Synthesis of new enzyme stabilisers inspired by compatible solutes of hyperthermophilic microorganisms

Dissertation presented to obtain the Ph.D degree in Engineering Sciences and Technology.

Land cover mapping analysis and urban growth modeling using remote sensing techniques

The rapid growth of big cities has been noticed since 1950s when the majority of world population turned to live in urban areas rather than villages, seeking better job opportunities and higher quality of services and lifestyle circumstances. This demographic transition from rural to urban is expected to have a continuous increase. Governments, especially in less developed countries, are going to face more challenges in different sectors, raising the essence of understanding the spatial pattern of the growth for an effective urban planning. The study aimed to detect, analyse and model the urban growth in Greater Cairo Region (GCR) as one of the fast growing mega cities in the world using remote sensing data. Knowing the current and estimated urbanization situation in GCR will help decision makers in Egypt to adjust their plans and develop new ones. These plans should focus on resources reallocation to overcome the problems arising in the future and to achieve a sustainable development of urban areas, especially after the high percentage of illegal settlements which took place in the last decades. The study focused on a period of 30 years; from 1984 to 2014, and the major transitions to urban were modelled to predict the future scenarios in 2025. Three satellite images of different time stamps (1984, 2003 and 2014) were classified using Support Vector Machines (SVM) classifier, then the land cover changes were detected by applying a high level mapping technique. Later the results were analyzed for higher accurate estimations of the urban growth in the future in 2025 using Land Change Modeler (LCM) embedded in IDRISI software. Moreover, the spatial and temporal urban growth patterns were analyzed using statistical metrics developed in FRAGSTATS software. The study resulted in an overall classification accuracy of 96%, 97.3% and 96.3% for 1984, 2003 and 2014’s map, respectively. Between 1984 and 2003, 19 179 hectares of vegetation and 21 417 hectares of desert changed to urban, while from 2003 to 2014, the transitions to urban from both land cover classes were found to be 16 486 and 31 045 hectares, respectively. The model results indicated that 14% of the vegetation and 4% of the desert in 2014 will turn into urban in 2025, representing 16 512 and 24 687 hectares, respectively.

Generalized Dasymetric Mapping Algorithm for Accessing Land-Use Change

Contém resumo

Mapping the Pasture Steppe in Bayankhongor, Mongolia: comparison of classification methods, using Landsat-8 and geophysical data

Grasslands in semi-arid regions, like Mongolian steppes, are facing desertification and degradation processes, due to climate change. Mongolia’s main economic activity consists on an extensive livestock production and, therefore, it is a concerning matter for the decision makers. Remote sensing and Geographic Information Systems provide the tools for advanced ecosystem management and have been widely used for monitoring and management of pasture resources. This study investigates which is the higher thematic detail that is possible to achieve through remote sensing, to map the steppe vegetation, using medium resolution earth observation imagery in three districts (soums) of Mongolia: Dzag, Buutsagaan and Khureemaral. After considering different thematic levels of detail for classifying the steppe vegetation, the existent pasture types within the steppe were chosen to be mapped. In order to investigate which combination of data sets yields the best results and which classification algorithm is more suitable for incorporating these data sets, a comparison between different classification methods were tested for the study area. Sixteen classifications were performed using different combinations of estimators, Landsat-8 (spectral bands and Landsat-8 NDVI-derived) and geophysical data (elevation, mean annual precipitation and mean annual temperature) using two classification algorithms, maximum likelihood and decision tree. Results showed that the best performing model was the one that incorporated Landsat-8 bands with mean annual precipitation and mean annual temperature (Model 13), using the decision tree. For maximum likelihood, the model that incorporated Landsat-8 bands with mean annual precipitation (Model 5) and the one that incorporated Landsat-8 bands with mean annual precipitation and mean annual temperature (Model 13), achieved the higher accuracies for this algorithm. The decision tree models consistently outperformed the maximum likelihood ones.

High-resolution spatial mapping of changes in the neurochemical profile after focal ischemia in mice.

After ischemic stroke, the ischemic damage to brain tissue evolves over time and with an uneven spatial distribution. Early irreversible changes occur in the ischemic core, whereas, in the penumbra, which receives more collateral blood flow, the damage is more mild and delayed. A better characterization of the penumbra, irreversibly damaged and healthy tissues is needed to understand the mechanisms involved in tissue death. MRSI is a powerful tool for this task if the scan time can be decreased whilst maintaining high sensitivity. Therefore, we made improvements to a (1) H MRSI protocol to study middle cerebral artery occlusion in mice. The spatial distribution of changes in the neurochemical profile was investigated, with an effective spatial resolution of 1.4 μL, applying the protocol on a 14.1-T magnet. The acquired maps included the difficult-to-separate glutamate and glutamine resonances and, to our knowledge, the first mapping of metabolites γ-aminobutyric acid and glutathione in vivo, within a metabolite measurement time of 45 min. The maps were in excellent agreement with findings from single-voxel spectroscopy and offer spatial information at a scan time acceptable for most animal models. The metabolites measured differed with respect to the temporal evolution of their concentrations and the localization of these changes. Specifically, lactate and N-acetylaspartate concentration changes largely overlapped with the T(2) -hyperintense region visualized with MRI, whereas changes in cholines and glutathione affected the entire middle cerebral artery territory. Glutamine maps showed elevated levels in the ischemic striatum until 8 h after reperfusion, and until 24 h in cortical tissue, indicating differences in excitotoxic effects and secondary energy failure in these tissue types. Copyright © 2011 John Wiley & Sons, Ltd.

Plasmodium falciparum merozoite surface protein 2: epitope mapping and biological activity analysis of specific antibodies

Background: Plasmodium falciparum(P. falciparum) merozoite surfaceprotein 2 (MSP-2) is one of bloodstage proteins that are associated withprotection from malaria. MSP-2 consistsof a highly polymorphic centralrepeat region flanked by a dimorphicregion that defines the two allelicfamilies, 3D7 and FC27; N- and Cterminalregions are conserved domains.Long synthetic peptides (LSP)representing the two allelic familiesof MSP-2 and constant regions arerecognized by sera from donors livingin endemic areas; and specific antibodies(Abs) are associated with protectionand active in antibody dependentcellular inhibition (ADCI) in vitro.However, the fine specificity ofAb response to the two allelic familiesof MSP-2 is unknown. Methods: Peptidesrepresenting dimorphic regionof 3D7 and FC27 families and theirC-terminal (common fragment to thetwo families) termed 3D7-D (88 aa),FC27-D (48 aa) and C (40 aa) respectivelywere synthesized. Overlapping20 mer peptides covering dimorphicand constant regions of two familieswere also synthesized for epitopemapping. Human sera were obtainedfrom donors living in malaria endemicareas. SpecificDand CregionsAbs were purified from single or poolhuman sera. Sera from mice were obtainedafter immunization with thetwo families LSP mixture in three differentadjuvants: alhydrogel (Alum),Glucopyranosyl Lipid Adjuvant-Stableoil-in-water Emulsion (GLA-SE)and Virosome. For ADCI, P. falciparum(strain 3D7) parasite wasmaintained in culture at 0.5% parasitemiaand 4% hematocrit in air tightbox at love oxygen (2%) and 37 ºC.Results: We identified several epitopesfrom the dimorphic and constantregions of both families of MSP-2, inmice and humans (adults and children).In human, most recognizedepitopes were the same in differentendemic regions for each domain ofthe two families of MSP-2. In mice,the differential recognition of epitopewas depending on the strain of mouseand interestingly on the adjuvantused. GLA-SE and alum as adjuvantswere more often associated with therecognition of multiple epitopes thanvirosomes. Epitope-specific Abs recognizednative merozoites of P.falciparum and were active in ADCIto block development of parasite.Conclusion: The delineation of a limitednumber of epitopes could be exploitedto develop MSP-2 vaccinesactive on both allelic families ofMSP-2.

Gene expression mapping in neuropathic pain : molecular plasticity in nociceptors and superficial dorsal horn

RESUME : La douleur neuropathique est le résultat d'une lésion ou d'un dysfonctionnement du système nerveux. Les symptômes qui suivent la douleur neuropathique sont sévères et leur traitement inefficace. Une meilleure approche thérapeutique peut être proposée en se basant sur les mécanismes pathologiques de la douleur neuropathique. Lors d'une lésion périphérique une douleur neuropathique peut se développer et affecter le territoire des nerfs lésés mais aussi les territoires adjacents des nerfs non-lésés. Une hyperexcitabilité des neurones apparaît au niveau des ganglions spinaux (DRG) et de la corne dorsale (DH) de la moelle épinière. Le but de ce travail consiste à mettre en évidence les modifications moléculaires associées aux nocicepteurs lésés et non-lésés au niveau des DRG et des laminae I et II de la corne dorsale, là où l'information nociceptive est intégrée. Pour étudier les changements moléculaires liés à la douleur neuropathique nous utilisons le modèle animal d'épargne du nerf sural (spared nerve injury model, SNI) une semaine après la lésion. Pour la sélection du tissu d'intérêt nous avons employé la technique de la microdissection au laser, afin de sélectionner une sous-population spécifique de cellules (notamment les nocicepteurs lésés ou non-lésés) mais également de prélever le tissu correspondant dans les laminae superficielles. Ce travail est couplé à l'analyse à large spectre du transcriptome par puce ADN (microarray). Par ailleurs, nous avons étudié les courants électriques et les propriétés biophysiques des canaux sodiques (Na,,ls) dans les neurones lésés et non-lésés des DRG. Aussi bien dans le système nerveux périphérique, entre les neurones lésés et non-lésés, qu'au niveau central avec les aires recevant les projections des nocicepteurs lésés ou non-lésés, l'analyse du transcriptome montre des différences de profil d'expression. En effet, nous avons constaté des changements transcriptionnels importants dans les nocicepteurs lésés (1561 gènes, > 1.5x et pairwise comparaison > 77%) ainsi que dans les laminae correspondantes (618 gènes), alors que ces modifications transcriptionelles sont mineures au niveau des nocicepteurs non-lésés (60 gènes), mais important dans leurs laminae de projection (459 gènes). Au niveau des nocicepteurs, en utilisant la classification par groupes fonctionnels (Gene Ontology), nous avons observé que plusieurs processus biologiques sont modifiés. Ainsi des fonctions telles que la traduction des signaux cellulaires, l'organisation du cytosquelette ainsi que les mécanismes de réponse au stress sont affectés. Par contre dans les neurones non-lésés seuls les processus biologiques liés au métabolisme et au développement sont modifiés. Au niveau de la corne dorsale de la moelle, nous avons observé des modifications importantes des processus immuno-inflammatoires dans l'aire affectée par les nerfs lésés et des changements associés à l'organisation et la transmission synaptique au niveau de l'aire des nerfs non-lésés. L'analyse approfondie des canaux sodiques a démontré plusieurs changements d'expression, principalement dans les neurones lésés. Les analyses fonctionnelles n'indiquent aucune différence entre les densités de courant tétrodotoxine-sensible (TTX-S) dans les neurones lésés et non-lésés même si les niveaux d'expression des ARNm des sous-unités TTX-S sont modifiés dans les neurones lésés. L'inactivation basale dépendante du voltage des canaux tétrodotoxine-insensible (TTX-R) est déplacée vers des potentiels positifs dans les cellules lésées et non-lésées. En revanche la vitesse de récupération des courants TTX-S et TTX-R après inactivation est accélérée dans les neurones lésés. Ces changements pourraient être à l'origine de l'altération de l'activité électrique des neurones sensoriels dans le contexte des douleurs neuropathiques. En résumé, ces résultats suggèrent l'existence de mécanismes différenciés affectant les neurones lésés et les neurones adjacents non-lésés lors de la mise en place la douleur neuropathique. De plus, les changements centraux au niveau de la moelle épinière qui surviennent après lésion sont probablement intégrés différemment selon la perception de signaux des neurones périphériques lésés ou non-lésés. En conclusion, ces modulations complexes et distinctes sont probablement des acteurs essentiels impliqués dans la genèse et la persistance des douleurs neuropathiques. ABSTRACT : Neuropathic pain (NP) results from damage or dysfunction of the peripheral or central nervous system. Symptoms associated with NP are severe and difficult to treat. Targeting NP mechanisms and their translation into symptoms may offer a better therapeutic approach.Hyperexcitability of the peripheral and central nervous system occurs in the dorsal root ganglia (DRG) and the dorsal horn (DH) of the spinal cord. We aimed to identify transcriptional variations in injured and in adjacent non-injured nociceptors as well as in corresponding laminae I and II of DH receiving their inputs.We investigated changes one week after the injury induced by the spared nerve injury model of NP. We employed the laser capture microdissection (LCM) for the procurement of specific cell-types (enrichment in nociceptors of injured/non-injured neurons) and laminae in combination with transcriptional analysis by microarray. In addition, we studied functionál properties and currents of sodium channels (Nav1s) in injured and neighboring non-injured DRG neurons.Microarray analysis at the periphery between injured and non-injured DRG neurons and centrally between the area of central projections from injured and non-injured neurons show significant and differential expression patterns. We reported changes in injured nociceptors (1561 genes, > 1.5 fold, >77% pairwise comparison) and in corresponding DH laminae (618 genes), while less modifications occurred in non-injured nociceptors (60 genes) and in corresponding DH laminae (459 genes). At the periphery, we observed by Gene Ontology the involvement of multiple biological processes in injured neurons such as signal transduction, cytoskeleton organization or stress responses. On contrast, functional overrepresentations in non-injured neurons were noted only in metabolic or developmentally related mechanisms. At the level of superficial laminae of the dorsal horn, we reported changes of immune and inflammatory processes in injured-related DH and changes associated with synaptic organization and transmission in DH corresponding to non-injured neurons. Further transcriptional analysis of Nav1s indicated several changes in injured neurons. Functional analyses of Nav1s have established no difference in tetrodotoxin-sensitive (TTX-S) current densities in both injured and non-injured neurons, despite changes in TTX-S Nav1s subunit mRNA levels. The tetrodotoxin-resistant (TTX-R) voltage dependence of steady state inactivation was shifted to more positive potentials in both injured and non-injured neurons, and the rate of recovery from inactivation of TTX-S and TTX-R currents was accelerated in injured neurons. These changes may lead to alterations in neuronal electrogenesis. Taken together, these findings suggest different mechanisms occurring in the injured neurons and the adjacent non-injured ones. Moreover, central changes after injury are probably driven in a different manner if they receive inputs from injured or non-injured neurons. Together, these distinct and complex modulations may contribute to NP.

Discovery and fine mapping of serum protein loci through transethnic meta-analysis.

Many disorders are associated with altered serum protein concentrations, including malnutrition, cancer, and cardiovascular, kidney, and inflammatory diseases. Although these protein concentrations are highly heritable, relatively little is known about their underlying genetic determinants. Through transethnic meta-analysis of European-ancestry and Japanese genome-wide association studies, we identified six loci at genome-wide significance (p < 5 × 10(-8)) for serum albumin (HPN-SCN1B, GCKR-FNDC4, SERPINF2-WDR81, TNFRSF11A-ZCCHC2, FRMD5-WDR76, and RPS11-FCGRT, in up to 53,190 European-ancestry and 9,380 Japanese individuals) and three loci for total protein (TNFRS13B, 6q21.3, and ELL2, in up to 25,539 European-ancestry and 10,168 Japanese individuals). We observed little evidence of heterogeneity in allelic effects at these loci between groups of European and Japanese ancestry but obtained substantial improvements in the resolution of fine mapping of potential causal variants by leveraging transethnic differences in the distribution of linkage disequilibrium. We demonstrated a functional role for the most strongly associated serum albumin locus, HPN, for which Hpn knockout mice manifest low plasma albumin concentrations. Other loci associated with serum albumin harbor genes related to ribosome function, protein translation, and proteasomal degradation, whereas those associated with serum total protein include genes related to immune function. Our results highlight the advantages of transethnic meta-analysis for the discovery and fine mapping of complex trait loci and have provided initial insights into the underlying genetic architecture of serum protein concentrations and their association with human disease.

Mapping Bias Overestimates Reference Allele Frequencies at the HLA Genes in the 1000 Genomes Project Phase I Data.

Next-generation sequencing (NGS) technologies have become the standard for data generation in studies of population genomics, as the 1000 Genomes Project (1000G). However, these techniques are known to be problematic when applied to highly polymorphic genomic regions, such as the human leukocyte antigen (HLA) genes. Because accurate genotype calls and allele frequency estimations are crucial to population genomics analyses, it is important to assess the reliability of NGS data. Here, we evaluate the reliability of genotype calls and allele frequency estimates of the single-nucleotide polymorphisms (SNPs) reported by 1000G (phase I) at five HLA genes (HLA-A, -B, -C, -DRB1, and -DQB1). We take advantage of the availability of HLA Sanger sequencing of 930 of the 1092 1000G samples and use this as a gold standard to benchmark the 1000G data. We document that 18.6% of SNP genotype calls in HLA genes are incorrect and that allele frequencies are estimated with an error greater than ±0.1 at approximately 25% of the SNPs in HLA genes. We found a bias toward overestimation of reference allele frequency for the 1000G data, indicating mapping bias is an important cause of error in frequency estimation in this dataset. We provide a list of sites that have poor allele frequency estimates and discuss the outcomes of including those sites in different kinds of analyses. Because the HLA region is the most polymorphic in the human genome, our results provide insights into the challenges of using of NGS data at other genomic regions of high diversity.

The genetics of glucosamine in yeast : cytoplasmic determinants conferring resistance

Two cytoplasmic, glucosamine resistant mutants of Saccharomyces cerevisiae, GR6 and GR10, were examined to determine whether or not the lesions involved were located on mitochondrial DNA. Detailed investigation of crosses of GR6 and GR10 or their derivatives to strains bearing known mitochondrial markers demonstrated that: 1. the frequency of glucos~~ine resistance in diploids was independent of factors influencing mitochondrial marker output. 2. upon tetrad analysis a variety of tetrad ratios was observed for glucosamine resistance whereas mitochondrial markers segregated 4:0 or 0:4 (resistant:sensitive). 3. glucosamine resistance and mitochondrial markers segregated differentially with time. 4. glucosamine resistance persisted following treatment of a GRIO derivative with ethidium bromide at concentrations high enough to eliminate all mitochondrial DNA. 5. haploid spore clones displayed two degrees of glucosamine resistance, weak and strong, while growth due to mitochondrial mutations was generally thick and confluent. 6. a number of glucosamine resistant diploids and haploids, which also possessed a mithchondrial resistance mutation, were unable to grow on medium containing both glucosamine and the particular drug involved. 3 These observations 1~ 6 provided strong evidence that the cytoplasmic glucosamine resistant mutations present in GR6 and GRiO were not situated on mitochondrial DNA. Comparison of the glucosamine resistance mutations to some other known cytoplasmic determinants revealed that: 7. glucosamine resistance and the expression of the killer phenotype were separate phenomena. 8. unlike yeast carrying resistance conferring episomes GR6 and GR10 were not resistant to venturicidin or oligomycin and the GR factor exhibited genetic behaviour different from that of the episomal determinants. These results 7--+8 suggested that glucosamine resistance was not associated with the killer determinant nor with alleged yeast episomes. It is therefore proposed that a yeast plasmid(s), previously undescribed, is responsible for glucosamine resistance. The evidence to date is compatible with the hypothesis that GR6 and GR10 carry allelic mutations of the same plasmid which is tentatively designated (GGM).

Mapping one university's response to internationalization : a case study

This study explored one university's response to the internationalization of higher education. Case study methodology was employed through a review of current and archival documents and interviews with key actors in the international spheres of the university. The historical, current, and future contexts were considered to situate the case study on a time line. Data analysis revealed that there were several points of division among the university community related to the response to internationalization, but also a major point of coherence in the centrality of inter-cultural understanding in efforts to internationalize. Other key findings included strengths, areas for improvement, and future directions of the university's response to internationalization. All of these findings were contextualized in findings related to the history of the university. In addition to these major findings, three themes in relation to the vision for internationalization at the institution were revealed: ( a) intercultural understanding, (b) the comprehensive status of the university, and (c) the financial benefits of internationalization. Recommendations are made for practice at the university in order to clarify this vision to develop a clear foundation from which to further build a response to internationalization that is solidly based on inter-cultural understanding, and recommendations for future research into the process of internationalization at the institutional level in Canada are suggested.

Letter (unsigned) to R.A. Lucas suggesting that the mapping of the marsh be put off until after the next meeting

Letter (unsigned) to R.A. Lucas suggesting that the mapping of the marsh be put off until after the next meeting. The rest of the letter deals with licenses, Nov. 29, 1882.