Characterisation of antimony-resistant Leishmania donovani isolates: biochemical and biophysical studies and interaction with host cells

Recent clinical isolates of Leishmania donovani from the hyperendemic zone of Bihar were characterised in vitro in terms of their sensitivity towards sodium stibogluconate in a macrophage culture system. The resulting half maximal effective concentration (EC(50)) values were compared with those of known sensitive isolates. Fifteen of the isolates showed decreased sensitivity towards SSG with an average EC(50) of 25.7 ± 4.5 μg/ml pentavalent antimony (defined as antimony resistant), whereas nine showed considerable sensitivity with an average EC(50) of 4.6 ± 1.7 μg/ml (defined as antimony sensitive). Out of those nine, seven were recent clinical isolates and the remaining two were known sensitive isolates. Compared with the antimony sensitive, resistant isolates showed enhanced expression of thiol metabolising enzymes in varying degrees coupled with increased intracellular non-protein thiol content, decreased fluorescence anisotropy (inversely proportional with membrane fluidity) and over-expression of the terminal glycoconjugates (N-acetyl-d-galactosaminyl residue). Macrophages infected with resistant but not with sensitive showed up-regulation of the ATP Binding Cassette transporter multidrug resistance protein 1 and permeability glycoprotein, while the supernatant contained abundant IL-10. The above results reinforce the notion that antimony resistant parasites have undergone a number of biochemical and biophysical changes as part of their adaptation to ensure their survival in the host.     

A phylogenetic approach to the identification of phosphoglucomutase genes

The expanding molecular database provides unparalleled opportunities for characterizing genes and for studying groups of related genes. We use sequences drawn from the database to construct an evolutionary framework for examining the important glycolytic enzyme phosphoglucomutase (PGM). Phosphoglucomutase plays a pivotal role in the synthesis and utilization of glycogen and is present in all organisms. In humans, there are three well-described isozymes, PGMI, PGM2, and PGM3. PGM1 was cloned 5 years ago; however, repeated attempts using both immunological approaches and molecular probes designed from PGM1 have failed to isolate either PGM2 or PGM3. Using a phylogenetic strategy, we first identified 47 highly divergent prokaryotic and eukaryotic PGM-like sequences from the database. Although overall amino acid identity often fell below 20%, the relative order, position, and sequence of three structural motifs, the active site and the magnesium-- and sugar-binding sites, were conserved in all 47 sequences. The phylogenetic history of these sequences was complex and marked by duplications and translocations; two instances of transkingdom horizontal gene transfer were identified. Nonetheless, the sequences fell within six well-defined evolutionary lineages, three of which contained only prokaryotes. Of the two prokaryotic/eukaryotic lineages, one contained bacterial, yeast, slimemold, invertebrate, and vertebrate homologs to human PGM1 and the second contained likely homologs to human PGM2. Indeed, an amino acid sequence, derived from a partial human cDNA, that fell within the second cross-kingdom lineage bears several characteristics expected for PGM2. A third lineage may contain homologs to human PGM3. On a general level, our phylogenetic-based approach shows promise for the further utilization of the extensive molecular database.      

Leech bite: a rare gynecologic emergency

Vaginal bleeding as the result of a leech bite is a rare occurrence. We report 2 cases of vaginal bleeding in young girls that resulted from a leech bite and required treatment. Clinical presentation and management for young girls is described. Health professionals working in rural areas where leech infestation is common should be aware that children are at risk for leech bites in the genital region; a high index of suspicion is of great help to make an early diagnosis and ensure prompt treatment.     

Does Colour of the Food Attract Ants?

Milky white, brown, yellow and pink sugar grains along with normal sugar grains in equal number were offered to the ants at different sites, in the foraging ground of a garden locating at Garia, Kolkata, India to note the role of colouration of the food in food selection, if any. It is revealed that the ants Anoplolepis gracilipes procured all the supplied colour sugar grains from the offered sites between 1 and 43 min and the normal sugar grains between 1 and 38 min from the offered sites. Results of statistical analyses clearly indicate that the colour of the food has no role to attract the ants A. gracilipes in respect to procurement times of the sugar grains noted. This suggests that the colour of the food does not act as an attractant for the ants A. gracilipes in food selection.     

Statistical properties of the variation at linked microsatellite loci: implications for the history of human Y chromosomes [published erratum appears in Mol Biol Evol 1997 Mar;14(3):354]

It has recently been suggested that observed levels of variation at microsatellite loci can be used to infer patterns of selection in genomes and to assess demographic history. In order to evaluate the feasibility of these suggestions it is necessary to know something about how levels of variation at microsatellite loci are expected to fluctuate due simply to stochasticity in the processes of mutation and inheritance (genetic sampling). Here we use recently derived properties of the stepwise mutation model to place confidence intervals around the variance in repeat score that is expected at mutation-drift equilibrium and outline a statistical test for whether an observed value differs significantly from expectation. We also develop confidence intervals for the time course of the buildup of variation following a complete elimination of variation, such as might be caused by a selective sweep or an extreme population bottleneck. We apply these methods to the variation observed at human Y-specific microsatellites. Although a number of authors have suggested the possibility of a very recent sweep, our analyses suggest that a sweep or extreme bottleneck is unlikely to have occurred anytime during the last approximately 74,000 years. To generate this result we use a recently estimated mutation rate for microsatellite loci of 5.6 x 10(-4) along with the variation observed at autosomal microsatellite loci to estimate the human effective population size. This estimate is 18,000, implying an effective number of 4,500 Y chromosomes. One important general conclusion to emerge from this study is that in order to reject mutation-drift equilibrium at a set of linked microsatellite loci it is necessary to have an unreasonably large number of loci unless the observed variance is far below that expected at mutation-drift equilibrium.      

Exploring dual inhibitory role of febrifugine analogues against Plasmodium utilizing structure-based virtual screening and molecular dynamic simulation

Malaria is an endemic disease caused by the protozoan parasite Plasomodium falciparum. Febrifugine analogues are natural compound obtained from the traditional Chinese herbs have shown significant antimalarial and anticancerous efficacy in experimental model. Development of resistance against the existing antimalarial drug has alarmed the scientific innovators to find a potential antimalarial molecule which can be further used by endemic countries for the elimination of this disease. In this study, structure-based virtual screening and molecular dynamics (MD) base approaches were used to generate potential antimalarial compound against plasmepsin II and prolyl-tRNA synthetase of Plasmodium. Here, we have docked series of febrifugine analogues (n = 11,395) against plasmepsin II in three different docking modes and then it was compared with previously reported target prolyl-tRNA synthetase. Extra precision docking resulted into 235 ligands having better docking score were subject for QikProp analysis. Better ligands (n = 39) obtained from QikProp analysis were subject for ADMET prediction and docking protocol validation through the estimation of receiver operator characteristics. In the later stage, 24 ligands obtained from ADMET study were subject for the estimation of binding energy through MM-GBSA and same were also docked against prolyl-tRNA synthetase to get compounds with dual inhibitor role. Finally, MD simulation and 2D fingerprint MACCS study of two best ligands have shown significant interaction with plasmepsin II and homology against known active ligand with noteworthy MACCS index, respectively. This study concludes that FA12 could be potential drug candidate to fight against Plasmodium falciparum parasites.