Origin and Evolution of Sulfadoxine Resistant Plasmodium falciparum

The Thailand-Cambodia border is the epicenter for drug-resistant falciparum malaria. Previous studies have shown that chloroquine (CQ) and pyrimethamine resistance originated in this region and eventually spread to other Asian countries and Africa. However, there is a dearth in understanding the origin and evolution of dhps alleles associated with sulfadoxine resistance. The present study was designed to reveal the origin(s) of sulfadoxine resistance in Cambodia and its evolutionary relationship to African and South American dhps alleles. We sequenced 234 Cambodian Plasmodium falciparum isolates for the dhps codons S436A/F, A437G, K540E, A581G and A613S/T implicated in sulfadoxine resistance. We also genotyped 10 microsatellite loci around dhps to determine the genetic backgrounds of various alleles and compared them with the backgrounds of alleles prevalent in Africa and South America. In addition to previously known highly-resistant triple mutant dhps alleles SGEGA and AGEAA (codons 436, 437, 540, 581, 613 are sequentially indicated), a large proportion of the isolates (19.3%) contained a 540N mutation in association with 437G/581G yielding a previously unreported triple mutant allele, SGNGA. Microsatellite data strongly suggest the strength of selection was greater on triple mutant dhps alleles followed by the double and single mutants. We provide evidence for at least three independent origins for the double mutants, one each for the SGKGA, AGKAA and SGEAA alleles. Our data suggest that the triple mutant allele SGEGA and the novel allele SGNGA have common origin on the SGKGA background, whereas the AGEAA triple mutant was derived from AGKAA on multiple, albeit limited, genetic backgrounds. The SGEAA did not share haplotypes with any of the triple mutants. Comparative analysis of the microsatellite haplotypes flanking dhps alleles from Cambodia, Kenya, Cameroon and Venezuela revealed an independent origin of sulfadoxine resistant alleles in each of these regions.     

Survey of susceptibility of clinical Clostridium diffiicile strains isolated from patients hospitalised in different departments of paediatric hospital to antimicrobial agents

This study was performed to determine the susceptibility of 50 C. difficile strains isolated from faecal samples of children suspected to antibiotic associated diarrhea (AAD) to antimicrobial agents: metronidazole, vancomycin, erythromycin, clindamycin, ciprofloxacin, moxifloksacin, gatifloksacin and imipenem. The all C. difficile strains were sensitived to metronidazole and vancomycin. Twenty six per cent of strains were resistant to erythromycin and clindamycin (MLS(B) type resistance). Resitance to ciprofloxacin, moxifloxacin, gatifloxacin and imipenem was detected in 98%, 8%, 8% and 30% of C. difficile strains, respectively.     

R-(+)-alpha-lipoic acid inhibits endothelial cell apoptosis and proliferation: involvement of Akt and retinoblastoma protein/E2F-1

Lipoic acid was recently demonstrated to improve endothelial dysfunction or retinopathy not only in rats but also in diabetic patients. We tested the hypothesis that R-(+)-alpha-lipoic acid (LA) directly affects human endothelial cell (EC) function (e.g., apoptosis, proliferation, and protein expression), independent of the cells' vascular origin. Macrovascular EC (macEC), isolated from umbilical (HUVEC) and adult saphenous veins and from aortae, as well as microvascular EC (micEC) from retinae, skin, and uterus, were exposed to LA (1 mumol/l-1 mmol/l) with/without different stimuli (high glucose, TNF-alpha, VEGF, wortmannin, LY-294002). Apoptosis, proliferation, cell cycle distribution, and protein expression were determined by DNA fragmentation assays, [(3)H]thymidine incorporation, FACS, and Western blot analyses, respectively. In macro- and microvascular EC, LA (1 mmol/l) reduced (P < 0.05) basal (macEC, -36 +/- 4%; micEC, -46 +/- 6%) and stimulus-induced (TNF-alpha: macEC, -75 +/- 11%; micEC, -68 +/- 13%) apoptosis. In HUVEC, inhibition of apoptosis by LA (500 mumol/l) was paralleled by reduction of NF-kappaB. LA's antiapoptotic activity was reduced by PI 3-kinase inhibitors (wortmannin, LY-294002), being in line with LA-induced Akt phosphorylation (Ser(437), +159 +/- 43%; Thr(308), +98 +/- 25%; P < 0.01). LA (500 mumol/l) inhibited (P < 0.001) proliferation of macEC (-29 +/- 3%) and micEC (-29 +/- 3%) by arresting the cells at the G(1)/S transition due to an increased ratio of cyclin E/p27(Kip) (4.2-fold), upregulation of p21(WAF-1/Cip1) (+104 +/- 21%), and reduction of cyclin A (-32 +/- 11%), of hyperphosphorylated retinoblastoma protein (macEC: -51 +/- 7%; micEC: -50 +/- 15%), and of E2F-1 (macEC: -48 +/- 3%; micEC: -31 +/- 10%). LA's ability to inhibit apoptosis and proliferation of ECs could beneficially affect endothelial dysfunction, which precedes manifestation of late diabetic vascular complications.     

Prenatal betamethasone exposure results in pituitary-adrenal hyporesponsiveness in adult sheep

Fetal exposure to synthetic glucocorticoids in sheep results in increased fetal hypothalamic-pituitary-adrenal (HPA) activity persisting to one year of age. We aimed to determine the effects of single or repeated maternal or fetal betamethasone injections on offspring HPA activity at 2 and 3 yr of age and whether changes in adrenal mediators of steroidogenesis contribute to changes in pituitary-adrenal function. Pregnant ewes or their fetuses received either repeated intramuscular saline or betamethasone injections (0.5 mg/kg) at 104, 111, 118, and 124 days of gestation (dG) or a single betamethasone injection at 104 dG followed by saline at 111, 118, and 124 dG. Offspring were catheterized at 2 and 3 yr of age and given corticotrophin-releasing hormone + arginine vasopressin challenges. Adrenal tissue was collected for quantitative RT-PCR mRNA determination at 3.5 yr of age. In 2-yr-old offspring, maternal betamethasone injections did not alter basal ACTH or cortisol levels, but repeated injections elevated ACTH responses. At 3 yr of age, basal ACTH was elevated, and both basal and stimulated cortisol levels were suppressed by repeated maternal injections. Basal and stimulated cortisol-to-ACTH ratios and basal cortisol-to-cytochrome P-450 17alpha-hydroxylase (P450c17) mRNA ratios were suppressed by repeated injections. Repeated fetal betamethasone injections attenuated basal ACTH and cortisol levels in offspring at 2 but not 3 yr of age. Plasma changes were not associated with altered adrenal P450c17, ACTH receptor, beta-hydroxysteroid dehydrogenase, or glucocorticoid receptor mRNA levels. These data suggest that maternal, but not fetal, betamethasone administration results in adrenal suppression in adulthood.     

Consequences of Melaleuca quinquenervia Invasion on Soil Nematodes in the Florida Everglades

The tree Melaleuca quinquenervia invades all types of habitats of South Florida leading to up to 80% loss of aboveground diversity. To examine impacts on the belowground ecosystem, we investigated the composition and diversity of nematodes from soils dominated by the invasive tree and compared them with soils supporting native plant communities at six locations across the Florida Everglades over three years. Despite the significant differences in soil type, hydrology, and native plant composition of the sites, there were consistent differences in nematode communities between soil environments under the native and invaded plant communities. The total abundance and diversity of nematodes in soils dominated by M. quinquenervia was 60% and 80% of adjacent soils under native plants. Fungal-feeding and plant-parasitic nematodes were twice as abundant under native plants as under M. quinquenervia. Nematode communities under M. quinquenervia were bacterivore-dominated, while under native vegetation plant-parasite dominated. The overall diversity of nematodes was 20% lower under the exotic than under native plants, with plant parasites being 36% and fungivores being 30% less diverse. Soil moisture, % of Ca, Mg, and clay particles and total soil C and N were greater in M. quinquenervia soils, but plant-available concentrations of P, K, Ca, and Mg as well as CEC were reduced. Overall, data suggests that the invasion process may modify soil biotic and abiotic conditions that in turn promote the advancement of the exotic M. quinquenervia and displacement of the native plants.     

DAT/SERT selectivity of flexible GBR 12909 analogs modeled using 3D-QSAR methods

The dopamine reuptake inhibitor GBR 12909 (1-{2-[bis(4-fluorophenyl)methoxy]ethyl}-4-(3-phenylpropyl)piperazine, 1) and its analogs have been developed as tools to test the hypothesis that selective dopamine transporter (DAT) inhibitors will be useful therapeutics for cocaine addiction. This 3D-QSAR study focuses on the effect of substitutions in the phenylpropyl region of 1. CoMFA and CoMSIA techniques were used to determine a predictive and stable model for the DAT/serotonin transporter (SERT) selectivity (represented by pK(i) (DAT/SERT)) of a set of flexible analogs of 1, most of which have eight rotatable bonds. In the absence of a rigid analog to use as a 3D-QSAR template, six conformational families of analogs were constructed from six pairs of piperazine and piperidine template conformers identified by hierarchical clustering as representative molecular conformations. Three models stable to y-value scrambling were identified after a comprehensive CoMFA and CoMSIA survey with Region Focusing. Test set correlation validation led to an acceptable model, with q(2)=0.508, standard error of prediction=0.601, two components, r(2)=0.685, standard error of estimate=0.481, F value=39, percent steric contribution=65, and percent electrostatic contribution=35. A CoMFA contour map identified areas of the molecule that affect pK(i) (DAT/SERT). This work outlines a protocol for deriving a stable and predictive model of the biological activity of a set of very flexible molecules.     

New bioinformatic tools for analysis of nucleotide modifications in eukaryotic rRNA

This report presents a valuable new bioinformatics package for research on rRNA nucleotide modifications in the ribosome, especially those created by small nucleolar RNA:protein complexes (snoRNPs). The interactive service, which is not available elsewhere, enables a user to visualize the positions of pseudouridines, 2'-O-methylations, and base methylations in three-dimensional space in the ribosome and also in linear and secondary structure formats of ribosomal RNA. Our tools provide additional perspective on where the modifications occur relative to functional regions within the rRNA and relative to other nearby modifications. This package of new tools is presented as a major enhancement of an existing but significantly upgraded yeast snoRNA database available publicly at http://people.biochem.umass.edu/sfournier/fournierlab/snornadb/. The other key features of the enhanced database include details of the base pairing of snoRNAs with target RNAs, genomic organization of the yeast snoRNA genes, and information on corresponding snoRNAs and modifications in other model organisms.     

Schistosoma mekongi and Schistosoma japonicum: Differences in the distribution of eggs in the viscera of mice

The difference in the distribution of Schistosoma eggs in the viscera has not been clearly elucidated in the two closely related species Schistosoma japonicum and Schistosoma mekongi. In this study, we quantitatively compared the distribution of eggs in mice infected with the two species. In S. mekongi-infected mice, 56.6% to 69.4% of total eggs were found in the distal small intestine 9 to 15 weeks after infection, while in S. japonicum-infected mice, 48.8% to 71.8% of eggs were found in the proximal small intestine during the same period. There were significantly more eggs in the liver in mice infected with S. japonicum than in those infected with S. mekongi. The number of adult worms recovered did not differ between the two species during the study period. The total number of eggs laid in the tissues also did not differ between the two species at 12 to 15 weeks postinfection, but in the earlier period the total number of eggs was significantly fewer in S. mekongi-infected than in S. japonicum-infected mice, suggesting the delayed maturation of the former compared with the latter. These results clearly show that S. japonicum and S. mekongi exhibit different oviposition behavior in their hosts.