of numerous mutations in these genes of interest in the Ghanaian isolates, some of which have been implicated in delayed parasite clearance is of great interest. The presence of these genotypes may account for the decline in the efficacies of ACT regimens being used to treat uncomplicated malaria in the country. The need for continuous monitoring of these genetic markers to give first-hand information on parasite susceptibility to antimalarial drugs to inform policy makers and stakeholders in malaria elimination in the country is further discussed.

An Optimized P. berghei Liver Stage-HepG2 Infection Model for Simultaneous Quantitative Bioimaging of Host and Parasite Nascent Proteomes.

The Plasmodium parasites that cause malaria undergo an obligate, asymptomatic developmental stage in the host liver before initiating the symptomatic blood-stage infection. The parasite liver stage is a key intervention point for antimalarial chemoprophylaxis: successful targeting of liver-stage parasites prevents disease development in individuals and can help to reduce parasite transmission in populations, as the gametocyte forms that transmit infection to mosquitos are exclusively found in the blood stage. Antimalarial drugs that can target multiple parasite stages are thus highly desirable, and one emerging cellular target for such multistage active compounds is the process of protein synthesis or translation. Quantitative study of liver stage translation, and thus mechanistic evaluation of translation inhibitors against liver stage parasites, is not amenable to the methods allowing quantification of asexual blood stage translation, such as radiolabeled amino acid incorporation or lysate-based translation of reporter transcripts. Here, we present a method using o-propargyl puromycin (OPP) labeling of host and parasite nascent proteomes in the P. berghei-HepG2 infection model, followed by automated confocal image acquisition and computational separation of P. berghei vs. H. sapiens nascent proteome signals to allow simultaneous readout of the effects of translation inhibitors on both host and parasite. This protocol details our HepG2 cell culture and infected monolayer handling optimized for microscopy, our OPP labeling workflow, and our approach to automated confocal imaging, image processing, and data analysis. Key features • Uses the o-propargyl puromycin labeling technique developed by Liu et al. to quantitatively analyze protein synthesis in Plasmodium berghei liver-stage parasites in actively translating hepatoma cells. • This quantitative approach should be adaptable for other puromycin-sensitive intracellular pathogens residing in actively translating host cells. • The P. berghei-infected HepG2 recovery and reseeding protocol presented here is of use in applications beyond nascent proteome labeling and quantification.

Proguanil Suppresses Breast Tumor Growth In Vitro and In Vivo by Inducing Apoptosis via Mitochondrial Dysfunction.

Breast cancer, ranking as the second leading cause of female cancer-related deaths in the U.S., demands the exploration of innovative treatments. Repurposing FDA-approved drugs emerges as an expedited and cost-effective strategy. Our study centered on proguanil, an antimalarial drug, reveals notable anti-proliferative effects on diverse breast cancer cell lines, including those derived from patients. Proguanil-induced apoptosis was associated with a substantial increase in reactive oxygen species (ROS) production, leading to reduced mitochondrial membrane potential, respiration, and ATP production. Proguanil treatment upregulated apoptotic markers (Bax, p-H2AX, cleaved-caspase 3, 9, cleaved PARP) and downregulated anti-apoptotic proteins (bcl-2, survivin) in breast cancer cell lines. In female Balb/c mice implanted with 4T1 breast tumors, daily oral administration of 20 mg/kg proguanil suppressed tumor enlargement by 55%. Western blot analyses of proguanil-treated tumors supported the in vitro findings, demonstrating increased levels of p-H2AX, Bax, c-PARP, and c-caspase3 as compared to controls. Our results collectively highlight proguanil's anticancer efficacy in vitro and in vivo in breast cancer, prompting further consideration for clinical investigations.

An In Silico and In Vitro Assessment of the Neurotoxicity of Mefloquine.

Mefloquine (MQ) is a quinoline-based anti-malarial drug used for chemoprophylaxis or as a treatment in combination with artesunate. Although MQ has clear anti-Plasmodium falciparum properties, it can induce neurotoxicity and undesired neuropsychiatric side effects in humans. Hence, this study aimed to characterize the neurotoxicity of MQ using human neuroblastoma SH-SY5Y cells. The effects of MQ on neuronal toxicity and cell viability were investigated over a concentration range of 1-100 µM using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays. The influence of MQ on cellular bioenergetics was examined by measuring cellular ATP levels and from the induction of reactive oxygen species (ROS). An in silico approach was used to assess the potential neurotoxicity of MQ mediated via binding to the active sites of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) and then experimentally validated via in vitro enzymatic assays. MQ was cytotoxic to neuronal cells in a concentration and exposure duration dependent manner and induced a significant reduction in viability at concentrations of ≥25 µM after a 24 h exposure. MQ adversely impacted cellular bioenergetics and significantly depleted ATP production at concentrations of ≥1 µM after 24 h. MQ-induced cellular ROS production, which was correlated with the induction of apoptosis, as revealed by flow cytometry. In silico studies suggested that MQ was a dual cholinesterase inhibitor and one with remarkably potent binding to BuChE. Modelling data were supported by in vitro studies which showed that MQ inhibited both human AChE and BuChE enzymes. In summary, MQ is an antimalarial drug that may induce neurotoxicity by impacting cellular bioenergetics and perturbing the activity of cholinesterases at exposure concentrations relevant to human dosage.

Progress towards malaria elimination in the Greater Mekong Subregion: perspectives from the World Health Organization.

Malaria remains a global health challenge, disproportionately affecting vulnerable communities. Despite substantial progress, the emergence of anti-malarial drug resistance poses a constant threat. The Greater Mekong Subregion (GMS), which includes Cambodia, China's Yunnan province, Lao People's Democratic Republic, Myanmar, Thailand, and Viet Nam has been the epicentre for the emergence of resistance to successive generations of anti-malarial therapies. From the perspective of the World Health Organization (WHO), this article considers the collaborative efforts in the GMS, to contain Plasmodium falciparum artemisinin partial resistance and multi-drug resistance and to advance malaria elimination. The emergence of artemisinin partial resistance in the GMS necessitated urgent action and regional collaboration resulting in the Strategy for Malaria Elimination in the Greater Mekong Subregion (2015-2030), advocating for accelerated malaria elimination interventions tailored to country needs, co-ordinated and supported by the WHO Mekong malaria elimination programme. The strategy has delivered substantial reductions in malaria across all GMS countries, with a 77% reduction in malaria cases and a 97% reduction in malaria deaths across the GMS between 2012 and 2022. Notably, China was certified malaria-free by WHO in 2021. Countries' ownership and accountability have been pivotal, with each GMS country outlining its priorities in strategic and annual work plans. The development of strong networks for anti-malarial drug resistance surveillance and epidemiological surveillance was essential. Harmonization of policies and guidelines enhanced collaboration, ensuring that activities were driven by evidence. Challenges persist, particularly in Myanmar, where security concerns have limited recent progress, though an intensification and acceleration plan aims to regain momentum. Barriers to implementation can slow progress and continuing innovation is needed. Accessing mobile and migrant populations is key to addressing remaining transmission foci, requiring effective cross-border collaboration. In conclusion, the GMS has made significant progress towards malaria elimination, particularly in the east where several countries are close to P. falciparum elimination. New and persisting challenges require sustained efforts and continued close collaboration. The GMS countries have repeatedly risen to every obstacle presented, and now is the time to re-double efforts and achieve the 2030 goal of malaria elimination for the region.

Bibliometric analysis of antimalarial drug resistance.

Background: Malaria has always been a serious infectious disease prevalent in the world. Antimalarial drugs such as chloroquine and artemisinin have been the main compounds used to treat malaria. However, the massive use of this type of drugs accelerates the evolution and spread of malaria parasites, leading to the development of resistance. A large number of related data have been published by researchers in recent years. CiteSpace software has gained popularity among us researchers in recent years, because of its ability to help us obtain the core information we want in a mass of articles. In order to analyze the hotspots and develop trends in this field through visual analysis, this study used CiteSpace software to summarize the available data in the literature to provide insights. Method: Relevant literature was collected from the Web of Science Core Collection (WOSCC) from 1 January 2015 to 29 March 2023. CiteSpace software and Microsoft Excel were used to analyze and present the data, respectively. Results: A total of 2,561 literatures were retrieved and 2,559 literatures were included in the analysis after the removal of duplicates. An irrefutable witness of the ever-growing interest in the topic of antimalarial drug resistance could be expressed by the exponentially increased number of publications and related citations from 2015 to 2022, and its sustained growth trend by 2023. During the past 7 years, USA, Oxford University, and David A Fidock are the country, institution, and author with the most publications in this field of research, respectively. We focused on the references and keywords from literature and found that the research and development of new drugs is the newest hotspot in this field. A growing number of scientists are devoted to finding new antimalarial drugs. Conclusion: This study is the first visual metrological analysis of antimalarial drug resistance, using bibliometric methods. As a baseline information, it is important to analyze research output published globally on antimalarial drug resistance. In order to better understand the current research situation and future research plan agenda, such baseline data are needed accordingly.

A computational strategy for systematic virtual screening of plasmodium falciparum heme detoxification protein inhibitors from the Drugbank database.

Antimalarial drug resistance poses one of the greatest threats to malaria treatment, resulting in increased morbidity and mortality. Heme Detoxification Protein (HDP) is among the essential hemoglobinases of P. falciparum (Pf), a vital molecular target for the treatment of malaria. In this study, we utilized the virtual screening workflow tool of the Schrodinger suite to find the best hits for the PfHDP from the DrugBank library. A total of 14,942 compounds were identified against the PfHDP. The top compounds with the highest docking scores and least energy scores were subjected to molecular simulations for 500 nanosecond to check the stability of the protein-drug complexes. The top three DrugBank compounds were found to be stable over 500 ns, namely DB09298 (silibinin), DB07426 (1-Hydroxy-2-(1,1':3',1''-Terphenyl-3-Yloxy) Ethane-1,1-Diyl] Bis (Phosphonic Acid), and DB07410 [(2-(3-Dibenzofuran-4-yl-Phenyl)-1-Hydroxy-1-Phosphono-Ethyl]-Phosphonic Acid). Overall analysis suggests that the top three compounds, DB09298, DB07426, and DB07410, have good stability for 500 ns. Their scaffolds can be used to design and develop new analogs of the target HDP protein. Silibinin, the anti-cancer drug, was found to be highly stable for the entire simulation period as compared to the other compounds. DB07426 shows its therapeutic effect on bones, especially in the treatment of osteoporosis, and DB07410 has anti-tumor, antibacterial, anti-oxidative, and anti-viral activities. All three compounds can be considered for repurposing as antimalarial drugs to evaluate the binding capacity or inhibition potential of these compounds. Further in-vivo and in-vitro analysis against the PfHDP protein should be conducted.Communicated by Ramaswamy H. Sarma.

A spatio-temporal model of multi-marker antimalarial resistance.

The emergence and spread of drug-resistant Plasmodium falciparum parasites have hindered efforts to eliminate malaria. Monitoring the spread of drug resistance is vital, as drug resistance can lead to widespread treatment failure. We develop a Bayesian model to produce spatio-temporal maps that depict the spread of drug resistance, and apply our methods for the antimalarial sulfadoxine-pyrimethamine. We infer from genetic count data the prevalences over space and time of various malaria parasite haplotypes associated with drug resistance. Previous work has focused on inferring the prevalence of individual molecular markers. In reality, combinations of mutations at multiple markers confer varying degrees of drug resistance to the parasite, indicating that multiple markers should be modelled together. However, the reporting of genetic count data is often inconsistent as some studies report haplotype counts, whereas some studies report mutation counts of individual markers separately. In response, we introduce a latent multinomial Gaussian process model to handle partially reported spatio-temporal count data. As drug-resistant mutations are often used as a proxy for treatment efficacy, point estimates from our spatio-temporal maps can help inform antimalarial drug policies, whereas the uncertainties from our maps can help with optimizing sampling strategies for future monitoring of drug resistance.

The problem of antimalarial resistance and its implications for drug discovery.

INTRODUCTION: Malaria remains a devastating infectious disease with hundreds of thousands of casualties each year. Antimalarial drug resistance has been a threat to malaria control and elimination for many decades and is still of concern today. Despite the continued effectiveness of current first-line treatments, namely artemisinin-based combination therapies, the emergence of drug-resistant parasites in Southeast Asia and even more alarmingly the occurrence of resistance mutations in Africa is of great concern and requires immediate attention. AREAS COVERED: A comprehensive overview of the mechanisms underlying the acquisition of drug resistance in Plasmodium falciparum is given. Understanding these processes provides valuable insights that can be harnessed for the development and selection of novel antimalarials with reduced resistance potential. Additionally, strategies to mitigate resistance to antimalarial compounds on the short term by using approved drugs are discussed. EXPERT OPINION: While employing strategies that utilize already approved drugs may offer a prompt and cost-effective approach to counter antimalarial drug resistance, it is crucial to recognize that only continuous efforts into the development of novel antimalarial drugs can ensure the successful treatment of malaria in the future. Incorporating resistance propensity assessment during this developmental process will increase the likelihood of effective and enduring malaria treatments.

Plasmodium falciparum ring-stage plasticity and drug resistance.

Malaria is a life-threatening tropical disease caused by parasites of the genus Plasmodium, of which Plasmodium falciparum is the most lethal. Malaria parasites have a complex life cycle, with stages occurring in both the Anopheles mosquito vector and human host. Ring stages are the youngest form of the parasite in the intraerythrocytic developmental cycle and are associated with evasion of spleen clearance, temporary growth arrest (TGA), and drug resistance. This formidable ability to survive and develop into mature, sexual, or growth-arrested forms demonstrates the inherent population heterogeneity. Here we highlight the role of the ring stage as a crossroads in parasite development and as a reservoir of surviving cells in the human host via TGA survival mechanisms.

Examining the disparities of anti-malarial drug consumption among children under the age of five: a study of 5 malaria-endemic countries.

BACKGROUND: Malaria is one of the most prominent illnesses affecting children, ranking as one of the key development concerns for many low- and middle-income countries (LMICs). There is not much information available on the use of anti-malarial drugs in LMICs in children under five. The study aimed to investigate disparities in anti-malarial drug consumption for malaria among children under the age of five in LMICs. METHODS: This study used recent available cross-sectional data from the Malaria Indicator Survey (MIS) datasets across five LMICs (Guinea, Kenya, Mali, Nigeria, and Sierra Leone), which covered a portion of sub-Saharan Africa. The study was carried out between January 2, 2023, and April 15, 2023, and included children under the age of five who had taken an anti-malarial drug for malaria 2 weeks before the survey date. The outcome variable was anti-malarial drug consumption, which was classified into two groups: those who had taken anti-malarial drugs and those who had not. RESULTS: In the study of LMICs, 32,397 children under five were observed, and among them, 44.1% had received anti-malarial drugs. Of the five LMICs, Kenya had the lowest (9.2%) and Mali had the highest (70.5%) percentages of anti-malarial drug consumption. Children under five with malaria are more likely to receive anti-malarial drugs if they are over 1 year old, live in rural areas, have mothers with higher education levels, and come from wealthier families. CONCLUSION: The study emphasizes the importance of developing universal coverage strategies for anti-malarial drug consumption at both the national and local levels. The study also recommends that improving availability and access to anti-malarial drugs may be necessary, as the consumption of these drugs for treating malaria in children under the age of five is shockingly low in some LMICs.

Low prevalence of antimalarial-resistance mutations in India during 2014-15: Impact of combining first-line therapy with primaquine.

BACKGROUND: Antimalarial drug resistance surveillance and containment are crucial for countries aiming to eliminate malaria. Monitoring resistance evolution through studies before and after treatment policy changes is crucial. METHOD: A total of 939 P. falciparum-positive blood samples were collected between 2014 and 2015 across ten sites in India, categorized into four geographic clusters. PCR-amplified products were sequenced to identify point mutations at drug-resistance-conferring genes (Pfdhfr, Pfdhps, Pfmdr1, Pfk13). RESULT: Triple Pfdhfr mutants were found only in northeast India bordering Myanmar, while the wildtype was dominant in central India. Pfdhps wildtypes were prevalent in all areas, and no double mutants were found. Except in Northwest India, Pfmdr1 wildtype was dominant in all clusters. Nonsynonymous double mutations were only found in northwest India. Only synonymous mutations occurred in Pfk13. These were found in Central India at low frequency. The pattern of linkage disequilibrium and principal component analysis reflects low pressure for drug resistance and heterogeneity between the geographic clusters. CONCLUSION: Resistance levels were highest in Northeast India, close to the Myanmar border, where resistance is common. Primaquine has been widely used as a gametocidal and schizonticidal drug, has likely contributed to maintaining low drug resistance levels and preventing strong selection for resistance.

The landscape of drug resistance in Plasmodium falciparum malaria in the Democratic Republic of Congo: a mapping systematic review.

CONTEXT: The Democratic Republic of Congo (DRC), one of the most malaria-affected countries worldwide, is a potential hub for global drug-resistant malaria. This study aimed at summarizing and mapping surveys of malaria parasites carrying molecular markers of drug-resistance across the country. METHODS: A systematic mapping review was carried out before July 2023 by searching for relevant articles through seven databases (PubMed, Embase, Scopus, African Journal Online, African Index Medicus, Bioline and Web of Science). RESULTS: We identified 1541 primary studies of which 29 fulfilled inclusion criteria and provided information related to 6385 Plasmodium falciparum clinical isolates (collected from 2000 to 2020). We noted the PfCRT K76T mutation encoding for chloroquine-resistance in median 32.1% [interquartile interval, IQR: 45.2] of analyzed malaria parasites. The proportion of parasites carrying this mutation decreased overtime, but wide geographic variations persisted. A single isolate had encoded the PfK13 R561H substitution that is invoked in artemisinin-resistance emergence in the Great Lakes region of Africa. Parasites carrying various mutations linked to resistance to the sulfadoxine-pyrimethamine combination were widespread and reflected a moderate resistance profile (PfDHPS A437G: 99.5% [IQR: 3.9]; PfDHPS K540E: 38.9% [IQR: 47.7]) with median 13.1% [IQR: 10.3] of them being quintuple IRN-GE mutants (i.e., parasites carrying the PfDHFR N51I-C59R-S108N and PfDHPS A437G-K540E mutations). These quintuple mutants tended to prevail in eastern regions of the country. Among circulating parasites, we did not record any parasites harboring mutations related to mefloquine-resistance, but we could suspect those with decreased susceptibility to quinine, amodiaquine, and lumefantrine based on corresponding molecular surrogates. CONCLUSIONS: Drug resistance poses a serious threat to existing malaria therapies and chemoprevention options in the DRC. This review provides a baseline for monitoring public health efforts as well as evidence for decision-making in support of national malaria policies and for implementing regionally tailored control measures across the country.

Unique Properties of Nutrient Channels on Plasmodium-Infected Erythrocytes.

Intracellular malaria parasites activate an ion and organic solute channel on their host erythrocyte membrane to acquire a broad range of essential nutrients. This plasmodial surface anion channel (PSAC) facilitates the uptake of sugars, amino acids, purines, some vitamins, and organic cations, but remarkably, it must exclude the small Na+ ion to preserve infected erythrocyte osmotic stability in plasma. Although molecular, biochemical, and structural studies have provided fundamental mechanistic insights about PSAC and advanced potent inhibitors as exciting antimalarial leads, important questions remain about how nutrients and ions are transported. Here, I review PSAC's unusual selectivity and conductance properties, which should guide future research into this important microbial ion channel.

Sampling for malaria molecular surveillance.

Strategic use of Plasmodium falciparum genetic variation has great potential to inform public health actions for malaria control and elimination. Malaria molecular surveillance (MMS) begins with a strategy to identify and collect parasite samples, guided by public-health priorities. In this review we discuss sampling design practices for MMS and point out epidemiological, biological, and statistical factors that need to be considered. We present examples for different use cases, including detecting emergence and spread of rare variants, establishing transmission sources and inferring changes in malaria transmission intensity. This review will potentially guide the collection of samples and data, serve as a starting point for further methodological innovation, and enhance utilization of MMS to support malaria elimination.