plasmodium falciparum
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2023 ◽  
Vol 83 ◽  
M. F. Nadeem ◽  
N. Zeeshan ◽  
A. A. Khattak ◽  
U. A. Awan ◽  
A. Yaqoob

Abstract Plasmodium falciparum resistance to Chloroquine (CQ) is a significant cause of mortality and morbidity worldwide. There is a paucity of documented data on the prevalence of CQ-resistant mutant haplotypes of Pfcrt and Pfmdr1 genes from malaria-endemic war effected Federally Administered Tribal Areas of Pakistan. The objective of this study was to investigate the prevalence of P. falciparum CQ-resistance in this area. Clinical isolates were collected between May 2017 and May 2018 from North Waziristan and South Waziristan agencies of Federally Administrated Trial Area. Subsequently, Giemsa-stained blood smears were examined to detect Plasmodium falciparum. Extraction of malarial DNA was done from microscopy positive P. falciparum samples, and P. falciparum infections were confirmed by nested PCR (targeting Plasmodium small subunit ribosomal ribonucleic acid (ssrRNA) genes). All PCR confirmed P. falciparum samples were sequenced by pyrosequencing to find out mutation in Pfcrt gene at codon K76T and in pfmdr1 at codons N86Y, Y184F, N1042D, and D1246Y. Out of 121 microscopies positive P. falciparum cases, 109 samples were positive for P. falciparum by nested PCR. Pfcrt K76T mutation was found in 96% of isolates, Pfmdr1 N86Y mutation was observed in 20%, and 11% harboured Y184F mutation. All samples were wild type for Pfmdr1 codon N1042D and D1246Y. In the FATA, Pakistan, the frequency of resistant allele 76T remained high despite the removal of CQ. However, current findings of the study suggest complete fixation of P. falciparum CQ-resistant genotype in the study area.

2022 ◽  
Vol 8 ◽  
Marion Louvois ◽  
Loïc Simon ◽  
Christelle Pomares ◽  
Pierre-Yves Jeandel ◽  
Elisa Demonchy ◽  

Malaria is still an endemic disease in Africa, with many imported cases in Europe. The standard treatment is intravenous artesunate for severe malaria and oral artemisinin-based combination therapy (ACT) for uncomplicated malaria. Delayed hemolytic anemia (DHA) after intravenous artesunate has been extensively described, and guidelines recommend biological monitoring until 1 month after the end of the treatment. A link with an autoimmune process is still unsure. Nevertheless, cases with positive direct antiglobulin test (DAT) have been reported. Conversely, DHA is not recognized as an adverse effect of oral ACT. Previously, only few cases of DHA occurring after oral ACT without intravenous artesunate administration have been reported. We report the case of a 42-year-old man returning from Togo. He was treated with dihydroartemisinin/piperaquine combination for uncomplicated Plasmodium falciparum malaria, with low parasitemia. Nine days after the end of the treatment, the patient developed hemolytic anemia with positive DAT. Eventually, the patient recovered after corticotherapy. After excluding common causes of autoimmune hemolytic anemia, we considered that dihydroartemisinin/piperaquine treatment was involved in this side effect.

2022 ◽  
Vol 21 (1) ◽  
Godfrey Manirakiza ◽  
Kennedy Kassaza ◽  
Ivan Mugisha Taremwa ◽  
Joel Bazira ◽  
Fredrick Byarugaba

Abstract Background The evolution of malaria infection has necessitated the development of highly sensitive diagnostic assays, as well as the use of dried blood spots (DBS) as a potential source of deoxyribonucleic acid (DNA) yield for polymerase chain reaction (PCR) assays. This study identified the different Plasmodium species in malaria-positive patients, and the anti-malarial drug resistance profile for Plasmodium falciparum using DBS samples collected from patients attending Kisoro Hospital in Kisoro district, Southwestern Uganda. Methods The blood samples were prospectively collected from patients diagnosed with malaria to make DBS, which were then used to extract DNA for real-time PCR and high-resolution melting (HRM) analysis. Plasmodium species were identified by comparing the control and test samples using HRM-PCR derivative curves. Plasmodium falciparum chloroquine (CQ) resistance transporter (pfcrt) and kelch13 to screen the samples for anti-malarial resistance markers. The HRM-PCR derivative curve was used to present a summary distribution of the different Plasmodium species as well as the anti-malarial drug profile. Results Of the 152 participants sampled, 98 (64.5%) were females. The average age of the participants was 34.9 years (range: 2 months–81 years). There were 134 samples that showed PCR amplification, confirming the species as Plasmodium. Plasmodium falciparum (N = 122), Plasmodium malariae (N = 6), Plasmodium ovale (N = 4), and Plasmodium vivax (N = 2) were the various Plasmodium species and their proportions. The results showed that 87 (71.3%) of the samples were sensitive strains/wild type (CVMNK), 4 (3.3%) were resistant haplotypes (SVMNT), and 31 (25.4%) were resistant haplotypes (CVIET). Kelch13 C580Y mutation was not detected. Conclusion The community served by Kisoro hospital has a high Plasmodium species burden, according to this study. Plasmodium falciparum was the dominant species, and it has shown that resistance to chloroquine is decreasing in the region. Based on this, molecular identification of Plasmodium species is critical for better clinical management. Besides, DBS is an appropriate medium for DNA preservation and storage for future epidemiological studies.

2022 ◽  
Viola Introini ◽  
Alejandro Marin-Menendez ◽  
Guilherme Nettesheim ◽  
Yen-Chun Lin ◽  
Silvia N Kariuki ◽  

Malaria parasites such as Plasmodium falciparum have exerted formidable selective pressures on the human genome. Of the human genetic variants associated with malaria protection, beta thalassaemia (a haemoglobinopathy) was the earliest to be associated with malaria prevalence. However, the malaria protective properties of beta thalassaemic erythrocytes remain unclear. Here we studied the mechanics and surface protein expression of beta thalassaemia heterozygous erythrocytes, measured their susceptibility to P. falciparum invasion, and calculated the energy required for merozoites to invade them. We found invasion-relevant differences in beta thalassaemic cells versus matched controls, specifically: elevated membrane tension, reduced bending modulus, and higher levels of expression of the major invasion receptor basigin. However, these differences acted in opposition to each other with respect to their likely impact on invasion, and overall we did not observe beta thalassaemic cells to have lower P. falciparum invasion efficiency for any of the strains tested.

Laís Pessanha de Carvalho ◽  
Sara Groeger-Otero ◽  
Andrea Kreidenweiss ◽  
Peter G. Kremsner ◽  
Benjamin Mordmüller ◽  

Boromycin is a boron-containing macrolide antibiotic produced by Streptomyces antibioticus with potent activity against certain viruses, Gram-positive bacteria and protozoan parasites. Most antimalarial antibiotics affect plasmodial organelles of prokaryotic origin and have a relatively slow onset of action. They are used for malaria prophylaxis and for the treatment of malaria when combined to a fast-acting drug. Despite the success of artemisinin combination therapies, the current gold standard treatment, new alternatives are constantly needed due to the ability of malaria parasites to become resistant to almost all drugs that are in heavy clinical use. In vitro antiplasmodial activity screens of tetracyclines (omadacycline, sarecycline, methacycline, demeclocycline, lymecycline, meclocycline), macrolides (oleandomycin, boromycin, josamycin, troleandomycin), and control drugs (chloroquine, clindamycin, doxycycline, minocycline, eravacycline) revealed boromycin as highly potent against Plasmodium falciparum and the zoonotic Plasmodium knowlesi. In contrast to tetracyclines, boromycin rapidly killed asexual stages of both Plasmodium species already at low concentrations (~ 1 nM) including multidrug resistant P. falciparum strains (Dd2, K1, 7G8). In addition, boromycin was active against P. falciparum stage V gametocytes at a low nanomolar range (IC50: 8.5 ± 3.6 nM). Assessment of the mode of action excluded the apicoplast as the main target. Although there was an ionophoric activity on potassium channels, the effect was too low to explain the drug´s antiplasmodial activity. Boromycin is a promising antimalarial candidate with activity against multiple life cycle stages of the parasite.

PLoS ONE ◽  
2022 ◽  
Vol 17 (1) ◽  
pp. e0262616
Swarnali Louha ◽  
Camelia Herman ◽  
Mansi Gupta ◽  
Dhruviben Patel ◽  
Julia Kelley ◽  

Sequencing large numbers of individual samples is often needed for countrywide antimalarial drug resistance surveillance. Pooling DNA from several individual samples is an alternative cost and time saving approach for providing allele frequency (AF) estimates at a population level. Using 100 individual patient DNA samples of dried blood spots from a 2017 nationwide drug resistance surveillance study in Haiti, we compared codon coverage of drug resistance-conferring mutations in four Plasmodium falciparum genes (crt, dhps, dhfr, and mdr1), for the same deep sequenced samples run individually and pooled. Samples with similar real-time PCR cycle threshold (Ct) values (+/- 1.0 Ct value) were combined with ten samples per pool. The sequencing success for samples in pools were higher at a lower parasite density than the individual samples sequence method. The median codon coverage for drug resistance-associated mutations in all four genes were greater than 3-fold higher in the pooled samples than in individual samples. The overall codon coverage distribution for pooled samples was wider than the individual samples. The sample pools with < 40 parasites/μL blood showed more discordance in AF calls for dhfr and mdr1 between the individual and pooled samples. This discordance in AF estimation may be due to low amounts of parasite DNA, which could lead to variable PCR amplification efficiencies. Grouping samples with an estimated ≥ 40 parasites/μL blood prior to pooling and deep sequencing yielded the expected population level AF. Pooling DNA samples based on estimates of > 40 parasites/μL prior to deep sequencing can be used for rapid genotyping of a large number of samples for these four genes and possibly other drug resistant markers in population-based studies. As Haiti is a low malaria transmission country with very few mixed infections and continued chloroquine sensitivity, the pooled sequencing approach can be used for routine national molecular surveillance of resistant parasites.

2022 ◽  
Samuel Pazicky ◽  
Arne Alder ◽  
Haydyn Mertens ◽  
Dmitri I. Svergun ◽  
Tim Gilberger ◽  

As the decline of malaria cases stalled over the last five years, novel targets in Plasmodium falciparum are necessary for the development of new drugs. Glycogen Synthase Kinase (PfGSK3) has been identified as a potential target, since its selective inhibitors were shown to disrupt the parasite's life cycle. In the uncanonical N‑terminal region of the parasite enzyme, we identified several autophosphorylation sites and probed their role in activity regulation of PfGSK3. By combining molecular modeling with experimental small-angle X-ray scattering data, we show that increased PfGSK3 activity is promoted by conformational changes in the PfGSK3 N‑terminus, triggered by N‑terminal phosphorylation. Our work provides novel insights into the structure and regulation of the malarial PfGSK3.

2022 ◽  
Miguel Silva ◽  
Carla Calçada ◽  
Nuno Osório ◽  
Vitória Baptista ◽  
Vandana Thathy ◽  

Abstract Adenosine triphosphate (ATP)-binding cassette (ABC) transporters play an important role in mediating solute or drug transport across cellular membranes. Although this class of transporters has been well characterized in diverse organisms little is known about the physiological roles in Plasmodium falciparum, the deadliest malaria parasite species. We studied the Plasmodium falciparum Multidrug Resistance-associated Protein 1 (PfMRP1; PF3D7_0112200), an ABC transporter localized to the parasite plasma membrane, generating genetic disrupted parasites. We demonstrate that parasites with disrupted pfmrp1 are resistant to folate analogs, methotrexate and aminopterin, with antimalarial activity. This phenotype occurs due to reduction in compound accumulation in the parasite cytoplasm. Phylogenetic analysis supports pfmrp1 being distantly related to ABC transporters in other eukaryotes, suggesting an unusual function. We propose that PfMRP1 can act as a solute importer, a function not previously observed in this organism.

Rebecca C. S. Edgar ◽  
Natalie A. Counihan ◽  
Sheena McGowan ◽  
Tania F. de Koning-Ward

Plasmodium falciparum malaria remains a global health problem as parasites continue to develop resistance to all antimalarials in use. Infection causes clinical symptoms during the intra-erythrocytic stage of the lifecycle where the parasite infects and replicates within red blood cells (RBC). During this stage, P. falciparum digests the main constituent of the RBC, hemoglobin, in a specialized acidic compartment termed the digestive vacuole (DV), a process essential for survival. Many therapeutics in use target one or multiple aspects of the DV, with chloroquine and its derivatives, as well as artemisinin, having mechanisms of action within this organelle. In order to better understand how current therapeutics and those under development target DV processes, techniques used to investigate the DV are paramount. This review outlines the involvement of the DV in therapeutics currently in use and focuses on the range of techniques that are currently utilized to study this organelle including microscopy, biochemical analysis, genetic approaches and metabolomic studies. Importantly, continued development and application of these techniques will aid in our understanding of the DV and in the development of new therapeutics or therapeutic partners for the future.

PLoS Biology ◽  
2022 ◽  
Vol 20 (1) ◽  
pp. e3001515
Maria L. Simões ◽  
Yuemei Dong ◽  
Godfree Mlambo ◽  
George Dimopoulos

Anopheles gambiae melanization-based refractoriness to the human malaria parasite Plasmodium falciparum has rarely been observed in either laboratory or natural conditions, in contrast to the rodent model malaria parasite Plasmodium berghei that can become completely melanized by a TEP1 complement-like system-dependent mechanism. Multiple studies have shown that the rodent parasite evades this defense by recruiting the C-type lectins CTL4 and CTLMA2, while permissiveness to the human malaria parasite was not affected by partial depletion of these factors by RNAi silencing. Using CRISPR/Cas9-based CTL4 knockout, we show that A. gambiae can mount melanization-based refractoriness to the human malaria parasite, which is independent of the TEP1 complement-like system and the major anti-Plasmodium immune pathway Imd. Our study indicates a hierarchical specificity in the control of Plasmodium melanization and proves CTL4 as an essential host factor for P. falciparum transmission and one of the most potent mosquito-encoded malaria transmission-blocking targets.

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