Les aptitudes du lait à la transformation en fromage sont étroitement liées à sa composition, notamment en protéines. Ces caractères, difficiles à mesurer directement, ont été prédits à partir des spectres moyen infrarouge (MIR) du lait pour la composition en protéines dans les 3 races bovines Montbéliarde, Normande et Holstein (projet PhénoFinlait) et pour 9 aptitudes fromagères et la composition fine du lait en race Montbéliarde (projet From’MIR). La méthode Partial Least Squares (PLS) a fourni des prédictions MIR plus précises que les méthodes bayésiennes testées.Une analyse génétique a été réalisée pour ces caractères prédits à partir de plus de six millions de spectres MIR de plus de 400 000 vaches.Les caractères fromagers et de composition du lait sont modérément à fortement héritables. Les corrélations génétiques entre caractères fromagers (rendements et coagulation) et avec la composition du lait (protéines, acides gras et minéraux) sont élevées et favorables.Les génotypes de 28 000 vaches ont été imputés jusqu’à la séquence complète grâce aux données du projet 1000 génomes bovins.Des analyses d’association (GWAS) révèlent de nombreux gènes et variants avec des effets forts sur la fromageabilité et la composition du lait. Un réseau de 736 gènes, par ailleurs associé à ces caractères, permet d’identifier des voies métaboliques et des gènes régulateurs fonctionnellement liés à ces caractères.Un prototype d’évaluation génomique a été mis en place en race Montbéliarde. Un modèle de type contrôles élémentaires, incluant les variants détectés par les GWAS et présumés causaux, donne les estimations des valeurs génomiques les plus précises. La simulation d’une sélection incluant les caractères fromagers montre qu’il est possible d’améliorer la fromageabilité du lait avec un impact limité sur le gain génétique des autres caractères sélectionnés.Les travaux présentés dans cette thèse ont abouti 1) à la détection de gènes (dont certains jamais décrits auparavant) et de variants candidats pour la composition et la fromageabilité du lait et 2) à la mise en place d’une évaluation génomique de la fromageabilité du lait en race Montbéliarde dans la zone AOP Comté. / The ability of milk to be processed into cheese is closely linked to its composition, in particular in proteins. These traits, which are difficult to measure directly, were predicted from milk mid-infrared (MIR) spectra for protein composition in 3 cattle breeds Montbéliarde, Normande and Holstein (PhénoFinlait project) and for 9 milk cheese-making properties (CMP) and composition traits in Montbéliarde cows (From’MIR project). The Partial Least Squares method provided more accurate predictions than the Bayesian methods tested.A genetic analysis was performed on these traits, predicted from more than six million MIR spectra of more than 400,000 cows.Milk CMP and composition traits are moderately to highly heritable. Genetic correlations between CMP (cheese yields and coagulation) and with milk composition (proteins, fatty acids and minerals) are high and favorable.The genotypes of 28,000 cows were imputed to whole genome sequences using the 1000 bovine genome reference population.Genome wide association studies (GWAS) reveal many genes and variants in these genes with strong effects on CMP and milk composition. A network of 736 genes, associated with these traits, enable the identification of metabolic pathways and regulatory genes functionally linked to these traits.A pilot genomic evaluation was set up in Montbéliarde cows. A test-day model, including variants detected by GWAS, provides the most accurate genomic value estimates. Simulation of a selection shows that it is possible to improve the cheesability of milk with a limited impact on the genetic gain of the traits that currently make up the breeding objective.The work presented in this thesis led to 1) the detection of genes (some of which have never been described before) and candidate variants for milk CMP and composition traits and 2) the implementation of a genomic evaluation of CMP predicted from MIR spectra in Montbéliarde cows of the Comté PDO area.

Les infections en biofilm, notamment de dispositifs médicaux, mettent fréquemment en échec les traitements antibiotiques, imposant le retrait du matériel. Pseudomonas aeruginosa s’est imposé comme le pathogène-type des infections en biofilm. Pour explorer les déterminants de l’échec du traitement antibiotique en biofilm, un modèle de biofilm in vitro à P. aeruginosa exposé à des doses supra-inhibitrices d’antibiotiques a été développé. En culture planctonique, une bithérapie deciprofloxacine et d’amikacine permettait de prévenir la sélection de mutants résistants pour des souches de P. aeruginosa de sensibilité diminuée à la ciprofloxacine ou à l’amikacine par surexpression d’efflux. En biofilm, l’association de la ciprofloxacine et de l’amikacine, administrées simultanément ou séquentiellement, n’était pas supérieure aux monothérapies, permettant une réduction bactérienne, mais pas d’éradication complète du biofilm. Quelles que soient les souches (sauvages ou exprimant un efflux) et l’antibiotique, l’échec microbiologique en biofilm était lié à la sélection de cellules persistantes, tolérantes aux antibiotiques. La ciprofloxacine induisait des modifications importantes de la structure du biofilm avec une réduction considérable des exopolysaccharides, composants majeurs de la matrice. L’étude transcriptomique de gènes potentiellement impliqués dans la persistance suggérait que l’activation précoce de la réponse stringente pourrait être une des voies principales de la tolérance en biofilm sous ciprofloxacine. Enfin, la présence de « small colony variants » au sein du biofilm, dotés d’une capacité accrue de formation de biofilm, témoignait de la diversité des populations en biofilm. Ces travaux participent ainsi à une meilleure compréhension des mécanismes d’échappement aux antibiotiques de P. aeruginosa en biofilm. / Biofilm device-related infections can lead to antibiotic failure requiring frequent removal of medical device. Pseudomonas aeruginosa has emerged as the typical pathogen for biofilm infections. To explore the determinants of antibiotic failure in biofilm, an in vitro P. aeruginosa biofilm model exposed to suprainhibitory antibiotic concentrations was developed. In planktonic culture, the ciprofloxacin and amikacin combination prevented the selection of resistant mutants in ciprofloxacin and amikacinlow-level resistant P. aeruginosa strains overexpressing efflux. In biofilm, the ciprofloxacin and amikacin combination, used simultaneously or sequentially, didn’t show superior effects compared to monotherapies. Despite an initial bacterial reduction, biofilm eradication was not obtained. Regardless of wild-type or efflux strains and antibiotic regimen used, antibiotic failure was related to the selection of antibiotic-tolerant cells named “persisters”. Ciprofloxacin induced significant alterations in the biofilm structure, notably a considerable reduction in the exopolysaccharides of the matrix. The transcriptomic analysis of genes, potentially involved in persistence, suggested that early activation of the stringent response might be one of the main pathways for ciprofloxacin tolerance in biofilm. Finally, the emergence of "small colony variants" within the biofilm, characterized by enhanced ability to form biofilm, attested to biofilm heterogeneity. This work therefore contributes to a better understanding of how P. aeruginosa biofilms escape antibiotic.

Any software development process deals with four main artifacts namely; requirement, design, implementation and test. Depending upon the functionality of a particular product there might be variants present in these artifacts. These variants influence all the artifacts involved in a software development process. Data in the higher level artifact affects the data present in the further artifacts and is also refined when we move towards the lower level of abstraction. This thesis deals with the handling of all the variant information present in all the artifacts. Verification and consistency checks on this information were to be automated for making the development process easier. The results achieved during this thesis discuss the solutions for the problem of inconsistent variant information present in all the artifacts. By defining the extension of the intermediate format to support the variant information at Vector Informatik GmbH this problem has been resolved. The data used during the development is the variant information. The generic intermediate format has been extended in a way so that it can further support a variety of use cases. Along with the formulation of a format, documentation of variant information and methods to extract variant information form C source code are also discussed.

Introduction: The continual increase of type 2 diabetes (T2D) prevalence is a global public health concern. The aetiology of T2D has not been fully elucidated and this is hampering the development of effective preventative and curative interventions to curb the T2D burden. Although much has been done to elucidate the environmental risk factors associated with T2D, little is known about the precise genetic risk factors that predispose people to it. There is limited knowledge about the common variants associated with T2D risk in the black South African population. However, evidence of shared common variants associated with T2D among people of different ethnicities has been documented. Nonetheless, the majority of the common variants that have been reported to be associated with T2D in other ethnicities are still yet to be evaluated in the black South African population. Objectives: The aim of this study was to evaluate the association of previously reported common genetic variants with T2D susceptibility, as indicated by impaired glucose tolerance (IGT), in a black South African population of Tswana descent. Methods: This study was a case-control study of 180 cases and 180 controls nested in the Prospective Urban Rural Epidemiology (PURE) study baseline data, which was collected in 2005. The DNA samples of the participants were genotyped for 77 single nucleotide polymorphisms (SNPs), using Illumina® VeraCode technology on the BeadXpress® platform. The gPlink software was used to evaluate the standard genetic models of disease penetrance for the association of the common variants with impaired glucose tolerance (IGT) while adjusting for age, sex and body mass index. Results: Four out of the 66 SNPs that were evaluated through the genetic association tests in this study were noted to be significantly associated with IGT (p< 0.05). Of the four SNPs, only rs1436955 was associated with an increase in T2D risk, while the other three variants, rs831571, rs8050136 and rs7542900, were noted to be associated with a decreased risk of T2D. However, none of the four SNPs was significantly associated with IGT after correcting for multiple testing (p <0.05). Conclusions: Black South Africans of Tswana descent might not share common variants associated with T2D risk, as indicated by IGT in other ethnicities. Wellpowered studies are required to evaluate the association of common variants with T2D risk in this population group. The results from this study emphasise the need for population-specific variants to assess the genetic susceptibility of complex diseases such as T2D in the black South African population. / MSc (Nutrition), North-West University, Potchefstroom Campus, 2014

Introduction: The continual increase of type 2 diabetes (T2D) prevalence is a global public health concern. The aetiology of T2D has not been fully elucidated and this is hampering the development of effective preventative and curative interventions to curb the T2D burden. Although much has been done to elucidate the environmental risk factors associated with T2D, little is known about the precise genetic risk factors that predispose people to it. There is limited knowledge about the common variants associated with T2D risk in the black South African population. However, evidence of shared common variants associated with T2D among people of different ethnicities has been documented. Nonetheless, the majority of the common variants that have been reported to be associated with T2D in other ethnicities are still yet to be evaluated in the black South African population. Objectives: The aim of this study was to evaluate the association of previously reported common genetic variants with T2D susceptibility, as indicated by impaired glucose tolerance (IGT), in a black South African population of Tswana descent. Methods: This study was a case-control study of 180 cases and 180 controls nested in the Prospective Urban Rural Epidemiology (PURE) study baseline data, which was collected in 2005. The DNA samples of the participants were genotyped for 77 single nucleotide polymorphisms (SNPs), using Illumina® VeraCode technology on the BeadXpress® platform. The gPlink software was used to evaluate the standard genetic models of disease penetrance for the association of the common variants with impaired glucose tolerance (IGT) while adjusting for age, sex and body mass index. Results: Four out of the 66 SNPs that were evaluated through the genetic association tests in this study were noted to be significantly associated with IGT (p< 0.05). Of the four SNPs, only rs1436955 was associated with an increase in T2D risk, while the other three variants, rs831571, rs8050136 and rs7542900, were noted to be associated with a decreased risk of T2D. However, none of the four SNPs was significantly associated with IGT after correcting for multiple testing (p <0.05). Conclusions: Black South Africans of Tswana descent might not share common variants associated with T2D risk, as indicated by IGT in other ethnicities. Wellpowered studies are required to evaluate the association of common variants with T2D risk in this population group. The results from this study emphasise the need for population-specific variants to assess the genetic susceptibility of complex diseases such as T2D in the black South African population. / MSc (Nutrition), North-West University, Potchefstroom Campus, 2014

Dans le but de mieux comprendre le mode d’action et de nous permettre de déterminer la cible de la tomatidine, nous avons dans un premier temps tenté de mieux circonscrire le spectre d’activité de la tomatidine. Grâce à ces travaux, nous sommes, en effet, maintenant en mesure de dire que la tomatidine possède une activité antibactérienne contre les espèces de la division des Firmicutes et plus précisément contre les bactéries de l’ordre des Bacillales dont font partie les genres Bacillus, Staphylococcus et Listeria. Nous avons également découvert, grâce à des expériences en collaboration avec le laboratoire d’Éric Marsault, qu’un analogue de la tomatidine (FC04-100) avait non seulement des propriétés similaires à la molécule naturelle, mais démontrait une activité par lui-même contre S. aureus à phénotype normal alors que la tomatidine possède uniquement une activité contre les « small colony variants ». De plus, alors que la tomatidine possède plutôt une activité bactériostatique contre la forme SCV de L. monocytogenes, le nouveau composé (FC04-100) démontre quant à lui, une forte activité bactéricide contre cette souche, tout comme contre la forme SCV des autres Bacillales. Parallèlement, et toujours dans le but de rechercher le mode d’action et la cible de la tomatidine, nous avons obtenu, par passages successifs dans un milieu avec antibiotiques, des mutants de S. aureus à phénotype normal et des SCV résistants à la tomatidine ou à la combinaison tomatidine et gentamicine. Après le séquençage de ces mutants, l’étude de la position de ces mutations, à l’aide de différents logiciels de bio-informatique, nous a permis d’émettre un modèle-hypothèse quant au mode d’action et à la cible de la tomatidine. Selon les résultats que nous avons à ce stade-ci, la cible de la tomatidine chez S. aureus serait la sous-unité c de l’ATP synthase. Cependant, son mode d’action serait également dépendant de la fonctionnalité de la chaine de transport des électrons et donc de la polarisation membranaire et de la production de ROS intracellulaire, ce qui expliquerait la différence d’activité entre les souches à phénotype normal et les SCV.

Background: The human Aldoketoreductase 1B10 gene (AKR1B10) encodes one of the enzymes belonging to the family of aldoketoreductases and may be involved in detoxification of nutrients during digestion. Further, AKR1B10 mRNA (messenger ribonucleic acid) expression was diminished in brain regions potentially involved in the regulation of eating behavior in rats which are more sensitive to cocaine and alcohol. We hypothesized that the human AKR1B10 gene may also play a role in the regulation of human eating behavior.

Breast cancer is a disease that afflicts a significant proportion of women globally. 5-10% of breast cancer cases are linked to inherited polymorphisms in critical genes such as BRCA1, a tumour suppressor essential for genomic stability. A dysfunctional BRCA1 gene can increase breast cancer risk by 60-80%. Previous microarray analysis established that differential gene expression in unperturbed Epstein-Barr virus transformed lymphocyte cell lines (EBV-LCL) was able to distinguish BRCA1 mutation carriers from controls with a high degree of accuracy. A 43-gene radiation-independent classifier for BRCA1 status was constructed. We hypothesize that this differential gene expression can be observed in a subset of these genes using quantitative PCR (qPCR) in both EBV-LCL and B-lymphocytes isolated from patients with known BRCA1 mutation carrier status. The 43-gene classifier was analyzed using gene ontology analysis and 4 target genes selected based on predictive value, expression intensity and gene ontology similarity. Genes selected were CXCR3, TBX21, MX2, and IFIT1, with GusB as an endogenous reference gene. EBV-LCL established from known BRCA1 mutation carriers and from BRCA1 wildtype individuals were obtained and RT-qPCR (reverse transcriptase qPCR) performed on isolated RNA. Our results showed significant downregulation of CXCR3 and TBX21 in BRCA1 mutation carriers (p=0.018 and p=0.003, respectively), as expected from previous microarray results. IFIT1, while showing a non-significant upregulation (p=0.183), agreed with the expected trend. MX2 did not show significant differential expression. These results indicate that differential gene expression has the potential to accurately distinguish pathogenic variants, even if it may require EBV immortalization of B-lymphocytes. To determine whether the assay could be extended to fresh blood samples, B-lymphocytes were isolated from patients with known BRCA1 mutation carrier status from North York General Hospital in Toronto, ON. An optimized protocol to enrich the B-lymphocyte population using magnetic separation was developed for this purpose. RT-qPCR using RNA isolated from these lymphocytes showed no significant differential gene expression in CXCR3 and TBX21. However, a low sample size, use of non-sequenced controls and a need for further qPCR optimization may call these results into question. In addition, problems with blood sample transportation from off-site sources resulted in an unacceptable drop in RNA integrity. While this gene expression assay may be limited to screening a small number of blood samples, results indicate that may still have clinical relevance that can be explored. This would necessitate further optimization of the qPCR methodology and resolution of the issues surrounding RNA integrity and sample transport. / Thesis (Master, Pathology & Molecular Medicine) -- Queen's University, 2013-09-26 13:13:50.809

No description available.

Genome wide association studies (GWAS) and their subsequent meta-analysis have identified a large number of susceptibility variants for Type 2 diabetes (T2D) risk. However, the familial aggregation seen in this disease is not yet fully explained. The sibling relative risk (λ_s) due to all known variants is ~1.104 which is well below the epidemiological estimates of λ_s of ~3.0. There has therefore been great interest in the potential role of variants that would have been largely invisible to the initial wave of GWAS and linkage approaches. Low frequency (minor allele frequency 1-5%), incompletely penetrant (odds ratio 2-4) variants (LFIP), are one such group of potential susceptibility variants. The overall objective of this project (designed and implemented in 2007-2010) was to evaluate the contribution of LFIP variants to the inherited susceptibility to T2D. I tested the specific hypothesis that genes already-implicated in diabetes pathogenesis (due to an established role in monogenic or multifactorial disease) also harbour LFIP variants, and that those variants may contribute appreciably to the prediction of disease risk. Mutations in exons only encoding isoform-A of HNF1A have been demonstrated to lead to a later age of diagnosis of HNF1A-MODY. This region was therefore felt to be auspicious for harbouring LFIP variants impacting on T2D risk. I have demonstrated that such variants impacting on T2D risk are unlikely to be present in this region by use of Sanger sequencing in a sample enriched for young onset, familial T2D. The role in T2D risk of candidate LFIP variants across 5 genes (HNF1A, HNF4A, PDX1, KCNJ15 and LARS2), was evaluated by large scale association studies. For one variant, T130I of HNF4A, a modest association (p=5x10^-4) with T2D was seen in UK samples and the strength of association was marginally improved by incorporation of all previous studies of this variant in T2D in a meta-analysis (p=2.1x10^-5). This study demonstrated the difficulties encountered in confirming the association of low frequency variants to complex diseases, especially for those with modest effect sizes. At the time of project design and inception “next-generation” sequencing platforms were in their infancy and the study design I planned (that of pooled, targeted sequencing) had not been widely applied. It was therefore necessary to design and optimise protocols for sample preparation for sequencing on this platform. I used the Genome Analyzer II platform to sequence ten genes previously implicated in T2D or monogenic diabetes pathogenesis in pooled DNA samples. This approach yielded in excess of 2900 variants, a large portion being novel. As part of this project I have highlighted heuristics that can be used in the follow-up of potential susceptibility variants discovered using high throughput sequencing. I have also established protocols and pathways for sample preparation that can be utilised across several next generation sequencing platforms for future studies in the host institution and beyond.