Artemisinin-resistant Malaria Research Articles

No evidence of amplified Plasmodium falciparum plasmepsin II gene copy number in an area with artemisinin-resistant malaria along the China\u2013Myanmar border

BackgroundThe emergence and spread of artemisinin resistance in Plasmodium falciparum poses a threat to malaria eradication, including China’s plan to eliminate malaria by 2020. Piperaquine (PPQ) resistance has emerged in Cambodia, compromising an important partner drug that is widely used in China in the form of dihydroartemisinin (DHA)-PPQ. Several mutations in a P. falciparum gene encoding a kelch protein on chromosome 13 (k13) are associated with artemisinin resistance and have arisen spread in the Great Mekong subregion, including the China–Myanmar border. Multiple copies of the plasmepsin II/III (pm2/3) genes, located on chromosome 14, have been shown to be associated with PPQ resistance.MethodsThe therapeutic efficacy of DHA-PPQ for the treatment of uncomplicated P. falciparum was evaluated along the China–Myanmar border from 2010 to 2014. The dry blood spots samples collected in the efficacy study prior DHA-PPQ treatment and from the local hospital by passive detection were used to amplify k13 and pm2. Polymorphisms within k13 were genotyped by capillary sequencing and pm2 copy number was quantified by relative-quantitative real-time polymerase chain reaction. Treatment outcome was evaluated with the World Health Organization protocol. A linear regression model was used to estimate the association between the day 3 positive rate and k13 mutation and the relationship of the pm2 copy number variants and k13 mutations.ResultsDHA-PPQ was effective for uncomplicated P. falciparum infection in Yunnan Province with cure rates > 95%. Twelve non synonymous mutations in the k13 domain were observed among the 268 samples with the prevalence of 44.0% and the predominant mutation was F446I with a prevalence of 32.8%. Only one sample was observed with multi-copies of pm2, including parasites with and without k13 mutations. The therapeutic efficacy of DHA-PPQ was > 95% along the China–Myanmar border, consistent with the lack of amplification of pm2.ConclusionDHA-PPQ for uncomplicated P. falciparum infection still showed efficacy in an area with artemisinin-resistant malaria along the China–Myanmar border. There was no evidence to show PPQ resistance by clinical study and molecular markers survey. Continued monitoring of the parasite population using molecular markers will be important to track emergence and spread of resistance in this region.

Molecular characterization of Plasmodium falciparum DNA-3-methyladenine glycosylase

BackgroundThe emergence of artemisinin-resistant malaria parasites highlights the need for novel drugs and their targets. Alkylation of purine bases can hinder DNA replication and if unresolved would eventually result in cell death. DNA-3-methyladenine glycosylase (MAG) is responsible for the repair of those alkylated bases. Plasmodium falciparum (Pf) MAG was characterized for its potential for development as an anti-malarial candidate.MethodsNative PfMAG from crude extract of chloroquine- and pyrimethamine-resistant P. falciparum K1 strain was partially purified using three chromatographic procedures. From bio-informatics analysis, primers were designed for amplification, insertion into pBAD202/D-TOPO and heterologous expression in Escherichia coli of recombinant PfMAG. Functional and biochemical properties of the recombinant enzyme were characterized.ResultsPfMAG activity was most prominent in parasite schizont stages, with a specific activity of 147 U/mg (partially purified) protein. K1 PfMAG contained an insertion of AAT (coding for asparagine) compared to 3D7 strain and 16% similarity to the human enzyme. Recombinant PfMAG (74 kDa) was twice as large as the human enzyme, preferred double-stranded DNA substrate, and demonstrated glycosylase activity over a pH range of 4–9, optimal salt concentration of 100–200 mM NaCl but reduced activity at 250 mM NaCl, no requirement for divalent cations, which were inhibitory in a dose-dependent manner.ConclusionPfMAG activity increased with parasite development being highest in the schizont stages. K1 PfMAG contained an indel AAT (asparagine) not present in 3D7 strain and the recombinant enzyme was twice as large as the human enzyme. Recombinant PfMAG had a wide range of optimal pH activity, and was inhibited at high (250 mM) NaCl concentration as well as by divalent cations. The properties of PfMAG provide basic data that should be of assistance in developing anti-malarials against this potential parasite target.

Optimal 10-Aminoartemisinins With Potent Transmission-Blocking Capabilities for New Artemisinin Combination Therapies-Activities Against Blood Stage P. falciparum Including PfKI3 C580Y Mutants and Liver Stage P. berghei Parasites.

We have demonstrated previously that amino-artemisinins including artemiside and artemisone in which an amino group replaces the oxygen-bearing substituents attached to C-10 of the current clinical artemisinin derivatives dihydroartemisinin (DHA), artemether and artesunate, display potent activities in vitro against the asexual blood stages of Plasmodium falciparum (Pf). In particular, the compounds are active against late blood stage Pf gametocytes, and are strongly synergistic in combination with the redox active drug methylene blue. In order to fortify the eventual selection of optimum amino-artemisinins for development into new triple combination therapies also active against artemisinin-resistant Pf mutants, we have prepared new amino-artemisinins based on the easily accessible and inexpensive DHA-piperazine. The latter was converted into alkyl- and aryl sulfonamides, ureas and amides. These derivatives were screened together with the comparator drugs DHA and the hitherto most active amino-artemisinins artemiside and artemisone against asexual and sexual blood stages of Pf and liver stage P. berghei (Pb) sporozoites. Several of the new amino-artemisinins bearing aryl-urea and -amide groups are potently active against both asexual, and late blood stage gametocytes (IC50 0.4-1.0 nM). Although the activities are superior to those of artemiside (IC50 1.5 nM) and artemisone (IC50 42.4 nM), the latter are more active against the liver stage Pb sporozoites (IC50 artemisone 28 nM). In addition, early results indicate these compounds tend not to display reduced susceptibility against parasites bearing the Pf Kelch 13 propeller domain C580Y mutation characteristic of artemisinin-resistant Pf. Thus, the advent of the amino-artemisinins including artemiside and artemisone will enable the development of new combination therapies that by virtue of the amino-artemisinin component itself will possess intrinsic transmission-blocking capabilities and may be effective against artemisinin resistant falciparum malaria.

Rubber plantations and drug resistant malaria: a cross-sectional survey in Cambodia

BackgroundThe ongoing spread of artemisinin resistant Plasmodium falciparum malaria is a major threat to global health. In response, countries in the Greater Mekong Sub-region, including Cambodia, have declared ambitious goals to eliminate malaria. Major challenges include the lack of information on the at-risk population-individuals who live or work in or near the forest where the malaria vectors are found, including plantation workers. This study aimed to address this knowledge gap through a cross-sectional survey conducted in rubber plantations in Cambodia in 2014.MethodsThe survey was conducted in two rounds in four provinces and included a malaria prevalence survey, analysis for the K13 genetic mutation, and a comprehensive behavioural questionnaire. Forty plantations were included in each round, and 4201 interviews were conducted. An additional 701 blood samples were collected from family members of plantation workers.ResultsOverall malaria prevalence was relatively low with adjusted PCR prevalence rate of 0.6% for P. falciparum and 0.3% for Plasmodium vivax, and was very heterogenous between plantations. There was little difference in risk between permanent residents and temporary workers, and between the two rounds. The main risk factors for P. falciparum infection were smaller plantations, age under 30 years, lack of self-reported use of a treated net and recent travel, especially to the Northeastern provinces. Proximity of plantations to the forest was also a risk factor for malaria in round one, while male gender was also a risk factor for malaria by either species.ConclusionsWith Cambodia’s P. falciparum elimination target on the horizon, identifying every single malaria case will become increasingly important. Plantations workers are relatively accessible compared to some other at-risk groups and will likely remain a high priority. Ongoing surveillance and adaptive strategies will be critical if malaria elimination is to be achieved in this setting.

Quantitative and rapid Plasmodium falciparum malaria diagnosis and artemisinin-resistance detection using a CMOS Lab-on-Chip platform

Early and accurate diagnosis of malaria and drug-resistance is essential to effective disease management. Available rapid malaria diagnostic tests present limitations in analytical sensitivity, drug-resistance testing and/or quantification. Conversely, diagnostic methods based on nucleic acid amplification stepped forwards owing to their high sensitivity, specificity and robustness. Nevertheless, these methods commonly rely on optical measurements and complex instrumentation which limit their applicability in resource-poor, point-of-care settings. This paper reports the specific, quantitative and fully-electronic detection of Plasmodium falciparum, the predominant malaria-causing parasite worldwide, using a Lab-on-Chip platform developed in-house. Furthermore, we demonstrate on-chip detection of C580Y, the most prevalent single-nucleotide polymorphism associated to artemisinin-resistant malaria. Real-time non-optical DNA sensing is facilitated using Ion-Sensitive Field-Effect Transistors, fabricated in unmodified complementary metal-oxide-semiconductor (CMOS) technology, coupled with loop-mediated isothermal amplification. This work holds significant potential for the development of a fully portable and quantitative malaria diagnostic that can be used as a rapid point-of-care test.

Declining responsiveness of childhood Plasmodium falciparum infections to artemisinin-based combination treatments ten years following deployment as first-line antimalarials in Nigeria

BackgroundThe development and spread of artemisinin-resistant Plasmodium falciparum malaria in Greater Mekong Subregion has created impetus for continuing global monitoring of efficacy of artemisinin-based combination therapies (ACTs). This post analyses is aimed to evaluate changes in early treatment response markers 10 years after the adoption of ACTs as first-line treatments of uncomplicated falciparum malaria in Nigeria.MethodsAt 14 sentinel sites in six geographical areas of Nigeria, we evaluated treatment responses in 1341 children under 5 years and in additional 360 children under 16 years with uncomplicated malaria enrolled in randomized trials of artemether-lumefantrine versus artesunate-amodiaquine at 5-year interval in 2009–2010 and 2014–2015 and at 2-year interval in 2009–2010 and 2012–2015, respectively after deployment in 2005.ResultsAsexual parasite positivity 1 day after treatment initiation (APPD1) rose from 54 to 62% and 2 days after treatment initiation from 5 to 26% in 2009–2010 to 2014–2015 (P = 0.002 and P < 0.0001, respectively). Parasite clearance time increased significantly from 1.6 days (95% confidence interval [CI]: 1.55–1.64) to 1.9 days (95% CI, 1.9–2.0) and geometric mean parasite reduction ratio 2 days after treatment initiation decreased significantly from 11 000 to 4700 within the same time period (P < 0.0001 for each). Enrolment parasitaemia > 75 000 μl− 1, haematocrit > 27% 1 day post-treatment initiation, treatment with artemether-lumefantrine and enrolment in 2014–2015 independently predicted APPD1. In parallel, Kaplan-Meier estimated risk of recurrent infections by day 28 rose from 8 to 14% (P = 0.005) and from 9 to 15% (P = 0.02) with artemether-lumefantrine and artesunate-amodiaquine, respectively. Mean asexual parasitaemia half-life increased significantly from 1.1 h to 1.3 h within 2 years (P < 0.0001).ConclusionsThese data indicate declining parasitological responses through time to the two ACTs may be due to emergence of parasites with reduced susceptibility or decrease in immunity to the infections in these children.Trial registrationPan African Clinical Trial Registration PACTR201508001188143, 3 July 2015; PACTR201508001191898, 7 July 2015 and PACTR201508001193368, 8 July 2015 PACTR201510001189370, 3 July 2015; PACTR201709002064150, 1 March 2017; https://www.pactr.samrca.ac.za

Pre-clinical study of iron oxide nanoparticles fortified artesunate for efficient targeting of malarial parasite.

BackgroundArtesunate the most potent antimalarial is widely used for the treatment of multidrug-resistant malaria. The antimalarial cytotoxicity of artesunate has been mainly attributed to its selective, irreversible and iron- radical-mediated damage of parasite biomolecules. In the present research, iron oxide nanoparticle fortified artesunate was tested in P. falciparum and in an experimental malaria mouse model for enhancement in the selectivity and toxicity of artesunate towards parasite. Artesunate was fortified with nontoxic biocompatible surface modified iron oxide nanoparticle which is specially designed and synthesized for the sustained pH-dependent release of Fe2+ within the parasitic food vacuole for enhanced ROS spurt.MethodsAntimalarial efficacy of Iron oxide nanoparticle fortified artesunate was evaluated in wild type and artemisinin-resistant Plasmodium falciparum (R539T) grown in O + ve human blood and in Plasmodium berghei ANKA infected swiss albino mice. Internalization of nanoparticles, the pH-dependent release of Fe2+, production of reactive oxygen species and parasite biomolecule damage by iron oxide nanoparticle fortified artesunate was studied using various biochemical, biophysical, ultra-structural and fluorescence microscopy. For determining the efficacy of ATA-IONP+ART on resistant parasite ring survival assay was performed.ResultsThe nanoparticle fortified artesunate was highly efficient in the 1/8th concentration of artesunate IC50 and led to retarded growth of P. falciparum with significant damage to macromolecules mediated via enhanced ROS production. Similarly, preclinical In vivo studies also signified a radical reduction in parasitemia with ~8–10-fold reduced dosage of artesunate when fortified with iron oxide nanoparticles. Importantly, the ATA-IONP combination was efficacious against artemisinin-resistant parasites.InterpretationSurface coated iron-oxide nanoparticle fortified artesunate can be developed into a potent therapeutic agent towards multidrug-resistant and artemisinin-resistant malaria in humans.FundThis study is supported by the Centre for Study of Complex Malaria in India funded by the National Institute of Health, USA.

Assessment of the risk posed to Singapore by the emergence of artemisinin-resistant malaria in the Greater Mekong Subregion.

ObjectiveTo assess the public health risk to Singapore posed by the emergence of artemisinin-resistant (ART-R) malaria in the Greater Mekong Subregion (GMS).MethodsWe assessed the likelihood of importation of drug-resistant malaria into Singapore and the impact on public health of its subsequent secondary spread in Singapore. Literature on the epidemiology and contextual factors associated with ART-R malaria was reviewed. The epidemiology of malaria cases in Singapore was analysed. The vulnerability and receptivity of Singapore were examined, including the connectivity with countries reporting ART-R malaria, as well as the preparedness of Singaporean health authorities. Sources of information include international journals, World Health Organization guidelines, data from the Singapore Ministry of Health and National Public Health Laboratory of the National Centre for Infectious Diseases, and the International Air Transport Association.ResultsThe importation of ART-R malaria into Singapore is possible given the close proximity and significant travel volume between Singapore and the GMS countries reporting artemisinin resistance. Singapore’s vulnerability is further enhanced by the presence of foreign workers from neighbouring endemic countries. Nonetheless, the overall likelihood of such an event is low based on the rarity and decreasing trend of imported malaria incidence.​With the presence of Anopheles vectors in Singapore, imported cases of drug-resistant malaria could cause secondary transmission. Nevertheless, the risk of sustained spread is likely to be mitigated by the comprehensive surveillance and control system in place for both infected vectors and human cases.DiscussionThis risk assessment highlights the need for a continued high degree of vigilance of ART-R malaria locally and globally to minimize the risk and public health impact of drug-resistant malaria in Singapore.

Assessment of the risk posed to Singapore by the emergence of artemisinin-resistant malaria

Purpose: To assess the public health risk to Singapore posed by the emergence of artemisinin-resistant malaria across the Greater Mekong Sub-region (GMS). Methods & Materials: The likelihood of importation of drug-resistant malaria into Singapore and the impact on public health of its subsequent secondary spread in Singapore were assessed to determine the overall risk. Epidemiology of malaria cases in Singapore was analysed. Vulnerability and receptivity of Singapore were examined, including the connectivity between Singapore and countries reporting artemisinin resistance (ART-R), as well as preparedness of the health authorities. Results: The importation of ART-R malaria in Singapore is possible given the close proximity and significant travel volume between Singapore and the GMS countries reporting ART-R. Singapore's vulnerability is further enhanced by its high dependency on foreign workers from neighbouring endemic countries. Nonetheless, the overall likelihood of such an event is low based on the rarity and decreasing trend of imported malaria incidence over the past few years. From 2008 to 2017, 12 out of 209 (5.7%) P. falciparum cases detected in Singapore had mutations associated with artemisinin resistance. All cases had recovered without complications. Conclusion: This risk assessment highlights the need for a high degree of vigilance over the local and global situation to be maintained to minimise the risk and severity of the public health threat of drug-resistant malaria to Singapore. Corrigendum to “Assessment of the risk posed to Singapore by the emergence of artemisinin-resistant malaria” [International Journal of Infectious Diseases 79(S1) (2019) 1–150]International Journal of Infectious DiseasesVol. 84PreviewThe authors regret an error to the Authorship list, which should appear as above. The authors apologise for any inconvenience caused. Full-Text PDF Open Access

Novel pfkelch13 Gene Polymorphism Associates With Artemisinin Resistance in Eastern India.

Artesunate-sulfadoxine-pyrimethamine (ASSP) is the frontline artemisinin combination therapy (ACT) in India. Random, irrational, subtherapeutic artemisinin doses and self-medication with ACT along with predominance of sulfadoxine-pyrimethamine resistance parasite invoked a strong possibility of emerging artemisinin-resistant malaria parasites. This study involved 226 patients with uncomplicated Plasmodium falciparum infection who had successfully completed the 42 days follow-up after ASSP combination therapy from April 2014 to January 2016. We assessed the ASSP treatment efficacy by evaluating parasite clearance half-life, pfkelch13, and other (pfdhfr, pfdhps, pfmdr1, pfcrt) gene mutations and survival of parasites as detected by an ex vivo ring-stage survival assay (RSA). Slow-clearing infections with longer parasite clearance half-lives (>5 hours) were observed in 12% isolates. Cure rate after ASSP treatment was declining to about 84.1%. ASSP failure was recorded in 15.9% (early treatment failure, 7.9%; late treatment failure, 7.9%) of isolates. In sum, 24 patients (10.6%) had parasite clearance half-lives greater than 5 hours with pfkelch13 polymorphism after 441 codon; in 15 of those patients (6.6%), parasites had not cleared by 72 hours after initiation of therapy. Median ex vivo ring-stage survival rate of these isolates was very high (12.2%; 95% confidence interval [CI], 10.9-13.8) from that of cured patients (0.9%; 95% CI, 0.09-1.07). Of these 15 patients, 13 patients had pfkelch13 G625R polymorphism, whereas 2 patients contained R539T polymorphism. As per the World Health Organization guideline, these 15 isolates were true artemisinin-resistant isolates. Identification of artemisinin-resistant isolates in India together with new mutations and increasing combination therapy failures blow alarms for urgent malaria control.

Heterogeneous distribution of k13 mutations in Plasmodium falciparum in Laos

BackgroundThe emergence and transnational spread of artemisinin resistance in Plasmodium falciparum in the Greater Mekong Sub-region (GMS) is a serious threat to malaria elimination in the region and could present a threat to malaria control in Africa. Recently, the Lao Government adopted the goal of malaria elimination by 2030, for which monitoring of artemisinin-resistant malaria within the country is indispensable. This study’s objectives were to assess the distribution of k13 mutations in Laos.MethodsPlasmodium falciparum isolates (n = 1151) were collected from five southern provinces in Laos between 2015 and 2016, and three isolates from the northernmost province bordering China in 2017. Polymorphisms of the k13 gene and two flanking regions were analysed to estimate relationship among the isolates.ResultsIn the five southern provinces, overall 55.5% of the isolates possessed artemisinin-resistant mutations of the k13 gene (C580Y, P574L, R539T, Y493H). The C580Y was the predominant mutation (87.2%). The frequencies of the k13 mutations were heterogeneous in the five southern provinces, but with a clear tendency showing the highest frequency in the south (72.5%) and to a lower degree when moving northward (28.0%). The three isolates from the Lao–Chinese border also possessed the C580Y mutation. Analysis of the flanking loci demonstrated that these three isolates were genetically very close to resistant strains originating from western Cambodia.ConclusionsArtemisinin resistance was observed to be rapidly increasing and spreading northwards through Laos and has now reached the Chinese border. The Lao and Chinese governments, as well as the international community, should make dedicated efforts to contain the spread of k13 mutations within Laos and in the GMS.

Implications of population-level immunity for the emergence of artemisinin-resistant malaria: a mathematical model

BackgroundArtemisinin-resistant Plasmodium falciparum has emerged in the Greater Mekong Subregion, an area of relatively low transmission, but has yet to be reported in Africa. A population-based mathematical model was used to investigate the relationship between P. falciparum prevalence, exposure-acquired immunity and time-to-emergence of artemisinin resistance. The possible implication for the emergence of resistance across Africa was assessed.MethodsThe model included human and mosquito populations, two strains of malaria (“wild-type”, “mutant”), three levels of human exposure-acquired immunity (none, low, high) with two types of immunity for each level (sporozoite/liver stage immunity and blood-stage/gametocyte immunity) and drug pressure based on per-capita treatment numbers.ResultsThe model predicted that artemisinin-resistant strains may circulate up to 10 years longer in high compared to low P. falciparum prevalence areas before resistance is confirmed. Decreased time-to-resistance in low prevalence areas was explained by low genetic diversity and immunity, which resulted in increased probability of selection and spread of artemisinin-resistant strains. Artemisinin resistance was estimated to be established by 2020 in areas of Africa with low (< 10%) P. falciparum prevalence, but not for 5 or 10 years later in moderate (10–25%) or high (> 25%) prevalence areas, respectively.ConclusionsAreas of low transmission and low immunity give rise to a more rapid expansion of artemisinin-resistant parasites, corroborating historical observations of anti-malarial resistance emergence. Populations where control strategies are in place that reduce malaria transmission, and hence immunity, may be prone to a rapid emergence and spread of artemisinin-resistant strains and thus should be carefully monitored.

Smartphones for community health in rural Cambodia: A feasibility study

Background: Village Malaria Workers (VMWs) are lay people trained to provide a valuable role in frontline testing and treatment of malaria in rural villages in Cambodia. Emergence of artemisinin-resistant malaria highlights the essential role of such VMWs in surveillance and early treatment of malaria. Smartphone technology offers huge potential to support VMWs in isolated and resource-poor settings. Methods: We investigated the feasibility of issuing established VMWs with a smartphone, bespoke Android application and solar charger to support their role. 27 VMWs in Kampong Cham and Kratie provinces participated. Results: 26/27 of the smartphones deployed were working well at study completion twelve months later. Interviews with VMWs using quantitative and qualitative methods revealed pride, ease of use and reports of faster communication with the smartphone. VMWs also expressed a strong wish to help people presenting with non-malarial fever, for which further potential supportive smartphone applications are increasingly available. Conclusions: As a result of this pilot study, two smartphone based reporting systems for malaria have been developed at the Cambodian National Malaria Center, and the programme is now being extended nationwide. The full code for the smartphone application is made available to other researchers and healthcare providers with this article. Smartphones represent a feasible platform for developing the VMW role to include other health conditions, thus maintaining the relevance of these important community health workers.

Smartphones for community health in rural Cambodia: A feasibility study.

Background: Village Malaria Workers (VMWs) are lay people trained to provide a valuable role in frontline testing and treatment of malaria in rural villages in Cambodia. Emergence of artemisinin-resistant malaria highlights the essential role of such VMWs in surveillance and early treatment of malaria. Smartphone technology offers huge potential to support VMWs in isolated and resource-poor settings. Methods: We investigated the feasibility of issuing established VMWs with a smartphone, bespoke Android application and solar charger to support their role. 27 VMWs in Kampong Cham and Kratie provinces participated. Results: 26/27 of the smartphones deployed were working well at study completion twelve months later. Interviews with VMWs using quantitative and qualitative methods revealed pride, ease of use and reports of faster communication with the smartphone. VMWs also expressed a strong wish to help people presenting with non-malarial fever, for which further potential supportive smartphone applications are increasingly available. Conclusions: As a result of this pilot study, two smartphone based reporting systems for malaria have been developed at the Cambodian National Malaria Center, and the programme is now being extended nationwide. The full code for the smartphone application is made available to other researchers and healthcare providers with this article. Smartphones represent a feasible platform for developing the VMW role to include other health conditions, thus maintaining the relevance of these important community health workers.

How "resistant" is artemisinin resistant malaria? - The risks of ambiguity using the term "resistant" malaria.

In scientific discourse, few would consider the widely used term resistance as ambiguous. The definition and usage of the term antimicrobial resistance revolves around the concept that microorganisms change in ways that render antimicrobial medications clinically ineffective. Because artemisinins have become the cornerstone for antimalarial therapy, the widely used term artemisinin resistance in scientific literature is highly alarming. Naturally, many people will assume that artemisinin resistance must essentially be the same as antimicrobial resistance, which means it is clinically ineffective. However, this is incorrect, and the WHO defines artemisinin resistance differently to antimicrobial resistance as "partial/relative resistance". This means that parasite clearance times are increased but does not automatically mean that artemisinins have become clinically inefficacious. Is the ambiguous use of the term resistance justified and appropriate, although it might be misleading biomedical researchers, the media, policy makers and possibly attending physicians? Science is also about clear and unambiguous use of terminology, so that a message is accurately communicated and understood. Ambiguity can lead to misunderstandings, and misunderstandings can cause wrong actions; unnecessarily so.

How to combat emerging artemisinin resistance: Lessons from "The Three Little Pigs".

It is rare to come across an Aesop’s fable in respectable journals. It might catch scientists outside the malaria field by surprise to learn that the famous story of “The Boy Who Cried Wolf” has been repeatedly compared to the threat from artemisinin-resistant malaria parasites, including the two latest reports on the rise of a specific haplotype in Cambodia and Thailand, sensationally dubbed “Super Malaria” by the media [1, 2]. The comparison to a children’s tale should not negate the fact that malaria drug resistance is one of the most pressing threats to the global public health community. Here, the findings leading to this contentious discourse will be delineated in order to provide a perspective. Possible solutions will be presented to stimulate further research and discussion to solve one of the greatest public health challenges of our lifetime.

Assessment of household ownership of bed nets in areas with and without artemisinin resistance containment measures in Myanmar

BackgroundMyanmar lies in the Greater Mekong Subregion where there is artemisinin-resistant Plasmodium falciparum malaria. As the artemisinin compound is the pillar of effective antimalarial therapies, containing the spread of artemisinin resistance is a national and global priority. The use of insecticide-treated bed nets/long-lasting insecticidal nets (ITNs/LLINs) is the key intervention for ensuring the reduction of malaria transmission and the spread of resistant strains, and for eventually eliminating malaria. This study aimed at assessing household ownership of, access to, and utilization of bed nets in areas of Myanmar with and without artemisinin resistance containment measures.MethodsSecondary data from a nationwide community-based malaria survey conducted by the National Malaria Control Program in 2014 were analyzed. Based on evidence of artemisinin resistance, Myanmar was divided into tiers 1, 2, and 3: townships in tiers 1 and 2 were aggregated as the Myanmar Artemisinin Resistance Containment (MARC) areas and were compared with tier 3 townships, which were defined as non-MARC areas. The chi-square test was used to compare groups, and the level of significance was set at P ≤ 0.05.ResultsOf the 6328 households assessed, 97.2% in both MARC and non-MARC areas had at least one bed net (any type), but only 63% of households had ITNs/LLINs. Only 44% of households in MARC areas and 24% in non-MARC areas had adequate numbers of ITNs/LLINs (one ITN/LLIN per two persons, P < 0.001). Nearly 44% of household members had access to ITNs/LLINs. Regarding the utilization of ITNs/LLINs, 45% of household members used them in MARC areas and 36% used them in non-MARC areas (P < 0.001, desired target = 100%). Utilization of ITNs/LLINs among children aged below five years and pregnant women (high malaria risk groups) was low, at 44% and 42%, respectively.ConclusionsThis study highlights the nationwide shortfalls in the ownership of, access to, and utilization of ITNs/LLINs in Myanmar, which is of particular concern in terms of containing the spread of artemisinin resistance. It highlights the need for priority attention to be paid and mobilization of resources in order to improve bed net coverage and utilization through bed net distribution and/or social marketing, information dissemination, and awareness-raising.

Population pharmacokinetic and pharmacodynamic properties of artesunate in patients with artemisinin sensitive and resistant infections in Southern Myanmar

BackgroundArtemisinins are the most effective anti-malarial drugs for uncomplicated and severe Plasmodium falciparum malaria. However, widespread artemisinin resistance in the Greater Mekong Region of Southeast Asia is threatening the possibility to control and eliminate malaria. This work aimed to evaluate the pharmacokinetic and pharmacodynamic properties of artesunate and its active metabolite, dihydroartemisinin, in patients with sensitive and resistant falciparum infections in Southern Myanmar. In addition, a simple nomogram previously developed to identify artemisinin resistant malaria infections was evaluated.MethodsFifty-three (n = 53) patients were recruited and received daily oral artesunate monotherapy (4 mg/kg) for 7 days. Frequent artesunate and dihydroartemisinin plasma concentration measurements and parasite microscopy counts were obtained and evaluated using nonlinear mixed-effects modelling.ResultsThe absorption of artesunate was best characterized by a transit-compartment (n = 3) model, followed by one-compartment disposition models for artesunate and dihydroartemisinin. The drug-dependent parasite killing effect of dihydroartemisinin was described using an Emax function, with a mixture model discriminating between artemisinin sensitive and resistant parasites. Overall, 56% of the studied population was predicted to have resistant malaria infections. Application of the proposed nomogram to identify artemisinin-resistant malaria infections demonstrated an overall sensitivity of 90% compared to 55% with the traditional day-3 positivity test.ConclusionThe pharmacokinetic-pharmacodynamic properties of artesunate and dihydroartemisinin were well-characterized with a mixture model to differentiate between drug sensitive and resistant infections in these patients. More than half of all patients recruited in this study had artemisinin-resistant infections. The relatively high sensitivity of the proposed nomogram highlights its potential clinical usefulness.

Adenovirus-prime and baculovirus-boost heterologous immunization achieves sterile protection against malaria sporozoite challenge in a murine model

With the increasing prevalence of artemisinin-resistant malaria parasites, a highly efficacious and durable vaccine for malaria is urgently required. We have developed an experimental virus-vectored vaccine platform based on an envelope-modified baculovirus dual-expression system (emBDES). Here, we show a conceptually new vaccine platform based on an adenovirus-prime/emBDES-boost heterologous immunization regimen expressing the Plasmodium falciparum circumsporozoite protein (PfCSP). A human adenovirus 5-prime/emBDES-boost heterologous immunization regimen consistently achieved higher sterile protection against transgenic P. berghei sporozoites expressing PfCSP after a mosquito-bite challenge than reverse-ordered or homologous immunization. This high protective efficacy was also achieved with a chimpanzee adenovirus 63-prime/emBDES-boost heterologous immunization regimen against an intravenous sporozoite challenge. Thus, we show that the adenovirus-prime/emBDES-boost heterologous immunization regimen confers sterile protection against sporozoite challenge by two individual routes, providing a promising new malaria vaccine platform for future clinical use.

A Novel Single-Nucleotide Polymorphism Loop Mediated Isothermal Amplification Assay for Detection of Artemisinin-Resistant Plasmodium falciparum Malaria.

BackgroundArtemisinin-resistant malaria (ARM) remains a significant threat to malaria elimination. In the Greater Mekong subregion, the prevalence of ARM in certain regions has reached greater than 90%. Artemisinin-resistant malaria is clinically identified by delayed parasite clearance and has been associated with mutations in the propeller domain of the kelch 13 gene. C580Y is the most prevalent mutation. The detection of ARM currently relies on labor-intensive and time-consuming methods such as clinical phenotyping or in vitro susceptibility testing.MethodsWe developed a novel single-nucleotide polymorphism loop mediated isothermal amplification (SNP-LAMP) test method for the detection of the C580Y mutation using a novel primer design strategy.ResultsThe SNP-LAMP was 90.0% sensitive (95% confidence interval [CI], 66.9–98.3) and 91.9% specific (95% CI, 82.6–96.7) without knowledge of the parasite load and was 100% sensitive (95% CI, 79.9–100) and 97.3% specific (95% CI, 89.7–99.5) when the parasitemia was within the assay dynamic range. Tests with potential application near-to-patient such as SNP-LAMP may be deployed in low- and middle-income and developed countries.ConclusionsSingle-nucleotide polymorphism LAMP can serve as a surveillance tool and guide treatment algorithms for ARM in a clinically relevant time frame, prevent unnecessary use of additional drugs that may drive additional resistance, and avoid longer treatment regimens that cause toxicity for the patient.