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International Journal of Antimicrobial... Dec 2023The clinical use of artemisinin-based combination therapies is threatened by increasing failure rates due to the emergence and spread of multiple drug resistance genes...
BACKGROUND
The clinical use of artemisinin-based combination therapies is threatened by increasing failure rates due to the emergence and spread of multiple drug resistance genes in most human Plasmodium strains. The aim of this study was to generate artemether-resistant (AM) parasites from Plasmodium berghei ANKA (AM), and determine their fitness cost.
METHODS
Artemether resistance was generated by increasing drug pressure doses gradually for 9 months. Effective doses (ED and ED) were determined using the 4-day suppressive test, and the indices of resistance (I) at 50% and 90% (I and I) were determined using the ratio of either ED or ED of AM to AM, respectively. The stability of the AM parasites was evaluated by: five drug-free passages (5DFPs), 3 months of cryopreservation (CP), and drug-free serial passages (DFSPs) for 4 months. Analysis of variance was used to compare differences in growth rates between AM and AM with 95% confidence intervals.
RESULTS
ED and ED of AM were 0.61 and 3.43 mg/kg/day respectively. I and I after 20 cycles of artemether selection pressure were 19.67 and 21.45, respectively; 5DFP values were 39.16 and 15.27, respectively; 3-month CP values were 29.36 and 10.79, respectively; and DFSP values were 31.34 and 12.29, respectively. The mean parasitaemia value of AM (24.70% ± 3.60) relative to AM (37.66% ± 3.68) at Day 7 post infection after DFSPs revealed a fitness cost of 34.41%.
CONCLUSION
A moderately stable AMP. berghei line was generated. Known and unknown mutations may be involved in modulating artemether resistance, and therefore molecular investigations are recommended.
Topics: Animals; Humans; Artemether; Antimalarials; Malaria; Parasites; Plasmodium berghei; Plasmodium falciparum; Drug Resistance
PubMed: 37865152
DOI: 10.1016/j.ijantimicag.2023.107012 -
The FEBS Journal Jan 2021The Apicomplexa phylum groups important human and animal pathogens that cause severe diseases, encompassing malaria, toxoplasmosis, and cryptosporidiosis. In common with... (Review)
Review
The Apicomplexa phylum groups important human and animal pathogens that cause severe diseases, encompassing malaria, toxoplasmosis, and cryptosporidiosis. In common with most organisms, apicomplexans rely on heme as cofactor for several enzymes, including cytochromes of the electron transport chain. This heme derives from de novo synthesis and/or the development of uptake mechanisms to scavenge heme from their host. Recent studies have revealed that heme synthesis is essential for Toxoplasma gondii tachyzoites, as well as for the mosquito and liver stages of Plasmodium spp. In contrast, the erythrocytic stages of the malaria parasites rely on scavenging heme from the host red blood cell. The unusual heme synthesis pathway in Apicomplexa spans three cellular compartments and comprises enzymes of distinct ancestral origin, providing promising drug targets. Remarkably given the requirement for heme, T. gondii can tolerate the loss of several heme synthesis enzymes at a high fitness cost, while the ferrochelatase is essential for survival. These findings indicate that T. gondii is capable of salvaging heme precursors from its host. Furthermore, heme is implicated in the activation of the key antimalarial drug artemisinin. Recent findings established that a reduction in heme availability corresponds to decreased sensitivity to artemisinin in T. gondii and Plasmodium falciparum, providing insights into the possible development of combination therapies to tackle apicomplexan parasites. This review describes the microeconomics of heme in Apicomplexa, from supply, either from de novo synthesis or scavenging, to demand by metabolic pathways, including the electron transport chain.
Topics: Animals; Anti-Infective Agents; Artemisinins; Cryptosporidium; Cytochromes; Erythrocytes; Ferrochelatase; Gene Expression; Heme; Host-Pathogen Interactions; Humans; Life Cycle Stages; Metabolic Networks and Pathways; Plasmodium berghei; Plasmodium falciparum; Protozoan Proteins; Toxoplasma
PubMed: 32530125
DOI: 10.1111/febs.15445 -
Vaccine Jan 2023Antigens expressed during the sexual development of malaria parasites are transmission-blocking vaccine (TBV) targets. Pb22, a protein expressed and localized to the...
Antigens expressed during the sexual development of malaria parasites are transmission-blocking vaccine (TBV) targets. Pb22, a protein expressed and localized to the plasma membrane of gametes and ookinetes in Plasmodium berghei, is an excellent TBV candidate. Here, we evaluated the TB potential of the Plasmodium vivax ortholog Pv22 using a transgenic P. berghei parasite line and P. vivax clinical isolates. The full-length recombinant Pv22 (rPv22) protein was produced and used to immunize mice and rabbits to obtain antibodies. We generated a transgenic P. berghei line (TrPv22Pb) by inserting the pv22 gene into the pb22 locus and showed that Pv22 expression completely rescued the defects in male gametogenesis of the pb22 deletion parasite. Since Pv22 in the transgenic parasite showed similar expression and localization patterns to Pb22, we used the TrPv22Pb parasite as a surrogate to evaluate the TB potential of Pv22. In mosquito feeding assays, mosquitoes feeding on rPv22-immunized mice infected with TrPv22Pb parasites showed a 49.3-53.3 % reduction in the oocyst density compared to the control group. In vitro assays showed that the rPv22 immune sera significantly inhibited exflagellation and ookinete formation of the TrPv22Pb parasites. In a direct membrane feeding assay using three clinical P. vivax isolates, the rabbit anti-rPv22 antibodies also significantly decreased the oocyst density by 53.7, 30.2, and 26.2 %, respectively. This study demonstrated the feasibility of using transgenic P. berghei parasites expressing P. vivax antigens as a potential tool to evaluate TBV candidates. However, the much weaker TB activity of Pv22 obtained from two complementary assays suggest that Pv22 may not be a promising TBV candidate for P. vivax.
Topics: Male; Animals; Mice; Rabbits; Malaria; Plasmodium vivax; Plasmodium berghei; Malaria Vaccines; Protozoan Proteins; Malaria, Vivax; Recombinant Proteins; Antibodies, Protozoan; Culicidae
PubMed: 36503858
DOI: 10.1016/j.vaccine.2022.11.058 -
Journal of Global Antimicrobial... Mar 2020The aim of this study was to evaluate the efficacy of pyrimethamine-loaded poloxamer 407 nanomicelles on Plasmodium berghei strain NICD in vivo.
OBJECTIVES
The aim of this study was to evaluate the efficacy of pyrimethamine-loaded poloxamer 407 nanomicelles on Plasmodium berghei strain NICD in vivo.
METHODS
Pyrimethamine-loaded nanomicelles were prepared and their zeta potential, particle size and polydispersity index were measured. For antiplasmodial assessment, 54 mice were randomly divided into six groups. Four groups were infected intraperitoneally with P. berghei, whereas the two remaining groups did not receive the parasite (negative controls). Three of the P. berghei-infected groups received treatment with either pyrimethamine-loaded nanomicelles (2 mg/kg), pyrimethamine (2 mg/kg) or empty nanomicelles (2 mg/kg); the fourth group remained untreated (positive control). The parasitaemia rate, survival rate and histopathological changes in the liver, spleen and kidneys were examined and were compared with the negative and positive control groups.
RESULTS
The mean parasitaemia rate differed significantly between the nanoformulated pyrimethamine group and each of the other groups (P<0.05). Moreover, the survival rate of mice in the nanoformulated pyrimethamine group (7/9; 78%) was significantly higher compared with each of the other groups (P<0.01). The main histopathological changes, including hepatic necrosis in the liver, lymphoid hypoplasia in the spleen, and tubular nephrosis and perivascular and interstitial lymphocytic infiltration in the kidneys, were considerably lower in the nanoformulated pyrimethamine group than in the pyrimethamine and positive control groups.
CONCLUSION
Pyrimethamine-loaded nanomicelles showed potent antimalarial activity and can be considered as a potential candidate for further examination of their suitability as an antimalarial drug.
Topics: Animals; Antimalarials; Disease Models, Animal; Drug Compounding; Liver; Malaria; Male; Mice; Micelles; Nanoparticles; Parasitemia; Particle Size; Plasmodium berghei; Poloxamer; Pyrimethamine; Random Allocation; Spleen; Survival Analysis; Treatment Outcome
PubMed: 31404680
DOI: 10.1016/j.jgar.2019.08.002 -
MBio Feb 2023Mature gametocytes of Plasmodium falciparum display a banana (falciform) shape conferred by a complex array of subpellicular microtubules (SPMT) associated with the...
Mature gametocytes of Plasmodium falciparum display a banana (falciform) shape conferred by a complex array of subpellicular microtubules (SPMT) associated with the inner membrane complex (IMC). Microtubule-associated proteins (MAPs) define MT populations and modulate interaction with pellicular components. Several MAPs have been identified in Toxoplasma gondii, and homologues can be found in the genomes of species, but the function of these proteins for asexual and sexual development of malaria parasites is still unknown. Here, we identified a novel subpellicular MAP, termed SPM3, that is conserved within the genus , especially within the subgenus , but absent in other Apicomplexa. Conditional knockdown and targeted gene disruption of in Plasmodium falciparum cause severe morphological defects during gametocytogenesis, leading to round, nonfalciform gametocytes with an aberrant SPMT pattern. In contrast, knockout in Plasmodium berghei, a species with round gametocytes, caused no defect in gametocytogenesis, but sporozoites displayed an aberrant motility and a dramatic defect in invasion of salivary glands, leading to a decreased efficiency in transmission. Electron microscopy revealed a dissociation of the SPMT from the IMC in knockout parasites, suggesting a function of SPM3 in anchoring MTs to the IMC. Overall, our results highlight SPM3 as a pellicular component with essential functions for malaria parasite transmission. A key structural feature driving the transition between different life cycle stages of the malaria parasite is the unique three-membrane pellicle, consisting of the parasite plasma membrane (PPM) and a double membrane structure underlying the PPM termed the inner membrane complex (IMC). Additionally, there are numerous linearly arranged intramembranous particles (IMPs) linked to the IMC, which likely link the IMC to the subpellicular microtubule cytoskeleton. Here, we identified, localized, and characterized a novel subpellicular microtubule-associated protein unique to the genus . The knockout of this protein in the human-pathogenic species P. falciparum resulted in malformed gametocytes and aberrant microtubules. We confirmed the microtubule association in the P. berghei rodent malaria homologue and show that its knockout results in a perturbed microtubule architecture, aberrant sporozoite motility, and decreased transmission efficiency.
Topics: Animals; Humans; Parasites; Microtubule-Associated Proteins; Plasmodium falciparum; Malaria; Plasmodium berghei; Sporozoites; Protozoan Proteins
PubMed: 36625655
DOI: 10.1128/mbio.03318-22 -
Experimental Parasitology May 2021The quest for the development of a novel antimalarial drug informed the decision to subject phytol to in vivo trials following a demonstration of therapeutic potential...
The quest for the development of a novel antimalarial drug informed the decision to subject phytol to in vivo trials following a demonstration of therapeutic potential against chloroquine sensitive strain of Plasmodium falciparum under in vitro condition. On this basis, the in vivo anti-Plasmodium berghei activity of phytol including the ameliorative effects of the compound on P. berghei-associated anaemia and organ damage were investigated. Mice were infected with chloroquine-sensitive strain of P. berghei and were treated with phytol at a dose of 10 and 20 mg/kg body weight (BW) for four days. The levels of parasitemia, packed cell volume and redox sensitive biomarkers of liver, brain and spleen tissues were determined. Our result revealed that phytol significantly (p < 0.05) suppressed the multiplication of P. berghei in a dose-dependent manner. Additionally, the phytol significantly (p < 0.05) ameliorated the P. berghei-induced anaemia and brain damage. Data from the present study demonstrated that phytol has suppressive effect on P. berghei and could ameliorate some P. berghei-induced pathological changes.
Topics: Analysis of Variance; Anemia; Animals; Antimalarials; Brain; Chloroquine; Dose-Response Relationship, Drug; Female; Hematocrit; Liver; Malaria; Male; Mice; Oxidation-Reduction; Parasitemia; Phytol; Plasmodium berghei; Random Allocation; Spleen
PubMed: 33736972
DOI: 10.1016/j.exppara.2021.108097 -
Infectious Disorders Drug Targets 2022Antimalarial prescription remains a challenge in pregnant women because of maternal and fetal complications. Recently, garlic and α-β-arteether combination treatment...
BACKGROUND
Antimalarial prescription remains a challenge in pregnant women because of maternal and fetal complications. Recently, garlic and α-β-arteether combination treatment in malariainfected mice conferred protection. The purpose of this study is to evaluate the efficacy of these drugs during malaria in pregnancy and its safety measures.
OBJECTIVE
The study evaluates the efficacy of arteether and garlic combination drugs in protection against malaria-infected pregnant mice.
METHODS
Plasmodium berghei-infected pregnant mouse model was used to assess the combination drug efficacy and the outcome of abnormalities of the disease after drug treatment. After optimizing the dose and gestation period, maternal protection was confirmed by parasite clearance in smear and mortality observation. In addition, maternal hematological parameters, different organ histopathology, and IgG levels were documented along with the fetal and infant outcomes.
RESULTS
Arteether monotherapy resulted in spontaneous fetal abortion or resorption, while dosage optimization and garlic combination resulted in pregnancy completion and malaria protection. The derangements observed in the histoarchitecture of organs and hematological parameters caused by malaria infection revealed improvement after drug treatment, and the smear observation confirms the clearance of malaria parasite in the peripheral blood, but IgG level was maintained at the same higher level as in malaria-infected mice.
CONCLUSION
The first report of an arteether and garlic combination demonstrating high efficacy in protecting against malaria-infected pregnant mice establishes its safety as a viable possible treatment for pregnancy-associated malaria.
Topics: Animals; Antimalarials; Artemisinins; Female; Garlic; Humans; Immunoglobulin G; Malaria; Mice; Plasmodium berghei; Pregnancy
PubMed: 35016599
DOI: 10.2174/1871526522666220110163945 -
Parasite Immunology Nov 2020Merozoite surface protein 8 (MSP-8) of Plasmodium parasites plays an important role in erythrocyte invasion and is a potential malaria vaccine candidate.
AIMS
Merozoite surface protein 8 (MSP-8) of Plasmodium parasites plays an important role in erythrocyte invasion and is a potential malaria vaccine candidate.
METHODS AND RESULTS
In this study, virus-like particles (VLPs) expressing MSP-8 of Plasmodium berghei on the surface of influenza virus matrix protein 1 (M1) core protein were generated for vaccine efficacy assessment. Mice were intramuscularly (IM) immunized with MSP-8 VLPs twice and challenge-infected with P. berghei. We found that VLP vaccination elicited higher levels of P. berghei-specific IgG antibody response in the sera, along with blood CD4 and CD8 T-cell response enhancement compared to the naïve control mice. CD4 and CD8 effector memory T-cell and memory B-cell responses in the spleen were found to be higher in VLP-immunized mice compared to control mice. VLP vaccination significantly reduced inflammatory cytokine (IFN-γ) response in the spleen and parasitemia levels in blood compared to naïve control mice.
CONCLUSIONS
These results indicate that MSP-8 containing virus-like particles could be a vaccine candidate for blood-stage vaccine design.
Topics: Animals; Antigens, Protozoan; Female; Immunization; Malaria; Malaria Vaccines; Mice; Mice, Inbred BALB C; Parasitemia; Plasmodium berghei; Protozoan Proteins
PubMed: 32738150
DOI: 10.1111/pim.12781 -
Open Biology Aug 2022Protein phosphatase 1 (PP1) is a key enzyme for development. However, the detailed mechanisms underlying its regulation remain to be deciphered. Here, we report the...
Protein phosphatase 1 (PP1) is a key enzyme for development. However, the detailed mechanisms underlying its regulation remain to be deciphered. Here, we report the functional characterization of the leucine-rich repeat protein 1 (PbLRR1), an orthologue of SDS22, one of the most ancient and conserved PP1 interactors. Our study shows that PbLRR1 is expressed during intra-erythrocytic development of the parasite, and up to the zygote stage in mosquitoes. PbLRR1 can be found in complex with PbPP1 in both asexual and sexual stages and inhibits its phosphatase activity. Genetic analysis demonstrates that PbLRR1 depletion adversely affects the development of oocysts. PbLRR1 interactome analysis associated with phospho-proteomics studies identifies several novel putative PbLRR1/PbPP1 partners. Some of these partners have previously been characterized as essential for the parasite sexual development. Interestingly, and for the first time, Inhibitor 3 (I3), a well-known and direct interactant of PP1, was found to be drastically hypophosphorylated in PbLRR1-depleted parasites. These data, along with the detection of I3 with PP1 in the LRR1 interactome, strongly suggest that the phosphorylation status of PbI3 is under the control of the PP1-LRR1 complex and could contribute (in)directly to oocyst development. This study provides new insights into previously unrecognized PbPP1 fine regulation of oocyst development through its interaction with PbLRR1.
Topics: Animals; Leucine-Rich Repeat Proteins; Oocysts; Phosphorylation; Plasmodium berghei; Protein Phosphatase 1
PubMed: 35920043
DOI: 10.1098/rsob.220015 -
Journal of Proteomics Sep 2020Passage of malaria parasites through mosquitoes involves multiple developmental transitions, from gametocytes that are ingested with the blood meal, through to...
Passage of malaria parasites through mosquitoes involves multiple developmental transitions, from gametocytes that are ingested with the blood meal, through to sporozoites that are transmitted by insect bite to the host. During the transformation from gametocyte to oocyst, the parasite forms a unique transient organelle named the crystalloid, which is involved in sporozoite formation. In Plasmodium berghei, a complex of six LCCL domain-containing proteins (LAPs) reside in the crystalloid and are required for its biogenesis. However, little else is known about the molecular mechanisms that underlie the crystalloid's role in sporogony. In this study, we have used transgenic parasites stably expressing LAP3 fused to GFP, combined with GFP affinity pulldown and high accuracy mass spectrometry, to identify an extended LAP interactome of some fifty proteins. We show that many of these are targeted to the crystalloid, including members of two protein families with CPW-WPC and pleckstrin homology-like domains, respectively. Our findings indicate that the LAPs are part of an intricate protein complex, the formation of which facilitates both crystalloid targeting and biogenesis. SIGNIFICANCE: Reducing malaria parasite transmission by mosquitoes is a key component of malaria eradication and control strategies. This study sheds important new light on the molecular composition of the crystalloid, an enigmatic parasite organelle that is essential for sporozoite formation and transmission from the insect to the vertebrate host. Our findings provide new mechanistic insight into how proteins are delivered to the crystalloid, and indicate that the molecular mechanisms that underlie crystalloid function are complex, involving several protein families unique to Plasmodium and closely related organisms. The new crystalloid proteins identified will form a useful starting point for studies aimed at unravelling how the crystalloid organelle facilitates sporogony and transmission.
Topics: Animals; Crystalloid Solutions; Humans; Malaria; Organelles; Plasmodium berghei; Protozoan Proteins
PubMed: 32736136
DOI: 10.1016/j.jprot.2020.103925