scholarly journals Differential Association of Plasmodium falciparum Na+/H+Exchanger Polymorphism and Quinine Responses in Field- and Culture-Adapted Isolates of Plasmodium falciparum

2011 ◽  
Vol 55 (12) ◽  
pp. 5834-5841 ◽  
Author(s):  
Stéphane Pelleau ◽  
Lionel Bertaux ◽  
Sébastien Briolant ◽  
Michael T. Ferdig ◽  
Véronique Sinou ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Drug Testing ◽  
Inhibitory Concentration ◽  
Cumulative Effect ◽  
Differential Association ◽  
Microsatellite Sequence ◽  
Content Type ◽  
Field Isolates ◽  
Selection Of

ABSTRACTPlasmodium falciparumisolates with decreased susceptibility to quinine are increasingly being found in malaria patients. Mechanisms involved in this resistance are not yet understood. Several studies claim that alongside mutations in the Pfcrtand Pfmdr1genes, the Pfnhe-1Na+/H+exchanger polymorphism plays a role in decreasing susceptibility. However, conflicting results on the link between the Pfnhe-1gene and quinine resistance arise from field- and culture-adapted isolates. We tested the association between Pfnhe-1, Pfcrt, and Pfmdr1polymorphisms in field- and culture-adapted isolates from various countries with theirin vitrosusceptibility to quinine. Field isolates presented a higher diversity of the Pfnhe-1microsatellite sequence than culture-adapted isolates. In culture-adapted isolates but not in field isolates, mutations in the Pfcrtand Pfmdr1genes, as well as a higher number of DNNND repeats in the Pfnhe-1gene, were associated with a higher 50% inhibitory concentration (IC50) of quinine. Furthermore, most of the culture-adapted isolates with more than one DNNND repeat in the Pfnhe-1gene also harbored mutated Pfcrtand Pfmdr1genes with an apparent cumulative effect on quinine susceptibility. This study supports the involvement of the Pfnhe-1gene in the modulation of thein vitroquinine response when associated with mutated Pfcrtand Pfmdr1genes. Culture adaptation could be responsible for selection of specific haplotypes of these three genes. Methods used for drug testing might thus influence the association between Pfnhe-1polymorphism and quinine susceptibility. However, we do not exclude the possibility that in particular settings, Pfnhe-1polymorphism can be used as a molecular marker for surveillance of quinine resistance.

scholarly journals ReducedIn VitroDoxycycline Susceptibility in Plasmodium falciparum Field Isolates from Kenya Is Associated with PfTetQ KYNNNN Sequence Polymorphism

2014 ◽  
Vol 58 (10) ◽  
pp. 5894-5899 ◽  
Author(s):  
Angela O. Achieng ◽  
Luiser A. Ingasia ◽  
Dennis W. Juma ◽  
Agnes C. Cheruiyot ◽  
Charles A. Okudo ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Amino Acid ◽  
Genetic Polymorphisms ◽  
Copy Number ◽  
Sequence Polymorphism ◽  
Limited Information ◽  
Content Type ◽  
Field Isolates ◽  
Potential Marker

ABSTRACTDoxycycline is widely used for malaria prophylaxis by international travelers. However, there is limited information on doxycycline efficacy in Kenya, and genetic polymorphisms associated with reduced efficacy are not well defined.In vitrodoxycycline susceptibility profiles for 96Plasmodium falciparumfield isolates from Kenya were determined. Genetic polymorphisms were assessed inP. falciparummetabolite drug transporter (Pfmdt) andP. falciparumGTPasetetQ(PftetQ) genes. Copy number variation of the gene and the number of KYNNNN amino acid motif repeats within the protein encoded by PftetQwere determined. Reducedin vitrosusceptibility to doxycycline was defined by 50% inhibitory concentrations (IC50s) of ≥35,000 nM. The odds ratio (OR) of having 2 PfTetQ KYNNNN amino acid repeats in isolates with IC50s of >35,000 nM relative to those with IC50s of <35,000 nM is 15 (95% confidence interval [CI], 3.0 to 74.3;Pvalue of <0.0002). Isolates with 1 copy of the Pfmdtgene had a median IC50of 6,971 nM, whereas those with a Pfmdtcopy number of >1 had a median IC50of 9,912 nM (P= 0.0245). Isolates with 1 copy of PftetQhad a median IC50of 6,370 nM, whereas isolates with a PftetQcopy number of >1 had a median IC50of 3,422 nM (P< 0.0007). Isolates with 2 PfTetQ KYNNNN motif repeats had a median IC50of 26,165 nM, whereas isolates with 3 PfTetQ KYNNNN repeats had a median IC50of 3,352 nM (P= 0.0023). PfTetQ sequence polymorphism is associated with a reduced doxycycline susceptibility phenotype in Kenyan isolates and is a potential marker for susceptibility testing.

scholarly journals In VitroSensitivities of Plasmodium falciparum Isolates from the China-Myanmar Border to Piperaquine and Association with Polymorphisms in Candidate Genes

2013 ◽  
Vol 57 (4) ◽  
pp. 1723-1729 ◽  
Author(s):  
Mingming Hao ◽  
Dandan Jia ◽  
Qing Li ◽  
Yongshu He ◽  
Lili Yuan ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Molecular Mechanisms ◽  
Inhibitory Concentration ◽  
Pearson Correlation ◽  
Geometric Mean ◽  
Content Type ◽  
Artemisinin Derivatives ◽  
Wide Range ◽  
High Prevalence

ABSTRACTThe recent reports of resistance inPlasmodium falciparumto artemisinin derivatives and their partner drugs demand intensive studies toward understanding the molecular mechanisms of resistance. In this study, we examined thein vitrosusceptibility of 63P. falciparumfield isolates collected from the China-Myanmar border area to chloroquine (CQ) and piperaquine (PPQ). Parasite isolates remained highly resistant to CQ, with the geometric mean 50% inhibitory concentration (IC50) of 252.7 nM and a range of 51.9 to 1,052.0 nM. In comparison, these parasites had a geometric mean IC50of 28.4 nM for PPQ, with a fairly wide range of 5.3 to 132.0 nM, suggesting that certain parasite isolates displayed relatively high levels of resistance to PPQ. Interestingly, within the 4 years of study, the parasites exhibited a continuous decline in susceptibilities to both CQ and PPQ, and there was a significant correlation between responses to CQ and PPQ (Pearson correlation coefficient = 0.79,P< 0.0001). Consistent with the CQ-resistant phenotype, all parasites carried thepfcrtK76T mutation, and most parasites had the CVIET type that is prevalent in Southeast Asia. In contrast,pfmdr1mutations were relatively rare, and no gene amplification was detected. Only thepfmdr1N1042D mutation was associated with resistance to CQ. For thepfmrp1gene, four substitutions reached relatively high prevalence of >22%, and the I876V mutation was associated with reduced sensitivity to CQ. However, we could not establish a link between PPQ responses and the polymorphisms in the three genes associated with quinoline drug resistance.

scholarly journals Repeat Polymorphisms in the Low-Complexity Regions of Plasmodium falciparum ABC Transporters and Associations withIn VitroAntimalarial Responses

2013 ◽  
Vol 57 (12) ◽  
pp. 6196-6204 ◽  
Author(s):  
John Okombo ◽  
Abdirahman I. Abdi ◽  
Steven M. Kiara ◽  
Leah Mwai ◽  
Lewa Pole ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Abc Transporters ◽  
Inhibitory Concentration ◽  
Low Complexity ◽  
Clinical Isolates ◽  
Inverse Association ◽  
Sequence Profile ◽  
Content Type ◽  
Functional Studies

ABSTRACTThePlasmodium falciparumgenome is rich in regions of low amino acid complexity which evolve with few constraints on size. To explore the extent of diversity in these loci, we sequenced repeat regions inpfmdr1,pfmdr5,pfmdr6,pfmrp2, and the antigenic locuspfmsp8in laboratory and cultured-adapted clinical isolates. We further assessed associations between the repeats and parasitein vitroresponses to 7 antimalarials to determine possible adaptive roles of these repeats in drug tolerance. Our results show extensive repeat variations in the reference and clinical isolates in all loci. We also observed a modest increase in dihydroartemisinin activity in parasites harboring thepfmdr1sequence profile 7-2-10 (reflecting the number of asparagine repeats, number of aspartate repeats, and number of asparagine repeats in the final series of the gene product) (P= 0.0321) and reduced sensitivity to chloroquine, mefloquine, quinine, and dihydroartemisinin in those with the 7-2-11 profile (P= 0.0051, 0.0068, 0.0011, and 0.0052, respectively). Interestingly, we noted an inverse association between two drugs whereby isolates with 6 asparagine repeats encoded bypfmdr6were significantly more susceptible to piperaquine than those with 8 (P= 0.0057). Against lumefantrine, those with 8 repeats were, however, more sensitive (P= 0.0144). Inpfmrp2, the 7-DNNNTS/NNNNTS (number of DNNNTS or NNNNTS motifs; underlining indicates dimorphism) repeat group was significantly associated with a higher lumefantrine 50% inhibitory concentration (IC50) (P= 0.008) than in those without. No associations were observed withpfmsp8. These results hint at the probable utility of some repeat conformations as markers ofin vitroantimalarial response; hence, biochemical functional studies to ascertain their role inP. falciparumare required.

scholarly journals In vitro and in vivo inhibition of malaria parasite infection by monoclonal antibodies against Plasmodium falciparum circumsporozoite protein (CSP)

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Merricka C. Livingstone ◽  
Alexis A. Bitzer ◽  
Alish Giri ◽  
Kun Luo ◽  
Rajeshwer S. Sankhala ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Monoclonal Antibodies ◽  
Disease Burden ◽  
Vaccine Candidate ◽  
Specific Binding ◽  
Circumsporozoite Protein ◽  
Target Epitope ◽  
Selection Of

AbstractPlasmodium falciparum malaria contributes to a significant global disease burden. Circumsporozoite protein (CSP), the most abundant sporozoite stage antigen, is a prime vaccine candidate. Inhibitory monoclonal antibodies (mAbs) against CSP map to either a short junctional sequence or the central (NPNA)n repeat region. We compared in vitro and in vivo activities of six CSP-specific mAbs derived from human recipients of a recombinant CSP vaccine RTS,S/AS01 (mAbs 317 and 311); an irradiated whole sporozoite vaccine PfSPZ (mAbs CIS43 and MGG4); or individuals exposed to malaria (mAbs 580 and 663). RTS,S mAb 317 that specifically binds the (NPNA)n epitope, had the highest affinity and it elicited the best sterile protection in mice. The most potent inhibitor of sporozoite invasion in vitro was mAb CIS43 which shows dual-specific binding to the junctional sequence and (NPNA)n. In vivo mouse protection was associated with the mAb reactivity to the NANPx6 peptide, the in vitro inhibition of sporozoite invasion activity, and kinetic parameters measured using intact mAbs or their Fab fragments. Buried surface area between mAb and its target epitope was also associated with in vivo protection. Association and disconnects between in vitro and in vivo readouts has important implications for the design and down-selection of the next generation of CSP based interventions.

scholarly journals Kenyan Plasmodium falciparum Field Isolates: Correlation between Pyrimethamine and Chlorcycloguanil Activity In Vitro and Point Mutations in the Dihydrofolate Reductase Domain

1998 ◽  
Vol 42 (1) ◽  
pp. 164-169 ◽  
Author(s):  
A. Nzila-Mounda ◽  
E. K. Mberu ◽  
C. H. Sibley ◽  
C. V. Plowe ◽  
P. A. Winstanley ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Dihydrofolate Reductase ◽  
Drug Response ◽  
Point Mutations ◽  
Distinct Group ◽  
Wild Type ◽  
Field Isolates ◽  
Vitro Data ◽  
In Vitro Data

ABSTRACT Sixty-nine Kenyan Plasmodium falciparum field isolates were tested in vitro against pyrimethamine (PM), chlorcycloguanil (CCG), sulfadoxine (SD), and dapsone (DDS), and their dihydrofolate reductase (DHFR) genotypes were determined. The in vitro data show that CCG is more potent than PM and that DDS is more potent than SD. DHFR genotype is correlated with PM and CCG drug response. Isolates can be classified into three distinct groups based on their 50% inhibitory concentrations (IC50s) for PM and CCG (P< 0.01) and their DHFR genotypes. The first group consists of wild-type isolates with mean PM and CCG IC50s of 3.71 ± 6.94 and 0.24 ± 0.21 nM, respectively. The second group includes parasites which all have mutations at codon 108 alone or also at codons 51 or 59 and represents one homogeneous group for which 25- and 6-fold increases in PM and CCG IC50s, respectively, are observed. Parasites with mutations at codons 108, 51, and 59 (triple mutants) form a third distinct group for which nine- and eightfold increases in IC50s, respectively, of PM and CCG compared to the second group are observed. Surprisingly, there is a significant decrease (P < 0.01) of SD and DDS susceptibility in these triple mutants. Our data show that more than 92% of Kenyan field isolates have undergone at least one point mutation associated with a decrease in PM activity. These findings are of great concern because they may indicate imminent PM-SD failure, and there is no affordable antimalarial drug to replace PM-SD (Fansidar).

scholarly journals Assessment of the Drug Susceptibility of Plasmodium falciparum Clinical Isolates from Africa by Using a Plasmodium Lactate Dehydrogenase Immunodetection Assay and an Inhibitory Maximum Effect Model for Precise Measurement of the 50-Percent Inhibitory Concentration

2006 ◽  
Vol 50 (10) ◽  
pp. 3343-3349 ◽  
Author(s):  
Halima Kaddouri ◽  
Serge Nakache ◽  
Sandrine Houzé ◽  
France Mentré ◽  
Jacques Le Bras
Keyword(s):  
Drug Resistance ◽  
Plasmodium Falciparum ◽  
Lactate Dehydrogenase ◽  
Active Metabolite ◽  
Inhibitory Concentration ◽  
Drug Susceptibility ◽  
Maximum Effect ◽  
Malaria Parasites ◽  
Effect Model

ABSTRACT The extension of drug resistance among malaria-causing Plasmodium falciparum parasites in Africa necessitates implementation of new combined therapeutic strategies. Drug susceptibility phenotyping requires precise measurements. Until recently, schizont maturation and isotopic in vitro assays were the only methods available, but their use was limited by technical constraints. This explains the revived interest in the development of replacement methods, such as the Plasmodium lactate dehydrogenase (pLDH) immunodetection assay. We evaluated a commercially controlled pLDH enzyme-linked immunosorbent assay (ELISA; the ELISA-Malaria antigen test; DiaMed AG, Cressier s/Morat, Switzerland) to assess drug susceptibility in a standard in vitro assay using fairly basic laboratory equipment to study the in vitro resistance of malaria parasites to major antimalarials. Five Plasmodium falciparum clones and 121 clinical African isolates collected during 2003 and 2004 were studied by the pLDH ELISA and the [8-3H]hypoxanthine isotopic assay as a reference with four antimalarials. Nonlinear regression with a maximum effect model was used to estimate the 50% inhibitory concentration (IC50) and its confidence intervals. The two methods were observed to have similar reproducibilities, but the pLDH ELISA demonstrated a higher sensitivity. The high correlation (r = 0.98) and the high phenotypic agreement (κ = 0.88) between the two methods allowed comparison by determination of the IC50s. Recently collected Plasmodium falciparum African isolates were tested by pLDH ELISA and showed drug resistance or decreased susceptibilities of 62% to chloroquine and 11.5% to the active metabolite of amodiaquine. No decreased susceptibility to lumefantrine or the active metabolite of artemisinin was detected. The availability of this simple and highly sensitive pLDH immunodetection assay will provide an easier method for drug susceptibility testing of malaria parasites.

scholarly journals Molecular Characterization of the CytochromebGene andIn VitroAtovaquone Susceptibility of Plasmodium falciparum Isolates from Kenya

2015 ◽  
Vol 59 (3) ◽  
pp. 1818-1821 ◽  
Author(s):  
Luicer A. Ingasia ◽  
Hoseah M. Akala ◽  
Mabel O. Imbuga ◽  
Benjamin H. Opot ◽  
Fredrick L. Eyase ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Genetic Polymorphism ◽  
Molecular Characterization ◽  
Mutant Allele ◽  
Inhibitory Concentration ◽  
Wild Type Allele ◽  
Wild Type ◽  
Western Kenya ◽  
Content Type

ABSTRACTThe prevalence of a genetic polymorphism(s) at codon 268 in the cytochromebgene, which is associated with failure of atovaquone-proguanil treatment, was analyzed in 227Plasmodium falciparumparasites from western Kenya. The prevalence of the wild-type allele was 63%, and that of the Y268S (denoting a Y-to-S change at position 268) mutant allele was 2%. There were no pure Y268C or Y268N mutant alleles, only mixtures of a mutant allele(s) with the wild type. There was a correlation between parasite 50% inhibitory concentration (IC50) and parasite genetic polymorphism; mutant alleles had higher IC50s than the wild type.

scholarly journals Nitro/Nitrosyl-Ruthenium Complexes Are Potent and Selective Anti-Trypanosoma cruzi Agents Causing Autophagy and Necrotic Parasite Death

2014 ◽  
Vol 58 (10) ◽  
pp. 6044-6055 ◽  
Author(s):  
Tanira M. Bastos ◽  
Marília I. F. Barbosa ◽  
Monize M. da Silva ◽  
José W. da C. Júnior ◽  
Cássio S. Meira ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Inhibitory Concentration ◽  
Content Type ◽  
X Ray ◽  
X Ray Crystallography ◽  
Ruthenium Nitrosyl ◽  
Index Analysis ◽  
Antiparasitic Drug ◽  
Drug Complex

ABSTRACTcis-[RuCl(NO2)(dppb)(5,5′-mebipy)] (complex 1),cis-[Ru(NO2)2(dppb)(5,5′-mebipy)] (complex 2),ct-[RuCl(NO)(dppb)(5,5′-mebipy)](PF6)2(complex 3), andcc-[RuCl(NO)(dppb)(5,5′-mebipy)](PF6)2(complex 4), where 5,5′-mebipy is 5,5′-dimethyl-2,2′-bipyridine and dppb is 1,4-bis(diphenylphosphino)butane, were synthesized and characterized. The structure of complex 2 was determined by X-ray crystallography. These complexes exhibited a higher anti-Trypanosoma cruziactivity than benznidazole, the current antiparasitic drug. Complex 3 was the most potent, displaying a 50% effective concentration (EC50) of 2.1 ± 0.6 μM against trypomastigotes and a 50% inhibitory concentration (IC50) of 1.3 ± 0.2 μM against amastigotes, while it displayed a 50% cytotoxic concentration (CC50) of 51.4 ± 0.2 μM in macrophages. It was observed that the nitrosyl complex 3, but not its analog lacking the nitrosyl group, releases nitric oxide into parasite cells. This release has a diminished effect on the trypanosomal protease cruzain but induces substantial parasite autophagy, which is followed by a series of irreversible morphological impairments to the parasites and finally results in cell death by necrosis. In infected mice, orally administered complex 3 (five times at a dose of 75 μmol/kg of body weight) reduced blood parasitemia and increased the survival rate of the mice. Combination index analysis of complex 3 indicated that itsin vitroactivity against trypomastigotes is synergic with benznidazole. In addition, drug combination enhanced efficacy in infected mice, suggesting that ruthenium-nitrosyl complexes are potential constituents for drug combinations.

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.

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