The usefulness of rapid diagnostic tests in the new context of low malaria transmission in zanzibar.

BACKGROUND\ud \ud We assessed if histidine-rich-protein-2 (HRP2) based rapid diagnostic test (RDT) remains an efficient tool for Plasmodium falciparum case detection among fever patients in Zanzibar and if primary health care workers continue to adhere to RDT results in the new epidemiological context of low malaria transmission. Further, we evaluated the performance of RDT within the newly adopted integrated management of childhood illness (IMCI) algorithm in Zanzibar.\ud \ud METHODS AND FINDINGS\ud \ud We enrolled 3890 patients aged ≥2 months with uncomplicated febrile illness in this health facility based observational study conducted in 12 primary health care facilities in Zanzibar, between May-July 2010. One patient had an inconclusive RDT result. Overall 121/3889 (3.1%) patients were RDT positive. The highest RDT positivity rate, 32/528 (6.1%), was found in children aged 5-14 years. RDT sensitivity and specificity against PCR was 76.5% (95% CI 69.0-83.9%) and 99.9% (95% CI 99.7-100%), and against blood smear microscopy 78.6% (95% CI 70.8-85.1%) and 99.7% (95% CI 99.6-99.9%), respectively. All RDT positive, but only 3/3768 RDT negative patients received anti-malarial treatment. Adherence to RDT results was thus 3887/3889 (99.9%). RDT performed well in the IMCI algorithm with equally high adherence among children <5 years as compared with other age groups.\ud \ud CONCLUSIONS\ud \ud The sensitivity of HRP-2 based RDT in the hands of health care workers compared with both PCR and microscopy for P. falciparum case detection was relatively low, whereas adherence to test results with anti-malarial treatment was excellent. Moreover, the results provide evidence that RDT can be reliably integrated in IMCI as a tool for improved childhood fever management. However, the relatively low RDT sensitivity highlights the need for improved quality control of RDT use in primary health care facilities, but also for more sensitive point-of-care malaria diagnostic tools in the new epidemiological context of low malaria transmission in Zanzibar.\ud \ud TRIAL REGISTRATION\ud \ud ClinicalTrials.gov NCT01002066

Rapid diagnostic tests for molecular surveillance of Plasmodium falciparum malaria -assessment of DNA extraction methods and field applicability

Background: The need for new malaria surveillance tools and strategies is critical, given improved global malaria control and regional elimination efforts. High quality Plasmodium falciparum DNA can reliably be extracted from malaria rapid diagnostic tests (RDTs). Together with highly sensitive molecular assays, wide scale collection of used RDTs may serve as a modern tool for improved malaria case detection and drug resistance surveillance. However, comparative studies of DNA extraction efficiency from RDTs and the field applicability are lacking. The aim of this study was to compare and evaluate different methods of DNA extraction from RDTs and to test the field applicability for the purpose of molecular epidemiological investigations. Methods: DNA was extracted from two RDT devices (Paracheck-PfW and SD Bioline Malaria Pf/Pan (R)), seeded in vitro with 10-fold dilutions of cultured 3D7 P. falciparum parasites diluted in malaria negative whole blood. The level of P. falciparum detection was determined for each extraction method and RDT device with multiple nested-PCR and real-time PCR assays. The field applicability was tested on 855 paired RDT (Paracheck-Pf) and filter paper (Whatman (R) 3MM) blood samples (734 RDT negative and 121 RDT positive samples) collected from febrile patients in Zanzibar 2010. RDT positive samples were genotyped at four key single nucleotide polymorphisms (SNPs) in pfmdr1 and pfcrt as well as for pfmdr1 copy number, all associated with anti-malarial drug resistance. Results: The P. falciparum DNA detection limit varied with RDT device and extraction method. Chelex-100 extraction performed best for all extraction matrixes. There was no statistically significant difference in PCR detection rates in DNA extracted from RDTs and filter paper field samples. Similarly there were no significant differences in the PCR success rates and genotyping outcomes for the respective SNPs in the 121 RDT positive samples. Conclusions: The results support RDTs as a valuable source of parasite DNA and provide evidence for RDT-DNA extraction for improved malaria case detection, molecular drug resistance surveillance, and RDT quality control.ACT Consortium through Bill and Melinda Gates Foundation; Swedish International Development Agency (SIDA) [SWE 2009-193]; Swedish Civil Contingencies Agency (MSB) [2010-7991]; Swedish Medical Research Council (VR) [2009-3785]; Goljes Foundationinfo:eu-repo/semantics/publishedVersio

Temporal trends of molecular markers associated with artemether- lumefantrine tolerance/resistance in Bagamoyo district, Tanzania

Background: Development and spread of Plasmodium falciparum resistance to artemisinin-based combination therapy (ACT) constitutes a major threat to recent global malaria control achievements. Surveillance of molecular markers could act as an early warning system of ACT-resistance before clinical treatment failures are apparent. The aim of this study was to analyse temporal trends of established genotypes associated with artemether-lumefantrine tolerance/resistance before and after its deployment as first-line treatment for uncomplicated malaria in Tanzania 2006. Methods: Single nucleotide polymorphisms in the P. falciparum multidrug resistance gene 1 (pfmdr1) N86Y, Y184F, D1246Y and P. falciparum chloroquine transporter gene (pfcrt) K76T were analysed from dried blood spots collected during six consecutive studies from children with uncomplicated P. falciparum malaria in Fukayosi village, Bagamoyo District, Tanzania, between 2004-2011. Results: There was a statistically significant yearly increase of pfmdr1 N86, 184F, D1246 and pfcrt K76 between 2006-2011 from 14% to 61% (yearly OR = 1.38 [95% CI 1.25-1.52] p \u3c 0.0001), 14% to 35% (OR = 1.17 [95% CI 1.07-1.30] p = 0.001), 54% to 85% (OR = 1.21 [95% CI 1.03-1.42] p = 0.016) and 49% to 85% (OR = 1.33 [95% CI 1.17-1.51] p \u3c 0.0001), respectively. Unlike for the pfmdr1 SNP, a significant increase of pfcrt K76 was observed already between 2004-2006, from 26% to 49% (OR = 1.68 [95% CI 1.17-2.40] p = 0.005). From 2006 to 2011 the pfmdr1 NFD haplotype increased from 10% to 37% (OR = 1.25 [95% CI 1.12-1.39] p \u3c 0.0001), whereas the YYY haplotype decreased from 31% to 6% (OR = 0.73 [95% CI 0.56-0.98] p = 0.018). All 390 successfully analysed samples had one copy of the pfmdr1 gene. Conclusion: The temporal selection of molecular markers associated with artemether-lumefantrine tolerance/resistance may represent an early warning sign of impaired future drug efficacy. This calls for stringent surveillance of artemether-lumefantrine efficacy in Tanzania and emphasizes the importance of molecular surveillance as a complement to standard in vivo trials. © 2013 Malmberg et al.; licensee BioMed Central Ltd

The Inviscid Limit and Boundary Layers for Navier-Stokes Flows

The validity of the vanishing viscosity limit, that is, whether solutions of the Navier-Stokes equations modeling viscous incompressible flows converge to solutions of the Euler equations modeling inviscid incompressible flows as viscosity approaches zero, is one of the most fundamental issues in mathematical fluid mechanics. The problem is classified into two categories: the case when the physical boundary is absent, and the case when the physical boundary is present and the effect of the boundary layer becomes significant. The aim of this article is to review recent progress on the mathematical analysis of this problem in each category.Comment: To appear in "Handbook of Mathematical Analysis in Mechanics of Viscous Fluids", Y. Giga and A. Novotn\'y Ed., Springer. The final publication is available at http://www.springerlink.co

Discovering and linking public omics data sets using the Omics Discovery Index.

Biomedical data are being produced at an unprecedented rate owing to the falling cost of experiments and wider access to genomics, transcriptomics, proteomics and metabolomics platforms1, 2. As a result, public deposition of omics data is on the increase. This presents new challenges, including finding ways to store, organize and access different types of biomedical data stored on different platforms. Here, we present the Omics Discovery Index (OmicsDI; http://www.omicsdi.org), an open-source platform that enables access, discovery and dissemination of omics data sets

Cellular characterisation of the GCKR P446L variant associated with type 2 diabetes risk

Aims/hypothesis Translation of genetic association signals into molecular mechanisms for diabetes has been slow. The glucokinase regulatory protein (GKRP; gene symbol GCKR) P446L variant, associated with inverse modulation of glucose- and lipid-related traits, has been shown to alter the kinetics of glucokinase (GCK) inhibition. As GCK inhibition is associated with nuclear sequestration, we aimed to determine whether this variant also alters the direct interaction between GKRP and GCK and their intracellular localisation. Methods Fluorescently tagged rat and human wild-type (WT)- or P446L-GCKR and GCK were transiently transfected into HeLa cells and mouse primary hepatocytes. Whole-cell and nuclear fluorescence was quantified in individual cells exposed to low- or high-glucose conditions (5.5 or 25 mmol/l glucose, respectively). Interaction between GCK and GKRP was measured by sensitised emission-based fluorescence resonance energy transfer (FRET) efficiency

Drug resistance associated genetic polymorphisms in Plasmodium falciparum and Plasmodium vivax collected in Honduras, Central America

Background: In Honduras, chloroquine and primaquine are recommended and still appear to be effective for treatment of Plasmodium falciparum and Plasmodium vivax malaria. The aim of this study was to determine the proportion of resistance associated genetic polymorphisms in P. falciparum and P. vivax collected in Honduras. Methods: Blood samples were collected from patients seeking medical attention at the Hospital Escuela in Tegucigalpa from 2004 to 2006 as well as three regional hospitals, two health centres and one regional laboratory during 2009. Single nucleotide polymorphisms in P. falciparum chloroquine resistance transporter (pfcrt), multidrug resistance 1 (pfmdr1), dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes and in P. vivax multidrug resistance 1 (pvmdr1) and dihydrofolate reductase (pvdhfr) genes were detected using PCR based methods. Results: Thirty seven P. falciparum and 64 P. vivax samples were collected. All P. falciparum infections acquired in Honduras carried pfcrt, pfmdr1, pfdhps and pfdhfr alleles associated with chloroquine, amodiaquine and sulphadoxine-pyrimethamine sensitivity only. One patient with parasites acquired on a Pacific Island had pfcrt 76 T and pfmdr1 86Y alleles. That patient and a patient infected in West Africa had pfdhfr 51I, 59 R and 108 N alleles. Pvmdr1 976 F was found in 7/37 and two copies of pvmdr1 were found in 1/37 samples. Pvdhfr 57 L + 58 R was observed in 2/57 samples. Conclusion: The results indicate that P. falciparum from Honduras remain sensitive to chloroquine and sulphadoxine-pyrimethamine. This suggests that chloroquine and sulphadoxine-pyrimethamine should be efficacious for treatment of uncomplicated P. falciparum malaria, supporting current national treatment guidelines. However, genetic polymorphisms associated with chloroquine and sulphadoxine-pyrimethamine tolerance were detected in local P. vivax and imported P. falciparum infections. Continuous monitoring of the prevalence of drug resistant/tolerant P. falciparum and P. vivax is therefore essential also in Honduras.Swedish International Development Cooperation Agency, Department for research Cooperation (Sida-SAREC) [75007082/03]info:eu-repo/semantics/publishedVersio

Paraoxonase 1 Polymorphism and Prenatal Pesticide Exposure Associated with Adverse Cardiovascular Risk Profiles at School Age

Background: Prenatal environmental factors might influence the risk of developing cardiovascular disease later in life. The HDL-associated enzyme paraoxonase 1 (PON1) has anti-oxidative functions that may protect against atherosclerosis. It also hydrolyzes many substrates, including organophosphate pesticides. A common polymorphism, PON1 Q192R, affects both properties, but a potential interaction between PON1 genotype and pesticide exposure on cardiovascular risk factors has not been investigated. We explored if the PON1 Q192R genotype affects cardiovascular risk factors in school-age children prenatally exposed to pesticides. Methods: Pregnant greenhouse-workers were categorized as high, medium, or not exposed to pesticides. Their children underwent a standardized examination at age 6-to-11 years, where blood pressure, skin folds, and other anthropometric parameters were measured. PON1-genotype was determined for 141 children (88 pesticide exposed and 53 unexposed). Serum was analyzed for insulin-like growth factor I (IGF-I), insulin-like growth factor binding protein 3 (IGFBP3), insulin and leptin. Body fat percentage was calculated from skin fold thicknesses. BMI results were converted to age and sex specific Z-scores. Results: Prenatally pesticide exposed children carrying the PON1 192R-allele had higher abdominal circumference, body fat content, BMI Z-scores, blood pressure, and serum concentrations of leptin and IGF-I at school age than unexposed children. The effects were related to the prenatal exposure level. For children with the PON1 192QQ genotype, none of the variables was affected by prenatal pesticide exposure. Conclusion: Our results indicate a gene-environment interaction between prenatal pesticide exposure and the PON1 gene. Only exposed children with the R-allele developed adverse cardiovascular risk profiles thought to be associated with the R-allele

Effectiveness of artemether-lumefantrine provided by community health workers in under-five children with uncomplicated malaria in rural Tanzania: an open label prospective study

\ud Home-management of malaria (HMM) strategy improves early access of anti-malarial medicines to high-risk groups in remote areas of sub-Saharan Africa. However, limited data are available on the effectiveness of using artemisinin-based combination therapy (ACT) within the HMM strategy. The aim of this study was to assess the effectiveness of artemether-lumefantrine (AL), presently the most favoured ACT in Africa, in under-five children with uncomplicated Plasmodium falciparum malaria in Tanzania, when provided by community health workers (CHWs) and administered unsupervised by parents or guardians at home. An open label, single arm prospective study was conducted in two rural villages with high malaria transmission in Kibaha District, Tanzania. Children presenting to CHWs with uncomplicated fever and a positive rapid malaria diagnostic test (RDT) were provisionally enrolled and provided AL for unsupervised treatment at home. Patients with microscopy confirmed P. falciparum parasitaemia were definitely enrolled and reviewed weekly by the CHWs during 42 days. Primary outcome measure was PCR corrected parasitological cure rate by day 42, as estimated by Kaplan-Meier survival analysis. This trial is registered with ClinicalTrials.gov, number NCT00454961. A total of 244 febrile children were enrolled between March-August 2007. Two patients were lost to follow up on day 14, and one patient withdrew consent on day 21. Some 141/241 (58.5%) patients had recurrent infection during follow-up, of whom 14 had recrudescence. The PCR corrected cure rate by day 42 was 93.0% (95% CI 88.3%-95.9%). The median lumefantrine concentration was statistically significantly lower in patients with recrudescence (97 ng/mL [IQR 0-234]; n = 10) compared with reinfections (205 ng/mL [114-390]; n = 92), or no parasite reappearance (217 [121-374] ng/mL; n = 70; p ≤ 0.046). Provision of AL by CHWs for unsupervised malaria treatment at home was highly effective, which provides evidence base for scaling-up implementation of HMM with AL in Tanzania.\u