The identification and differentiation of the Candida parapsilosis complex species by polymerase chain reaction-restriction fragment length polymorphism of the internal transcribed spacer region of the rDNA

Currently, it is accepted that there are three species that were formerly grouped under Candida parapsilosis: C. para- psilosis sensu stricto, Candida orthopsilosis, andCandida metapsilosis. In fact, the antifungal susceptibility profiles and distinct virulence attributes demonstrate the differences in these nosocomial pathogens. An accurate, fast, and economical identification of fungal species has been the main goal in mycology. In the present study, we searched sequences that were available in the GenBank database in order to identify the complete sequence for the internal transcribed spacer (ITS)1-5.8S-ITS2 region, which is comprised of the forward and reverse primers ITS1 and ITS4. Subsequently, an in silico polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed to differentiate the C. parapsilosis complex species. Ninety-eight clinical isolates from patients with fungaemia were submitted for analysis, where 59 isolates were identified as C. parapsilosis sensu stricto, 37 were identified as C. orthopsilosis, and two were identified as C. metapsilosis. PCR-RFLP quickly and accurately identified C. parapsilosis complex species, making this method an alternative and routine identification system for use in clinical mycology laboratories.

Complete genome sequence of Deltapapillomavirus 4 (bovine papillomavirus 2) from a bovine papillomavirus lesion in Amazon Region, Brazil

The complete genome sequence of bovine papillomavirus 2 (BPV2) from Brazilian Amazon Region was determined using multiple-primed rolling circle amplification followed by Illumina sequencing. The genome is 7,947 bp long, with 45.9% GC content. It encodes seven early (E1, E2,E4, E5, E6,E7, and E8) and two late (L1 and L2) genes. The complete genome of a BPV2 can help in future studies since this BPV type is highly reported worldwide although the lack of complete genome sequences available.

Impaired adipose tissue expandability and lipogenic capacities as ones of the main causes of metabolic disorders.

Obesity is considered a major health problem. However, mechanisms involved and its comorbidities are not elucidated. Recent theories concerning the causes of obesity have focused on a limit to the functional capacity of adipose tissue, comparing it with other vital organs. This assumption has been the central point of interest in our laboratory. We proposed that the failure of adipose tissue is initiated by the difficulty of this tissue to increase its cellularity due to excess in fat contribution, owing to genetic or environmental factors. Nevertheless, why the adipose tissue reduces its capacity to make new adipocytes via mesenchymal cells of the stroma has not yet been elucidated. Thus, we suggest that this tissue ceases fulfilling its main function, the storage of excess fat, thereby affecting some of the key factors involved in lipogenesis, some of which are reviewed in this paper (PPARγ, ROR1, FASN, SCD1, Rab18, BrCa1, ZAG, and FABP4). On the other hand, mechanisms involved in adipose tissue expandability are also impaired, predominating hypertrophy via an increase in apoptosis and a decrease in adipogenesis and angiogenesis. However, adipose tissue failure is only part of this great orchestra, only a chapter of this nightmare.

Nucleotide sequence of the 5' noncoding region and part of the gag gene of mouse mammary tumor virus; identification of the 5' splicing site for subgenomic mRNAs.

We have determined the sequence of the first 1371 nucleotides at the 5' end of the genome of mouse mammary tumor virus using molecularly cloned proviral DNA of the GR virus strain. The most likely initiation codon used for the gag gene of mouse mammary tumor virus is the first one, located 312 nucleotides from the 5' end of the viral RNA. The 5' splicing site for the subgenomic mRNA's is located approximately 288 nucleotides downstream from the 5' end of the viral RNA. From the DNA sequence the amino acid sequence of the N-terminal half of the gag precursor protein, including p10 and p21, was deduced (353 amino acids).

Rate of gene sequence evolution and species diversification in flowering plants: a re-evaluation

Barraclough and co-workers (in a paper published in 1996) observed that there was a significant positive correlation between the rate of evolution of the rbcL chloroplast gene within families of flowering plants and the number of species in those families. We tested three additional data sets of our own (based on both plastid and nuclear genes) and used methods designed specifically for the comparison of sister families (based on random speciation and extinction). We show that, over all sister groups, the correlation between the rate of gene evolution and an increased diversity is not always present. Despite tending towards a positive association, the observation of individual probabilities presents a U-shaped distribution of association (i.e. it can be either significantly positive or negative). We discuss the influence of both phylogenetic sampling and applied taxonomies on the results.

Advances in Sequence Analysis: Theory, Methods, Applications

This book gives a general view of sequence analysis, the statistical study of successions of states or events. It includes innovative contributions on life course studies, transitions into and out of employment, contemporaneous and historical careers, and political trajectories. The approach presented in this book is now central to the life-course perspective and the study of social processes more generally. This volume promotes the dialogue between approaches to sequence analysis that developed separately, within traditions contrasted in space and disciplines. It includes the latest developments in sequential concepts, coding, atypical datasets and time patterns, optimal matching and alternative algorithms, survey optimization, and visualization. Field studies include original sequential material related to parenting in 19th-century Belgium, higher education and work in Finland and Italy, family formation before and after German reunification, French Jews persecuted in occupied France, long-term trends in electoral participation, and regime democratization. Overall the book reassesses the classical uses of sequences and it promotes new ways of collecting, formatting, representing and processing them. The introduction provides basic sequential concepts and tools, as well as a history of the method. Chapters are presented in a way that is both accessible to the beginner and informative to the expert.

Cytotoxic T cells deficient in both functional fas ligand and perforin show residual cytolytic activity yet lose their capacity to induce lethal acute graft-versus-host disease.

Graft-versus-host disease (GVHD) is the main complication after allogeneic bone marrow transplantation. Although the tissue damage and subsequent patient mortality are clearly dependent on T lymphocytes present in the grafted inoculum, the lethal effector molecules are unknown. Here, we show that acute lethal GVHD, induced by the transfer of splenocytes from C57BL/6 mice into sensitive BALB/c recipients, is dependent on both perforin and Fas ligand (FasL)-mediated lytic pathways. When spleen cells from mutant mice lacking both effector molecules were transferred to sublethally irradiated allogeneic recipients, mice survived. Delayed mortality was observed with grafted cells deficient in only one lytic mediator. In contrast, protection from lethal acute GVHD in resistant mice was exclusively perforin dependent. Perforin-FasL-deficient T cells failed to lyse most target cells in vitro. However, they still efficiently killed tumor necrosis factor alpha-sensitive fibroblasts, demonstrating that cytotoxic T cells possess a third lytic pathway.

Quantitative sequence analysis of FBN1 premature termination codons provides evidence for incomplete NMD in leukocytes.

We improved, evaluated, and used Sanger sequencing for quantification of single nucleotide polymorphism (SNP) variants in transcripts and gDNA samples. This improved assay resulted in highly reproducible relative allele frequencies (e.g., for a heterozygous gDNA 50.0+/-1.4%, and for a missense mutation-bearing transcript 46.9+/-3.7%) with a lower detection limit of 3-9%. It provided excellent accuracy and linear correlation between expected and observed relative allele frequencies. This sequencing assay, which can also be used for the quantification of copy number variations (CNVs), methylations, mosaicisms, and DNA pools, enabled us to analyze transcripts of the FBN1 gene in fibroblasts and blood samples of patients with suspected Marfan syndrome not only qualitatively but also quantitatively. We report a total of 18 novel and 19 known FBN1 sequence variants leading to a premature termination codon (PTC), 26 of which we analyzed by quantitative sequencing both at gDNA and cDNA levels. The relative amounts of PTC-containing FBN1 transcripts in fresh and PAXgene-stabilized blood samples were significantly higher (33.0+/-3.9% to 80.0+/-7.2%) than those detected in affected fibroblasts with inhibition of nonsense-mediated mRNA decay (NMD) (11.0+/-2.1% to 25.0+/-1.8%), whereas in fibroblasts without NMD inhibition no mutant alleles could be detected. These results provide evidence for incomplete NMD in leukocytes and have particular importance for RNA-based analyses not only in FBN1 but also in other genes.

Didactique de la géographie et formation initiale des enseignants spécialistes : conception et première évaluation du nouveau dispositif de formation initiale des enseignants de géographie du Secondaire supérieur à la HEP Vaud

Geography as a school subject is specifically thought for and by the schools. The contents of the school subject, nowadays, do not reflect the concerns and the evolution of the discipline as such. Nevertheless, official curricula set school objectives that address issues affecting the world and people's lives. These issues are coherent with the ones addressed by geography as a social science, that is to say the study of how people and their environment interact and how societies are interconnected through space. On an every day basis, Geography as a school subject is most of the time reduced to accumulating knowledge outside any given context. This knowledge may even be partially untrue or old and the related activities focus on low cognitive tensions. These practices do not contribute to the learners' understanding of the world because it does not allow them to build a geographical competence, which they. will need as future citizens in order to make responsible choices when they are confronted to questions related to how the locations of human and physical features are influenced by each other and how they interact across space. The central part of the text relies on the ideas and the processes discussed in the publications, which constitute the published file; it is divided into two parts. The first part (chapter 4) presents a didactic approach, which gives meaningful insights into Geography as a school subject and shows a brief account of the theoretical background that supports it. This socio-constructivist approach relies on the main following features: a priming stage (élément déclencheur), which presents geographical knowledge as an issue to be explored, discussed or solved; the issue is given to learners;. the planning of the teaching-learning sequence in small units launched by the main issue in the priming stage ; the interconnections of geographical knowledge with integrative concepts ; the synthetic stage or reporting stage where final concepts and knowledge are put together in order to be learned. Such an approach allows learners to re-invest the knowledge they have built themselves. This knowledge is organised by geographical integrative concepts, which represent true thinking operative tools and with which key issues in the geographical thinking are associated. The second part of the text (chapter 5) displays the didactic principles that governed the conception of the new initial training course for the future upper secondary school teachers at the HEP Vaud. The ambition of this course is to prepare future teachers to plan and realize the teaching of geography that provides pupils with the tools to understand better how people and their environment interact and how societies are interconnected through space. One of the tools for the teachers is the conceptual framework, whose most salient interest is to be relevant at every stage of the preparation and planning of the teaching, including the necessary epistemological reflection that should always be present. The synthesis of the text starts with a short account of the first evaluation of the new course. Various reflections on the future concerns and issues, that the didactics and methodology of Geography will be confronted with, constitute the synthesis.

Structural and metamorphic evolution of the northern Himachal Himalaya, NW India - (Spiti-eastern Lahul-Parvati valley traverse)

The Himalayan orogen is the result of the collision between the Indian and Asian continents that began 55-50 Ma ago, causing intracontinental thrusting and nappe formation. Detailed mapping as well as structural and microfabric analyses on a traverse from the Tethyan Himalaya southwestward through the High Himalayan Crystalline and the Main Central Thrust zone (MCT zone) to the Lesser Himalayan Sequence in the Spiti-eastern Lahul-Parvati valley area reveal eight main phases of deformation, a series of late stage phases and five stages of metamorphic crystallization. This sequence of events is integrated into a reconstruction of the tectonometamorphic evolution of the Himalayan orogen in northern Himachal Pradesh. The oldest phase D-1 is preserved as relies in the High Himalayan Crystalline. Its deformational conditions are poorly known, but the metamorphic evolution is well documented by a prograde metamorphism reaching peak conditions within the upper amphibolite facies. This indicates that D-1 was an important tectonometamorphic event including considerable crustal thickening. The structural, metamorphic and sedimentary record suggest that D-1 most probably represents an early stage of continental collision. The first event clearly attributed to the collision between India and Asia is documented by two converging nappe systems, the NE-verging Shikar Beh Nappe and the SW-verging north Himalayan nappes. The D-2 Shikar Beh Nappe is characterized by isoclinal folding and top-to-the NE shearing, representing the main deformation in the High Himalayan Crystalline. D-2 also caused the main metamorphism in the High Himalayan Crystalline that was of a Barrovian-type, reaching upper amphibolite facies peak conditions. The Shikar Beh Nappe is interpreted to have formed within the Indian crust SW of the subduction zone. Simultaneously with NE-directed nappe formation, incipient subduction of India below Asia caused stacking of the SW-verging north Himalayan Nappes, that were thrust from the northern edge of the subducted continent toward the front of the Shikar Beh Nappe. As a result, the SW-verging folds of the D-3 Main Fold Zone formed in the Tethyan Himalaya below the front of the north Himalayan nappes. D-3 represents the main deformation in the Tethyan Himalaya, associated with a greenschist facies metamorphism. Folding within the Main Fold Zone subsequently propagated toward SW into the High Himalayan Crystalline, where it overprinted the preexisting D-2 structures. After subduction at the base of the north Himalayan nappes, the subduction zone stepped to the base of the High Himalayan Crystalline, where D-3 folds were crosscut by SW-directed D-4 thrusting. During D-4, the Crystalline Nappe, comprising the Main Fold Zone and relies of the Shikar Beh Nappe was thrust toward SW over the Lesser Himalayan Sequence along the 4 to 5 kms thick Main Central Thrust zone. Thrusting was related to a retrograde greenschist facies overprint at the base of the Crystalline Nappe and to pro-grade greenschist facies conditions in the Lesser Himalayan Sequence. Simultaneously with thrusting at the base of the Crystalline Nappe, higher crustal levels were affected by NE-directed D-5 normal extensional shearing and by dextral strike-slip motion, indicating that the high-grade metamorphic Crystalline Nappe was extruded between the low-grade metamorphic Lesser Himalayan Sequence at the base and the north Himalayan nappes at the top. The upper boundary of the Crystalline Nappe is not clearly delimited and passes gradually into the low-grade rocks at the front of the north Himalayan nappes. Extrusion of the Crystalline Nappe was followed by the phase D-6, characterized by large-scale, upright to steeply inclined, NE-verging folds and by another series of normal and extensional structures D-7+D-8 that may be related to ongoing extrusion of the Crystalline Nappe. The late stage evolution is represented by the phases D-A and D-B that indicate shortening parallel to the axis of the mountain chain and by D-C that is interpreted to account for the formation of large-scale domes with NNW-SSE-trending axes, an example of which is exposed in the Larji-Kullu-Rampur tectonic window.

Neuroprotection in cerebral ischemia with XG-102, a new peptide inhibitor of JNK

Abstract Stroke or cerebrovascular accident, whose great majority is of ischemic nature, is the third leading cause of mortality and long lasting disability in industrialised countries. Resulting from the loss of blood supply to the brain depriving cerebral tissues of oxygen and glucose, it induces irreversible neuronal damages. Despite the large amount of research carried out into the causes and pathogenic features of cerebral ischemia the progress toward effective treatments has been poor. Apart the clot-busting drug tissue-type plasminogen activator (tPA) as effective therapy for acute stroke (reperfusion by thrombolysis) but limited to a low percentage of patients, there are currently no other approved medical treatments. The need for new therapy strategies is therefore imperative. Neuronal death in cerebral ischemia is among others due to excitotoxic mechanisms very early after stroke onset. One of the main involved molecular pathways leading to excitotoxic cell death is the c-Jun NH2-terminal kinase (JNK) pathway. Several studies have already shown the efficacy of a neuroprotective agent of a new type, a dextrogyre peptide synthesized in the retro inverso form (XG102, formerly D-JNKI1), which is protease-resistant and cell-penetrating and that selectively and strongly blocks the access of JNK to many of its targets. A powerful protection was observed with this compound in several models of ischemia (Borsello et al. 2003;Hirt et al. 2004). This chimeric compound, made up of a 10 amino acid TAT transporter sequence followed by a 20 amino acids JNK binding domain (JBD) sequence from JNK inhibitor protein (JIP) molecule, induced both a major reduction in lesion size and improved functional outcome. Moreover it presents a wide therapeutic window. XG-102 has proved its powerful efficacy in an occlusion model of middle cerebral artery in mice with intracérebroventricular (i.c.v.) injection but in order to be able to consider the development of this drug for human ischemic stroke it was therefore necessary to determine the feasibility of its systemic administration. The studies being the subject of this thesis made it possible to show a successful neuroprotection with XG-102 administered systemically after transient mouse middle cerebral artery occlusion (MCAo). Moreover our data. provided information about the feasibility to combine XG-102 with tPA without detrimental action on cell survival. By combining the benefits from a reperfusion treatment with the effects of a neuroprotective compound, it would represent the advantage of bringing better chances to protect the cerebral tissue. Résumé L'attaque cérébrale ou accident vasculaire cérébral, dont la grande majorité est de nature ischémique, constitue la troisième cause de mortalité et d'infirmité dans les pays industrialisés. Résultant de la perte d'approvisionnement de sang au cerveau privant les tissus cérébraux d'oxygène et de glucose, elle induit des dommages neuronaux irréversibles. En dépit du nombre élevé de recherches effectuées pour caractériser les mécanismes pathogènes de l'ischémie. cérébrale, les progrès vers des traitements efficaces restent pauvres. Excepté l'activateur tissulaire du plasminogène (tPA) dont le rôle est de désagréger les caillots sanguins et employé comme thérapie efficace contre l'attaque cérébrale aiguë (reperfusion par thrombolyse) mais limité à un faible pourcentage de patients, il n'y a actuellement aucun autre traitement médical approuvé. Le besoin de nouvelles stratégies thérapeutiques est par conséquent impératif. La mort neuronale dans l'ischémie cérébrale est entre autres due à des mécanismes excitotoxiques survenant rapidement après le début de l'attaque cérébrale. Une des principales voies moléculaires impliquée conduisant à la mort excitotoxique des cellules est la voie de la c-Jun NH2terminal kinase (JNK). Plusieurs études ont déjà montré l'efficacité d'un agent neuroprotecteur d'un nouveau type, un peptide dextrogyre synthétisé sous la forme retro inverso (XG-102, précédemment D-JNKI1) résistant aux protéases, capable de pénétrer dans les cellules et de bloquer sélectivement et fortement l'accès de JNK à plusieurs de ses cibles. Une puissante protection a été observée avec ce composé dans plusieurs modèles d'ischémie (Borsello et al. 2003;Hirt et al. 2004). Ce composé chimérique, construit à partir d'une séquence TAT de 10 acides aminés suivie par une séquence de 20 acides aminés d'un domaine liant JNK (JBD) issu de la molécule JNK protéine inhibitrice. (JIP), induit à la fois une réduction importante de la taille de lésion et un comportement fonctionnel amélioré. De plus il présente une fenêtre thérapeutique étendue. XG-102 a prouvé sa puissante efficacité dans un modèle d'occlusion de l'artère cérébrale moyenne chez la souris avec injection intracerebroventriculaire (i.c.v.) mais afin de pouvoir envisager le développement de ce composé pour l'attaque cérébrale chez l'homme, il était donc nécessaire de déterminer la faisabilité de son administration systémique. Les études faisant l'objet de cette thèse ont permis de montrer une neuroprotection importante avec XG-102 administré de façon systémique après l'occlusion transitoire de l'artère cérébrale moyenne chez la souris (MCAo). De plus nos données ont fourni des informations quant à la faisabilité de combiner XG-102 et tPA, démontrant une protection efficace par XG-102 malgré l'action nuisible du tPA sur la survie des cellules. En combinant les bénéfices de la reperfusion avec les effets d'un composé neurooprotecteur, cela représenterait l'avantage d'apporter des meilleures chances de protéger le tissu cérébral.

Main Street Messenger, Summer 2001

Newsletter for Economic Development

Main Street Messenger, Spring, 2002

Newsletter for Economic Development