Underpinning Sustainable Vector Control through Informed Insecticide Resistance Management

Background: There has been rapid scale-up of malaria vector control in the last ten years. Both of the primary control strategies, long-lasting pyrethroid treated nets and indoor residual spraying, rely on the use of a limited number of insecticides. Insecticide resistance, as measured by bioassay, has rapidly increased in prevalence and has come to the forefront as an issue that needs to be addressed to maintain the sustainability of malaria control and the drive to elimination. Zambia’s programme reported high levels of resistance to the insecticides it used in 2010, and, as a result, increased its investment in resistance monitoring to support informed resistance management decisions. Methodology/Principal Findings: A country-wide survey on insecticide resistance in Zambian malaria vectors was performed using WHO bioassays to detect resistant phenotypes. Molecular techniques were used to detect target-site mutations and microarray to detect metabolic resistance mechanisms. Anopheles gambiae s.s. was resistant to pyrethroids,DDT and carbamates, with potential organophosphate resistance in one population. The resistant phenotypes were conferred by both target-site and metabolic mechanisms. Anopheles funestus s.s. was largely resistant to pyrethroids and carbamates, with potential resistance to DDT in two locations. The resistant phenotypes were conferred by elevated levels of cytochrome p450s. Conclusions/Significance: Currently, the Zambia National Malaria Control Centre is using these results to inform their vector control strategy. The methods employed here can serve as a template to all malaria-endemic countries striving to create a sustainable insecticide resistance management pla

Failure of artesunate-mefloquine combination therapy for uncomplicated Plasmodium falciparum malaria in southern Cambodia

<p>Abstract</p> <p>Background</p> <p>Resistance to anti-malarial drugs hampers control efforts and increases the risk of morbidity and mortality from malaria. The efficacy of standard therapies for uncomplicated <it>Plasmodium falciparum </it>and <it>Plasmodium vivax </it>malaria was assessed in Chumkiri, Kampot Province, Cambodia.</p> <p>Methods</p> <p>One hundred fifty-one subjects with uncomplicated falciparum malaria received directly observed therapy with 12 mg/kg artesunate (over three days) and 25 mg/kg mefloquine, up to a maximum dose of 600 mg artesunate/1,000 mg mefloquine. One hundred nine subjects with uncomplicated vivax malaria received a total of 25 mg/kg chloroquine, up to a maximum dose of 1,500 mg, over three days. Subjects were followed for 42 days or until recurrent parasitaemia was observed. For <it>P. falciparum </it>infected subjects, PCR genotyping of <it>msp1</it>, <it>msp2</it>, and <it>glurp </it>was used to distinguish treatment failures from new infections. Treatment failure rates at days 28 and 42 were analyzed using both per protocol and Kaplan-Meier survival analysis. Real Time PCR was used to measure the copy number of the <it>pfmdr1 </it>gene and standard 48-hour isotopic hypoxanthine incorporation assays were used to measure IC₅₀for anti-malarial drugs.</p> <p>Results</p> <p>Among <it>P. falciparum </it>infected subjects, 47.0% were still parasitemic on day 2 and 11.3% on day 3. The PCR corrected treatment failure rates determined by survival analysis at 28 and 42 days were 13.1% and 18.8%, respectively. Treatment failure was associated with increased <it>pfmdr1 </it>copy number, higher initial parasitaemia, higher mefloquine IC₅₀, and longer time to parasite clearance. One <it>P. falciparum </it>isolate, from a treatment failure, had markedly elevated IC₅₀for both mefloquine (130 nM) and artesunate (6.7 nM). Among <it>P. vivax </it>infected subjects, 42.1% suffered recurrent <it>P. vivax </it>parasitaemia. None acquired new <it>P. falciparum </it>infection.</p> <p>Conclusion</p> <p>The results suggest that artesunate-mefloquine combination therapy is beginning to fail in southern Cambodia and that resistance is not confined to the provinces at the Thai-Cambodian border. It is unclear whether the treatment failures are due solely to mefloquine resistance or to artesunate resistance as well. The findings of delayed clearance times and elevated artesunate IC₅₀suggest that artesunate resistance may be emerging on a background of mefloquine resistance.</p

Genetic Determination and Linkage Mapping of Plasmodium falciparum Malaria Related Traits in Senegal

Plasmodium falciparum malaria episodes may vary considerably in their severity and clinical manifestations. There is good evidence that host genetic factors contribute to this variability. To date, most genetic studies aiming at the identification of these genes have used a case/control study design for severe malaria, exploring specific candidate genes. Here, we performed a family-based genetic study of falciparum malaria related phenotypes in two independent longitudinal survey cohorts, as a first step towards the identification of genes and mechanisms involved in the outcome of infection. We studied two Senegalese villages, Dielmo and Ndiop that differ in ethnicity, malaria transmission and endemicity. We performed genome-scan linkage analysis of several malaria-related phenotypes both during clinical attacks and asymptomatic infection. We show evidence for a strong genetic contribution to both the number of clinical falciparum malaria attacks and the asymptomatic parasite density. The asymptomatic parasite density showed linkage to chromosome 5q31 (LOD = 2.26, empirical p = 0.0014, Dielmo), confirming previous findings in other studies. Suggestive linkage values were also obtained at three additional chromosome regions: the number of clinical malaria attacks on chromosome 5p15 (LOD = 2.57, empirical p = 0.001, Dielmo) and 13q13 (LOD = 2.37, empirical p = 0.0014 Dielmo), and the maximum parasite density during asymptomatic infection on chromosome 12q21 (LOD = 3.1, empirical p<10−4, Ndiop). While regions of linkage show little overlap with genes known to be involved in severe malaria, the four regions appear to overlap with regions linked to asthma or atopy related traits, suggesting that common immune related pathways may be involved

Opsonising antibodies to P. falciparum Merozoites associated with immunity to clinical malaria

Naturally acquired humoral immunity to the malarial parasite Plasmodium falciparum can protect against disease, although the precise mechanisms remain unclear. Although antibody levels can be measured by ELISA, few studies have investigated functional antibody assays in relation to clinical outcomes. In this study we applied a recently developed functional assay of antibody-mediated opsonisation of merozoites, to plasma samples from a longitudinal cohort study conducted in a malaria endemic region of Papua New Guinea (PNG). Phagocytic activity was quantified by flow cytometry using a standardized and high-throughput protocol, and was subsequently evaluated for association with protection from clinical malaria and high-density parasitemia. Opsonising antibody responses were found to: i) increase with age, ii) be enhanced by concurrent infection, and iii) correlate with protection from clinical episodes and high-density parasitemia. Stronger protective associations were observed in individuals with no detectable parasitemia at baseline. This study presents the first evidence for merozoite phagocytosis as a correlate of acquired immunity and clinical protection against P. falciparum malaria

Diversity of Plasmodium falciparum Chloroquine Resistance Transporter (pfcrt) Exon 2 Haplotypes in the Pacific from 1959 to 1979

Nearly one million deaths are attributed to malaria every year. Recent reports of multi-drug treatment failure of falciparum malaria underscore the need to understand the molecular basis of drug resistance. Multiple mutations in the Plasmodium falciparum chloroquine resistance transporter (pfcrt) are involved in chloroquine resistance, but the evolution of complex haplotypes is not yet well understood. Using over 4,500 archival human serum specimens collected from 19 Pacific populations between 1959 and 1979, the period including and just prior to the appearance of chloroquine treatment failure in the Pacific, we PCR-amplified and sequenced a portion of the pfcrt exon 2 from 771 P. falciparum-infected individuals to explore the spatial and temporal variation in falciparum malaria prevalence and the evolution of chloroquine resistance. In the Pacific, the prevalence of P. falciparum varied considerably across ecological zones. On the island of New Guinea, the decreases in prevalence of P. falciparum in coastal, high-transmission areas over time were contrasted by the increase in prevalence during the same period in the highlands, where transmission was intermittent. We found 78 unique pfcrt haplotypes consisting of 34 amino acid substitutions and 28 synonymous mutations. More importantly, two pfcrt mutations (N75D and K76T) implicated in chloroquine resistance were present in parasites from New Hebrides (now Vanuatu) eight years before the first report of treatment failure. Our results also revealed unexpectedly high levels of genetic diversity in pfcrt exon 2 prior to the historical chloroquine resistance selective sweep, particularly in areas where disease burden was relatively low. In the Pacific, parasite genetic isolation, as well as host acquired immune status and genetic resistance to malaria, were important contributors to the evolution of chloroquine resistance in P. falciparum

Vivax malaria in Mauritania includes infection of a Duffy-negative individual

<p>Abstract</p> <p>Background</p> <p>Duffy blood group polymorphisms are important in areas where <it>Plasmodium vivax </it>is present because this surface antigen is thought to act as a key receptor for this parasite. In the present study, Duffy blood group genotyping was performed in febrile uninfected and <it>P. vivax</it>-infected patients living in the city of Nouakchott, Mauritania.</p> <p>Methods</p> <p><it>Plasmodium vivax </it>was identified by real-time PCR. The Duffy blood group genotypes were determined by standard PCR followed by sequencing of the promoter region and exon 2 of the Duffy gene in 277 febrile individuals. Fisher's exact test was performed in order to assess the significance of variables.</p> <p>Results</p> <p>In the Moorish population, a high frequency of the <it>FYB^ES/FYB^ES</it>genotype was observed in uninfected individuals (27.8%), whereas no <it>P. vivax</it>-infected patient had this genotype. This was followed by a high level of <it>FYA/FYB</it>, <it>FYB/FYB</it>, <it>FYB/FYB^ES</it>and <it>FYA/FYB^ES</it>genotype frequencies, both in the <it>P. vivax</it>-infected and uninfected patients. In other ethnic groups (Poular, Soninke, Wolof), only the <it>FYB^ES/FYB^ES</it>genotype was found in uninfected patients, whereas the <it>FYA/FYB^ES</it>genotype was observed in two <it>P. vivax</it>-infected patients. In addition, one patient belonging to the Wolof ethnic group presented the <it>FYB^ES/FYB^ES</it>genotype and was infected by <it>P. vivax</it>.</p> <p>Conclusions</p> <p>This study presents the Duffy blood group polymorphisms in Nouakchott City and demonstrates that in Mauritania, <it>P. vivax </it>is able to infect Duffy-negative patients. Further studies are necessary to identify the process that enables this Duffy-independent <it>P. vivax </it>invasion of human red blood cells.</p