Identification and functional validation of the novel antimalarial resistance locus PF10_0355 in Plasmodium falciparum

The Plasmodium falciparum parasite's ability to adapt to environmental pressures, such as the human immune system and antimalarial drugs, makes malaria an enduring burden to public health. Understanding the genetic basis of these adaptations is critical to intervening successfully against malaria. To that end, we created a high-density genotyping array that assays over 17,000 single nucleotide polymorphisms (～ 1 SNP/kb), and applied it to 57 culture-adapted parasites from three continents. We characterized genome-wide genetic diversity within and between populations and identified numerous loci with signals of natural selection, suggesting their role in recent adaptation. In addition, we performed a genome-wide association study (GWAS), searching for loci correlated with resistance to thirteen antimalarials; we detected both known and novel resistance loci, including a new halofantrine resistance locus, PF10_0355. Through functional testing we demonstrated that PF10_0355 overexpression decreases sensitivity to halofantrine, mefloquine, and lumefantrine, but not to structurally unrelated antimalarials, and that increased gene copy number mediates resistance. Our GWAS and follow-on functional validation demonstrate the potential of genome-wide studies to elucidate functionally important loci in the malaria parasite genome.     

Harnessing genomics and genome biology to understand malaria biology

Malaria is an important human disease and is the target of a global eradication campaign. New technological and informatics advancements in population genomics are being leveraged to identify genetic loci under selection in the malaria parasite and to find variants that are associated with key clinical phenotypes, such as drug resistance. This article provides a timely Review of how population-genetics-based strategies are being applied to Plasmodium falciparum both to identify genetic loci as key targets of interventions and to develop monitoring and surveillance tools that are crucial for the successful elimination and eradication of malaria.     

Different regulatory mechanisms underlie similar transposable element profiles in pufferfish and fruitflies

Comparative analysis of recently sequenced eukaryotic genomes has uncovered extensive variation in transposable element (TE) abundance, diversity, and distribution. The TE profile in the sequenced pufferfish genomes is more similar to that of Drosophila melanogaster than to human or mouse, in that pufferfish TEs exhibit low overall abundance, high family diversity, and localization in the heterochromatin. It has been suggested that selection against the deleterious effects of ectopic recombination between TEs has structured the TE profile in Drosophila and pufferfish but not in humans. We test this hypothesis by measuring the sample frequency of 48 euchromatic TE insertions in the genome of the green spotted pufferfish (Tetraodon nigroviridis). We estimate the strength of selection acting on recent insertions by analyzing the site frequency spectrum using a maximum-likelihood approach. We show that in contrast to Drosophila, euchromatic TE insertions in Tetraodon are selectively neutral and that the low copy number and compartmentalized distribution of TEs in the Tetraodon genome must be caused by regulation by means other than purifying selection acting on recent insertions. Inference of regulatory processes governing TE profiles should take into account factors such as effective population size, incidence of inbreeding/outcrossing, and other species-specific traits.     

Differential Cytotoxicities of N-Methyl-β-Carbolinium Analogues of MPP+ in PC12 Cells: Insights into Potential Neurotoxicants in Parkinson's Disease

N-Methylated β-carbolinium cations that can form in vivo from environmental or endogenous β-carbolines are putative neurotoxic factors in Parkinson's disease. The cytotoxicities of 11 N-methylated β-carbolinium cations and N-methyl-4-phenylpyridinium cation (MPP⁺), the experimental parkinsonian neurotoxicant which the carbolinium cations structurally resemble, were examined using rat pheochromocytoma (PC12) cells cultured in “low energy” N-5 medium; cell death was estimated by released lactate dehydrogenase activity and viable cell protein. Of the eight N-monomethylated β-carbolinium cations utilized, only 2-methyl-harmalinium (harmaline-2- methiodide) was as cytotoxic as MPP⁺. Also, three N²(β), N⁸(indole)-dimethylated β-carbolinium cations displayed cytotoxic effects, with the simplest, 2,9-dimethylnorhar- manium, approaching the effectiveness of MPP⁺ in PC12 cells cultured in N-5 medium. However, when PC12 cells grown in higher energy Dulbecco's modified Eagle's medium were utilized with selected effective cations, it was observed that the cultures were relatively resistant to MPP⁺ and 2,9-dimethylnorharmanium, but remained vulnerable to 2-methylharmalinium. The results are interpreted to mean that different cytotoxic mechanisms exist for the two most potent β-carbolinium cations—namely, a mechanism for the 2,9-dimethyl-β-carbolinium species that, as with MPP⁺, is conditional on mitochondrial ATP depletion, but a different (or additional) mechanism for 2- methylharmalinium that is independent of mitochondrial inhibition. The possible accumulation of these cytotoxic cations in Parkinson's disease is discussed in the context of these findings.     

Waist Circumference as a Screening Tool for Cardiovascular Risk Factors: Evaluation of Receiver Operating Characteristics (ROC)

To evaluate the receiver operating characteristics (ROC) to determine the cutoffs of waist circumference as a potential population directed screening tool for hypercholesterolaemia (≥6.5 mmol/L), low high density lipoprotein cholesterol (<0.9 mmol/L), and hypertension (treated and/or systolic ≥160 and/or diastolic blood pressure ≥95 mmHg), in 2183 men and 2698 women aged 20 to 59 years selected at random from Dutch civil registries. Main outcome measures: Height, weight, body mass index (BMI), waist circumference, total plasma cholesterol and high density lipoprotein cholesterol concentrations, and blood pressure. Results: ROC curves showed that sensitivity equalled specificity at waist circumferences between 93–95 cm in men and 81–84 cm in women for identifying individual risk factors, and 92 cm in men and 81 cm in women for identifying those with at least one risk factor. Sensitivity and specificity were equal at levels between 61% to 69% for identifying individual risk factors, with positive predictions (56.8% in men and 37.8% in women) within 2% of those using previously defined ‘Action Level 1’ of waist circumference 94 cm in men and 80 cm in women (58.8% in men and 37.4% in women). Risk prediction by anthropometric methods was relatively low: ROC areas for identifying each risk factor by waist varied from 55% to 60%, and reached about 65% for identifying at least one risk factor. Height accounted for less than 03% of variance in waist circumference. Using BMI at 25 kg/m² gave similar prediction to waist, but its combination with waist did not improve predictive values. Conclusions: Measurement of waist circumference ‘Action Level 1’ at 94 cm (37 inches) in men and 80 cm (32 inches) in women could be adopted as a simpler valid alternative to BMI for health promotion, to alert those at risk of cardiovascular disease, and as a guide to risk avoidance by self-weight management     

Novel S-Adenosylmethionine-Dependent Indole-N-Methylation of β-Carbolines in Brain Particulate Fractions

Guinea pig brain S-adenosylmethionine (SAM)-dependent N-methyltransferase activity toward physiologically relevant beta-carboline (BC) substrates was examined with reverse-phase HPLC and radiochemical detection. Representative BCs, norharman and harmine, were enzymatically methylated on the 2[beta]-nitrogen by [3H]CH3-SAM in undialyzed homogenates to yield 2[beta]-methylated BCs and subsequently on the 9[indole]-nitrogen to generate 2,9-dimethylated BC products. This may be the first account of mammalian indole N-methyl transfer. There was no HPLC evidence for 9-methyl BC or (from carbon methylation) 2,6-dimethyl BC products. Capillary gas chromatography-mass spectrometry analysis confirmed the structures of the 2,9-dimethyl and 2-methyl products of norharman. The 2[beta]- and 9[indole]-N-methylation activities were mainly in the nuclear fractions and were negligible in undialyzed cytosol. This differs from the cytosolic SAM-dependent N-methylations reported with other azaheterocyclics, including 1,2,3,4-tetrahydro-BCs. The involvement of a single enzyme was suggested because the two N-methyl transfers with BC substrate had similar subcellular activity patterns, regional brain distributions, and Km and Vmax values. Sequential N-methylation of various BCs that have been observed in vivo may be a unique route to centrally retained N2,N9-dimethylated beta-carbolinium ions. Because they resemble the synthetic parkinsonian toxicant, N-methyl-4-phenylpyridinium, with respect to structure and neurotoxic activity, such "bioactivated" carbolinium ions could be endogenous causative factors in Parkinson's disease.     

Peatland vascular plant functional types affect dissolved organic matter chemistry

Aims

The readily available global rock phosphate (P) reserves may be depleted within the next 50–130 years warranting careful use of this finite resource. We develop a model that allows us to assess a range of P fertiliser and soil management strategies for Barley in order to find which one maximises plant P uptake under certain climate conditions.

Methods

Our model describes the development of the P and water profiles within the soil. Current cultivation techniques such as ploughing and reduced till gradient are simulated along with fertiliser options to feed the top soil or the soil right below the seed.

Results

Our model was able to fit data from two barley field trials, achieving a good fit at early growth stages but a poor fit at late growth stages, where the model underestimated plant P uptake. A well-mixed soil (inverted and 25 cm ploughing) is important for optimal plant P uptake and provides the best environment for the root system.

Conclusions

The model is sensitive to the initial state of P and its distribution within the soil profile; experimental parameters which are sparsely measured. The combination of modelling and experimental data provides useful agricultural predictions for site specific locations.

     

The predictive value of transcranial duplex sonography for the clinical diagnosis in undiagnosed parkinsonian syndromes: comparison with SPECT scans

Background  

Transcranial duplex sonography (TCD) of the substantia nigra has emerged as a promising, non-invasive tool to diagnose idiopathic Parkinson's disease (IPD). However, its diagnostic accuracy in patients with undefined parkinsonism remains to be determined.     

Comparative evolutionary genomics of human malaria parasites

The parasites Plasmodium falciparum and Plasmodium vivax are responsible for the majority of human malaria cases worldwide. Despite many similarities in their biology, they frequently are studied in isolation. With the completion of the P. vivax genome and the generation of an initial P. falciparum genetic diversity map, attempts are being made to infer inter- and intra-species genome evolution. Here, we briefly review our current knowledge of comparative evolutionary genomics of the two species in the light of several presentations at the Molecular Approaches to Malaria 2008 meeting in Lorne, Australia and ask the question: can evolutionary genomics of one species inform the other?