Serological markers of sand fly exposure to evaluate insecticidal nets against visceral leishmaniasis in India and Nepal: a cluster-randomized trial

Background

Visceral leishmaniasis is the world'' second largest vector-borne parasitic killer and a neglected tropical disease, prevalent in poor communities. Long-lasting insecticidal nets (LNs) are a low cost proven vector intervention method for malaria control; however, their effectiveness against visceral leishmaniasis (VL) is unknown. This study quantified the effect of LNs on exposure to the sand fly vector of VL in India and Nepal during a two year community intervention trial.

Methods

As part of a paired-cluster randomized controlled clinical trial in VL-endemic regions of India and Nepal we tested the effect of LNs on sand fly biting by measuring the antibody response of subjects to the saliva of Leishmania donovani vector Phlebotomus argentipes and the sympatric (non-vector) Phlebotomus papatasi. Fifteen to 20 individuals above 15 years of age from 26 VL endemic clusters were asked to provide a blood sample at baseline, 12 and 24 months post-intervention.

Results

A total of 305 individuals were included in the study, 68 participants provided two blood samples and 237 gave three samples. A random effect linear regression model showed that cluster-wide distribution of LNs reduced exposure to P. argentipes by 12% at 12 months (effect 0.88; 95% CI 0.83–0.94) and 9% at 24 months (effect 0.91; 95% CI 0.80–1.02) in the intervention group compared to control adjusting for baseline values and pair. Similar results were obtained for P. papatasi.

Conclusions

This trial provides evidence that LNs have a limited effect on sand fly exposure in VL endemic communities in India and Nepal and supports the use of sand fly saliva antibodies as a marker to evaluate vector control interventions.     

Diagnosis of Indian visceral leishmaniasis by nucleic acid detection using PCR

Background

PCR based diagnosis for Visceral Leishmaniasis (VL), despite numerous published primers, remains far from being applied in the field. The present study was planned to design a Leishmania specific diagnostic assay and to evaluate its sensitivity and specificity on a sample size, which to the best of our knowledge is the largest ever screened in one study.

Methods

Leishmania specific primers were developed using 18S rRNA gene and their sensitivity was evaluated on 500 parasitologically confirmed patients with VL and 25 Post Kala-azar Dermal Leishmaniasis (PKDL) patients. Specificity was calculated on 250 healthy endemic controls, 250 healthy non endemic controls and 250 non leishmanial diseases like malaria.

Results

Our PCR assay had a sensitivity of 87.8% (95%CI: 84.1–89.8) using 200 µL of patient''s peripheral-blood. Specificity was absolute in non-endemic healthy controls and in subjects with different diseases while in endemic controls it was 84% (95%CI: 78.9–88.0). Its overall specificity was 94.6% (95%CI-92.8–96.1).

Conclusions

The PCR assay developed is sensitive enough to detect the 18S rRNA gene in an amount equivalent to a single parasite or less in a one million human cell environment. The high sensitivity of this PCR diagnostic test with relatively non-invasive peripheral blood sampling method opens up the possibility of its deployment in field for the routine diagnosis of VL.     

Tocotrienols confer resistance to ischemia in hypercholesterolemic hearts: insight with genomics

Most clinical trials with vitamin E could not lower cholesterol and thus, have been deemed unsuccessful. Recently, tocotrienols, isomers of vitamin E have been found to lower LDL levels. To explore if tocotrienols could be the drug target for vitamin E, rabbits were kept on cholesterol diet for 60 days supplemented with tocotrienol-α, tocotrienol-δ, and tocotrienol-γ for the last 30 days. The serum cholesterol levels (in mmol/l) were 24.4 (tocotrienol-α), 34.9 (tocotrienol-δ), 19.8 (tocotrienol-γ) vs. 39.7 (control). Left ventricular function including aortic flow and developed pressure exhibited significantly improved recovery with tocotrienol-γ and -α, but not with tocotrienol-δ. The myocardial infarct size showed a similar pattern: 33% (tocotrienol-α), 23% (tocotrienol-γ), and 47% (tocotrienol-δ). To examine the molecular mechanisms of cardioprotective effects, gene expression profile was determined using Atlas 1.2/1.2II followed by determination of gene profiles using PedQuest 8.3 software. Based on genomic profiles, the following cholesterol-related proteins were examined: FABP, TGF-β (cholesterol suppresses TGF-β), ET-1 (increased by hypercholesterolemia), SPOT 14 (linked with hypercholesterolemia), and matrix metalloproteinase (MMP) 2 and MMP9 (cholesterol regulates MMP2 and MMP9 expression) in the heart. Consistent with the cardioprotective effects of tocotrienol-α and -γ, these two isomers reduced ET-1, decreased MMP2 and MM9, increased TGF-β and reduced SPOT 14, while tocotrienol-δ had no effects. The results of the present study demonstrate that the two isomers of tocotrienols, α and γ, render the hypercholesterolemic hearts resistant to ischemic reperfusion injury by lowering several hypercholesterolemic proteins including MMP2, MMP9, ET-1, and SPOT 14 and upregulating TGF-β.     

Construction of multiple herbicide resistant ammonia excreting strains of cyanobacteriumNostoc muscorum

Summary Machete resistant (Mat ^r), basalin resistant (Bas ^r), 3(3,4 dichlorophenyl)-1,1-dimethyl urea resistant (DCMU ^r), atrazine resistant (Atr ^r) and propanil resistant (Prp ^r) phenotypes ofGloeocapsa sp. were cotransformed toNostoc muscorum at high frequency. Spontaneously occurring mutants of the multiple herbicide resistant transformant containing L-methionine-DL-sulfoximine resistant (Msx ^r), ethylene diamine resistant (Eda ^r), or phosphinothricin resistant (Ppt ^r) glutamine synthetase (GS) showed extracellular liberation of ammonia resulting from fixation of N₂ under photosynthetic conditions. Results suggest a definite role of GS activity in regulation of extracellular ammonia.     

PELP1 is a reader of histone H3 methylation that facilitates oestrogen receptor‐α target gene activation by regulating lysine demethylase 1 specificity

Histone methylation has a key role in oestrogen receptor (ERα)‐mediated transactivation of genes. Proline glutamic acid and leucine‐rich protein 1 (PELP1) is a new proto‐oncogene that functions as an ERα co‐regulator. In this study, we identified histone lysine demethylase, KDM1, as a new PELP1‐interacting protein. These proteins, PELP1 and KDM1, were both recruited to ERα target genes, and PELP1 depletion affected the dimethyl histone modifications at ERα target genes. Dimethyl‐modified histones H3K4 and H3K9 are recognized by PELP1, and PELP1 alters the substrate specificity of KDM1 from H3K4 to H3K9. Effective demethylation of dimethyl H3K9 by KDM1 requires a KDM1–ERα–PELP1 functional complex. These results suggest that PELP1 is a reader of H3 methylation marks and has a crucial role in modulating the histone code at the ERα target genes.     

Polycystic ovary syndrome in Indian women: a mass spectrometry based serum metabolomics approach

Introduction

Polycystic ovary syndrome (PCOS) is a complex, heterogeneous endocrinological disorder with uncertain pathogenesis and is very common in women of reproductive age. There are few reports of utilizing metabolomics approach to understand the complex pathophysiology of PCOS. However, excluding one previous NMR-based metabolomics study, none of the study was conducted in Indian population.

Objective

The study aims to compare the serum metabolomic profile of PCOS women with controls from the Eastern region of India.

Methods

PCOS women (n?=?35) and healthy control women (n?=?30) undergoing tubal ligation were recruited for this study. Serum metabolic profiles were generated using liquid chromatography–tandem mass spectrometry (LC-MS/MS) and gas chromatography–mass spectrometry (GC-MS). Multivariate statistical analysis was applied to spectral data obtained from both the LC-MS/MS and GC/MS.

Results

Nine metabolites were identified to be most significantly dysregulated in sera of PCOS women; however, few other identified metabolites were also altered but with lesser significance. Amongst these metabolites, riboflavin, sucrose, adenine and N-acetyl glycine, phosphoric acid and cortisol were down-regulated, whereas, thymine, cystathionine, and phenylalanine were up-regulated in PCOS when compared with controls. The observed changes in metabolite expression suggested alterations in aminoacyl-tRNA biosynthesis, metabolism of nitrogen, alanine-aspartate-glutamate, galactose, glycine-serine-threonine, and pyrimidine-purine among several metabolic pathways possibly implicated in these PCOS women.

Conclusion

The altered metabolites identified in PCOS women of Eastern Indian population, provide insight into current perceptive of the disease pathology, metabolic involvements, and may be considered as putative markers of PCOS.

     

Indole-based heterocyclic inhibitors of p38alpha MAP kinase: designing a conformationally restricted analogue

p38alpha Mitogen Activated Protein Kinase (MAP kinase) is an intracellular soluble serine threonine kinase. p38alpha kinase is activated in response to cellular stresses, growth factors and cytokines such as interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF-alpha). The central role of p38alpha activation in settings of both chronic and acute inflammation has led efforts to find inhibitors of this enzyme as possible therapies for diseases such as rheumatoid arthritis, where p38alpha activation is thought to play a causal role. Herein, we report structure-activity relationship studies on a series of indole-based heterocyclic inhibitors that led to the design and identification of a new class of p38alpha inhibitors.     

A procedure for detection and quantitation of cavity volumes proteins. Application to measure the strength of the hydrophobic driving force in protein folding

Accurate identification of cavities is important in the study of protein structure, stability, design, and ligand binding. Identification and quantitation of cavities is a nontrivial problem because most cavities are connected to the protein exterior. We describe a computational procedure for quantitating cavity volumes and apply this to derive an estimate of the hydrophobic driving force in protein folding. A grid-based Monte Carlo procedure is used to position water molecules on the surface of a protein. A Voronoi procedure is used to identify and quantitate empty space within the solvated protein. Additional cavities not detected by other existing procedures can be identified. Most of these are close to surface concavities. Residue volumes for both the interior and the surface residues as well as cavity volumes are in good agreement with volumes calculated from fully hydrated protein structures obtained from molecular dynamic simulations. We show that the loss of stability because of cavity-creating mutations correlates better with cavity volumes determined by this procedure than with cavity volumes determined by other methods. Available structural and thermodynamic data for a number of cavity-containing mutants were analyzed to obtain estimates of 26.1 cal x mol(-1) x A(-3) and 18.5 cal x mol(-1) x A(-2) for the relative contributions of cavity formation and the hydrophobic effect to the observed stability changes. The present estimate for the hydrophobic driving force is at the lower end of estimates derived from model compound studies and considerably lower than previous estimates of approximately 50 cal x mol(-1) x A(-2) derived from protein mutational data. In the absence of structural rearrangement, on average, deletion of a single methylene group is expected to result in losses in stability of 0.41 and 0.70 kcal x mol(-1) resulting from decrease in hydrophobicity and packing, respectively.     

Elucidation of factors responsible for enhanced thermal stability of proteins: a structural genomics based study

Understanding the molecular basis for the enhanced stability of proteins from thermophiles has been hindered by a lack of structural data for homologous pairs of proteins from thermophiles and mesophiles. To overcome this difficulty, complete genome sequences from 9 thermophilic and 21 mesophilic bacterial genomes were aligned with protein sequences with known structures from the protein data bank. Sequences with high homology to proteins with known structures were chosen for further analysis. High quality models of these chosen sequences were obtained using homology modeling. The current study is based on a data set of models of 900 mesophilic and 300 thermophilic protein single chains and also includes 178 templates of known structure. Structural comparisons of models of homologous proteins allowed several factors responsible for enhanced thermostability to be identified. Several statistically significant, specific amino acid substitutions that occur going from mesophiles to thermophiles are identified. Most of these are at solvent-exposed sites. Salt bridges occur significantly more often in thermophiles. The additional salt bridges in thermophiles are almost exclusively in solvent-exposed regions, and 35% are in the same element of secondary structure. Helices in thermophiles are stabilized by intrahelical salt bridges and by an increase in negative charge at the N-terminus. There is an approximate decrease of 1% in the overall loop content and a corresponding increase in helical content in thermophiles. Previously overlooked cation-pi interactions, estimated to be twice as strong as ion-pairs, are significantly enriched in thermophiles. At buried sites, statistically significant hydrophobic amino acid substitutions are typically consistent with decreased side chain conformational entropy.