High frequency of artemisinin partial resistance mutations in the Great Lakes region revealed through rapid pooled deep sequencing

The Journal of Infectious Diseases, Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 5, 2024

Abstract Background In Africa, the first Plasmodium falciparum artemisinin partial resistance mutation was Kelch13 (K13) 561H, detected and validated at appreciable frequency in Rwanda 2014. Surveillance to better define extent of emergence neighboring countries is critical. Methods We used novel liquid blood drop preservation with pooled sequencing provide cost-effective rapid assessment frequencies multiple collection sites across regions Uganda, Tanzania, Democratic Republic Congo. Malaria-positive samples (N = 5465) from 39 health facilities collected between May 2022 March 2023 were sequenced 199 pools. Results Rwanda, K13 561H 675V 90% 65% sites, an average 19.0% (range, 0%–54.5%) 5.0% (0%–35.5%), respectively. had high sites. appeared 1.6% Uganda. absent Congo, although seen low frequency. Concerningly, candidate mutations observed: 441L, 449A, 469F co-occurred mutations, suggesting that they are arising under same pressures. Other markers for decreased susceptibility artemether-lumefantrine common: P multidrug protein 1 N86 98.0% 63.3%–100%) 184F 47.0% (0%–94.3%) chloroquine transporter 76T 14.7% (0%–58.6%). Additionally, sulfadoxine-pyrimethamine–associated show frequencies. Conclusions rapidly expanding region, further endangering control efforts potential engendering partner drug resistance.

Language: Английский

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Prevalence of mutations associated with artemisinin partial resistance and sulfadoxine–pyrimethamine resistance in 13 regions in Tanzania in 2021: a cross-sectional survey

The Lancet Microbe, Journal Year: 2024, Volume and Issue: 5(10), P. 100920 - 100920

Published: Aug. 16, 2024

The emergence of the artemisinin partial resistance (ART-R) mutation in Plasmodium falciparum kelch13 gene (k13), Arg561His, Rwanda and regional presence polymorphisms affecting sulfadoxine-pyrimethamine have raised concern neighbouring Tanzania. goal this study was to assess status antimalarial Tanzania, with a focus on border Rwanda, understand distribution Arg561His mutation, partner drug resistance, chemoprevention drugs. In cross-sectional survey, capillary dried blood spots were collected from malaria positive asymptomatic individuals community symptomatic health facilities aged 6 months older, 13 regions mainland Tanzania Jan 31 June 26, 2021. Exclusion criteria included residence areas other than target sites, presenting facility for care treatment conditions malaria, not providing informed consent. Samples assessed genetic relatedness using molecular inversion probes targeting P short-read whole-genome sequencing. primary outcome prevalence markers at region level, as well district level Kagera, northwest country Rwanda. 6855 (88·1%) 7782 spot samples successfully genotyped. overall k13 Kagera 7·7% (90% CI 6·0-9·4; 50 649), highest districts near Rwandan (22·8% [31 136] Karagwe, 14·4% [17 118]) Kyerwa, 1·4% [two 144] Ngara). uncommon regions. Haplotype analysis suggested that some these parasites are related isolates 2015, supporting spread Arg561His. However, novel haplotype observed, potentially indicating second origin region. Other validated (one Arg622Ile two Ala675Val isolates) also identified. A prevalent dihydrofolate reductase Ile164Leu associated identified (15·2% [12·6-17·8%]; 80 526). mutant crt Lys76Thr chloroquine amodiaquine uncommon, occurring only 75 2861 genotyped isolates, whereases wild-type mdr1 Asn86Tyr allele, reduced sensitivity lumefantrine, found 99·7% (3819 3830) countrywide. These findings show is common multiple emergences ART-R, similar what seen southeast Asia, occurred. Mutations high levels common. results raise concerns about long-term efficacy antimalarials Understanding how interact drivers essential combating ART-R Africa. This funded by Bill & Melinda Gates Foundation National Institutes Health.

Language: Английский

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Sensitive, Highly Multiplexed Sequencing of Microhaplotypes From thePlasmodium falciparumHeterozygome

The Journal of Infectious Diseases, Journal Year: 2020, Volume and Issue: 225(7), P. 1227 - 1237

Published: Aug. 24, 2020

Abstract Background Targeted next-generation sequencing offers the potential for consistent, deep coverage of information-rich genomic regions to characterize polyclonal Plasmodium falciparum infections. However, methods identify and sequence these are currently limited. Methods A bioinformatic pipeline multiplex were developed simultaneously 100 targets applied dried blood spot (DBS) controls field isolates from Mozambique. For comparison, whole-genome data generated same controls. Results Using publicly available genomes, 4465 high-diversity suited targeted identified, representing P. heterozygome. this study, 93 microhaplotypes with high diversity (median expected heterozygosity = 0.7) selected along 7 drug resistance loci. The method achieved very 99%), specificity (99.8%), sensitivity (90% haplotypes 5% within sample frequency in spots parasites/µL). In silico analyses revealed that provided much higher resolution discriminate related unrelated infections than biallelic single-nucleotide polymorphism barcodes. Conclusions laboratory outlined here provide a flexible tool efficient, low-cost, high-throughput interrogation genome, can be tailored address multiple questions interest various epidemiological settings.

Language: Английский

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Malaria protection due to sickle haemoglobin depends on parasite genotype

Nature, Journal Year: 2021, Volume and Issue: 602(7895), P. 106 - 111

Published: Dec. 9, 2021

Abstract Host genetic factors can confer resistance against malaria 1 , raising the question of whether this has led to evolutionary adaptation parasite populations. Here we searched for association between candidate host and variants in 3,346 Gambian Kenyan children with severe caused by Plasmodium falciparum . We identified a strong sickle haemoglobin (HbS) three regions genome, which is not explained population structure or other covariates, replicated additional samples. The HbS-associated alleles include nonsynonymous gene acyl-CoA synthetase family member 2–4 PfACS8 on chromosome 2, second region containing structural variation 11. are linkage disequilibrium have frequencies that covary frequency HbS across populations, particular being much more common Africa than parts world. estimated protective effect malaria, as determined comparison cases controls, varies greatly according genotype at these loci. These findings open up new avenue enquiry into biological epidemiological significance polymorphisms genome forces their high African P.

Language: Английский

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Describing the current status of Plasmodium falciparum population structure and drug resistance within mainland Tanzania using molecular inversion probes

Molecular Ecology, Journal Year: 2020, Volume and Issue: 30(1), P. 100 - 113

Published: Oct. 27, 2020

High-throughput Plasmodium genomic data is increasingly useful in assessing prevalence of clinically important mutations and malaria transmission patterns. Understanding parasite diversity for identification specific human or populations that can be targeted by control programmes, to monitor the spread associated with drug resistance. An up-to-date understanding regional population dynamics also critical impact efforts. However, this largely absent from high-burden nations Africa, date, no such analysis has been conducted parasites Tanzania countrywide. To end, over 1,000 P. falciparum clinical isolates were collected 2017 13 sites seven administrative regions across Tanzania, genotyped at 1,800 variable positions genome-wide using molecular inversion probes. Population structure was detectable among Tanzanian parasites, approximately separating northern southern districts identifying genetically admixed north. Isolates nearby more likely related compared sampled distant districts. Known resistance seen increased frequency (including two infections carrying pfk13-R561H), additional variants undetermined significance antimalarial varied geography. Malaria Indicator Survey (2017) corresponded genetic findings, including average region-level complexity-of-infection estimates. The identified here provide information on extant spatial patterns which future surveys relatedness compared.

Language: Английский

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Drug susceptibility of Plasmodium falciparum in eastern Uganda: a longitudinal phenotypic and genotypic study

The Lancet Microbe, Journal Year: 2021, Volume and Issue: 2(9), P. e441 - e449

Published: June 18, 2021

Treatment and control of malaria depends on artemisinin-based combination therapies (ACTs) is challenged by drug resistance, but thus far resistance to artemisinins partner drugs has primarily occurred in southeast Asia. The aim this study was characterise antimalarial susceptibility Plasmodium falciparum isolates from Tororo Busia districts Uganda. In prospective longitudinal study, P were collected patients aged 6 months or older presenting at the District Hospital (Tororo district, a site with relatively low incidence) Masafu General (Busia high-incidence site) eastern Uganda clinical symptoms malaria, positive Giemsa-stained blood film for falciparum, no signs severe disease. Ex-vivo susceptibilities ten measured using 72-h microplate growth inhibition assay SYBR Green detection. Relevant genetic polymorphisms characterised molecular methods. We compared results those earlier studies region searched associations between parasite genotypes. From June 10, 2016, July 29, 2019, 361 district 79 440 participants. Of total isolates, 392 (89%) successfully grew culture showed excellent chloroquine (median half-maximal inhibitory concentration [IC50] 20·0 nM [IQR 12·0-26·0]), monodesethylamodiaquine (7·1 [4·3-8·9]), pyronaridine (1·1 [0·7-2·3]), piperaquine (5·6 [3·3-8·6]), ferroquine (1·8 [1·5-3·3]), AQ-13 (24·0 [17·0-32·0]), lumefantrine (5·1 [3·2-7·7]), mefloquine (9·5 [6·6-13·0]), dihydroartemisinin (1·5 [1·0-2·0]), atovaquone (0·3 [0·2-0·4]). Compared our 2010-13, significant improvements seen IC50 288·0 122·0-607·0]; p<0·0001), (76·0 [44·0-137]; (21·0 [7·6-43·0]; small decrease (3·0 [1·1-7·6]; change (1·3 [0·8-2·5]; p=0·64). Chloroquine (IC50>100 nM) more common (11 [15%] 71), (12 [4%] 320; p=0·0017). increases 2010-12 2016-19 prevalences wild-type multidrug protein 1 (PfMDR1) Asn86Tyr 60% (391 653) 99% (418 422; PfMDR1 Asp1246Tyr (390 650) 90% (371 419; transporter (PfCRT) Lys76Thr 7% (44 675) 87% (364 417; p<0·0001). Our show marked changes phenotypes genotypes during past decade. These suggest that additional will be over time continued surveillance key ACT components warranted. National Institutes Health Medicines Malaria Venture.

Language: Английский

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Multiplexed ddPCR-amplicon sequencing reveals isolated Plasmodium falciparum populations amenable to local elimination in Zanzibar, Tanzania

Nature Communications, Journal Year: 2023, Volume and Issue: 14(1)

Published: June 22, 2023

Abstract Zanzibar has made significant progress toward malaria elimination, but recent stagnation requires novel approaches. We developed a highly multiplexed droplet digital PCR (ddPCR)-based amplicon sequencing method targeting 35 microhaplotypes and drug-resistance loci, successfully sequenced 290 samples from five districts covering both main islands. Here, we elucidate fine-scale Plasmodium falciparum population structure infer relatedness connectivity of infections using an identity-by-descent (IBD) approach. Despite high genetic diversity, observe pronounced spatial temporal parasite structure. Clusters near-clonal on Pemba indicate persistent local transmission with limited importation, presenting opportunity for elimination efforts. Furthermore, admixed Unguja detect substantial fraction (2.9%) significantly related infection pairs between the mainland, suggesting importation. Our study provides high-resolution view across archipelago actionable insights prioritizing

Language: Английский

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Drug resistance and vaccine target surveillance of Plasmodium falciparum using nanopore sequencing in Ghana

Nature Microbiology, Journal Year: 2023, Volume and Issue: 8(12), P. 2365 - 2377

Published: Nov. 23, 2023

Malaria results in over 600,000 deaths annually, with the highest burden of young children living sub-Saharan Africa. Molecular surveillance can provide important information for malaria control policies, including detection antimalarial drug resistance. However, genome sequencing capacity malaria-endemic countries is limited. We designed and implemented an end-to-end workflow to detect Plasmodium falciparum resistance markers diversity vaccine target circumsporozoite protein (csp) using nanopore Ghana. analysed 196 clinical samples showed that our method rapid, robust, accurate straightforward implement. Importantly, could be applied dried blood spot samples, which are readily collected endemic settings. report P. parasites Ghana mostly susceptible chloroquine, persistent sulfadoxine-pyrimethamine no evidence artemisinin Multiple single nucleotide polymorphisms were identified csp, but their significance uncertain. Our study demonstrates feasibility genomic countries.

Language: Английский

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Using a mobile nanopore sequencing lab for end-to-end genomic surveillance of Plasmodium falciparum: A feasibility study

PLOS Global Public Health, Journal Year: 2024, Volume and Issue: 4(2), P. e0002743 - e0002743

Published: Feb. 1, 2024

Genomic epidemiology holds promise for malaria control and elimination efforts, example by informing on Plasmodium falciparum genetic diversity prevalence of mutations conferring anti-malarial drug resistance. Limited sequencing infrastructure in many malaria-endemic areas prevents the rapid generation genomic data. To address these issues, we developed validated assays P . nanopore endemic sites using a mobile laboratory, targeting key antimalarial resistance markers microhaplotypes. Using two multiplexed PCR reactions, amplified six highly polymorphic microhaplotypes ten markers. We bioinformatics workflow that allows genotyping polyclonal infections, including minority clones. panels mock dried blood spot (DBS) diagnostic test (RDT) samples archived DBS, demonstrating even, high read coverage across amplicons (range: 580x to 3,212x median coverage), haplotype calling accuracy, ability explore within-sample infections. field-tested feasibility Zanzibar close collaboration with local program DBS routinely collected RDTs as sample inputs. Our assay identified haplotypes known confer antimalarials dhfr , dhps mdr1 genes, but no evidence artemisinin partial Most infections (60%) were polyclonal, microhaplotype (median H E = 0.94). In conclusion, our generated actionable data within few days, current challenges implementing countries accelerate elimination.

Language: Английский

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Population genomic evidence of structured and connected Plasmodium vivax populations under host selection in Latin America

Ecology and Evolution, Journal Year: 2024, Volume and Issue: 14(3)

Published: March 1, 2024

Abstract Pathogen genomic epidemiology has the potential to provide a deep understanding of population dynamics, facilitating strategic planning interventions, monitoring their impact, and enabling timely responses, thereby supporting control elimination efforts parasitic tropical diseases. Plasmodium vivax , responsible for most malaria cases outside Africa, shows high genetic diversity at level, driven by factors like sub‐patent infections, hidden reservoir hypnozoites, early transmission mosquitoes. While Latin America made significant progress in controlling falciparum it faces challenges with residual P. . To characterize structure we have analyzed largest collection genomes date, including 1474 high‐quality from 31 countries across Asia, Oceania, America. globally, American isolates form distinctive population, which is further divided into sub‐populations occasional clonal pockets. Genetic within continent was associated intensity transmission. Population differentiation exists between Central North Coast South America, vs. Amazon Basin, gene flow but limited connectivity Northwest Basin. Shared regions these parasite populations indicate adaptive evolution, particularly genes related DNA replication, RNA processing, invasion, motility – crucial parasite's survival diverse environments. Understanding population‐level adaptations effective efforts, offering insights mechanisms behind drug resistance, immune evasion, dynamics.

Language: Английский

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