This thesis is an investigation of the genetic basis of tocopherol accumulation in soybean (Glycine max [L.] Merrill) seeds. Soybean is the world’s most widely grown protein and oilseed crop and the principle source of vitamin E (tocopherols) as a supplement. Tocopherols (α-, β-, γ- and δ-isomers) are powerful antioxidants that contain human health benefits, including a decrease in the risk of lung cancer, heart disease and osteoporosis. The purpose of this research was to identify genetic and biochemical components affecting tocopherol accumulation in soybean seeds. The objectives were to: 1) investigate location and year effects on soybean seed tocopherol levels in the field; 2) determine environmental factors affecting soybean seed tocopherol levels under controlled conditions; 3) identify simple sequence repeat (SSR) markers that tag quantitative trail loci (QTL) for individual and total tocopherols; and 4) evaluate the potential role of VTE1, VTE3 and VTE4 genes in tocopherol accumulation using the candidate gene approach. Seventy nine recombinant inbred lines (RILs) derived from the cross between OAC Bayfield and OAC Shire were grown in the field at Elora, Woodstock and St. Pauls, ON, in 2009 and 2010. The tocopherol components were quantified using high performance liquid chromatography (HPLC). The results showed a significant (p < 0.001) genotype, environment and genotype x environment effect for each tocopherol component. It was discovered that a 2 x phosphate fertilizer (K2SO4 at 1.0M/150mL) and 30 ˚C temperature treatment increased each tocopherol component, whereas drought had no effects. Single marker analysis identified 42 QTL and interval mapping identified 26 QTL across 17 chromosomes. Significant two-locus epistatic interactions were found with a total of 122 and 152 in the 2009 and 2010 field seasons, respectively. The multiple locus models explained 18.4% to 72.2% with an average of 45.7% of the total phenotypic variation. The candidate gene approach using nucleotide sequences from the coding regions identified two single nucleotide polymorphisms (SNPs) in VTE1, five SNPs in VTE4 and none in VTE3. The SNPs were predicted to cause functional protein changes and the genes co-localized with some of the identified QTL. The results of this study provide a better understanding of the environmental factors and genetic mechanisms that influence the accumulation of tocopherols in soybean seeds. / Grain Farmers of Ontario, Vitamin Research Award

La sténose valvulaire aortique (SVA) est une valvulopathie résultant en l'ouverture incomplète de la valve aortique. La calcification des feuillets associée au vieillissement est la cause la plus importante de la SVA. Sa pathogénèse implique des dépôts de lipoprotéines, de l'inflammation et de la calcification des feuillets. Notre étude vise à identifier les gènes associés à une prédisposition à la SVA afin de mieux comprendre les mécanismes sous-jacents à cette maladie et potentiellement identifier de nouvelles cibles thérapeutiques. Pour ce faire, nous avons recruté 190 patients avec SVA dégénérative et 192 témoins, appariés pour l'âge et le sexe, puis effectué une étude d’association par gènes candidats en utilisant des marqueurs génétiques polymorphiques (SNP). Les gènes candidats choisis incluent (1) ceux dont les polymorphismes ont été présumés associés à la SVA dans des études antérieures (APOB, APOE, ESR1, PTH et VDR) (2) des gènes dont les polymorphismes ont été significativement associés et validés pour quelques maladies inflammatoires (IL-10, TNFAIP3) ou pour le métabolisme lipidique (PCSK9, LDLR) dans des études d’association pangénomiques, et (3) des gènes impliqués dans la pathogénie de la SVA à partir d’études faites sur des modèles animaux en lien avec la calcification (BMP2, CCR5, CTGF, LRP5, MSX2, WNT3), le remodelage tissulaire (CTSS, MMP9) ou le métabolisme lipidique (SMPD1). Pour les gènes des groupes (1) et (2), nous avons utilisé les SNPs rapportés dans la littérature comme étant significativement associés. Pour le groupe (3), nous avons effectué une approche par «tagSNP» qui consiste à sélectionner un groupe de SNP capturant la variabilité génétique dans la région ciblée. Au total, 81 SNPs dans 18 gènes ont été testés. Nous avons trouvé une association nominale avec les gènes BMP2 (OR = 1.55, IC95%: 1.14-2.10, p = 0.004) et LRP5 (OR = 1.47, IC95%: 1.06-2.03, p = 0.023) après ajustement pour la maladie coronarienne. Les gènes BMP2 et LRP5, impliqués dans la calcification selon certains modèles expérimentaux, sont donc associés à la SVA. Ce travail devrait être validé dans une cohorte indépendante plus large dans un avenir rapproché et il pourrait être étendu à d’autres gènes. / Aortic valve stenosis (AVS) is a valvular heart disease caused by calcification leading to incomplete opening of the aortic valve. Calcification of valve leaflets associated with aging is the most common cause of AVS. AVS pathogenesis involves lipoprotein deposits, chronic inflammation and calcification of the aortic valve leaflets. Our study aims to identify genes associated with AVS in order to better understand its mechanisms and potentially identify new therapeutic targets. We recruited 190 cases with AVS of different severity and 192 controls matched for age and sex. Then we conducted a candidate gene association study using single nucleotide polymorphisms (SNPs). The candidate genes selected include: (1) those with polymorphisms putatively implicated in previous genetic association studies of AVS (APOB, APOE, ESR1, PTH and VDR); (2) those with validated associations to inflammatory diseases (IL-10, TNFAIP3) or lipid metabolism (LDLR ,PCSK9) in genome-wide association studies and, (3) genes impliated in AVS pathogenesis from studies with animal models and thought to be involved in calcification (BMP2, CCR5, CTGF, LRP5, MXS2, WNT3); tissue remodeling (CTSS, MMP9) or lipid metabolism (SMPD1). For the first two categories of genes, we tested the SNPs reported to be associated in the literature and, in the third category we used a tag-SNP approach which consists of selecting a subset of SNPs to capture variability in the target region. Finally, 81 SNPs in 18 genes were tested. We found a nominal association of BMP2 (OR=1.55, CI: 1.14 – 2.10, p=0.004) and LRP5 (OR=1.47, CI: 1.06 – 2.03, p=0.023) with presence of AVS after adjustment for coronary heart disease. The genes BMP2 and LRP5, which are known to be involved in calcification based on animal models, are associated with AVS. The result of the current study should be validated in a larger independent cohort in the near future and then, it could also be extended to the study of other genes.

Escherichia coli diarrhoea is the most important source of mortality in piglets. The most frequently isolated strain in enterotoxigenic E. coli diarrhoea is F4ab/ac. Recent studies in South Africa reported non-fimbrial strains such as PAA and EAST-1 to be prevalent. The objective of the study was to determine whether there are breed differences among pigs with respect to E. coli adhesion phenotypes and correlate them to polymorphisms at selected candidate genes in the South African population. A total of 225 pigs aged 3-12 weeks of the imported (Large White, Landrace and Duroc), local and crossbreds, were sampled from the Eastern Cape and Limpopo provinces of South Africa and genotyped for PCR-RFLP polymorphisms at four candidate genes associated with E. coli F4ab/ac resistance/susceptibility. These genes were Mucin 4 (MUC4), Mucin 13, (MUC13), Mucin 20 (MUC20) and Transferrin Receptor (TFRC). The TFRC and MUC13 genes were less polymorphic, the C allele was close to fixation and the homozygous CC genotype was the most frequent in all three pig populations. There was a significant difference (P <0.05) in allelic and genotypic distribution amongst breeds for the TFRC locus. The g.8227G>C polymorphism in MUC4 segregated in all three breeds and the marker was moderately polymorphic. There was a significant difference (P <0.05) in genotypic distribution amongst breeds for MUC4.The g.191C>T polymorphism in MUC20 segregated in the local and crossbred pigs and was close to fixation in the imported pigs. There was a significant difference (P <0.05) in allelic and genotypic distribution amongst breeds for MUC20, which was moderately polymorphic. There was a reduction in heterozygosity in both the TFRC and MUC13 loci, although MUC4 and MUC20 genes had higher heterozygosity levels. The MUC4 gene had a negative FIS value, indicating outbreeding at this locus. The MUC20, MUC13 and TFRC genes had a positive FIS value, indicating inbreeding at these loci. Overall, the studied population was outbred. Imported pigs in TFRC and MUC20 deviated from Hardy-Weinberg equilibrium (HWE). All breeds were in HWE at the MUC4 and MUC13 genes. There was no linkage disequilibrium observed amongst the analysed loci. iv A total of 109 piglets of three breeds (Large White, indigenous and crossbred) aged 3-5 weeks, were investigated for the susceptibility to E. coli F4, PAA strains and EAST-1 toxin. Adhesion tests were conducted on pig intestinal cells, which were viewed under a phase contrast microscope. Three phenotypes were identified as, adhesive, weakly adhesive and non-adhesive. There was a significant association (P <0.05) between breed and level of adherence of the F4 and PAA strains. Highest frequencies of adhesion phenotypes were observed in the indigenous pigs for both F4 and PAA E. coli strains. Large White pigs had the lowest frequency of non-adhesion in F4 and PAA E. coli strains. The F4 strain had a higher (P <0.05) level of adherence compared to PAA and EAST-1 in Large White pigs. Age of pigs had a significant effect on the level of E. coli adherence in indigenous and crossbred pigs (P <0.05). Adhesion of F4 and EAST-1 was higher in weaned indigenous and crossbred pigs, respectively, than in suckling piglets. There was no significant difference between F4 adhesion and the genotypes at all four candidate genes genotypes. The study showed that both imported and local pig populations carry receptors and are susceptible to F4, PAA and EAST-1 E. coli infections. Indigenous pigs were less susceptible than Large White to E. coli infection. Although polymorphic and segregating in the populations, the MUC4 g.8227G>C and MUC20 g.191C>T mutations were not associated with the adhesion phenotypes and cannot be used in the selection of susceptible animals. / M.Sc.Agric. University of KwaZulu-Natal, Pietermaritzburg 2013.

This thesis analyses the process of women&rsquo / s legislative recruitment in Turkey by focusing on the interaction among the certain features of the Turkish political system including the attitudes of the party elite toward the enhancement of women&rsquo / s political representation. It has been demonstrated in earlier studies that one of the reasons behind women&rsquo / s low level of parliamentary representation in Turkey is the fact that the selectors in the political parties fail to support women candidates adequately in the elections. Related to that, women&rsquo / s legislative recruitment is also likely to be impeded by the unsupportive nature of the main dynamics of political system such as political culture, party system and the v electoral system. Considering the fact that these institutional and attitudinal factors are highly inter-related with each other, the insufficient number of women candidates nominated by the major social democratic party, the Republican People&rsquo / s Party in the 1990s, including the latest national elections on November 3rd, 2002 calls for an analysis of not only the attitudes of the RPP selectors toward positive discrimination mechanisms such as the quotas for women but also of the relevant party institutional factors rooted in the Turkish political system. The study contends that the RPP&rsquo / s women-friendly party culture and selectors&rsquo / positive perspectives towards women&rsquo / s political integration are not sufficient factors for the promotion of women&rsquo / s legislative recruitment within the party. The weakness of the intra-party democracy in the RPP which is particularly the result of the oligarchic structure and the problems with institutionalization significantly impede women&rsquo / s legislative recruitment within the party. The study has found that the low level of women&rsquo / s legislative recruitment in the RPP is a consequence of the interaction between the institutional and attitudinal factors within the RPP which perpetuates patriarchal elite oligarchy in the nomination process. The documented gap between the RPP&rsquo / s party rhetoric and practice in this study is found as the reflection of the party&rsquo / s problems in internalizing social democratic values.

In the late 20th century, the proportions of women and men elected into European national parliaments became increasingly equal. Political parties shape these outcomes by selecting and fielding candidates in elections. Scholars recognise that parties' actions do not occur in isolation; yet there is little systematically comparative research about the configurations of conditions in which these actions occur. Previous research also often relies on studies of West European parties. This doctoral thesis investigates how conditions inside and outside parties combine to create gender-equal parliamentary presence. The thesis examines the extent to which Western European experiences apply to Central and East European parties, and explores the conditions that stand in the way of progress towards gender balance. It presents three empirical studies. The first is a qualitative comparative analysis of 57 West European parties during the late 1980s, a period in which the trend towards equality accelerated. The second study applies the knowledge produced in the first analysis to cases in Central and Eastern Europe. It uses an original dataset covering six parties in four EU member states in a structured focused comparison. Finally, the thesis presents an in-depth case study of an unexpectedly gender-balanced Latvian party. The analyses show that gender-equal parliamentary presence is achieved when conditions inside and outside parties combine, and that no condition is necessary or singularly sufficient. The absence of gender-equal parliaments is sustained by combinations other than the absence of those that lead to gender-balance. Operationalisations from Western Europe turn out to be largely applicable to cases in Central and Eastern Europe. These latter cases also demonstrate that organisational instability need not impede women’s presence in elected office. / Under sent 1900-tal har andelen kvinnor och män i nationella demokratiska parlament i Europa blivit alltmer jämstora. Politiska partier formar politisk representation genom att välja egna kandidater till val. Forskare har visat att partiers beteende på den punkten inte sker i isolering, men det finns ändå en brist på systematiskt jämförande studier om vilka kombinationer av villkor som leder till jämn könsrepresentation i nationella parlament. Dessutom vilar tidigare studier ofta på kunskap om situationen i Västeuropa. Den här doktorsavhandlingen undersöker hur villkor i och utanför politiska partier kombineras för att uppnå jämställd parlamentarisk representation. Den utforskar i vilken grad de västeuropeiska erfarenheterna är användbara i Öst- och Centraleuropa, och studerar villkoren som upprätthåller manlig dominans i parlamentariska partier. Den presenterar tre empiriska studier. Den första är en kvalitativt jämförande studie (fsQCA) av 57 Västeuropeiska partier under sent 1980-tal, en period då andelen kvinnor i nationella parlament ökade. Den andra studien tillämpar kunskapen från den första studien på fall i Öst- och Centraleuropa. Studien bygger på ett unikt dataset med sex partier från fyra EU-stater, i en strukturerad fokuserad jämförelse. Slutligen presenteras en fallstudie av ett ovanligt jämställt parti i Lettland. Analyserna visar att lika andelar kvinnor och män i nationella parlament åstadkoms när villkor i och utanför partier kombineras, och att inget villkor är nödvändigt eller ensamt tillräckligt. Frånvaro av jämn representation upprätthålls av andra kombinationer än de som leder till jämn representation. Operationaliseringarna som utvecklades för Västeuropa är applicerbara i Öst- och Centraleuropa. De senare fallen visar också att organisatorisk instabilitet inte behöver hindra en jämställd parlamentarisk närvaro.

Background: Primary aldosteronism (PAL) has emerged as the commonest specifically treatable and potentially curable form of secondary hypertension. With its propensity towards cardiovascular complications above that expected from hypertension alone, PAL is a potentially highly detrimental state which should be detected as early as possible in the course of the disease and treated appropriately. The detection of earlier, milder, normokalaemic forms of PAL using the aldosterone/renin ratio (ARR) as a screening test has significantly enlarged the clinical spectrum of PAL and facilitated identification of a new familial variety (familial hyperaldosteronism type II, FH-II). Unlike familial hyperaldosteronism type I (FH-I), FH-II is not glucocorticoid remediable and is not caused by the CYP11B1/CYP11B2 “hybrid” gene mutation. The genetic defects underlying FH-II have not yet been elucidated and hence, detection of FH-II still involves complicated and time-consuming biochemical screening by ARR testing and confirmation by carefully performed suppression testing such as fludrocortisone suppression testing. Diagnosing PAL by currently available biochemical methods is tedious. Finding a simple and reliable genetic test for FH-II which could be applied to all members of a family with known FH-II and also to apparently sporadic PAL would simplify patient management. A genome-wide search has already demonstrated linkage of FH-II to chromosome 7p22, consistent with this locus harbouring the causative gene/s for FH-II. Three candidate genes [retinoblastoma-associated Kruppel-associated box gene (RBaK, involved in tumorigenesis and cell cycle control), postmeiotic segregation increased 2 (PMS2, involved in DNA mismatch repair and tumour predisposition) and guanine nucleotide-binding protein alpha-12 (GNA12, a transforming oncogene)] within this linked locus have been examined in an attempt to find the causative mutations for FH-II, but no clear causative mutations have so far been found. PAL continues to be a challenging yet rewarding disease to manage. Although much has been learnt about PAL, there are still many areas which have not been explored. PAL considered due to bilateral autonomous production of aldosterone is usually treated medically because unilateral adrenalectomy has been considered ineffective. Since medical treatment may cause adverse effects or fail to control hypertension, defining the role of unilateral adrenalectomy in bilateral PAL is an important clinical issue, but quality outcome data are lacking. The candidate therefore peformed a retrospective study of the efficacy of unilateral adrenalectomy in patients with bilateral PAL. In patients with unilateral PAL, unilateral laparoscopic adrenalectomy has been shown to correct hypokalaemia and lead to cure or improvement in hypertension control. While most studies have focused on clinical and biochemical outcomes, to the candidate’s knowledge, there are no data on the effects of adrenalectomy on quality of life (QOL). Assessing the QOL in patients with unilateral PAL before and after unilateral laparoscopic adrenalectomy (which cured hypokalaemia in all and hypertension in the majority) provided an insight into the degree to which the disease process and/or its treatment affects the life of an individual with PAL. Aims: The overall aims of this thesis were to further explore the genetic basis of FH-II, to examine the role of adrenalectomy in patients with bilateral PAL and the effects of unilateral adrenalectomy on QOL in unilateral PAL as a first step in dissecting out the effects of medical and surgical treatment on QOL in the more common bilateral PAL. In order to address the overall aims, the specific aims of the thesis were (1) to narrow the linked region at 7p22 by phenotyping and genotyping additional FH-II families from Italy, using more closely spaced microsatellite markers at 7p22, and then assess the combined multipoint logarithm of odds (LOD) score for these Italian as well as two Australian and one South American families; (2) to sequence candidate genes in the narrowed linked region for FH-II associated mutations; (3) to assess the role of unilateral adrenalectomy in bilateral PAL and identify predictive parameters; and (4) to assess the quality of life following unilateral adrenalectomy in patients with unilateral PAL. Methods and Results: Two Italian families with FH-II were genotyped using seven closely spaced microsatellite markers at 7p22. All known affected individuals from each of the two Italian families were found to share identical haplotypes for the seven markers, consistent with linkage of the disease locus with the 7p22 region. The multipoint LOD score of the now five known families with FH-II which demonstrate linkage at 7p22, calculated using MERLIN linkage analysis was highly significant at 5.22. Three candidate genes in this linked region were then examined for mutations causing FH-II; the replication protein A 3 (RPA3), zinc finger protein 12 (ZNF12) and glucocorticoid induced transcripts 1 (GLCCI1) genes were selected as they are involved in cell cycle control, and adrenal hyperplasia and adenomas are common in FH-II. Using the method of polymerase chain reaction-sequencing, coding regions, splice sites, proximal promoter, 5’ and 3’ untranslated regions (UTR) were sequenced in affected and unaffected subjects from the 7p22-linked FH-II families. Identified single nucleotide polymorphisms (SNPs) were genotyped to assess significance. For RPA3, four different SNPs were initially found to segregate with the affectation status, that is, they were present in the two affected and not the two unaffected subjects from the largest Australian family (family 1, eight affecteds) with FH-II. However, only two SNPs (rs2024374 G/C and rs17169194 T/G) were further genotyped as that they were in functionally important positions of the gene (that is, in regulatory regions within the promoter and 5’ UTR) and because of the relatively low allele frequency reported in the literature for these two SNPs in controls. Further genotyping of these SNPs was carried out in another six affecteds and four unaffecteds from the same family and a complete segregation of these two SNPs with affectation status was seen in family 1. The G/C mutation rs2024374 in the RPA3 promoter results in the loss of three transcription factor binding sites and creation of one new site. The factors for which the binding sites in the RPA3 promoter and 5’UTR were altered by these two SNPs were involved in regulation of cell differentiation, proliferation and apoptosis. Hence, it is possible that altered activity of the RPA3 promoter and 5’UTR in family 1 could result in predisposition to adrenal hyperplasia or neoplasia, altered ARR and/or hypertension. Genotyping of these SNPs was then carried out in another two pedigrees (families 2 and 3) that showed linkage to 7p22, and in 75 normotensive, non-PAL control subjects. However, neither of these two SNPs segregated with the affectation status in family 2 and 3, and they were present in 30% and 20% of controls, respectively. For ZNF12 and GLCCI1, no evidence of causative mutations was found in the coding regions, splice sites, proximal promoter region and proximal 5’ and 3’ UTR. Between 1984 and 2004, 51 of 684 patients diagnosed with bilateral PAL underwent unilateral adrenalectomy. Forty patients fulfilled the inclusion criteria and were followed for at least 12 (median 56.4) months. Hypertension was cured in 15% and improved in 20%, usually within one year of unilateral adrenalectomy. The proportion with controlled hypertension was significantly (p<0.001) higher after adrenalectomy (65%) than before (25%). Mean systolic (p<0.001) and diastolic (p<0.001) blood pressure, left ventricular mass index (p<0.05) and aldosterone/renin ratio (p<0.001) fell. Serum creatinine independently predicted hypertension cure. From 2007 through 2008, QOL was evaluated prospectively using the internationally validated SF-36 questionnaire before and 3 and 6 months post-operatively in 22 patients [14 males, 8 females; mean age 50.0 ± 2.0 (range 27-69) years] with unilateral PAL who underwent adrenalectomy within the Endocrine Hypertension Centre, Greenslopes and Princess Alexandra Hospitals. Pre-operatively, the SF36 score for each QOL domain was lower for PAL patients than reported for the Australian general population, significantly so for physical functioning (p<0.05), role physical (p<0.001), vitality (p<0.001) and general health (p<0.05). Compared with pre-adrenalectomy, there were significant increments in mean scores at 3 months for physical functioning (p<0.05), role physical (p<0.05), general health (p<0.001), role emotional (p<0.05), social functioning (p<0.05), mental health (p<0.001) and vitality (p<0.001); and at 6 months for physical functioning (p<0.05), role physical (p<0.05), general health (p<0.05), role emotional (p<0.05), mental health (p<0.05) and vitality (p<0.001). Mean SBP and DBP improved significantly (p<0.001), with 86% of these patients cured (BP≤140/90, no drugs) and the remaining 14 % improved. Mean plasma potassium (p<0.001) and renin concentration rose (p<0.01), whereas mean upright plasma aldosterone (p<0.001), aldosterone/renin ratio (p<0.001) and number of antihypertensive agents fell (p<0.001). Conclusion: In the Italian families with FH-II available for study, work by the candidate included in this thesis confirmed linkage of FH-II to chromosome 7p22. The combined multipoint LOD score of 5.22 for the five families showing linkage at 7p22 was highly significant. Linkage to 7p22 in Italian families with FH-II extends the previous positive findings to a third geographical area, bringing greater certainty regarding the importance of this locus in identifying causative mutations. Although no clear causative mutations were found in the three 7p22 candidate genes examined, it is conceivable that the rs2024374 G/C and/or rs17169194 T/G SNPs in RPA3 could act in conjunction with another 7p22 mutation in family 1, resulting in the FH-II phenotype. Examination of the outcome of unilateral adrenalectomy in patients with bilateral PAL suggests that this surgical approach can be beneficial in certain carefully selected patients and should not be automatically excluded as a treatment option. Unilateral adrenalectomy in patients with unilateral PAL has positive impacts not only on clinical and biochemical parameters but also on QOL. The findings of this thesis provide new insights into the genetic basis and therapeutic options and treatment outcomes of PAL and further highlight its importance as a common, genetically based, specifically treatable and potentially curable cause of hypertension and cardiovascular disease. It also points the way to potentially very useful studies in future by exploring longer term effects of unilateral laparoscopic adrenalectomy as treatment for PAL on QOL, to compare unilateral adrenalectomy in those with unilateral versus bilateral PAL, and to compare surgery with specific medical treatment.

Introdução: As fissuras de lábio e/ou palato (FL/P) são malformações congênitas comuns na espécie humana, apresentando prevalência de 1/700 recém nascidos vivos, variando de acordo com os diferentes grupos étnicos e fatores sócio-econômicos. As FL/P apresentam padrão complexo de herança, estando envolvidos fatores genéticos e ambientais. Entre os fatores ambientais deficiências de vitaminas já foram descritas, e diversos estudos sugerem que o uso de ácido fólico periconcepcional pode prevenir a recorrência das fissuras orais. Objetivos: Estudar características bioquímicas e polimorfismos em genes da rota metabólica do folato em FL/P não sindrômicas (NS). Métodos: Foram incluídas 140 mulheres (113 mães não afetadas de crianças com fissuras e 27 mulheres afetadas). Todas as mulheres realizaram dosagens bioquímicas (B12, folato sérico e eritrocitário, hematócrito, hemoglobina e homocisteína). Foi realizada extração de DNA destas mulheres e seus familiares, assim como de trios adicionais (mãe, pai e filho) num total de 428 indivíduos de 231 famílias. 28 polimorfismos de 14 genes da rota metabólica de folato foram genotipados usando TaqMan (Applied Biosystems) ou reação em cadeia de polimerase (PCR). Resultados: Não foi encontrada associação entre os dados bioquímicos nos dois grupos de mulheres (afetadas e não afetadas). O teste de desequilíbrio de transmissão (TDT) revelou significância para os seguintes polimorfismos nos genes BHMTrs651852 (p=0.04), MTRRrs1532268 (p=0.04) e NNMTrs694539 (p=0.03). A interação gene-gene demonstrou significância entre MTRRrs1532268 versus MTRrs10925235 (p=0.03), MTRRrs1532268 versus MTRRrs1801394 (p=0.003), MTRRrs1532268 versus NNMTrs2852447 (p=0.008), NNMTrs694539 versus DHFRrs1643638 (p=<0.0001), NNMTrs694539 versus SHMT1rs921986 (p=0.0001), NNMTrs694539 versus SHMT1rs2168781 (p=0.03). Conclusão: Polimorfismos em genes envolvendo o metabolismo do ácido fólico podem contribuir para a ocorrência de FL/P. Os genes BHMT, MTRR e NNMT mostraram associação com FL/P. Este estudo foi o primeiro a encontrar associação entre o gene NNMT e fissuras orais. Estes achados, portando, devem ser confirmados por estudos adicionais. Estes dados são importantes para o entendimento dos fatores que predispõem às FL/P, e para ser realizada de maneira mais adequada e individualizada a prevenção desta anomalia congênita com ácido fólico. / Introduction: Cleft lip and/or palate (CL/P) are common congenital anomalies with prevalence of 1/700 live births affecting different ethnic groups and social economic status. CL/P has a complex inheritance involving environmental and genetic factors. Among the environmental factors, deficiency of vitamins were reported and several studies have suggested that the use of periconcepcional folic acid might prevent oral clefts. Objective: The aim of this study is to evaluate the biochemical and polymorphisms in genes of the folic acid pathway in non-syndromic CL/P. Methods: 140 women were included, (113) unaffected mothers with CL/P children and 27 affected women. In all women a biochemical measurement (B12, serum folate and eritrocyte, hematocrit, hemoglobin and homocysteine) was performed. We also had DNA extraction of these women and their families, as well as additional trio of (mother, father and son) in a total of 428 individuals of 231 unrelated families. 28 polymorphisms of 14 genes of the folate pathway were genotyped using the TaqMan (Applied Biosystem) or Polymerase Chain Reaction (PCR). Results: Among the biochemical data in the two groups of women (affected and unaffected with cleft lip children) no association was found. The transmission desequilibrium test (TDT) has showed significance for the following polymorphisms in the genes such as BHMTrs651852 (p=0.04), MTRRrs1532268 (p=0.04) and NNMTrs694539 (p=0.03). The gene-gene interaction has showed significance between MTRRrs1532268 versus MTRrs10925235 (p=0.03), MTRRrs1532268 versus NNMTrs2852447 (p=0.008), NNMTrs694539 versus DHFRrs1643638 (p=<0.0001), NNMTrs694539 versus SHMT1rs921986 (p=0.0001), NNMTrs694539 versus SHMT1rs2168781 (p=0.03). Conclusion: Polymorphisms in genes involving the folic acid metabolism might contribute to the occurrence of CL/P. The genes BHMT, MTRR and NNMT have showed association with CL/P. This was the first study to find association between the NNMT and oral clefts. Thus, additional studies are important to these results. These data are important to understand the causes of CL/P as well as to prevent this congenital anomaly with folic acid.

Milk fat is the recipient of numerous functions. It affects the biological, chemical, sensory and technological properties of milk. The Czech Republic is consumed and processed, especially cows, goats and ewes milk. The aim was to focus on the description of selected fatty acids in milk fat of dairy cows and goats because of their importance to human health and the factors that influence their composition. The results show that the content of short-chain fatty acids was compared to dairy cows higher in goat breeds. It was also found that the proportion of saturated fatty acids was influenced mainly breed and individuality, while the composition of unsaturated fatty acids was affected more nutrition and feeding. The work describes the biological factors such as breed, individuality, parity and stage of lactation, of the external factors described nutrition and feeding. Chapter biological factors mainly deals with the influence of genetics and contains a description of selected candidate genes affecting milk fat and fatty acids.

Made available in DSpace on 2016-06-02T20:21:37Z (GMT). No. of bitstreams: 1 6501.pdf: 1672444 bytes, checksum: 64754c3f12e26620a22bf55af9f8d5ff (MD5) Previous issue date: 2014-08-13 / The Brazilian cattle industry is presented as highlighted on the world stage and the significant participation of this productive sector in the economy means that there is concern with production losses, among which stands out those caused by infestation of Rhipicephalus (Boophilus) microplus, main ectoparasite vector cattle and various diseases. The genetic variability for resistance to the cattle tick shows that this trait can be genetically improved. For the execution of this work, it was used a study of genome wide association (GWAS) for resistance to Rhipicephalus (Boophilus) microplus, performed by Dr. Fernando Flores Cardoso, with 260 Hereford and 500 Braford animals. The monitoring of the infestation was accomplished by counting tick females larger than 4.5 mm from one of the animal's body side, and the degree of infestation was evaluated for each animal by averaging at least two consecutive counts, with intervals of approximately thirty days, in the months of highest incidence of the parasite. The animals were genotyped using a 50K SNP chip, and it was found a total of 37,346 SNPs that passed in quality test. Among these markers, 178 showed significant effects and allowed the mining of 175 genes in these regions, at an interval of 200 Kb (100 Kb for each side of each marker). Most of these polymorphisms associated with the trait is located in regions without defined functions (intronic and intergenic), and only one of them is located in the splicing region. The most significant regions of the GWAS were identified on chromosomes 7, 21 and 23, which were found 72 genes in linkage disequilibrium with the molecular markers. Therefore, a functional annotation of the genes on these 3 chromosomes was performed, allowing the choice of 11 candidate genes for the study of various metabolic pathways in which they are inserted. Among these pathways, the most important are those related to immune responses, secretion and intracellular transport, calcium influx and epidermal growth and differentiation. / A bovinocultura brasileira apresenta-se como destaque no cenário mundial e a expressiva participação deste setor produtivo na economia faz com que haja preocupação com as perdas produtivas, dentre as quais destaca-se aquelas causadas pela infestação do carrapato Rhipicephalus (Boophilus) microplus, principal ectoparasita de bovinos e vetor de diversas doenças. A variabilidade genética observada para a resistência dos bovinos ao carrapato permite que essa característica seja melhorada geneticamente, como forma alternativa de controle desses ectoparasitos. Para a execução do presente trabalho, foi utilizado um estudo de associação genômica ampla (GWAS) para a resistência ao carrapato R. microplus, o qual foi realizado pela equipe do Dr. Fernando Flores Cardoso (Embrapa Pecuária Sul), com 260 animais da raça Hereford e 500 animais da raça Braford. O monitoramento das infestações foi realizado por meio da contagem de fêmeas do carrapato com tamanho superior a 4,5 mm em um dos lados do corpo do animal, e o grau de infestação de cada animal foi avaliado pela média de pelo menos duas contagens consecutivas, com intervalos de aproximadamente trinta dias, conduzidas no sobreano, nos meses de maior incidência do parasito. Os animais foram genotipados com utilização de um chip de SNPs de 50 K e, após a realização do GWAS, verificou-se que um total de 37.346 SNPs passou nos teste de qualidade. Dentre esses marcadores, 178 SNPs apresentaram efeitos significativos e permitiram a mineração de 175 genes nessas regiões, em um intervalo de 200 Kb (100 Kb para cada lado de cada marcador). A maioria dos polimorfismos associados com a característica está localizada em regiões sem funções determinadas (intergênicas e intrônicas), apenas um deles encontra-se em região de splicing. Sendo assim, estes marcadores podem constituir mutações não causais que se encontram em desequilíbrio de ligação com mutações funcionais. As regiões mais significativas do GWAS foram identificadas nos cromossomos 7, 21 e 23, onde foram identificados 72 genes em desequilíbrio de ligação com os marcadores moleculares. Portanto, foi realizada uma anotação funcional dos genes localizados nesses 3 cromossomos, o que permitiu a seleção de 11 genes candidatos para um estudo mais aprofundado das vias metabólicas nas quais eles estão inseridos. Verificou-se que esses genes participam de processos importantes em vias já relacionadas com a resistência a carrapatos, tais como apresentação de antígenos, transporte e secreção intracelular e diferenciação da epiderme.

Introdução: As fissuras de lábio e/ou palato (FL/P) são malformações congênitas comuns na espécie humana, apresentando prevalência de 1/700 recém nascidos vivos, variando de acordo com os diferentes grupos étnicos e fatores sócio-econômicos. As FL/P apresentam padrão complexo de herança, estando envolvidos fatores genéticos e ambientais. Entre os fatores ambientais deficiências de vitaminas já foram descritas, e diversos estudos sugerem que o uso de ácido fólico periconcepcional pode prevenir a recorrência das fissuras orais. Objetivos: Estudar características bioquímicas e polimorfismos em genes da rota metabólica do folato em FL/P não sindrômicas (NS). Métodos: Foram incluídas 140 mulheres (113 mães não afetadas de crianças com fissuras e 27 mulheres afetadas). Todas as mulheres realizaram dosagens bioquímicas (B12, folato sérico e eritrocitário, hematócrito, hemoglobina e homocisteína). Foi realizada extração de DNA destas mulheres e seus familiares, assim como de trios adicionais (mãe, pai e filho) num total de 428 indivíduos de 231 famílias. 28 polimorfismos de 14 genes da rota metabólica de folato foram genotipados usando TaqMan (Applied Biosystems) ou reação em cadeia de polimerase (PCR). Resultados: Não foi encontrada associação entre os dados bioquímicos nos dois grupos de mulheres (afetadas e não afetadas). O teste de desequilíbrio de transmissão (TDT) revelou significância para os seguintes polimorfismos nos genes BHMTrs651852 (p=0.04), MTRRrs1532268 (p=0.04) e NNMTrs694539 (p=0.03). A interação gene-gene demonstrou significância entre MTRRrs1532268 versus MTRrs10925235 (p=0.03), MTRRrs1532268 versus MTRRrs1801394 (p=0.003), MTRRrs1532268 versus NNMTrs2852447 (p=0.008), NNMTrs694539 versus DHFRrs1643638 (p=<0.0001), NNMTrs694539 versus SHMT1rs921986 (p=0.0001), NNMTrs694539 versus SHMT1rs2168781 (p=0.03). Conclusão: Polimorfismos em genes envolvendo o metabolismo do ácido fólico podem contribuir para a ocorrência de FL/P. Os genes BHMT, MTRR e NNMT mostraram associação com FL/P. Este estudo foi o primeiro a encontrar associação entre o gene NNMT e fissuras orais. Estes achados, portando, devem ser confirmados por estudos adicionais. Estes dados são importantes para o entendimento dos fatores que predispõem às FL/P, e para ser realizada de maneira mais adequada e individualizada a prevenção desta anomalia congênita com ácido fólico. / Introduction: Cleft lip and/or palate (CL/P) are common congenital anomalies with prevalence of 1/700 live births affecting different ethnic groups and social economic status. CL/P has a complex inheritance involving environmental and genetic factors. Among the environmental factors, deficiency of vitamins were reported and several studies have suggested that the use of periconcepcional folic acid might prevent oral clefts. Objective: The aim of this study is to evaluate the biochemical and polymorphisms in genes of the folic acid pathway in non-syndromic CL/P. Methods: 140 women were included, (113) unaffected mothers with CL/P children and 27 affected women. In all women a biochemical measurement (B12, serum folate and eritrocyte, hematocrit, hemoglobin and homocysteine) was performed. We also had DNA extraction of these women and their families, as well as additional trio of (mother, father and son) in a total of 428 individuals of 231 unrelated families. 28 polymorphisms of 14 genes of the folate pathway were genotyped using the TaqMan (Applied Biosystem) or Polymerase Chain Reaction (PCR). Results: Among the biochemical data in the two groups of women (affected and unaffected with cleft lip children) no association was found. The transmission desequilibrium test (TDT) has showed significance for the following polymorphisms in the genes such as BHMTrs651852 (p=0.04), MTRRrs1532268 (p=0.04) and NNMTrs694539 (p=0.03). The gene-gene interaction has showed significance between MTRRrs1532268 versus MTRrs10925235 (p=0.03), MTRRrs1532268 versus NNMTrs2852447 (p=0.008), NNMTrs694539 versus DHFRrs1643638 (p=<0.0001), NNMTrs694539 versus SHMT1rs921986 (p=0.0001), NNMTrs694539 versus SHMT1rs2168781 (p=0.03). Conclusion: Polymorphisms in genes involving the folic acid metabolism might contribute to the occurrence of CL/P. The genes BHMT, MTRR and NNMT have showed association with CL/P. This was the first study to find association between the NNMT and oral clefts. Thus, additional studies are important to these results. These data are important to understand the causes of CL/P as well as to prevent this congenital anomaly with folic acid.