Plasmodium falciparum histidine-rich proteins 2 (PfHRP2) based RDTs are advocated in falciparum malaria-endemic regions, particularly when quality microscopy is not available. However, diversity and any deletion in the pfhrp2 and pfhrp3 genes can affect the performance of PfHRP2-based RDTs. A total of 400 samples collected from uncomplicated malaria cases from Kenya were investigated for the amino acid repeat profiles in exon 2 of pfhrp2 and pfhrp3 genes. In addition, PfHRP2 levels were measured in 96 individuals with uncomplicated malaria. We observed a unique distribution pattern of amino acid repeats both in the PfHRP2 and PfHRP3. 228 PfHRP2 and 124 PfHRP3 different amino acid sequences were identified. Of this, 214 (94%) PfHRP2 and 81 (65%) PfHRP3 amino acid sequences occurred only once. Thirty-nine new PfHRP2 and 20 new PfHRP3 amino acid repeat types were identified. PfHRP2 levels were not correlated with parasitemia or the number of PfHRP2 repeat types. This study shows the variability of PfHRP2, PfHRP3 and PfHRP2 concentration among uncomplicated malaria cases. These findings will be useful to understand the performance of PfHRP2-based RDTs in Kenya.

BackgroundPlasmodium falciparum resistance to chloroquine (CQ) denied healthcare providers access to a cheap and effective anti-malarial drug. Resistance has been proven to be due to point mutations on the parasite’s pfcrt gene, particularly on codon 76, resulting in an amino acid change from lysine to threonine. This study sought to determine the prevalence of the pfcrt K76T mutation 13 years after CQ cessation in Msambweni, Kenya.MethodsFinger-prick whole blood was collected on 3MM Whatman® filter paper from 99 falciparum malaria patients. Parasite DNA was extracted via the Chelex method from individual blood spots and used as template in nested PCR amplification of pfcrt. Apo1 restriction enzyme was used to digest the amplified DNA to identify the samples as wild type or sensitive at codon 76. Prevalence figures of the mutant pfcrt 76T gene were calculated by dividing the number of samples bearing the mutant gene with the total number of samples multiplied by 100 %. Chi square tests were used to test the significance of the findings against previous prevalence figures.ResultsOut of 99 clinical samples collected in 2013, prevalence of the mutant pfcrt 76T gene stood at 41 %.ConclusionThe results indicate a significant [χ2 test, P ≤ 0.05 (2006 vs 2013)] reversal to sensitivity by the P. falciparum population in the study site compared to the situation reported in 2006 at the same study site. This could primarily be driven by diminished use of CQ in the study area in line with the official policy. Studies to establish prevalence of the pfcrt 76T gene could be expanded countrywide to establish the CQ sensitivity status and predict a date when CQ may be re-introduced as part of malaria chemotherapy.

Abstract:A new triterpenoid, 3-oxo-12β-hydroxy-oleanan-28,13β-olide (1), and six known triterpenoids 2-7 were isolated from the root bark of Ekebergia capensis, an African medicinal plant. A limonoid 8 and two glycoflavonoids 9-10 were found in its leaves. The metabolites were identified by NMR and MS analyses, and their cytotoxicity was evaluated against the mammalian African monkey kidney (vero), mouse breast cancer (4T1), human larynx carcinoma (HEp2) and human breast cancer (MDA-MB-231) cell lines. Out of the isolates, oleanonic acid (2) showed the highest cytotoxicity, i.e., IC50's of 1.4 and 13.3 µM against the HEp2 and 4T1 cells, respectively. Motivated by the higher cytotoxicity of the crude bark extract as compared to the isolates, the interactions of oleanonic acid (2) with OPEN ACCESSMolecules 2014, 19 14236 five triterpenoids 3-7 were evaluated on vero cells. In an antiplasmodial assay, seven of the metabolites were observed to possess moderate activity against the D6 and W2 strains of P. falciparum (IC50 27.1-97.1 µM), however with a low selectivity index (IC50(vero)/IC50(P. falciparum-D6) < 10). The observed moderate antiplasmodial activity may be due to general cytotoxicity of the isolated triterpenoids.

No abstract

Detection of Plasmodium species by microscopy has been the gold standard for diagnosis of malaria for more than a century. Despite the fact that there is a significant decline in the number of positive cases reported from microscopy, antimalarial drugs prescriptions are on continuous increase as patients present with symptoms of malaria. This makes it difficult to establish accuracy, sensitivity and specificity of light microscopy in diagnosis of malaria in epidemic areas. This study was designed to compare microscopy with polymerase chain reaction as diagnostic methods for malaria in three epidemic areas in Kenya. A total of 356 patients presenting with malaria symptoms were diagnosed by microscopy and dried blood filter paper spots were collected from patient in Kisii, West Pokot and Narok districts. Plasmodium falciparum DNA was extracted from the dried blood filter samples. Primers specific for the Plasmodium Species were designed and used in a two step amplification of the Pfmdr gene. The PCR products were analyzed in ethidium bromide stained 1.5% agarose gel. It was found that 72 out of 350 specimens diagnosed as negative were positive for P. falciparum by nested PCR, while 6 which were microscopy positive were confirmed so by nested PCR. This study demonstrates that there is a high level of misdiagnosis which may either lead to denial for deserved treatment or undeserved treatment. Nested PCR detection of malaria parasites is a very useful complement to microscopy although it is expensive and takes long time. Additionally, smear negative patients suspected to have malaria should be subjected to PCR diagnosis to improve rational drug use. The economic burden of misdiagnosis and mistreatment of malaria outweighs that of PCR diagnosis, hence this diagnostic mode could be tenable in the long run even in rural areas.