High frequency of Plasmodium falciparum chloroquine resistance marker (pfcrt T76 mutation) in Yemen: An urgent need to re-examine malaria drug policy

<p>Abstract</p> <p>Background</p> <p>Malaria remains a significant health problem in Yemen with <it>Plasmodium falciparum </it>being the predominant species which is responsible for 90% of the malaria cases. Despite serious concerns regarding increasing drug resistance, chloroquine is still used for the prevention and treatment of malaria in Yemen. This study was carried out to determine the prevalence of choloroquine resistance (CQR) of <it>P. falciparum </it>isolated from Yemen based on the <it>pfcrt </it>T76 mutation.</p> <p>Methods</p> <p>A cross-sectional study was carried out among 511 participants from four governorates in Yemen. Blood samples were screened using microscopic and species-specific nested PCR based on the 18S rRNA gene to detect and identify <it>Plasmodium </it>species. Blood samples positive for <it>P. falciparum </it>were used for detecting the <it>pfcrt </it>T76 mutation using nested-PCR.</p> <p>Results</p> <p>The prevalence of <it>pfcrt </it>T76 mutation was 81.5% (66 of 81 isolates). Coastal areas/foothills had higher prevalence of <it>pfcrt </it>T76 mutation compared to highland areas (90.5% <it>vs </it>71.8%) (p = 0.031). The <it>pfcrt </it>T76 mutation had a significant association with parasitaemia (p = 0.045). Univariate analysis shows a significant association of <it>pfcrt </it>T76 mutation with people aged > 10 years (OR = 9, 95% CI = 2.3 - 36.2, p = 0.001), low household income (OR = 5, 95% CI = 1.3 - 19.5, p = 0.027), no insecticide spray (OR = 3.7, 95% CI = 1.16 - 11.86, p = 0.025) and not sleeping under insecticide treated nets (ITNs) (OR = 4.8, 95% CI = 1.38 - 16.78, p = 0.01). Logistic regression model confirmed age > 10 years and low household income as predictors of <it>pfcrt </it>T76 mutation in Yemen <it>P. falciparum </it>isolates.</p> <p>Conclusions</p> <p>The high prevalence of <it>pfcrt </it>T76 mutation in Yemen could be a predictive marker for the prevalence of <it>P. falciparum </it>CQR. This finding shows the necessity for an in-vivo therapeutic efficacy test for CQ.<it> P. falciparum </it>CQR should be addressed in the national strategy to control malaria.</p

Therapeutic efficacy of chloroquine for treatment of Plasmodium vivax malaria cases in Halaba district, South Ethiopia

<p>Abstract</p> <p>Background</p> <p>Chloroquine is an anti-malarial drug being used to treat <it>Plasmodium vivax </it>malaria cases in Ethiopia. However, emergence of chloroquine resistant strains of the parasite has challenged the current efficacy of the drug. Therefore, the aim of this study was to assess the effectiveness of chloroquine against <it>P. vivax </it>strains in one of the malaria endemic areas of Ethiopia, namely Halaba district, located in South Nations and Nationalities Peoples Region (SNNPR) of South Ethiopia</p> <p>Results</p> <p>Among 87 malaria patients enrolled in the study, only 80 of them completed the 28-days follow-up. Seven of them dropped from the study for different reasons. Among those study participants that completed their follow-up, 69 were classified under the category of adequate clinical and parasitological response (ACPR). However, the remaining 11 cases were considered as under treatment failure mainly due to recurrence of parasitemia on day 7 (four patients), day 14 (six patients), and day 21 (one patient). The age of all cases of treatment failures was found to be less than 20 years. The load of parasitemia of patients with treatment failure on day of admission (4709.4/μl) was higher than day of recurrence (372.37/μl). Parasite reduction ratio (PRR) of treatment failure cases was 12.6/μl.</p> <p>Conclusion</p> <p>This report revealed the rise in treatment failure (13% [95% CI = 0.074 - 0.217]) as compared to earlier reports from Ethiopia. It signals the spreading of chloroquine resistant <it>P. vivax </it>(CRPv) strains to malaria endemic areas of Ethiopia. It is recommended that all concerned bodies should act aggressively before further expansion of the current drug resistant malaria.</p

Host candidate gene polymorphisms and clearance of drug-resistant Plasmodium falciparum parasites

Resistance to anti-malarial drugs is a widespread problem for control programmes for this devastating disease. Molecular tests are available for many anti-malarial drugs and are useful tools for the surveillance of drug resistance. However, the correlation of treatment outcome and molecular tests with particular parasite markers is not perfect, due in part to individuals who are able to clear genotypically drug-resistant parasites. This study aimed to identify molecular markers in the human genome that correlate with the clearance of malaria parasites after drug treatment, despite the drug resistance profile of the protozoan as predicted by molecular approaches

A thirteen-year analysis of Plasmodium falciparum populations reveals high conservation of the mutant pfcrt haplotype despite the withdrawal of chloroquine from national treatment guidelines in Gabon

<p>Abstract</p> <p>Background</p> <p>Chloroquine resistance (CR) decreased after the removal of chloroquine from national treatment guidelines in Malawi, Kenia and Tanzania. In this investigation the prevalence of the chloroquine resistance (CQR) conferring mutant <it>pfcrt </it>allele and its associated chromosomal haplotype were determined before and after the change in Gabonese national treatment guidelines from chloroquine (CQ) to artesunate plus amodiaquine (AQ) in 2003.</p> <p>Methods</p> <p>The prevalence of the wild type <it>pfcrt </it>allele was assessed in 144 isolates from the years 2005 - 07 by PCR fragment restriction digest and direct sequencing. For haplotype analysis of the chromosomal regions flanking the <it>pfcrt </it>locus, microsatellite analysis was done on a total of 145 isolates obtained in 1995/96 (43 isolates), 2002 (47 isolates) and 2005 - 07 (55 isolates).</p> <p>Results</p> <p>The prevalence of the mutant <it>pfcrt </it>allele decreased from 100% in the years 1995/96 and 2002 to 97% in 2005 - 07. Haplotype analysis showed that in 1995/96 79% of the isolates carried the same microsatellite alleles in a chromosomal fragment spanning 39 kb surrounding the <it>pfcrt </it>locus. In 2002 and 2005 - 07 the prevalence of this haplotype was 62% and 58%, respectively. <it>Pfcrt </it>haplotype analysis showed that all wild type alleles were CVMNK.</p> <p>Conclusion</p> <p>Four years after the withdrawal of CQ from national treatment guidelines the prevalence of the mutant <it>pfcrt </it>allele remains at 97%. The data suggest that the combination of artesunate plus AQ may result in continued selection for the mutant <it>pfcrt </it>haplotype even after discontinuance of CQ usage.</p

Molecular markers of anti-malarial drug resistance in Lahj Governorate, Yemen: baseline data and implications

<p>Abstract</p> <p>Background</p> <p>This is an investigation of anti-malarial molecular markers coupled with a therapeutic efficacy test of chloroquine (CQ) against falciparum malaria in an area of unstable malaria in Lahj Governorate, Yemen. The study was aimed at assessment of therapeutic response to CQ and elucidation of baseline information on molecular markers for <it>Plasmodium falciparum </it>resistance against CQ and sulphadoxine/pyrimethamine (SP).</p> <p>Methods</p> <p>Between 2002 and 2003 the field test was conducted according to the standard WHO protocol to evaluate the therapeutic efficacy of CQ in 124 patients with falciparum malaria in an endemic area in Lahj Governorate in Yemen. Blood samples collected during this study were analysed for <it>P. falciparum </it>chloroquine resistance transporter gene (<it>pfcrt</it>)-76 polymorphisms, mutation <it>pfcrt-</it>S163R and the antifolate resistance-associated mutations dihydrofolate reductase (<it>dhfr</it>)-C59R and dihydropteroate synthase (<it>dhps</it>)-K540E. Direct DNA sequencing of the <it>pfcrt </it>gene from three representative field samples was carried out after DNA amplification of the 13 exons of the <it>pfcrt </it>gene.</p> <p>Results</p> <p>Treatment failure was detected in 61% of the 122 cases that completed the 14-day follow-up. The prevalence of mutant <it>pfcrt </it>T76 was 98% in 112 amplified pre-treatment samples. The presence of <it>pfcrt </it>T76 was poorly predictive of <it>in vivo </it>CQ resistance (PPV = 61.8%, 95% CI = 52.7-70.9). The prevalence of <it>dhfr </it>Arg-59 mutation in 99 amplified samples was 5%, while the <it>dhps </it>Glu-540 was not detected in any of 119 amplified samples. Sequencing the <it>pfcrt </it>gene confirmed that Yemeni CQ resistant <it>P. falciparum </it>carry the old world (Asian and African) CQ resistant haplotype CVIETSESI at positions 72,73,74,75,76,220,271, 326 and 371.</p> <p>Conclusion</p> <p>This is the first study to report baseline information on the characteristics and implications of anti-malarial drug resistance markers in Yemen. It is also the first report of the haplotype associated with CQR <it>P. falciparum </it>parasites from Yemen. Mutant <it>pfcrt</it>T76 is highly prevalent but it is a poor predictor of treatment failure in the study population. The prevalence of mutation <it>dhfr</it>Arg59 is suggestive of emerging resistance to SP, which is currently a component of the recommended combination treatment of falciparum malaria in Yemen. More studies on these markers are recommended for surveillance of resistance in the study area.</p

In silico characterisation of putative Plasmodium falciparum vaccine candidates in African malaria populations.

Genetic diversity of surface exposed and stage specific Plasmodium falciparum immunogenic proteins pose a major roadblock to developing an effective malaria vaccine with broad and long-lasting immunity. We conducted a prospective genetic analysis of candidate antigens (msp1, ama1, rh5, eba175, glurp, celtos, csp, lsa3, Pfsea, trap, conserved chrom3, hyp9, hyp10, phistb, surfin8.2, and surfin14.1) for malaria vaccine development on 2375 P. falciparum sequences from 16 African countries. We described signatures of balancing selection inferred from positive values of Tajima's D for all antigens across all populations except for glurp. This could be as a result of immune selection on these antigens as positive Tajima's D values mapped to regions with putative immune epitopes. A less diverse phistb antigen was characterised with a transmembrane domain, glycophosphatidyl anchors between the N and C- terminals, and surface epitopes that could be targets of immune recognition. This study demonstrates the value of population genetic and immunoinformatic analysis for identifying and characterising new putative vaccine candidates towards improving strain transcending immunity, and vaccine efficacy across all endemic populations

Falciparum malaria and HIV-1 in hospitalized adults in Maputo, Mozambique: does HIV-infection obscure the malaria diagnosis?

<p>Abstract</p> <p>Background</p> <p>The potential impact of HIV-1 on falciparum malaria has been difficult to determine because of diagnostic problems and insufficient epidemiological data.</p> <p>Methods</p> <p>In a prospective, cross-sectional study, clinical and laboratory data was registered consecutively for all adults admitted to a medical ward in the Central Hospital of Maputo, Mozambique, during two months from 28^thOctober 2006. Risk factors for fatal outcome were analysed. The impact of HIV on the accuracy of malaria diagnosis was assessed, comparing "Presumptive malaria", a diagnosis assigned by the ward clinicians based on fever and symptoms suggestive of malaria in the absence of signs of other infections, and "Verified malaria", a malaria diagnosis that was not rejected during retrospective review of all available data.</p> <p>Results</p> <p>Among 333 included patients, fifteen percent (51/333) had "presumptive malaria", ten percent (28 of 285 tested persons) had positive malaria blood slides, while 69.1% (188/272) were HIV positive. Seven percent (n = 23) had "verified malaria", after the diagnosis was rejected in patients with neck stiffness or symptom duration longer than 2 weeks (n = 5) and persons with negative (n = 19) or unknown malaria blood slide (n = 4). Clinical stage of HIV infection (CDC), hypotension and hypoglycaemia was associated with fatal outcome. The "presumptive malaria" diagnosis was rejected more frequently in HIV positive (20/31) than in HIV negative patients (2/10, p = 0.023).</p> <p>Conclusion</p> <p>The study suggests that the fraction of febrile illness attributable to malaria is lower in HIV positive adults. HIV testing should be considered early in evaluation of patients with suspected malaria.</p

Artemisinin-based combinations versus amodiaquine plus sulphadoxine-pyrimethamine for the treatment of uncomplicated malaria in Faladje, Mali

<p>Abstract</p> <p>Background</p> <p>Because of the emergence of chloroquine resistance in Mali, artemether-lumefantrine (AL) or artesunate-amodiaquine (AS+AQ) are recommended as first-line therapy for uncomplicated malaria, but have not been available in Mali until recently because of high costs.</p> <p>Methods</p> <p>From July 2005 to January 2006, a randomized open-label trial of three oral antimalarial combinations, namely AS+AQ, artesunate plus sulphadoxine-pyrimethamine (AS+SP), and amodiaquine plus sulphadoxine-pyrimethamine (AQ+SP), was conducted in Faladje, Mali. Parasite genotyping by polymerase chain reaction (PCR) was used to distinguish new from recrudescent <it>Plasmodium falciparum </it>infections.</p> <p>Results</p> <p>397 children 6 to 59 months of age with uncomplicated <it>Plasmodium falciparum </it>malaria were enrolled, and followed for 28 days to assess treatment efficacy. Baseline characteristics were similar in all three treatment groups. The uncorrected rates of adequate clinical and parasitologic response (ACPR) were 55.7%, 90.8%, and 97.7% in AS+AQ, AS+SP, and AQ+SP respectively (p < 0.001); after PCR correction ACPR rates were similar among treatment groups: 95.4%, 96.9%, and 99.2% respectively (p = 0.17). Mean haemoglobin concentration increased across all treatment groups from Day 0 (9.82 ± 1.68 g/dL) to Day 28 (10.78 ± 1.49 g/dL) (p < 0.001), with the greatest improvement occurring in children treated with AQ+SP. On Day 2, the prevalence of parasitaemia was significantly greater among children treated with AQ+SP (50.8%) than in children treated with AS+AQ (10.5%) or AS+SP (10.8%) (p < 0.001). No significant difference in gametocyte carriage was found between groups during the follow-up period.</p> <p>Conclusion</p> <p>The combination of AQ+SP provides a potentially low cost alternative for treatment of uncomplicated <it>P. falciparum </it>infection in Mali and appears to have the added value of longer protective effect against new infection.</p

A database of antimalarial drug resistance

A large investment is required to develop, license and deploy a new antimalarial drug. Too often, that investment has been rapidly devalued by the selection of parasite populations resistant to the drug action. To understand the mechanisms of selection, detailed information on the patterns of drug use in a variety of environments, and the geographic and temporal patterns of resistance is needed. Currently, there is no publically-accessible central database that contains information on the levels of resistance to antimalaria drugs. This paper outlines the resources that are available and the steps that might be taken to create a dynamic, open access database that would include current and historical data on clinical efficacy, in vitro responses and molecular markers related to drug resistance in Plasmodium falciparum and Plasmodium vivax. The goal is to include historical and current data on resistance to commonly used drugs, like chloroquine and sulfadoxine-pyrimethamine, and on the many combinations that are now being tested in different settings. The database will be accessible to all on the Web. The information in such a database will inform optimal utilization of current drugs and sustain the longest possible therapeutic life of newly introduced drugs and combinations. The database will protect the valuable investment represented by the development and deployment of novel therapies for malaria