NS2 Proteins of GB Virus B and Hepatitis C Virus Share Common Protease Activities and Membrane Topologies.

GB virus B (GBV-B), which is hepatotropic in experimentally infected small New World primates, is a member of the Hepacivirus genus but phylogenetically relatively distant from hepatitis C virus (HCV). To gain insights into the role and specificity of hepaciviral nonstructural protein 2 (NS2), which is required for HCV polyprotein processing and particle morphogenesis, we investigated whether NS2 structural and functional features are conserved between HCV and GBV-B. We found that GBV-B NS2, like HCV NS2, has cysteine protease activity responsible for cleavage at the NS2/NS3 junction, and we experimentally confirmed the location of this junction within the viral polyprotein. A model for GBV-B NS2 membrane topology was experimentally established by determining the membrane association properties of NS2 segments fused to green fluorescent protein (GFP) and their nuclear magnetic resonance structures using synthetic peptides as well as by applying an N-glycosylation scanning approach. Similar glycosylation studies confirmed the HCV NS2 organization. Together, our data show that despite limited amino acid sequence similarity, GBV-B and HCV NS2 proteins share a membrane topology with 3 N-terminal transmembrane segments, which is also predicted to apply to other recently discovered hepaciviruses. Based on these data and using trans-complementation systems, we found that intragenotypic hybrid NS2 proteins with heterologous N-terminal membrane segments were able to efficiently trans-complement an assembly-deficient HCV mutant with a point mutation in the NS2 C-terminal domain, while GBV-B/HCV or intergenotypic NS2 chimeras were not. These studies indicate that virus- and genotype-specific intramolecular interactions between N- and C-terminal domains of NS2 are critically involved in HCV morphogenesis. IMPORTANCE: Nonstructural protein 2 (NS2) of hepatitis C virus (HCV) is a multifunctional protein critically involved in polyprotein processing and virion morphogenesis. To gain insights into NS2 mechanisms of action, we investigated whether NS2 structural and functional features are conserved between HCV and GB virus B (GBV-B), a phylogenetically relatively distant primate hepacivirus. We showed that GBV-B NS2, like HCV NS2, carries cysteine protease activity. We experimentally established a model for GBV-B NS2 membrane topology and demonstrated that despite limited sequence similarity, GBV-B and HCV NS2 share an organization with three N-terminal transmembrane segments. We found that the role of HCV NS2 in particle assembly is genotype specific and relies on critical interactions between its N- and C-terminal domains. This first comparative analysis of NS2 proteins from two hepaciviruses and our structural predictions of NS2 from other newly identified mammal hepaciviruses highlight conserved key features of the hepaciviral life cycle.

Student's watcher : comparison between ASP.NET and PHP+CSS

"Student’s Watcher” is a small Web application which wants to show in a visual, simple and fast way, the evolution of the students. The main project table displays such things as marks and comments about students. We can add a comment for each mark to explain why this mark. The objective is to be able to know if some student has a problem, how is going his year, marks in other courses, or even, to know if he has a bad week in a different subjects. We can see the evolution of students in past years to do an objective comparison. It also allows inserting global comments of student, we have a list of these, and all professors can add new ones, where we can see more general valuations. “Student’s Watcher” was begun in ASP.net, but finally my project would be developed in PHP, HTML and CSS. This project wants to be a comparison between two of most important languages used nowadays, ASPX and PHP

Aider les étudiants en médecine à approcher la mort [Helping medical students to approach death].

A project recently launched by the Faculty of biology and medicine of Lausanne introduces the approach of facing death during both the dissection and the course of clinical activities. Existential questions relating to mortality are bound to arise sooner or later during the course of the study. For the sake of humanized clinical practice, these questions must be confronted. In response to a request by a student association, an accompanying curriculum with active student's contribution through encounters with death in anatomy and clinical situations was created in Lausanne. Students will benefit from this new program throughout their curriculum. This program is the first of its kind in Switzerland.

The sandfly fauna, anthropophily and the seasonal activities of Pintomyia spinicrassa (Diptera: Psychodidae: Phlebotominae) in a focus of cutaneous leishmaniasis in northeastern Colombia

This study was conducted to identify the sandfly fauna and the anthropophilic species in a coffee-growing area of Villanueva, Norte de Santander, Colombia, a focus of American cutaneous leishmaniasis, and to analyse the relationship between the most frequent species and rainfall, relative humidity and temperature, with the aim of contributing to epidemiological surveillance in the area. Sandfly collections were performed fortnightly between February 2006-September 2007 using automatic light traps, Shannon traps, protected human bait and aspiration in resting places. A total of 7,051 sandflies belonging to 12 species were captured. Pintomyia spinicrassa (95.7%) predominated. Pintomyia oresbia and Lutzomyia sp. of Pichinde were found in the state of Norte de Santander for the first time. Pi. spinicrassa, Pintomyia nuneztovari, Micropygomyia venezuelensis, Lutzomyia (Helcocyrtomyia) scorzai and Lu. (Helcocyrtomyia) sp. were captured on the protected human bait. A significant association between Pi. spinicrassa abundance and the total rainfall and the average temperature and humidity 10 days before the collection was observed. The dominance of Pi. spinicrassa, a recognised vector of Leishmania braziliensis, especially during the dry periods, indicates that the risk of parasite transmission may increase.

Using free data mining software and clustering algorithms to find predictors from student qualifications

In this project a research both in finding predictors via clustering techniques and in reviewing the Data Mining free software is achieved. The research is based in a case of study, from where additionally to the KDD free software used by the scientific community; a new free tool for pre-processing the data is presented. The predictors are intended for the e-learning domain as the data from where these predictors have to be inferred are student qualifications from different e-learning environments. Through our case of study not only clustering algorithms are tested but also additional goals are proposed.

Characterization of a plant-derived peptide displaying water clarifying and antimicrobial activities

SUMMARY Drinking water is currently a scarce world resource, the preparation of which requires complex treatments that include clarification of suspended particles and disinfection. Seed extracts of Moringa oleifera Lam., a tropical tree, have been proposed as an environment- friendly alternative, due to their traditional use for the clarification of drinking water. However, the precise nature of the active components was unknown. Here, we show that recombinant or synthetic forms of a cationic seed polypeptide mediate efficient sedimentation of suspended mineral particles and bacteria. Unexpectedly, the polypeptide was also found to possesses a bactericidal activity capable of disinfecting heavily contaminated water. Furthermore, the polypeptide has been shown to efficiently kill several pathogenic bacteria, including antibiotic-resistant isolates of Pseudomona, Streptococcus and Legionella species. Structural modeling of the peptide coupled to the functional analysis of synthetic peptide derivatives delineated distinct structural determinants for the flocculation and antibacterial activities. Our results suggest that a glutamine-rich portion of the polypeptide is involved in the sedimentation process; alternatively, the antibacterial activity depends on a amphiphilic loop. Assembly of multiple copies of this loop into a branched peptide derivative strongly enhances antibacterial activity without displaying hemolytic effect. In conclusion, this polypeptide displays the unprecedented feature of combining efficient water purification and disinfectant properties indicating different molecular mechanisms involved in each case. This work not only identified the features responsible for these activities but also provides useful information that has implications for the further development of this cationic polypeptide as a potent antibacterial agent. RESUME L'eau potable est actuellement une ressource limitée dans le monde. La production d'eau propre à la consommation exige des traitements complexes, incluant la clarification des particules en suspension ainsi que sa désinfection par des additifs chimiques. Les extraits de la graine d'un arbre tropical, Moringa oleifera, sont utilisés traditionnellement en Afrique afin de clarifier l'eau. Quoique la nature exacte des composants actifs était inconnue, on a pu mettre en évidence un polypeptide cationique contenu dans ces graines, capable de sédimenter de manière efficace des particules minérales en suspension ainsi que des bactéries. Ce travail a aussi mis en évidence que ce polypeptide a une activité bactéricide, permettant une désinfection d'eau fortement contaminée. De plus, nous avons démontré que ce polypeptide est efficace contre de nombreuses souches bactériennes pathogènes, également celles résistantes aux antibiotiques comme Pseudomonas, Streptococcus et Legionella. L'analyse de la structure moléculaire de ce polypeptide, couplée à son analyse fonctionnelle a mis en évidence deux domaines structuraux distinct, un pour l'activité de floculation et l'autre pour l'activité antibactérienne. Nos résultats suggèrent que le domaine riche en glutamine est impliqué dans le processus de sédimentation et que l'activité antimicrobienne dépend d'un domaine formant une boucle amphiphilique. En ramifiant plusieurs copies de cette boucle on a pu augmenter de manière significative l'activité antibactérienne. En conclusion, nous avons pu démontrer que ce polypeptide à la capacité unique de combiner des propriétés de purification et de désinfection de l'eau, ce qui implique des mécanismes moléculaires distincts pour ces deux activités. Ce travail a permis d'identifier les domaines du polypeptide qui sont responsables de ses activités et offre une perspective pour le développement d'un nouvel agent antimicrobien.

Anthropophilic biting behaviour of Anopheles (Kerteszia) neivai Howard, Dyar & Knab associated with Fishermen’s activities in a malaria-endemic area in the Colombian Pacific

On the southwest Pacific Coast of Colombia, a field study was initiated to determine the human-vector association between Anopheles (Kerteszia) neivai and fishermen, including their nearby houses. Mosquitoes were collected over 24-h periods from mangrove swamps, marshlands and fishing vessels in three locations, as well as in and around the houses of fishermen. A total of 6,382 mosquitoes were collected. An. neivai was most abundant in mangroves and fishing canoes (90.8%), while Anopheles albimanus was found indoors (82%) and outdoors (73%). One An. neivai and one An. albimanus collected during fishing activities in canoes were positive for Plasmodium vivax , whereas one female An. neivai collected in a mangrove was positive for P. vivax . In the mangroves and fishing canoes, An. neivai demonstrated biting activity throughout the day, peaking between 06:00 pm-07:00 pm and there were two minor peaks at dusk and dawn. These peaks coincided with fishing activities in the marshlands and mangroves, a situation that places the fishermen at risk of contracting malaria when they are performing their daily activities. It is recommended that protective measures be implemented to reduce the risk that fishermen will contract malaria.

Megazol and its bioisostere 4H-1,2,4-triazole: comparing the trypanocidal, cytotoxic and genotoxic activities and their in vitro and in silico interactions with the Trypanosoma brucei nitroreductase enzyme

Megazol (7) is a 5-nitroimidazole that is highly active against Trypanosoma cruzi and Trypanosoma brucei, as well as drug-resistant forms of trypanosomiasis. Compound 7 is not used clinically due to its mutagenic and genotoxic properties, but has been largely used as a lead compound. Here, we compared the activity of 7 with its 4H-1,2,4-triazole bioisostere (8) in bloodstream forms of T. brucei and T. cruzi and evaluated their activation by T. brucei type I nitroreductase (TbNTR) enzyme. We also analysed the cytotoxic and genotoxic effects of these compounds in whole human blood using Comet and fluorescein diacetate/ethidium bromide assays. Although the only difference between 7 and 8 is the substitution of sulphur (in the thiadiazole in 7) for nitrogen (in the triazole in 8), the results indicated that 8 had poorer antiparasitic activity than 7 and was not genotoxic, whereas 7 presented this effect. The determination of Vmax indicated that although 8 was metabolised more rapidly than 7, it bounds to the TbNTR with better affinity, resulting in equivalent kcat/KM values. Docking assays of 7 and 8 performed within the active site of a homology model of the TbNTR indicating that 8 had greater affinity than 7.

The effects of CYP2D6 and CYP3A activities on the pharmacokinetics of immediate release oxycodone.

BACKGROUND AND PURPOSE: There is high interindividual variability in the activity of drug-metabolizing enzymes catalysing the oxidation of oxycodone [cytochrome P450 (CYP) 2D6 and 3A], due to genetic polymorphisms and/or drug-drug interactions. The effects of CYP2D6 and/or CYP3A activity modulation on the pharmacokinetics of oxycodone remains poorly explored. EXPERIMENTAL APPROACH: A randomized crossover double-blind placebo-controlled study was performed with 10 healthy volunteers genotyped for CYP2D6 [six extensive (EM), two deficient (PM/IM) and two ultrarapid metabolizers (UM)]. The volunteers randomly received on five different occasions: oxycodone 0.2 mg x kg(-1) and placebo; oxycodone and quinidine (CYP2D6 inhibitor); oxycodone and ketoconazole (CYP3A inhibitor); oxycodone and quinidine+ketoconazole; placebo. Blood samples for plasma concentrations of oxycodone and metabolites (oxymorphone, noroxycodone and noroxymorphone) were collected for 24 h after dosing. Phenotyping for CYP2D6 (with dextromethorphan) and CYP3A (with midazolam) were assessed at each session. KEY RESULTS: CYP2D6 activity was correlated with oxymorphone and noroxymorphone AUCs and C(max) (-0.71 < Spearman correlation coefficient rhos < -0.92). Oxymorphone C(max) was 62% and 75% lower in PM than EM and UM. Noroxymorphone C(max) reduction was even more pronounced (90%). In UM, oxymorphone and noroxymorphone concentrations increased whereas noroxycodone exposure was halved. Blocking CYP2D6 (with quinidine) reduced oxymorphone and noroxymorphone C(max) by 40% and 80%, and increased noroxycodone AUC(infinity) by 70%. Blocking CYP3A4 (with ketoconazole) tripled oxymorphone AUC(infinity) and reduced noroxycodone and noroxymorphone AUCs by 80%. Shunting to CYP2D6 pathway was observed after CYP3A4 inhibition. CONCLUSIONS AND IMPLICATIONS: Drug-drug interactions via CYP2D6 and CYP3A affected oxycodone pharmacokinetics and its magnitude depended on CYP2D6 genotype.

Blood shizonticidal activities of phenazines and naphthoquinoidal compounds against Plasmodium falciparum in vitro and in mice malaria studies

Due to the recent advances of atovaquone, a naphthoquinone, through clinical trials as treatment for malarial infection, 19 quinone derivatives with previously reported structures were also evaluated for blood schizonticide activity against the malaria parasite Plasmodium falciparum. These compounds include 2-hydroxy-3-methylamino naphthoquinones (2-9), lapachol (10), nor-lapachol (11), iso-lapachol (12), phthiocol (13) and phenazines (12-20). Their cytotoxicities were also evaluated against human hepatoma and normal monkey kidney cell lines. Compounds 2 and 5 showed the highest activity against P. falciparum chloroquine-resistant blood-stage parasites (clone W2), indicated by their low inhibitory concentration for 50% (IC50) of parasite growth. The therapeutic potential of the active compounds was evaluated according to the selectivity index, which is a ratio of the cytotoxicity minimum lethal dose which eliminates 50% of cells and the in vitro IC50. Naphthoquinones 2 and 5, with activities similar to the reference antimalarial chloroquine, were also active against malaria in mice and suppressed parasitaemia by more than 60% in contrast to compound 11 which was inactive. Based on their in vitro and in vivo activities, compounds 2 and 5 are considered promising molecules for antimalarial treatment and warrant further study.

BDNF overexpression in mouse hippocampal astrocytes promotes local neurogenesis and elicits anxiolytic-like activities.

The therapeutic activity of selective serotonin (5-HT) reuptake inhibitors (SSRIs) relies on long-term adaptation at pre- and post-synaptic levels. The sustained administration of SSRIs increases the serotonergic neurotransmission in response to a functional desensitization of the inhibitory 5-HT1A autoreceptor in the dorsal raphe. At nerve terminal such as the hippocampus, the enhancement of 5-HT availability increases brain-derived neurotrophic factor (BDNF) synthesis and signaling, a major event in the stimulation of adult neurogenesis. In physiological conditions, BDNF would be expressed at functionally relevant levels in neurons. However, the recent observation that SSRIs upregulate BDNF mRNA in primary cultures of astrocytes strongly suggest that the therapeutic activity of antidepressant drugs might result from an increase in BDNF synthesis in this cell type. In this study, by overexpressing BDNF in astrocytes, we balanced the ratio between astrocytic and neuronal BDNF raising the possibility that such manipulation could positively reverberate on anxiolytic-/antidepressant-like activities in transfected mice. Our results indicate that BDNF overexpression in hippocampal astrocytes produced anxiolytic-/antidepressant-like activity in the novelty suppressed feeding in relation with the stimulation of hippocampal neurogenesis whereas it did not potentiate the effects of the SSRI fluoxetine on these parameters. Moreover, overexpressing BDNF revealed the anxiolytic-like activity of fluoxetine in the elevated plus maze while attenuating 5-HT neurotransmission in response to a blunted downregulation of the 5-HT1A autoreceptor. These results emphasize an original role of hippocampal astrocytes in the synthesis of BDNF, which can act through neurogenesis-dependent and -independent mechanisms to regulate different facets of anxiolytic-like responses.

Sandfly species diversity in association with human activities in the Kani tribe settlements of the Western Ghats, Thiruvananthapuram, Kerala, India

Sandfly prevalence in the Kani tribe settlements of Western Ghats in India was investigated. A total of 1,279 sandflies comprising 17 species was obtained. Sandfly abundance showed a negative correlation (r = -0.97, p = 0.003) with increase in altitudinal ranges from 0-1,000 m. When sandfly samples were grouped according to landscape characteristics of the location, the estimated Shannon-Weiner index (H) and species richness index (S) were high and species evenness index (J) was low in settlements located at 0-300 m altitudinal range. On the contrary, the values of H and J were high, while S was low at 301-600 m altitudinal range. With further increase in altitude, species diversity, S and J were low. Though the relative abundance of sandflies decreased with increase in altitude, the influence of altitudinal variation could not be attributed to determine sandfly diversity, since the number of sampling units were not uniform at all the altitudinal gradients due to nonavailability of suitable resting shelters. Sandfly species showed great aggregation at 0-300 m altitude interval, where not only the number of settlements were maximum (n = 19), but also the environmental conditions favoured sandfly abundance due to the concentration of tribal settlements, human dwellings and his activities.