Application of knowledge discovery and data mining methods in livestock genomics for hypothesis generation and identification of biomarker candidates influencing meat quality traits in pigs

Recent advancements in genomics and genome profiling technologies have lead to an increase in the amount of data available in livestock genomics. Yet, most of the studies done in livestock genomics have been following a reductionist approach and very few studies have either followed data mining or knowledge discovery concepts or made use of the wealth of information available in the public domain to gain new knowledge. The goals of this thesis were: (i) the adoption of existing analysis strategies or the development of novel approaches in livestock genomics for integrative data analysis following the principles of data mining and knowledge discovery and (ii) demonstrating the application of such approaches in livestockgenomics for hypothesis generation and biomarker discovery. A pig meat quality trait termed androstenone measurement in backfat was selected as the target phenotype for the experiments. Two experiments were performed as a part of this thesis. The first one followed a knowledge driven approach merging high-throughput expression data with metabolic interaction network. Based on the results from this experiment, several novel biomarker candidates and a hypothesis regarding different mechanisms regulating androstenone synthesis in porcine testis samples with divergent androstenone measurements in back fat were proposed. The model proposed that the elevated levels of androstenone synthesis in sample population could be due to the combined effect of cAMP/PKA signaling, elevated levels of fatty acid metabolism and anti lipid peroxidation activity of members of glutathione metabolic pathway. The second experiment followed a data driven approach and integrated gene expression data from multiple porcine populations to identify similarities in gene expression patterns related to hepatic androstenone metabolism. The results indicated that one of the low androstenone phenotype specific co-expression cluster was functionally enriched in pathways related to androgen and androstenone metabolism and that the members of this cluster exhibited weak co-expression in high androstenone phenotype. Based on the results from this experiment, this co-expression cluster was proposed as a signature cluster for hepatic androstenone metabolism in boars with low androstenone content in back fat. The results from these experiments indicate that integrative analysis approaches following data mining and knowledge discovery concepts can be used for the generation of new knowledge from existing data in livestock genomics. But, limited data availability in livestock genomics is a hindrance to the extensive use such analysis methods in livestock genomics field for gaining new knowledge. In conclusion, this study was aimed at demonstrating the capabilities of data mining and knowledge discovery methods and integrative analysis approaches to generate new knowledge in livestock genomics using existing datasets. The results from the experiments hint the possibilities of further exploring such methods for knowledge generation in this field. Although the application of such methods is limited in livestock genomics due to data availability issues at present, the increase in data availability due to evolving high throughput technologies and decrease in data generation costs would aid in the wide spread use of such methods in livestock genomics in the coming future

DETERMINATION AND TOXICOLOGICAL EVALUATION OF MICROCYSTINS IN TROPICAL RESERVOIRS.

Ph.DDOCTOR OF PHILOSOPH

Understanding how high stocking densities and concurrent limited oxygen availability drive social cohesion and adaptive features in regulatory growth, antioxidant defense and lipid metabolism in farmed gilthead sea bream (Sparus aurata)

The study combined the use of biometric, behavioral, physiological and external tissue damage scoring systems to better understand how high stocking densities drive schooling behavior and other adaptive features during the finishing growing phase of farmed gilthead sea bream in the Western Mediterranean. Fish were grown at three different final stocking densities (LD, 8.5 kg/m3; MD, 17 kg/m3; HD, 25 kg/m3). Water oxygen concentration varied between 5 and 6 ppm in LD fish to 3–4 ppm in HD fish with the summer rise of water temperature from 19°C to 26°C (May–July). HD fish showed a reduction of feed intake and growth rates, but they also showed a reinforced social cohesion with a well-defined endogenous swimming activity rhythm with feeding time as a main synchronization factor. The monitored decrease of the breathing/swimming activity ratio by means of the AEFishBIT data-logger also indicated a decreased energy partitioning for growth in the HD environment with a limited oxygen availability. Plasma glucose and cortisol levels increased with the rise of stocking density, and the close association of glycaemia with the expression level of antioxidant enzymes (mn-sod, gpx4, prdx5) in liver and molecular chaperones (grp170, grp75) in skeletal muscle highlighted the involvement of glucose in redox processes via rerouting in the pentose-phosphate-pathway. Other adaptive features included the depletion of oxidative metabolism that favored lipid storage rather than fatty acid oxidation to decrease the oxygen demand as last electron acceptor in the mitochondrial respiratory chain. This was coincident with the metabolic readjustment of the Gh/Igf endocrine-growth cascade that promoted the regulation of muscle growth at the local level rather than a systemic action via the liver Gh/Igf axis. Moreover, correlation analyses within HD fish displayed negative correlations of hepatic transcripts of igf1 and igf2 with the data-logger measurements of activity and respiration, whereas the opposite was found for muscle igf2, ghr1 and ghr2. This was indicative of a growth-regulatory transition that supported a proactive instead of a reactive behavior in HD fish, which was considered adaptive to preserve an active and synchronized feeding behavior with a minimized risk of oxidative stress and epidermal skin damage

Statistical modelling of masked gene regulatory pathway changes across microarray studies of interferon gamma activated macrophages

Interferon gamma (IFN-γ) regulation of macrophages plays an essential role in innate immunity and pathogenicity of viral infections by directing large and small genome-wide changes in the transcriptional program of macrophages. Smaller changes at the transcriptional level are difficult to detect but can have profound biological effects, motivating the hypothesis of this thesis that responses of macrophages to immune activation by IFN-γ include small quantitative changes that are masked by noise but represent meaningful transcriptional systems in pathways against infection. To test this hypothesis, statistical meta-analysis of microarray studies is investigated as a tool to obtain the necessary increase in analysis sensitivity. Three meta-analysis models (Effect size model, Rank Product model, Fisher’s sum of logs) and three further modified versions were applied to a heterogeneous set of four microarray studies on the effect of IFN-γ on murine macrophages. Performance assessments include recovery of known biology and are followed by development of novel biological hypotheses through secondary analysis of meta-analysis outcomes in context of independent biological data sources. A separate network analysis of a microarray time course study investigate s if gene sets with coordinated time-dependent relationships overlap can also identify subtle IFN-γ related transcriptional changes in macrophages that match those identified through meta-analysis. It was found that all meta-analysis models can identify biologically meaningful transcription at enhanced sensitivity levels, with slightly improved performance advantages for a non-parametric model (Rank Product meta-analysis). Meta-analysis yielded consistently regulated genes, hidden in individual microarray studies, related to sterol biosynthesis (Stard3, Pgrmc1, Galnt6, Rab11a, Golga4, Lrp10), implicated in cross-talk between type II and type I interferon or IL-10 signalling (Tbk1, Ikbke, Clic4, Ptpre, Batf), and circadian rhythm (Csnk1e). Further network analysis confirms that meta-analysis findings are highly concentrated in a distinct immune response cluster of co-expressed genes, and also identifies global expression modularisation in IFN-γ treated macrophages, pointing to Trafd1 as a central anti-correlated node topologically linked to interactions with down-regulated sterol biosynthesis pathway members. Outcomes from this thesis suggest that small transcriptional changes in IFN-γ activated macrophages can be detected by enhancing sensitivity through combination of multiple microarray studies. Together with use of bioinformatical resources, independent data sets and network analysis, further validation assigns a potential role for low or variable transcription genes in linking type II interferon signalling to type I and TLR signalling, as well as the sterol metabolic network

Histological, cellular, and molecular abnormalities in forebrain and spinal cord of three distinct mouse models of Down syndrome

Down syndrome (DS) is a developmental disorder caused by a triplication of human chromosome 21, which contains approximately 550 genes. DS is the most common autosomal aneuploidy occurring with an incidence of 1 in 793 live births. Hallmarks of DS include abnormal central nervous system (CNS) development and function resulting in intellectual disability (ID), motor dysfunction, and early onset Alzheimer’s neuropathology. Studies have elucidated widespread neurohistological abnormalities in brains of fetuses with DS as early as 20 weeks of gestation, suggesting that early dysfunction in neural development may set the stage for exacerbated CNS abnormalities throughout life. Additionally, the complex constellation of symptoms associated with DS changes over the lifespan, particularly in adolescence and in middle to old age. Thus, these periods may represent opportune windows for age-specific therapeutic interventions. Due to ethical and practical constraints, use of human samples is alone insufficient to characterize the etiological underpinnings of DS phenotypes across the lifespan. Furthermore, while human data are instructive for drug development, preclinical trials are necessary for target validation, to establish dosage, and to prove safety and efficacy of any proposed therapeutic. With the advent of mouse models of DS, informative studies on the neurobiology of DS as well as preclinical testing of proposed therapies are possible. Here, we use a multi-pronged approach to assess molecular, neuroanatomical, and behavioral phenotypes indicative of brain and SC function in three distinct mouse models of DS: Ts1Cje, Ts65Dn, and Dp16. We identify neurodevelopment phenotypes, cytoarchitectural aberrations, bioenergetic abnormalities, myelination deficits, and motor/cognitive dysfunction at multiple ages spanning the period between embryonic day 12.5 and 6-7 months in trisomic mice. Additionally, we show that while Ts65Dn mice recapitulate all known phases of histological, functional, and behavioral phenotypes typical of DS starting from prenatal development and into middle age, this is not true for the Ts1Cje or Dp16 models. Lastly, we present promising outcomes of two possible therapies for cognitive and motor dysfunction in Ts65Dn mice. Altogether our findings provide insights into the underlying neurobiology of ID and motor dysfunction in DS and elucidate molecular changes that can be targeted for future therapeutic intervention.2018-07-09T00:00:00

Role of monounsaturated fatty acids in the improvement of non-alcoholic fatty liver disease and type 2 diabetes mellitus

Programa de Doctorado en Biotecnología, Ingeniería y Tecnología QuímicaLínea de Investigación: Biotecnología, Biomedicina y Ciencias de la SaludClave Programa: DBICódigo Línea: 110En los últimos años, debido a la creciente incidencia de obesidad, estilos de vida sedentarios y dietas poco saludables en todo el mundo, ha aumentado la prevalencia del síndrome metabólico y sus comorbilidades, como la diabetes mellitus tipo dos (T2D) y la enfermedad del hígado graso no alcohólico (EGHNA). Europa ha sido testigo de entre el 20 % y el 30 % de los casos. Por lo tanto, el síndrome metabólico se considera como un importante problema de salud pública. En un intento por comprender estas enfermedades y descubrir los mecanismos moleculares implicados para encontrar posibles dianas terapéuticas, en esta Tesis Doctoral se han estudiado varios modelos animales que recapitulan todas las características de estas enfermedades de la forma más parecida posible a los humanos ya que a pesar de la alta incidencia tanto del síndrome metabólico como de sus comorbilidades, no existe un tratamiento farmacológico efectivo. En general, una dieta hipercalórica, y en particular una rica en grasas trans, grasas saturadas, colesterol y bebidas endulzadas con fructosa, parece aumentar la adiposidad visceral y estimular la acumulación de lípidos en el hígado, así como la progresión de EGHNA y resistencia a la insulina. Sin embargo, la reducción de la ingesta calórica y la suplementación con ácidos grasos monoinsaturados omega-3 parece tener efectos preventivos y terapéuticos. Hay muchos estudios que demuestran que las dietas con grasas que representan al menos el 40% de la ingesta calórica conducen a la aparición de síndrome metabólico, T2D y EGHNA. Por el contrario, varios estudios indican que las grasas monoinsaturadas y poliinsaturadas tienen efectos beneficiosos sobre las enfermedades metabólicas. Por lo tanto, estos hallazgos contradictorios nos llevaron a considerar si una dieta alta en grasas con un alto consumo de ácidos grasos monoinsaturados y compuestos polifenólicos presentes en el aceite de oliva virgen extra puede tener efectos beneficiosos sobre estas enfermedades. Es por ello que la siguiente Tesis Doctoral que se presenta mediante la modalidad de compendio de publicaciones y en la que se aportan distintas contribuciones científicas en revistas con alto índice de impacto en el Journal of Citation Reports (JCR), se han evaluado los siguientes aspectos: (i) El papel de la ingesta de una dieta alta en grasas enriquecida en grasas monoinsaturadas en la etiopatogenia de la EGHNA y T2D. (ii) La optimización de diferentes modelos de ratones para el estudio de trastornos hepáticos relacionados con EGHNA. (iii) El estudio de los mecanismos responsables de los efectos beneficiosos del aceite de oliva virgen extra en tejidos metabólicos activos (hígado y páncreas). (iv) La comprensión de las rutas y redes de señalización molecular, para obtener una visión integradora necesaria para explicar las mejoras producidas a nivel fisiopatológico, en animales alimentados con dietas altas en grasas enriquecidas con ácidos grasos monoinsaturados provenientes del aceite de oliva virgen extra.Universidad Pablo de Olavide de Sevilla. Departamento de Biología Molecular e Ingeniería Bioquímic

Food Bioactives

A full awareness of the role played by a healthy diet, as part of a healthy lifestyle, in countering or slowing-down chronic and degenerative diseases has strongly increased the interest in food bioactives and the return of ancient foods that are nowadays considered functional. In fact, these dietary substances, to which nutraceutical attributes are increasingly entrusted, could display disease-preventing effects on animals and humans. In this context, polyphenols, which are widespread and mostly copious in dietary plant sources, have gained a lot of attention thanks to their potential ability to halt or reverse oxidative stress-related diseases. Indeed, food could contain, beyond health-promoting compounds, toxicants which are naturally occurring or process-induced dietary compounds with adverse effects on human health. The presence and abundance of bioactives are strictly related to their food source. Edible plant components largely contain beneficial secondary metabolites, but understanding them fully is still an important challenge as complex biotic and abiotic interactions are involved in their biosynthesis. Analytical methods, which are increasingly powerful, could enhance our knowledge of food bioactives, whereas the deep investigation of their bioactivity and bioavailability could make them particularly useful