scholarly journals Functional Analysis of the Exported Type IV HSP40 Protein PfGECO in Plasmodium falciparum Gametocytes

2011 ◽  
Vol 10 (11) ◽  
pp. 1492-1503 ◽  
Author(s):  
Belinda J. Morahan ◽  
Carolyn Strobel ◽  
Uzma Hasan ◽  
Beata Czesny ◽  
Pierre-Yves Mantel ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Heat Shock ◽  
Protective Role ◽  
Targeted Disruption ◽  
Content Type ◽  
Type Iv ◽  
Rbc Membrane ◽  
Host Defense Response ◽  
Hsp40 Protein

ABSTRACTDuringPlasmodium falciparuminfection, host red blood cell (RBC) remodeling is required for the parasite's survival. Such modifications are mediated by the export of parasite proteins into the RBC that alter the architecture of the RBC membrane and enable cytoadherence. It is probable that some exported proteins also play a protective role against the host defense response. This may be of particular importance for the gametocyte stage of the life cycle that is responsible for malaria transmission, since the gametocyte remains in contact with blood as it proceeds through five morphological stages (I to V) during its 12-day maturation. Using microarray analysis, we identified several genes with encoded secretory or export sequences that were differentially expressed during early gametocytogenesis. One of these,PfGECO, encodes a predicted type IV heat shock protein 40 (HSP40) that we show is expressed in gametocyte stages I to IV and is exported to the RBC cytoplasm. HSPs are traditionally induced under stressful conditions to maintain homeostasis, butPfGECOexpression was not increased upon heat shock, suggesting an alternate function. Targeted disruption ofPfGECOindicated that the gene is not essential for gametocytogenesisin vitro, and quantitative reverse transcriptase PCR (RT-PCR) showed that there was no compensatory expression of the other type IV HSP40 genes. AlthoughP. falciparumHSP40 members are implicated in the trafficking of proteins to the RBC surface, removal of PfGECO did not affect the targeting of other exported gametocyte proteins. This work has expanded the repertoire of known gametocyte-exported proteins to include a type IV HSP40, PfGECO.

scholarly journals Effect of Fluorescent Dyes onIn Vitro-Differentiated, Late-Stage Plasmodium falciparum Gametocytes

2014 ◽  
Vol 58 (12) ◽  
pp. 7398-7404 ◽  
Author(s):  
Tamirat Gebru ◽  
Benjamin Mordmüller ◽  
Jana Held
Keyword(s):  
Plasmodium Falciparum ◽  
Late Stage ◽  
Fluorescent Dyes ◽  
Clinical Symptoms ◽  
Laboratory Strain ◽  
Clinical Isolates ◽  
Synthetic Dyes ◽  
Content Type ◽  
Highly Active

ABSTRACTPlasmodium falciparumgametocytes are not associated with clinical symptoms, but they are responsible for transmitting the pathogen to mosquitoes. Therefore, gametocytocidal interventions are important for malaria control and resistance containment. Currently available drugs and vaccines are not well suited for that purpose. Several dyes have potent antimicrobial activity, but their use against gametocytes has not been investigated systematically. The gametocytocidal activity of nine synthetic dyes and four control compounds was tested against stage V gametocytes of the laboratory strain 3D7 and three clinical isolates ofP. falciparumwith a bioluminescence assay. Five of the fluorescent dyes had submicromolar 50% inhibitory concentration (IC50) values against mature gametocytes. Three mitochondrial dyes, MitoRed, dihexyloxacarbocyanine iodide (DiOC6), and rhodamine B, were highly active (IC50s < 200 nM). MitoRed showed the highest activity against gametocytes, with IC50s of 70 nM against 3D7 and 120 to 210 nM against clinical isolates. All compounds were more active against the laboratory strain 3D7 than against clinical isolates. In particular, the endoperoxides artesunate and dihydroartemisinin showed a 10-fold higher activity against 3D7 than against clinical isolates. In contrast to all clinically used antimalarials, several fluorescent dyes had surprisingly highin vitroactivity against late-stage gametocytes. Since they also act against asexual blood stages, they shall be considered starting points for the development of new antimalarial lead compounds.

scholarly journals Plasmodium falciparum Line-Dependent Association ofIn VitroGrowth-Inhibitory Activity and Risk of Malaria

2012 ◽  
Vol 80 (5) ◽  
pp. 1900-1908 ◽  
Author(s):  
Josea Rono ◽  
Anna Färnert ◽  
Daniel Olsson ◽  
Faith Osier ◽  
Ingegerd Rooth ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Inhibitory Activity ◽  
Content Type ◽  
Phenotypic Differences ◽  
Erythrocyte Invasion ◽  
Growth Inhibitory ◽  
Growth Inhibitory Activity ◽  
Control Study

ABSTRACTPlasmodium falciparum's ability to invade erythrocytes is essential for its survival within the human host. Immune mechanisms that impair this ability are therefore expected to contribute to immunity against the parasite. Plasma of humans who are naturally exposed to malaria has been shown to have growth-inhibitory activity (GIA)in vitro. However, the importance of GIA in relation to protection from malaria has been unclear. In a case-control study nested within a longitudinally followed population in Tanzania, plasma samples collected at baseline from 171 individuals (55 cases and 116 age-matched controls) were assayed for GIA using threeP. falciparumlines (3D7, K1, and W2mef) chosen based on their erythrocyte invasion phenotypes. Distribution of GIA differed between the lines, with most samples inhibiting the growth of 3D7 and K1 and enhancing the growth of W2mef. GIA to 3D7 was associated with a reduced risk of malaria within 40 weeks of follow-up (odds ratio, 0.45; 95% confidence interval [CI], 0.21 to 0.96;P= 0.04), whereas GIA to K1 and W2mef was not. These results show that GIA, as well as its association with protection from malaria, is dependent on theP. falciparumline and can be explained by differences in erythrocyte invasion phenotypes between parasite lines. Our study contributes knowledge on the biological importance of growth inhibition and the potential influence ofP. falciparumerythrocyte invasion phenotypic differences on its relationship to protective immunity against malaria.

scholarly journals A Chemical Rescue Screen Identifies a Plasmodium falciparum Apicoplast Inhibitor Targeting MEP Isoprenoid Precursor Biosynthesis

2014 ◽  
Vol 59 (1) ◽  
pp. 356-364 ◽  
Author(s):  
Wesley Wu ◽  
Zachary Herrera ◽  
Danny Ebert ◽  
Katie Baska ◽  
Seok H. Cho ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Antimalarial Drug ◽  
Drug Targets ◽  
Specific Activity ◽  
Drug Candidate ◽  
Content Type ◽  
Chemical Rescue ◽  
Growth Inhibitory ◽  
Parasite Populations

ABSTRACTThe apicoplast is an essential plastid organelle found inPlasmodiumparasites which contains several clinically validated antimalarial-drug targets. A chemical rescue screen identified MMV-08138 from the “Malaria Box” library of growth-inhibitory antimalarial compounds as having specific activity against the apicoplast. MMV-08138 inhibition of blood-stagePlasmodium falciparumgrowth is stereospecific and potent, with the most active diastereomer demonstrating a 50% effective concentration (EC50) of 110 nM. Whole-genome sequencing of 3 drug-resistant parasite populations from two independent selections revealed E688Q and L244I mutations inP. falciparumIspD, an enzyme in the MEP (methyl-d-erythritol-4-phosphate) isoprenoid precursor biosynthesis pathway in the apicoplast. The active diastereomer of MMV-08138 directly inhibited PfIspD activityin vitrowith a 50% inhibitory concentration (IC50) of 7.0 nM. MMV-08138 is the first PfIspD inhibitor to be identified and, together with heterologously expressed PfIspD, provides the foundation for further development of this promising antimalarial drug candidate lead. Furthermore, this report validates the use of the apicoplast chemical rescue screen coupled with target elucidation as a discovery tool to identify specific apicoplast-targeting compounds with new mechanisms of action.

scholarly journals Computational Chemogenomics Drug Repositioning Strategy Enables the Discovery of Epirubicin as a New Repurposed Hit for Plasmodium falciparum and P. vivax

2020 ◽  
Vol 64 (9) ◽  
Author(s):  
Letícia Tiburcio Ferreira ◽  
Juliana Rodrigues ◽  
Gustavo Capatti Cassiano ◽  
Tatyana Almeida Tavella ◽  
Kaira Cristina Peralis Tomaz ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
In Silico ◽  
Antimalarial Activity ◽  
Drug Repositioning ◽  
Dna Gyrase ◽  
Plasmodium Yoelii ◽  
Computer Assisted ◽  
Content Type

ABSTRACT Widespread resistance against antimalarial drugs thwarts current efforts for controlling the disease and urges the discovery of new effective treatments. Drug repositioning is increasingly becoming an attractive strategy since it can reduce costs, risks, and time-to-market. Herein, we have used this strategy to identify novel antimalarial hits. We used a comparative in silico chemogenomics approach to select Plasmodium falciparum and Plasmodium vivax proteins as potential drug targets and analyzed them using a computer-assisted drug repositioning pipeline to identify approved drugs with potential antimalarial activity. Among the seven drugs identified as promising antimalarial candidates, the anthracycline epirubicin was selected for further experimental validation. Epirubicin was shown to be potent in vitro against sensitive and multidrug-resistant P. falciparum strains and P. vivax field isolates in the nanomolar range, as well as being effective against an in vivo murine model of Plasmodium yoelii. Transmission-blocking activity was observed for epirubicin in vitro and in vivo. Finally, using yeast-based haploinsufficiency chemical genomic profiling, we aimed to get insights into the mechanism of action of epirubicin. Beyond the target predicted in silico (a DNA gyrase in the apicoplast), functional assays suggested a GlcNac-1-P-transferase (GPT) enzyme as a potential target. Docking calculations predicted the binding mode of epirubicin with DNA gyrase and GPT proteins. Epirubicin is originally an antitumoral agent and presents associated toxicity. However, its antiplasmodial activity against not only P. falciparum but also P. vivax in different stages of the parasite life cycle supports the use of this drug as a scaffold for hit-to-lead optimization in malaria drug discovery.

scholarly journals Ex VivoDrug Susceptibility of Ferroquine against Chloroquine-Resistant Isolates of Plasmodium falciparum and P. vivax

2011 ◽  
Vol 55 (9) ◽  
pp. 4461-4464 ◽  
Author(s):  
Jutta Marfurt ◽  
Ferryanto Chalfein ◽  
Pak Prayoga ◽  
Frans Wabiser ◽  
Enny Kenangalem ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Combination Therapy ◽  
Drug Class ◽  
Ex Vivo ◽  
Multidrug Resistant ◽  
Drug Action ◽  
Drug Uptake ◽  
Content Type ◽  
Excellent Activity

ABSTRACTFerroquine (FQ; SSR97193), a ferrocene-containing 4-aminoquinoline derivate, has potentin vitroefficacy against chloroquine (CQ)-resistantPlasmodium falciparumand CQ-sensitiveP. vivax. In the current study,ex vivoFQ activity was tested in multidrug-resistantP. falciparumandP. vivaxfield isolates using a schizont maturation assay. Although FQ showed excellent activity against CQ-sensitive and -resistantP. falciparumandP. vivax(median 50% inhibitory concentrations [IC50s], 9.6 nM and 18.8 nM, respectively), there was significant cross-susceptibility with the quinoline-based drugs chloroquine, amodiaquine, and piperaquine (forP. falciparum,r= 0.546 to 0.700,P< 0.001; forP. vivax,r= 0.677 to 0.821,P< 0.001). The observedex vivocross-susceptibility is likely to reflect similar mechanisms of drug uptake/efflux and modes of drug action of this drug class. However, the potent activity of FQ against resistant isolates of bothP. falciparumandP. vivaxhighlights a promising role for FQ as a lead antimalarial against CQ-resistantPlasmodiumand a useful partner drug for artemisinin-based combination therapy.

scholarly journals Delayed Parasite Clearance after Treatment with Dihydroartemisinin-Piperaquine in Plasmodium falciparum Malaria Patients in Central Vietnam

2014 ◽  
Vol 58 (12) ◽  
pp. 7049-7055 ◽  
Author(s):  
Kamala Thriemer ◽  
Nguyen Van Hong ◽  
Anna Rosanas-Urgell ◽  
Bui Quang Phuc ◽  
Do Manh Ha ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Clearance Rate ◽  
Artemisinin Resistance ◽  
Parasite Clearance ◽  
Clearance Time ◽  
Sensitivity Testing ◽  
Content Type ◽  
Parasite Clearance Time ◽  
Central Vietnam

ABSTRACTReduced susceptibility ofPlasmodium falciparumtoward artemisinin derivatives has been reported from the Thai-Cambodian and Thai-Myanmar borders. Following increasing reports from central Vietnam of delayed parasite clearance after treatment with dihydroartemisinin-piperaquine (DHA-PPQ), the current first-line treatment, we carried out a study on the efficacy of this treatment. Between September 2012 and February 2013, we conducted a 42-dayin vivoandin vitroefficacy study in Quang Nam Province. Treatment was directly observed, and blood samples were collected twice daily until parasite clearance. In addition, genotyping, quantitative PCR (qPCR), andin vitrosensitivity testing of isolates was performed. The primary endpoints were parasite clearance rate and time. The secondary endpoints included PCR-corrected and uncorrected cure rates, qPCR clearance profiles,in vitrosensitivity results (for chloroquine, dihydroartemisinin, and piperaquine), and genotyping for mutations in the Kelch 13 propeller domain. Out of 672 screened patients, 95 were recruited and 89 available for primary endpoint analyses. The median parasite clearance time (PCT) was 61.7 h (interquartile range [IQR], 47.6 to 83.2 h), and the median parasite clearance rate had a slope half-life of 6.2 h (IQR, 4.4 to 7.5 h). The PCR-corrected efficacy rates were estimated at 100% at day 28 and 97.7% (95% confidence interval, 91.2% to 99.4%) at day 42. At day 3, theP. falciparumprevalence by qPCR was 2.5 times higher than that by microscopy. The 50% inhibitory concentrations (IC50s) of isolates with delayed clearance times (≥72 h) were significantly higher than those with normal clearance times for all three drugs. Delayed parasite clearance (PCT, ≥72 h) was significantly higher among day 0 samples carrying the 543 mutant allele (47.8%) than those carrying the wild-type allele (1.8%;P= 0.048). In central Vietnam, the efficacy of DHA-PPQ is still satisfactory, but the parasite clearance time and rate are indicative of emerging artemisinin resistance. (This study has been registered at ClinicalTrials.gov under registration no. NCT01775592.)

scholarly journals Kelch Mutations in Plasmodium falciparum Protein K13 Do Not Modulate Dormancy after Artemisinin Exposure and Sorbitol Selection In Vitro

2018 ◽  
Vol 62 (5) ◽  
Author(s):  
Kimberly F. Breglio ◽  
Rifat S. Rahman ◽  
Juliana M. SÃ ◽  
Amanda Hott ◽  
David J. Roberts ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Single Site ◽  
Ring Stage ◽  
Content Type ◽  
Isogenic Lines ◽  
Falciparum Protein

ABSTRACT Some Kelch mutations of the Plasmodium falciparum K13 protein confer increased survival to dihydroartemisinin (DHA)-treated ring-stage parasites. Here, we asked if K13 mutations affect a dormancy phenotype allowing parasites to survive DHA exposure and then sorbitol selection. Although recrudescence from dormancy differed between two distinct parasite lines, it was similar for isogenic lines carrying single-site substitutions in K13. Therefore, K13 mutations do not alter the DHA-sorbitol combined dormancy phenotype; rather, traits from other loci likely determine this phenotype.

scholarly journals Activity of Ceftaroline and Epidemiologic Trends in Staphylococcus aureus Isolates Collected from 43 Medical Centers in the United States in 2009

2011 ◽  
Vol 55 (9) ◽  
pp. 4154-4160 ◽  
Author(s):  
Sandra S. Richter ◽  
Kristopher P. Heilmann ◽  
Cassie L. Dohrn ◽  
Fathollah Riahi ◽  
Andrew J. Costello ◽  
...  
Keyword(s):  
United States ◽  
Staphylococcus Aureus ◽  
The United States ◽  
Surveillance Program ◽  
Content Type ◽  
Type Iv ◽  
Medical Centers ◽  
High Level ◽  
Resistance Rates

ABSTRACTAStaphylococcus aureussurveillance program was initiated in the United States to examine thein vitroactivity of ceftaroline and epidemiologic trends. Susceptibility testing by Clinical and Laboratory Standards Institute broth microdilution was performed on 4,210 clinically significant isolates collected in 2009 from 43 medical centers. All isolates were screened formecAby PCR and evaluated by pulsed-field gel electrophoresis. Methicillin-resistantS. aureus(MRSA) were analyzed for Panton-Valentine leukocidin (PVL) genes and the staphylococcal cassette chromosomemec(SCCmec) type. All isolates had ceftaroline MICs of ≤2 μg/ml with an MIC50of 0.5 and an MIC90of 1 μg/ml. The overall resistance rates, expressed as the percentages of isolates that were intermediate and resistant (or nonsusceptible), were as follows: ceftaroline, 1.0%; clindamycin, 30.2% (17.4% MIC ≥ 4 μg/ml; 12.8% inducible); daptomycin, 0.2%; erythromycin, 65.5%; levofloxacin, 39.9%; linezolid, 0.02%; oxacillin, 53.4%; tetracycline, 4.4%; tigecycline, 0%; trimethoprim-sulfamethoxazole, 1.6%; vancomycin, 0%; and high-level mupirocin, 2.2%. ThemecAPCR was positive for 53.4% of the isolates. The ceftaroline MIC90s were 0.25 μg/ml for methicillin-susceptibleS. aureusand 1 μg/ml for MRSA. Among the 2,247 MRSA isolates, 51% were USA300 (96.9% PVL positive, 99.7% SCCmectype IV) and 17% were USA100 (93.4% SCCmectype II). The resistance rates for the 1,137 USA300 MRSA isolates were as follows: erythromycin, 90.9%; levofloxacin, 49.1%; clindamycin, 7.6% (6.2% MIC ≥ 4 μg/ml; 1.4% inducible); tetracycline, 3.3%; trimethoprim-sulfamethoxazole, 0.8%; high-level mupirocin, 2.7%; daptomycin, 0.4%; and ceftaroline and linezolid, 0%. USA300 is the dominant clone causing MRSA infections in the United States. Ceftaroline demonstrated potentin vitroactivity against recentS. aureusclinical isolates, including MRSA, daptomycin-nonsusceptible, and linezolid-resistant strains.

scholarly journals Unexpected Stress-Reducing Effect of PhaP, a Poly(3-Hydroxybutyrate) Granule-Associated Protein, in Escherichia coli

2011 ◽  
Vol 77 (18) ◽  
pp. 6622-6629 ◽  
Author(s):  
Alejandra de Almeida ◽  
Mariela V. Catone ◽  
Virgil A. Rhodius ◽  
Carol A. Gross ◽  
M. Julia Pettinari
Keyword(s):  
Escherichia Coli ◽  
Heat Shock ◽  
Stress Proteins ◽  
Protective Role ◽  
Pcr Analysis ◽  
Content Type ◽  
E Coli ◽  
Protein Levels ◽  
Stress Related Genes ◽  
Heat Stress Proteins

ABSTRACTPhasins (PhaP) are proteins normally associated with granules of poly(3-hydroxybutyrate) (PHB), a biodegradable polymer accumulated by many bacteria as a reserve molecule. These proteins enhance growth and polymer production in natural and recombinant PHB producers. It has been shown that the production of PHB causes stress in recombinantEscherichia coli, revealed by an increase in the concentrations of several heat stress proteins. In this work, quantitative reverse transcription (qRT)-PCR analysis was used to study the effect of PHB accumulation, and that of PhaP fromAzotobactersp. strain FA8, on the expression of stress-related genes in PHB-producingE. coli. While PHB accumulation was found to increase the transcription ofdnaKandibpA, the expression of these genes and ofgroES,groEL,rpoH,dps, andyfiDwas reduced, when PhaP was coexpressed, to levels even lower than those detected in the non-PHB-accumulating control. These results demonstrated the protective role of PhaP in PHB-synthesizingE. coliand linked the effects of the protein to the expression of stress-related genes, especiallyibpA. The effect of PhaP was also analyzed in non-PHB-synthesizing strains, showing that expression of this heterologous protein has an unexpected protective effect inE. coli, under both normal and stress conditions, resulting in increased growth and higher resistance to both heat shock and superoxide stress by paraquat. In addition, PhaP expression was shown to reduce RpoH protein levels during heat shock, probably by reducing or titrating the levels of misfolded proteins.

scholarly journals pfmdr1Amplification Is Related to Increased Plasmodium falciparum In Vitro Sensitivity to the Bisquinoline Piperaquine

2012 ◽  
Vol 56 (7) ◽  
pp. 3615-3619 ◽  
Author(s):  
M. I. Veiga ◽  
P. E. Ferreira ◽  
M. Malmberg ◽  
L. Jörnhagen ◽  
A. Björkman ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Copy Number ◽  
Statistical Significance ◽  
Resistance Locus ◽  
Content Type ◽  
Enhanced Sensitivity ◽  
In Vitro Sensitivity ◽  
Artemisinin Combination Therapies

ABSTRACTThe 4-aminoquinoline bisquinoline piperaquine is an important partner drug in one of the presently recommended artemisinin combination therapies. Recent clinical trials have confirmed its high efficacy in combination with dihydroartemisinin. Resistance to piperaquine alone has, however, been documented. Amplification in copy number of thePlasmodium falciparummultidrug resistance locus on chromosome 5, containing thepfmdr1gene, has been shown to confer resistance to structurally unrelated antimalarials. Through the determination of the 50% inhibitory concentrations (IC50s) and IC90s for piperaquine and chloroquine in a set of 46 adaptedP. falciparumcultures originating from the Thai-Burmese border, we have characterized the regions around thepfmdr1gene and identified a significant association between the presence ofpfmdr1duplications and enhanced sensitivity to piperaquine (P= 0.005 for IC50andP= 0.002 for IC90) and chloroquine, reaching statistical significance at IC90s (P= 0.026). These results substantiate the potential importance ofpfmdr1copy number amplifications in the efficacy of the combination therapy piperaquine-dihydroartemisinin. It supports the rational use of 4-aminoquinolines and artemisinin-based compounds, as they independently select for mutually incompatible combinations of mutations.

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