Prevalence of chloroquine and antifolate drug resistance alleles in Plasmodium falciparum clinical isolates from three areas in Ghana

Background: The emergence and spread of resistance in Plasmodium falciparum to chloroquine (CQ) necessitated the change from CQ to artemisinin-based combination therapies (ACTs) as first-line drug for the management of uncomplicated malaria in Ghana in 2005. Sulphadoxine-pyrimethamine (SP) which was the second line antimalarial drug in Ghana, was now adopted for intermittent preventive treatment of malaria in pregnancy (IPTp). Methods: To examine the prevalence of molecular markers associated with CQ and antifolate drug resistance in Ghana, we employed restriction fragment length polymorphism polymerase chain reaction to genotype and compare single nucleotide polymorphisms (SNPs) in the P. falciparum chloroquine resistance transporter ( pfcrt, PF3D7_0709000), multidrug resistance ( pfmdr1, PF3D7_0523000), bifunctional dihydrofolate reductase-thymidylate synthase ( pfdhfr, PF3D7_0417200) and dihydropteroate synthase ( pfdhps, PF3D7_0810800) genes. Parasites were collected from children with malaria reporting to hospitals in three different epidemiological areas of Ghana (Accra, Kintampo and Navrongo) in 2012-2013 and 2016-2017. Results: The overall prevalence of the CQ resistance-associated pfcrt 76T allele was 8%, whereas pfmdr1 86Y and 184F alleles were present in 10.2% and 65.1% of infections, respectively. The majority of the isolates harboured the antifolate resistance-associated pfdhfr alleles 51I (83.4%), 59R (85.9 %) and 108N (90.5%). Pfdhps 437G and 540E were detected in 90.6% and 0.7% of infections, respectively. We observed no significant difference across the three study sites for all the polymorphisms except for pfdhps 437G, which was more common in Accra compared to Kintampo for the 2016-2017 isolates. Across both pfdhfr and pfdhps genes, a large proportion (61%) of the isolates harboured the quadruple mutant combination (I 51 R 59 N 108/ G 437). CQ resistance alleles decreased during the 12 years after CQ withdrawal, but an mediate SP resistance alleles increased. Conclusion: Surveillance of the prevalence of resistance alleles is necessary in monitoring the efficacy of antimalarial drugs.

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Prevalence of chloroquine and antifolate drug resistance alleles in Plasmodium falciparum clinical isolates from three areas in Ghana

AAS Open Research ◽

10.12688/aasopenres.12825.1 ◽

2018 ◽

Vol 1 ◽

pp. 1 ◽

Cited By ~ 2

Author(s):

James Abugri ◽

Felix Ansah ◽

Kwaku P. Asante ◽

Comfort N. Opoku ◽

Lucas A. Amenga-Etego ◽

...

Keyword(s):

Drug Resistance ◽

Plasmodium Falciparum ◽

Chloroquine Resistance ◽

Published Data ◽

Nucleotide Polymorphisms ◽

Triple Mutant ◽

Chloroquine Resistance Transporter ◽

Significant Difference ◽

Quadruple Mutant ◽

Study Sites

Background: The emergence and spread of resistance in Plasmodium falciparum to chloroquine (CQ) and the antifolate drug sulfadoxine-pyrimethamine (SP) necessitated the change from CQ to artemisinin-based combination therapies (ACTs) as first-line drug for the management of uncomplicated malaria in Ghana in 2005. Methods: To examine the prevalence of molecular markers associated with CQ and antifolate drug resistance in Ghana, we genotyped single nucleotide polymorphisms (SNPs) in the P. falciparum chloroquine resistance transporter (pfcrt, PF3D7_0709000), multidrug resistance (pfmdr1, PF3D7_0523000), bifunctional dihydrofolate reductase-thymidylate synthase (pfdhfr, PF3D7_0417200) and dihydropteroate synthase (pfdhps, PF3D7_0810800) genes in children with malaria reporting to hospitals in three different epidemiological areas of Ghana (Accra, Kintampo and Navrongo) between 2012 and 2017. Results: The overall prevalence of the CQ resistance-associated pfcrt 76T allele was 8%, whereas pfmdr1 86Y and 184F alleles were present in 10% and 65% of infections respectively. Most of the isolates harboured the antifolate resistance-associated pfdhfr 51I, 59R and 108N alleles, including 68% of them with the triple mutant pfdhfr I51R59N108 combination. Pfdhps 437G and 540E were detected in 90.6% and 0.7% of infections, respectively. We observed no significant difference across the three study sites for all the polymorphisms except for pfdhps 437G, which was more common in Accra than at the other sites. Across both pfdhfr and pfdhps genes, a large proportion (61%) of the isolates harboured the quadruple mutant combination (I51R59N108/G437). Conclusion: Comparison of the present results to previously published data shows a significant decrease in the prevalence of CQ resistance alleles during the 12 years after CQ withdrawal, but an increase in the alleles that mediate SP resistance, which could be due to the continuous use of antifolate drugs for prophylaxis.

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Low prevalence of highly sulfadoxine‐resistant dihydropteroate synthase alleles in Plasmodium falciparum isolates in Benin

Malaria Journal ◽

10.1186/s12936-021-03605-5 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Samaly Souza Svigel ◽

Adicath Adeothy ◽

Augustin Kpemasse ◽

Ernest Houngbo ◽

Antoine Sianou ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Intermittent Preventive Treatment ◽

Preventive Treatment ◽

Nucleotide Polymorphisms ◽

Triple Mutant ◽

Single Nucleotide ◽

Dihydropteroate Synthase ◽

Seasonal Malaria Chemoprevention ◽

Study Sites ◽

Low Prevalence

Abstract Background In 2004, in response to high levels of treatment failure associated with sulfadoxine-pyrimethamine (SP) resistance, Benin changed its first-line malaria treatment from SP to artemisinin-based combination therapy for treatment of uncomplicated Plasmodium falciparum malaria. Resistance to SP is conferred by accumulation of single nucleotide polymorphisms (SNPs) in P. falciparum genes involved in folate metabolism, dihydrofolate reductase (Pfdhfr) and dihydropteroate synthase (Pfdhps), targeted by pyrimethamine and sulfadoxine, respectively. Because SP is still used for intermittent preventive treatment in pregnant women (IPTp) and seasonal malaria chemoprevention (SMCP) in Benin, the prevalence of Pfdhfr and Pfdhps SNPs in P. falciparum isolates collected in 2017 were investigated. Methods This study was carried out in two sites where the transmission of P. falciparum malaria is hyper-endemic: Klouékanmey and Djougou. Blood samples were collected from 178 febrile children 6–59 months old with confirmed uncomplicated P. falciparum malaria and were genotyped for SNPs associated with SP resistance. Results The Pfdhfr triple mutant IRN (N51I, C59R, and S108N) was the most prevalent (84.6%) haplotype and was commonly found with the Pfdhps single mutant A437G (50.5%) or with the Pfdhps double mutant S436A and A437G (33.7%). The quintuple mutant, PfdhfrIRN/PfdhpsGE (A437G and K540E), was rarely observed (0.8%). The A581G and A613S mutant alleles were found in 2.6 and 3.9% of isolates, respectively. Six isolates (3.9%) were shown to harbour a mutation at codon I431V, recently identified in West African parasites. Conclusions This study showed that Pfdhfr triple IRN mutants are near fixation in this population and that the highly sulfadoxine-resistant Pfdhps alleles are not widespread in Benin. These data support the continued use of SP for chemoprevention in these study sites, which should be complemented by periodic nationwide molecular surveillance to detect emergence of resistant genotypes.

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Prevalence and factors associated with carriage of Pfmdr1 polymorphisms among pregnant women receiving intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP) and artemether-lumefantrine for malaria treatment in Burkina Faso

Malaria Journal ◽

10.1186/s12936-020-03473-5 ◽

2020 ◽

Vol 19 (1) ◽

Author(s):

Hamtandi Magloire Natama ◽

Rouamba Toussaint ◽

Djamina Line Cerine Bazié ◽

Sékou Samadoulougou ◽

Maminata Coulibaly-Traoré ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Burkina Faso ◽

Pregnant Women ◽

Mutant Allele ◽

Intermittent Preventive Treatment ◽

Preventive Treatment ◽

Dried Blood Spots ◽

Nucleotide Polymorphisms ◽

High Coverage ◽

Factors Associated

Abstract Background Single nucleotide polymorphisms occurring in the Plasmodium falciparum multidrug resistant gene 1 (pfmdr1) are known to be associated with aminoquinoline resistance and, therefore, represent key P. falciparum markers for monitoring resistance both in susceptible groups (children under 5 years old and pregnant women) and in the general population. This study aimed to determine prevalence and factors associated with the carriage of pfmdr1 N86Y, Y184F and D1246Y polymorphisms among pregnant women in a setting of high malaria transmission in Burkina Faso. Methods Plasmodium falciparum isolates were collected at the first antenatal care visit (ANC-1) as well as at delivery from pregnant women participating in the COSMIC trial (NTC01941264), which assessed malaria preventive interventions during pregnancy in the Nanoro Health District. Here, pregnant women received intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP) and malaria infections and/or diseases were treated using artemether-lumefantrine (AL) during the trial. Parasite DNA was extracted from dried blood spots and the presence of pfmdr1 mutations at positions 86, 184 and 1246 was determined using nested PCR, followed by restriction fragment length polymorphism (RFLP) analysis. Results A prevalence of 13.2% (20/151) and 12.1% (14/116) of the pfmdr1 86Y mutant allele was found at ANC-1 and at delivery, respectively, while no mutant allele was observed for Y184F and D1246Y codons at both ANC-1 and at delivery. There were no significant factors associated with pfmdr1 86Y mutant allele carriage at ANC-1. However, malaria infections at delivery with a parasite density above the median (2237.2 (IQR: 613.5–11,425.7) parasites/µl) was associated with an increase risk of pfmdr1 86Y mutant allele carriage (AOR = 5.5 (95% CI 1.07–28.0); P = 0.04). In contrast, both three or more IPTp-SP doses (AOR = 0.25 (95% CI 0.07–0.92); P = 0.04) and one or more AL treatment (AOR = 0.25 (95% CI 0.07–0.89); P = 0.03) during pregnancy were associated with a significant reduce risk of pfmdr1 86Y mutant allele carriage at delivery. Conclusion These findings suggest that both high coverage of IPTp-SP and the use of AL for the treatment of malaria infection/disease during pregnancy select for pfmdr1 N86 wild-type allele at delivery.

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Increasing Prevalence of a Novel Triple-Mutant Dihydropteroate Synthase Genotype in Plasmodium falciparum in Western Kenya

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.04961-14 ◽

2015 ◽

Vol 59 (7) ◽

pp. 3995-4002 ◽

Cited By ~ 4

Author(s):

Naomi W. Lucchi ◽

Sheila Akinyi Okoth ◽

Franklin Komino ◽

Philip Onyona ◽

Ira F. Goldman ◽

...

Keyword(s):

Drug Resistance ◽

Plasmodium Falciparum ◽

Mutant Allele ◽

Double Mutant ◽

Intermittent Preventive Treatment ◽

Preventive Treatment ◽

Triple Mutant ◽

Western Kenya ◽

Content Type ◽

Dihydropteroate Synthase

ABSTRACTThe molecular basis of sulfadoxine-pyrimethamine (SP) resistance lies in a combination of single-nucleotide polymorphisms (SNPs) in two genes coding forPlasmodium falciparumdihydrofolate reductase (Pfdhfr) andP. falciparumdihydropteroate synthase (Pfdhps), targeted by pyrimethamine and sulfadoxine, respectively. The continued use of SP for intermittent preventive treatment in pregnant women in many African countries, despite SP's discontinuation as a first-line antimalarial treatment option due to high levels of drug resistance, may further increase the prevalence of SP-resistant parasites and/or lead to the selection of new mutations. An antimalarial drug resistance surveillance study was conducted in western Kenya between 2010 and 2013. A total of 203 clinical samples from children with uncomplicated malaria were genotyped for SNPs associated with SP resistance. The prevalence of the triple-mutantPfdhfrC50I51R59N108I164genotype and the double-mutantPfdhpsS436G437E540A581A613genotype was high. Two triple-mutantPfdhpsgenotypes, S436G437E540G581A613andH436G437E540A581A613, were found, with the latter thus far being uniquely found in western Kenya. The prevalence of the S436G437E540G581A613genotype was low. However, a steady increase in the prevalence of thePfdhpstriple-mutantH436G437E540A581A613genotype has been observed since its appearance in early 2000. Isolates with these genotypes shared substantial microsatellite haplotypes with the most common double-mutant allele, suggesting that this triple-mutant allele may have evolved locally. Overall, these findings show that the prevalence of theH436G437E540A581A613triple mutant may be increasing in this population and could compromise the efficacy of SP for intermittent preventive treatment in pregnant women if it increases the resistance threshold further.

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Prevalence of Plasmodium falciparum Chloroquine Resistance Transporter (Pfcrt76T) Mutation Associated with Antimalarial Drug Resistance in Two Different Epidemiological Setting (Banfora and Saponé) in Burkina Faso Few Years after the Implementation of Artemisnine Based Combination Therapy (ACTs)

International Journal of Pathogen Research ◽

10.9734/ijpr/2019/v3i330094 ◽

2020 ◽

pp. 1-11

Author(s):

Séni Nikiema ◽

Samuel Sindié Sermé ◽

Salif Sombié ◽

Amidou Diarra ◽

Noelie Bere Henry ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Burkina Faso ◽

Public Health Problem ◽

Antimalarial Drugs ◽

Sub Saharan Africa ◽

Chloroquine Resistance ◽

Chloroquine Resistance Transporter ◽

Significant Difference ◽

Sub Saharan ◽

Biology Laboratory

Introduction: In spite of considerable progress, malaria remains a public health problem in many areas, particularly in sub-Saharan Africa. One major complexity of malaria disease is caused by the development and the spread of vector and parasite resistance to insecticides and antimalarial drugs respectively. The Pfcrt76T gene mutation has been validated as a marker conferring resistance to chloroquine and other antimalarial drugs. The extension of Plasmodium falciparum resistance to commonly used antimalarial drugs (chloroquine, sulfadoxine-pyrimethamine) led to the adoption and the use of artemisinin-based combinations in Burkina Faso since 2005. Aims: The present study was initiated to assess the prevalence of the Pfcrt76T mutation in two different malaria epidemiological setting after a decade of introduction of artemisinin-based combination therapies (ACTs) in Burkina Faso. Methodology: The study population consisted of 181 uncomplicated malaria patients recruited in Banfora and Saponé health districts in 2012 and 2013. Blood samples were collected from finger prick on filter paper, dried and sent to the Molecular Biology Laboratory at Centre National de Recherche et de Formation sur le Paludisme (CNRFP) for molecular analyzes. DNA of Plasmodium falciparum was extracted with DNA extraction kit (Qiagen®) and the Pfcrt76T mutation was determined based on Polymerase Chain Reaction / Restriction Fragment Length Polymorphism technique (RFLP). Results: The results of this study showed that the frequency of the pfcrt76T mutant allele (33.7%) was statistically lower than the Pfcrt76K wild-type allele (57.4%) in the study area. Moreover, the prevalence of Pfcrt76T mutation was neither associated with the patient age nor with the parasite density while a significant difference was observed between the two epidemiological setting, Banfora and Saponé. Conclusion: The findings of this study has shown a drop in the prevalence of mutant parasites Pfcrt76T in both the study area eight years after the introduction of ACTs compared to previous studies.

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En-route to the ‘elimination’ of genotypic chloroquine resistance in Western and Southern Zambia, 14 years after chloroquine withdrawal

Malaria Journal ◽

10.1186/s12936-019-3031-4 ◽

2019 ◽

Vol 18 (1) ◽

Cited By ~ 2

Author(s):

Lungowe Sitali ◽

Mulenga C. Mwenda ◽

John M. Miller ◽

Daniel J. Bridges ◽

Moonga B. Hawela ◽

...

Keyword(s):

Drug Resistance ◽

Malaria Elimination ◽

Household Survey ◽

Chloroquine Resistance ◽

Multi Drug Resistance ◽

High Resolution Melt ◽

Cross Sectional ◽

Chloroquine Resistance Transporter ◽

Significant Difference ◽

Elimination Programme

Abstract Background Anti-malarial resistance is, and continues to be a significant challenge in the fight against malaria and a threat to achieving malaria elimination. In Zambia, chloroquine (CQ), a safe, affordable and well-tolerated drug, was removed from use in 2003 due to high levels of resistance evidenced with treatment failure. This study sought to investigate the prevalence of chloroquine resistance markers in Southern and Western Provinces of Zambia 14 years after the withdrawal of CQ. Methods Data from a cross-sectional, all-age household survey, conducted during the peak malaria transmission season (April–May 2017) was analysed. During the all-age survey, socio-demographic information and coverage of malaria interventions were collected. Consenting individuals were tested for malaria with a rapid diagnostic test and a spot of blood collected on filter paper to create a dried blood spot (DBS). Photo-induced electronic transfer–polymerase chain reaction (PET–PCR) was used to analyse the DBS for the presence of all four malaria species. Plasmodium falciparum positive samples were analysed by high resolution melt (HRM) PCR to detect the presence of genotypic markers of drug resistance in the P. falciparum chloroquine resistance transporter (Pfcrt) and P. falciparum multi-drug resistance (Pfmdr) genes. Results A total of 181 P. falciparum positive samples were examined for pfcrt K76T and MDR N86. Of the 181 samples 155 successfully amplified for Pfcrt and 145 for Pfmdr N86. The overall prevalence of CQ drug-resistant parasites was 1.9% (3/155), with no significant difference between the two provinces. No N86Y/F mutations in the Pfmdr gene were observed in any of the sample. Conclusion This study reveals the return of CQ sensitive parasites in Southern and Western Provinces of Zambia 14 years after its withdrawal. Surveillance of molecular resistant markers for anti-malarials should be included in the Malaria Elimination Programme so that resistance is monitored country wide.

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3-Halo Chloroquine Derivatives Overcome Plasmodium falciparum Chloroquine Resistance Transporter-Mediated Drug Resistance in P. falciparum

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01139-15 ◽

2015 ◽

Vol 59 (12) ◽

pp. 7891-7893 ◽

Cited By ~ 2

Author(s):

Sonia Edaye ◽

Dagobert Tazoo ◽

D. Scott Bohle ◽

Elias Georges

Keyword(s):

Drug Resistance ◽

Plasmodium Falciparum ◽

Chloroquine Resistance ◽

Content Type ◽

Chloroquine Resistance Transporter ◽

Highly Effective

ABSTRACTPolymorphism in thePlasmodium falciparumchloroquine resistance transporter (PfCRT) was shown to cause chloroquine resistance. In this report, we examined the antimalarial potential of novel 3-halo chloroquine derivatives (3-chloro, 3-bromo, and 3-iodo) against chloroquine-susceptible and -resistantP. falciparum. All three derivatives inhibited the proliferation ofP. falciparum; with 3-iodo chloroquine being most effective. Moreover, 3-iodo chloroquine was highly effective at potentiating and reversing chloroquine toxicity of drug-susceptible and -resistantP. falciparum.

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Detection of antimalarial drug resistance polymorphisms in Plasmodium falciparum chloroquine resistance transporter and Plasmodium falciparum multidrug resistance 1 genes of Plasmodium falciparum found in Kano State, Nigeria

Calabar Journal of Health Sciences ◽

10.25259/cjhs_14_2020 ◽

2021 ◽

Vol 5 ◽

pp. 8-14

Author(s):

Al-Mukhtar Yahuza Adamu ◽

Olayeni Stephen Olonitola ◽

Helen Ileigo Inabo ◽

Ahmad Babangida Suleiman

Keyword(s):

Drug Resistance ◽

Plasmodium Falciparum ◽

Multidrug Resistance ◽

Antimalarial Drug ◽

Artemisinin Combination Therapy ◽

Chloroquine Resistance ◽

Antimalarial Drug Resistance ◽

Antimalarial Agents ◽

Chloroquine Resistance Transporter ◽

Two Samples

Objectives: In 2018, malaria claimed an estimated 380,000 lives in African region, with Nigeria accounting for 24.0% (91,368) of malaria deaths from the region. Mutations in Plasmodium falciparum chloroquine resistance transporter (Pfcrt) and P. falciparum multidrug resistance 1 (Pfmdr-1) genes had reduced the effective use of artemisinin combination therapy through the development of resistance to these antimalarial agents. Our study set out to determine the antimalarial drug resistance polymorphisms in Pfcrt and Pfmdr-1 genes of P. falciparum isolates among patients in Kano State, Nigeria. Material and Methods: Malaria positive samples were collected across the three senatorial districts of Kano State. The samples were amplified using nested polymerase chain reaction to detect the Pfcrt and Pfmdr-1 genes. The amplicons were sequenced and bioinformatic analysis was done using CLC Sequence viewer 8.0 and BioEdit sequence alignment editor to detect the single-nucleotide polymorphisms. Results: In the Pfcrt gene, CVIET haplotype was seen in 26.2% of the samples while only two samples showed the 86Y mutation in the Pfmdr-1 gene. All the 86Y mutations and majority of the CVIET haplotypes were detected in the patients from rural settings where some of them noted that they consumed modern and traditional (herbs) antimalarial agents. One sample was observed to have the CVIET haplotype and N86Y mutation while the other five CVIET haplotypes were seen in five separate samples. A new mutation V62A was found in the Pfmdr-1 gene as observed in one of the sample. Conclusion: It is imperative to ensure the rational use of the right antimalarial agents and employ continuous resistance surveillance/mapping to ensure synergy in malaria containment and elimination strategies.

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