Search Results (15)

Malaria remains a health problem, with Plasmodium falciparum accounting for 96% of cases in Africa and 15% in Brazil. The growing threat of drug resistance to artemisinin-based combination therapies (ACTs) jeopardizes progress toward elimination. This study examined P. falciparum samples collected from 141 patients in Brazil (2013–2023) by PCR and DNA sequencing to identify single-nucleotide polymorphisms in the pfcrt, pfmdr1, and pfk13 genes. Half of the samples carried the SVMNTMCGI haplotype in pfcrt, and none of the samples showed C350R mutations. In pfmdr1, the NYCDY haplotype was dominant (70%), with low occurrences of N86Y (4%) and no Y184F polymorphisms. No mutations linked to artemisinin partial resistance were detected in pfk13. Only one Amazonas sample exhibited wild-type haplotypes across all genes. Genetic diversity was more pronounced in pfcrt than pfmdr1, reflecting selective drug pressure. Significant linkage disequilibrium (LD) was observed within pfcrt (C72S and K76T) and pfmdr1 (S1034C and N1042D), but not between the two genes. The absence of pfk13-resistant mutations and the low prevalence of key pfmdr1 markers support the efficacy of ACTs. The persistence of diverse haplotypes and intragenic LD reflects ongoing drug pressure, underscoring the need for continuous genetic surveillance to anticipate emerging resistance. Full article

Malaria has been and remains a significant challenge in Africa and other endemic settings. Roughly, 95% of global morbidity and mortality due to malaria occurs within African populations and affects millions of individuals, especially those living in sub-Saharan countries, predominantly due to disease complications. Cultural factors such as unawareness of and disinterest in using recommended preventive tools and combating the primary host (i.e., the female Anopheles mosquito) play a significant role. This host transmits the malaria-causing Plasmodium parasite by biting an infected individual and spreading it to humans. The current overview focuses on the molecular markers associated with antimalarial drug resistance in Plasmodium falciparum (P. falciparum) in Rwanda, considered an exemplar of sub-Saharan countries where malaria is prevalent and effective policies on the development of malaria treatment, approved recently by WHO in 2025, have been adopted. The prevalence of mutations in key resistance genes, including pfcrt, pfmdr1, and pfdhfr/pfdhps, are linked to resistance against common antimalarial drugs such as chloroquine and sulfadoxine-pyrimethamine (SP). In addition, the Plasmodium falciparum kelch13 (pfk13) gene is linked to resistance against artemisinin, as its mutations can cause delayed parasite clearance and treatment failure. Despite changes in therapeutic use policies owing to high prevalence of variant alleles, which reduce the drug’s efficacy resistance to SP, the gene persists in Rwanda. Malaria parasites are becoming more resistant to chloroquine, leading to diminished effectiveness and slower recovery or treatment failure. Surveillance data reported from several studies provide crucial insights into the evolving trends of resistance markers and are vital for guiding treatment protocols and informing therapeutic use policy decisions. It is important that we continue to maintain and develop the effectiveness of malaria prevention strategies and treatments, due to the multiple types of resistance found in the population. Full article

Malaria continues to be a significant public health burden in many tropical and subtropical regions. Mozambique ranks among the top countries affected by malaria, where it is a leading cause of morbidity and mortality, accounting for 29% of all hospital deaths in the general population and 42% of deaths amongst children under five. This review presents a comparative analysis of data on five critical genes associated with antimalarial drug resistance: pfmdr1, pfcrt, pfk13, pfdhfr, and pfdhps, along with the copy number variation (CNV) in genes pfmdr1 and pfpm2/3. These are genes associated with parasite response to antimalarials currently used to treat uncomplicated P. falciparum malaria in Mozambique. The review synthesizes data collected from published studies conducted in Mozambique after the introduction of artemisinin-based combination therapies (ACTs) (2006) up to June 2024, highlighting the presence or absence of mutations in these genes across Mozambique. We aimed at mapping the prevalence and distribution of these molecular markers across the country in order to contribute to the development of targeted interventions to sustain the efficacy of malaria treatments in Mozambique. Four databases were used to access the articles: PubMed, Science Direct, Scopus, and Google scholar. The search strategy identified 132 studies addressing malaria and antimalarial resistance. Of these, 112 were excluded for various reasons, leaving 20 studies to be included in this review. Children and pregnant women represent the majority of target groups in studies on all types of antimalarials. Most studies (87.5%) were conducted in the provinces of Maputo and Gaza. The primary alleles reported were pfcrt CVMNK, and in the most recent data, its wild-type form was found in the majority of patients. A low prevalence of mutations in the pfk13 gene was identified reflecting the effectiveness of ACTs. In pfk13, only mutation A578S was reported in Niassa and Tete. CNVs were observed in studies carried out in the south of Mozambique, with a frequency of 1.1–5.1% for pfmdr1 and a frequency of 1.1–3.4% for pfpm2. This review indicates that molecular markers linked to malaria resistance show considerable variation across provinces in Mozambique, with most up-to-date data accessible for Maputo and Gaza. In contrast, provinces such as Zambezia and Inhambane have limited data on several genes, while Nampula lacks data on all drug resistance markers. Full article

Djibouti is confronted with malaria resurgence, with malaria having been occurring in epidemic proportions since a decade ago. The current epidemiology of drug-resistant Plasmodium falciparum is not well known. Molecular markers were analyzed by targeted sequencing in 79 P. falciparum clinical isolates collected in Djibouti city in 2023 using the Miseq Illumina platform newly installed in the country. The objective of the study was to analyze the key codons in these molecular markers associated with antimalarial drug resistance. The prevalence of the mutant Pfcrt CVIET haplotype (92%) associated with chloroquine resistance and mutant Pfdhps-Pfdhfr haplotypes (7.4% SGEA and 53.5% IRN, respectively) associated with sulfadoxine-pyrimethamine resistance was high. By contrast, Pfmdr1 haplotypes associated with amodiaquine (YYY) or lumefantrine (NFD) resistance were not observed in any of the isolates. Although the “Asian-type” PfK13 mutations associated with artemisinin resistance were not observed, the “African-type” PfK13 substitution, R622I, was found in a single isolate (1.4%) for the first time in Djibouti. Our genotyping data suggest that most Djiboutian P. falciparum isolates are resistant to chloroquine and sulfadoxine-pyrimethamine but are sensitive to amodiaquine, lumefantrine, and artemisinin. Nonetheless, the presence of an isolate with the R622I PfK13 substitution is a warning signal that calls for a regular surveillance of molecular markers of antimalarial drug resistance. Full article

The objective of this study was to analyze the relationship between the frequency of artemisinin-based combination (ACT) drug resistance molecular markers and clinical forms of P. falciparum malaria and parasitemia. A cross-sectional study was carried out between January and April 2014 at the Operational Clinical Research Unit of Melen in febrile children aged 12 to 240 months with a Plasmodium sp. infection. A total of 3 mL of peripheral blood collected from an EDTA tube was used for leukocyte depletion. DNA mutation detection was performed by next generation sequencing (NGS). A total of 1075 patients were screened for malaria. Among them, 384 had a Plasmodium infection. P. falciparum mono-infection was found in 98.9% of the patients. Pfcrt-326T mutation was found in all isolates, while 37.9% had Pfmdr2-484I mutant allele. The highest median parasite densities were found in patients infected by parasites carrying the CVIET haplotype of the Pfcrt gene. The different genetic profiles found here, and their variations according to clinical and biological signs of severe malaria, are additional arguments for the surveillance of P. falciparum strains. Full article

The Greater Mekong Subregion (GMS) is the epicenter of antimalarial drug resistance. We determined in vitro susceptibilities to 11 drugs of culture-adapted Plasmodium falciparum isolates from adjacent areas (Laiza and Muse) along the China–Myanmar border. Parasites from this region were highly resistant to chloroquine and pyrimethamine but relatively sensitive to other antimalarial drugs. Consistently, the Dd2-like pfcrt mutations were fixed or almost fixed in both parasite populations, and new mutations mediating piperaquine resistance were not identified. Similarly, several mutations related to pfdhfr and pfdhps were also highly prevalent. Despite their geographical proximity, malaria parasites from Laiza showed significantly higher in vitro resistance to artemisinin derivatives, naphthoquine, pyronaridine, lumefantrine, and pyrimethamine than parasites from Muse. Likewise, the pfdhfr N51I, pfdhps A581G, pfmrp1 H785N, and pfk13 F446I mutations were significantly more frequent in Laiza than in Muse (p < 0.05). For the pfmdr1 mutations, Y184F was found only in Laiza (70%), whereas F1226Y was identified only in Muse (31.8%). Parasite isolates from Laiza showed a median RSA value of 5.0%, significantly higher than the 2.4% in Muse. Altogether, P. falciparum parasite populations from neighboring regions in the GMS may diverge substantially in their resistance to several antimalarial drugs. This information about different parasite populations will guide antimalarial treatment policies to effectively manage drug resistance during malaria elimination. Full article

Emergence and spreading of antimalarial drug resistant malaria parasites are great hurdles to combating malaria. Although approaches to investigate antimalarial drug resistance status in Myanmar malaria parasites have been made, more expanded studies are necessary to understand the nationwide aspect of antimalarial drug resistance. In the present study, molecular epidemiological analysis for antimalarial drug resistance genes in Plasmodium falciparum and P. vivax from the Mandalay region of Myanmar was performed. Blood samples were collected from patients infected with P. falciparum and P. vivax in four townships around the Mandalay region, Myanmar in 2015. Partial regions flanking major mutations in 11 antimalarial drug resistance genes, including seven genes (pfdhfr, pfdhps, pfmdr-1, pfcrt, pfk13, pfubp-1, and pfcytb) of P. falciparum and four genes (pvdhfr, pvdhps, pvmdr-1, and pvk12) of P. vivax were amplified, sequenced, and overall mutation patterns in these genes were analyzed. Substantial levels of mutations conferring antimalarial drug resistance were detected in both P. falciparum and P. vivax isolated in Mandalay region of Myanmar. Mutations associated with sulfadoxine-pyrimethamine resistance were found in pfdhfr, pfdhps, pvdhfr, and pvdhps of Myanmar P. falciparum and P. vivax with very high frequencies up to 90%. High or moderate levels of mutations were detected in genes such as pfmdr-1, pfcrt, and pvmdr-1 associated with chloroquine resistance. Meanwhile, low frequency mutations or none were found in pfk13, pfubp-1, pfcytb, and pvk12 of the parasites. Overall molecular profiles for antimalarial drug resistance genes in malaria parasites in the Mandalay region suggest that parasite populations in the region have substantial levels of mutations conferring antimalarial drug resistance. Continuous monitoring of mutations linked with antimalarial drug resistance is necessary to provide useful information for policymakers to plan for proper antimalarial drug regimens to control and eliminate malaria in the country. Full article

The effectiveness of artemisinin-based combination therapies (ACTs) depends not only on that of artemisinin but also on that of partner molecules. This study aims to evaluate the prevalence of mutations in the Pfdhfr, Pfdhps, and Pfmdr1 genes from isolates collected during a clinical study. Plasmodium genomic DNA samples extracted from symptomatic malaria patients from Dogondoutchi, Niger, were sequenced by the Sanger method to determine mutations in the Pfdhfr (codons 51, 59, 108, and 164), Pfdhps (codons 436, 437, 540, 581, and 613), and Pfmdr1 (codons 86, 184, 1034, and 1246) genes. One hundred fifty-five (155) pre-treatment samples were sequenced for the Pfdhfr, Pfdhps, and Pfmdr1 genes. A high prevalence of mutations in the Pfdhfr gene was observed at the level of the N51I (84.97%), C59R (92.62%), and S108N (97.39%) codons. The key K540E mutation in the Pfdhps gene was not observed. Only one isolate was found to harbor a mutation at codon I431V. The most common mutation on the Pfmdr1 gene was Y184F in 71.43% of the mutations found, followed by N86Y in 10.20%. The triple-mutant haplotype N51I/C59R/S108N (IRN) was detected in 97% of the samples. Single-mutant (ICS and NCN) and double-mutant (IRS, NRN, and ICN) haplotypes were prevalent at 97% and 95%, respectively. Double-mutant haplotypes of the Pfdhps (581 and 613) and Pfmdr (86 and 184) were found in 3% and 25.45% of the isolates studied, respectively. The study focused on the molecular analysis of the sequencing of the Pfdhfr, Pfdhps, and Pfmdr1 genes. Although a high prevalence of mutations in the Pfdhfr gene have been observed, there is a lack of sulfadoxine pyrimethamine resistance. There is a high prevalence of mutation in the Pfmdr184 codon associated with resistance to amodiaquine. These data will be used by Niger’s National Malaria Control Program to better monitor the resistance of Plasmodium to partner molecules in artemisinin-based combination therapies. Full article

Drug resistance often emerges from mutations in solute transporters. Single amino acid exchanges may alter functionality of transporters with ‘de novo’ ability to transport drugs away from their site of action. The PfMDR1 transporter (or P-glycoprotein 1) is located in the membrane of the digestive vacuole (DV), functions as an ATP-dependent pump, and transports substrates into the DV. In this study, four strains of Plasmodium falciparum, carrying various pfmdr1 gene mutations, were analysed for their transport characteristics of Fluo-4 in isolated DVs of parasites. To obtain quantitative estimates for PfMDR1 DV surface expression, PfMDR1 protein amounts on each strain’s DV membrane were evaluated by quantitative ELISA. Fluo-4, acting as a substrate for PfMDR1, was applied in DV uptake assays (‘reverse Ca²⁺ imaging’). Viable DVs were isolated from trophozoite stages with preserved PfMDR1 activity. This newly developed assay enabled us to measure the number of Fluo-4 molecules actively transported into isolated DVs per PfMDR1 molecule. The drug-resistant strain Dd2 presented the highest transport rates, followed by K1 and the drug-sensitive strain 3D7, compatible with their copy numbers. With this assay, an evaluation of the probability of resistance formation for newly developed drugs can be implemented in early stages of drug development. Full article

Malaria is one of the ‘big three’ killer infectious diseases, alongside tuberculosis and HIV. In non-endemic areas, malaria may occur in travelers who have recently been to or visited endemic regions. The number of imported malaria cases in Portugal has increased in recent years, mostly due to the close relationship with the community of Portuguese language countries. Samples were collected from malaria-infected patients attending Centro Hospitalar Lisboa Ocidental (CHLO) or the outpatient clinic of Instituto de Higiene e Medicina Tropical (IHMT-NOVA) between March 2014 and May 2021. Molecular characterization of Plasmodium falciparum pfk13 and pfmdr1 genes was performed. We analyzed 232 imported malaria cases. The majority (68.53%) of the patients came from Angola and only three patients travelled to a non-African country; one to Brazil and two to Indonesia. P. falciparum was diagnosed in 81.47% of the cases, P. malariae in 7.33%, P. ovale 6.47% and 1.72% carried P. vivax. No mutations were detected in pfk13. Regarding pfmdr1, the wild-type haplotype (N86/Y184/D1246) was also the most prevalent (64.71%) and N86/184F/D1246 was detected in 26.47% of the cases. The typical imported malaria case was middle-aged male, traveling from Angola, infected with P. falciparum carrying wild type pfmdr1 and pfk13. Our study highlights the need for constant surveillance of malaria parasites imported into Portugal as an important pillar of public health. Full article

Background: Artemether-lumefantrine is a highly effective artemisinin-based combination therapy that was adopted in Mali as first-line treatment for uncomplicated Plasmodium falciparum malaria. This study was designed to measure the efficacy of artemether-lumefantrine and to assess the selection of the P. falciparum chloroquine resistance transporter (pfcrt) and P. falciparum multi-drug resistance 1 (pfmdr1) genotypes that have been associated with drug resistance. Methods: A 28-day follow-up efficacy trial of artemether-lumefantrine was conducted in patients aged 6 months and older suffering from uncomplicated falciparum malaria in four different Malian areas during the 2009 malaria transmission season. The polymorphic genetic markers MSP2, MSP1, and Ca1 were used to distinguish between recrudescence and reinfection. Reinfection and recrudescence were then grouped as recurrent infections and analyzed together by PCR-restriction fragment length polymorphism (RFLP) to identify candidate markers for artemether-lumefantrine tolerance in the P. falciparum chloroquine resistance transporter (pfcrt) gene and the P. falciparum multi-drug resistance 1 (pfmdr1) gene. Results: Clinical outcomes in 326 patients (96.7%) were analyzed and the 28-day uncorrected adequate clinical and parasitological response (ACPR) rate was 73.9%. The total PCR-corrected 28-day ACPR was 97.2%. The pfcrt 76T and pfmdr1 86Y population prevalence decreased from 49.3% and 11.0% at baseline (n = 337) to 38.8% and 0% in patients with recurrent infection (n = 85); p = 0.001), respectively. Conclusion: Parasite populations exposed to artemether-lumefantrine in this study were selected toward chloroquine-sensitivity and showed a promising trend that may warrant future targeted reintroduction of chloroquine or/and amodiaquine. Full article

Half the human population is exposed to malaria. Plasmodium falciparum antimalarial drug resistance monitoring and development of new drugs are major issues related to the control of malaria. Methylene blue (MB), the oldest synthetic antimalarial, is again a promising drug after the break of its use as an antimalarial drug for more than 80 years and a potential partner for triple combination. Very few data are available on the involvement of polymorphisms on genes known to be associated with standard antimalarial drugs and parasite in vitro susceptibility to MB (cross-resistance). In this context, MB susceptibility was evaluated against 482 isolates of imported malaria from Africa by HRP2-based ELISA chemosusceptibility assay. A total of 12 genes involved in antimalarial drug resistance (Pfcrt, Pfdhfr, Pfmdr1, Pfmdr5, Pfmdr6, PfK13, Pfubq, Pfcarl, Pfugt, Pfact, Pfcoronin, and copy number of Pfpm2) were sequenced by Sanger method and quantitative PCR. On the Pfmdr1 gene, the mutation 86Y combined with 184F led to more susceptible isolates to MB (8.0 nM vs. 11.6 nM, p = 0.03). Concerning Pfmdr6, the isolates bearing 12 Asn repetitions were more susceptible to MB (4.6 nM vs. 11.6 nM, p = 0.005). None of the polymorphisms previously described as involved in antimalarial drug resistance was shown to be associated with reduced susceptibility to MB. Some genes (particularly PfK13, Pfugt, Pfact, Pfpm2) did not present enough genetic variability to draw conclusions about their involvement in reduced susceptibility to MB. None of the polymorphisms analyzed by multiple correspondence analysis (MCA) had an impact on the MB susceptibility of the samples successfully included in the analysis. It seems that there is no in vitro cross-resistance between MB and commonly used antimalarial drugs. Full article

Suspicion of failure in the effectiveness of artemisinin-based combination therapies (currently the first-line treatment of malaria, worldwide) is leading to the unofficial use of alternative antimalarials, including chloroquine and sulfadoxine/pyrimethamine, across northern Nigeria. To facilitate evidence-based resistance management, antimalarial resistance mutations were investigated in Plasmodium falciparum multidrug resistance-1 (pfmdr1) and chloroquine resistance transporter (pfcrt), in isolates from Kano, northwestern Nigeria. Out of the 88 samples genotyped for pfmdr1 N86Y mutation using PCR/restriction fragment length polymorphism, one sample contained the 86Y mutation (86Y_frequency = 1.14%). The analysis of 610 bp fragments of pfmdr1 from 16 isolates revealed two polymorphic sites and low haplotype diversity (H_d = 0.492), with only 86 Y mutations in one isolate, and 184 F replacements in five isolates (184F_frequency = 31.25%). The analysis of 267 bp fragments of pfcrt isolates revealed high polymorphism (H_d = 0.719), with six haplotypes and seven non-synonymous polymorphic sites. Eleven isolates (61.11%) were chloroquine-resistant, CQR (C₇₂V₇₃I₇₄E₇₅T₇₆ haplotype), two of which had an additional mutation, D⁵⁷E. An additional sequence was CQR, but of the C₇₂V₇₃M₇₄E₇₅T₇₆ haplotype, while the rest of the sequences (33.33%) were chloroquine susceptible (C₇₂V₇₃M₇₄N₇₅K₇₆ haplotype). The findings of these well characterized resistance markers should be considered when designing resistance management strategies in the northwestern Nigeria. Full article

The emergence and spread of drug resistance is a problem hindering malaria elimination in Southeast Asia. In this study, genetic variations in drug resistance markers of Plasmodium falciparum were determined in parasites from asymptomatic populations located in three geographically dispersed townships of Myanmar by PCR and sequencing. Mutations in dihydrofolate reductase (pfdhfr), dihydropteroate synthase (pfdhps), chloroquine resistance transporter (pfcrt), multidrug resistance protein 1 (pfmdr1), multidrug resistance-associated protein 1 (pfmrp1), and Kelch protein 13 (k13) were present in 92.3%, 97.6%, 84.0%, 98.8%, and 68.3% of the parasites, respectively. The pfcrt K76T, pfmdr1 N86Y, pfmdr1 I185K, and pfmrp1 I876V mutations were present in 82.7%, 2.5%, 87.5%, and 59.8% isolates, respectively. The most prevalent haplotypes for pfdhfr, pfdhps, pfcrt and pfmdr1 were 51I/59R/108N/164L, 436A/437G/540E/581A, 74I/75E/76T/220S/271E/326N/356T/371I, and 86N/130E/184Y/185K/1225V, respectively. In addition, 57 isolates had three different point mutations (K191T, F446I, and P574L) and three types of N-terminal insertions (N, NN, NNN) in the k13 gene. In total, 43 distinct haplotypes potentially associated with multidrug resistance were identified. These findings demonstrate a high prevalence of multidrug-resistant P. falciparum in asymptomatic infections from diverse townships in Myanmar, emphasizing the importance of targeting asymptomatic infections to prevent the spread of drug-resistant P. falciparum. Full article

Malaria is a significant public health problem in Ghana. Seasonal Malaria Chemoprevention (SMC) using a combination of sulfadoxine-pyrimethamine and amodiaquine has been implemented since 2015 in northern Ghana where malaria transmission is intense and seasonal. In this study, we estimated the prevalence of asymptomatic P. falciparum carriers in three ecological zones of Ghana, and compared the sensitivity and specificity of different molecular methods in identifying asymptomatic infections. Moreover, we examined the frequency of mutations in pfcrt, pfmdr1, pfdhfr, and pfdhps that relate to the ongoing SMC. A total of 535 asymptomatic schoolchildren were screened by microscopy and PCR (18s rRNA and TARE-2) methods. Among all samples, 28.6% were detected as positive by 18S nested PCR, whereas 19.6% were detected by microscopy. A high PCR-based asymptomatic prevalence was observed in the north (51%) compared to in the central (27.8%) and south (16.9%). The prevalence of pfdhfr-N51I/C59R/S108N/pfdhps-A437G quadruple mutant associated with sulfadoxine-pyrimethamine resistance was significantly higher in the north where SMC was implemented. Compared to 18S rRNA, TARE-2 serves as a more sensitive molecular marker for detecting submicroscopic asymptomatic infections in high and low transmission settings. These findings establish a baseline for monitoring P. falciparum prevalence and resistance in response to SMC over time. Full article