Submitted by Aline Batista (alinehb.ufpel@gmail.com) on 2017-06-08T15:25:06Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação - Fernanda Peres Lopes.pdf: 3142979 bytes, checksum: 5720a00dae8803fe8625b52b9c76c001 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-06-09T14:37:22Z (GMT) No. of bitstreams: 2 Dissertação - Fernanda Peres Lopes.pdf: 3142979 bytes, checksum: 5720a00dae8803fe8625b52b9c76c001 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-06-09T14:37:29Z (GMT). No. of bitstreams: 2 Dissertação - Fernanda Peres Lopes.pdf: 3142979 bytes, checksum: 5720a00dae8803fe8625b52b9c76c001 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-02-24 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Este estudo tem por objetivo analisar o processo variável de apagamento das vogais átonas [a, i, u] em posição final na fala de indivíduos da cidade de Pelotas (RS), conforme verificado em tarif[a] ~ tarif∅, equip[e] ~ equip∅, serviço ~ serviç∅, por exemplo. Para isso, partiu-se dos pressupostos da Fonologia de Uso (BYBEE, 2001, 2006, 2010), da Teoria de Exemplares (PIERREHUMBERT, 2001, 2003) e da Sociofonética (THOMAS, 2011; FOULKES; SCOBBIE; WATT, 2010). A amostra sob análise é constituída por oito informantes (quatro homens e quatro mulheres) entre 18 e 50 anos de idade e de dois níveis de escolaridade – sujeitos com até seis anos de escolaridade e sujeitos com, no mínimo, nove anos de escolaridade. A taxa de aplicação do apagamento na amostra analisada foi de 53% (N = 242) para a vogal [i], de 41% (N = 196) para a vogal [u] e de 0,8% (N = 4) para a vogal [a]. Os resultados indicaram que fatores como tipo de vogal, contexto precedente, ordem de produção e frequência lexical favorecem o apagamento, revelando a natureza predominantemente linguística do fenômeno. A única variável extralinguística que teve influência no apagamento foi a variável indivíduo. A análise acústica revelou que as vogais postônicas [i] e [u] produzidas pelos pelotenses tendem a abaixar enquanto a vogal [a] tende a elevar-se. Além disso, percebe-se uma centralização de [u], que ocupa, entre os homens, quase o mesmo espaço acústico que a vogal [a]. [ɐ], [ɪ] e [ʊ] são as vogais que representam o sistema encontrado na amostra. Com relação à duração, comparando-se os valores encontrados com os dados de Quintanilha-Azevedo (2016), percebe-se que tanto homens quanto mulheres produziram vogais mais curtas. Por fim, conclui-se que o apagamento representa o ponto final de uma trajetória que se inicia com a realização plena da vogal, passa pela redução de sua duração e pelo seu desvozeamento. / This thesis aims to analyze the variable process of deletion of the final unstressed vowels [a, i, u] by Brazilian Portuguese native speakers from the city of Pelotas, in the Southern state of Rio Grande do Sul, as noticed in words such as tarif[a] ~ tarif∅, equip[e] ~ equip∅, serviço ~ serviç∅, for instance. In order to achieve said aim, the concepts addressed by Usage-Based Phonology (BYBEE, 2001, 2006, 2010), the Exemplar Theory (PIERREHUMBERT, 2001, 2003) and Sociophonetics (THOMAS, 2011; FOULKES; SCOBBIE; WATT, 2010) were used as this research project's theoretical bases. The sample analyzed in this study consists of 8 informants (4 men and 4 women) from two different educational backgrounds (one group including subjects with up to 6 years of formal learning and the other including subjects with 9+ years of formal learning) with ages ranging from eighteen to fifty. The rate of deletion in the sample was of 53% (N = 242) for the vowel [i], of 41% (N = 196) for the vowel [u], and of 0,8% (N = 4) for the vowel [a]. The results indicate that such factors as type of vowel, preceding context, vowel production order and lexical frequency favor deletion, revealing the predominantly linguistic nature of the phenomenon. The variable "subject" was the only extralinguistic variable to influence deletion. Acoustic analysis showed that the posttonic vowels [i] and [u] produced by the subjects tend to lower as the vowel [a] tends to rise. In addition, the study also revealed a centralization of [u], which occupies among men almost the same acoustic vowel space as does the vowel [a]. Vowels [ɐ], [ɪ] and [ʊ] are representative of the system found in the sample. Regarding duration, when comparing the values included in this research project with those presented by Quintanilha-Azevedo (2016), it can be noticed that both men and women produced shorter vowels. Lastly, it was found that deletion represents the final stage of a process that begins with the production of a full-quality vowel, moves on to its reduction, and ultimately reaches the devoicing of said vowel.

Objetivos: Investigar a ocorrência de queixa respiratória nos pacientes com sinais clínicos da Síndrome Velocardiofacial (SVCF) submetidos à cirurgia para a correção da Disfunção Velofaríngea (DVF), comparativamente àqueles com fissura isolada de palato sem sinais da SVCF, além de verificar se a condição respiratória pré-cirúrgica interfere na escolha do tratamento cirúrgico para correção da DVF. Material e Método: Estudo retrospectivo e prospectivo com 30 indivíduos de ambos os sexos, que realizaram procedimento cirúrgico para a correção da DVF (veloplastia intravelar ou retalho faríngeo), sendo 15 com sinais clínicos da SVCF (grupo estudo) e 15 com fissura isolada de palato sem sinais clínicos da SVCF, pareado por sexo e idade (grupo controle). Um levantamento de sintomas respiratórias foi realizado utilizando-se três questionários, um aplicado antes e após a cirurgia (Caouette-Laberge et al 1992) e dois aplicados apenas após a cirurgia (Petry et al 2008, Berlin, proposto por Netzer et al 1999). As comparações foram realizadas por meio do Teste Exato de Fisher, considerando-se nível de significância de 5%. Resultados: Sintomas como respiração oral e ronco estavam presentes nos períodos pré e pós-cirúrgico em ambos os grupos estudados, não sendo detectada diferença entre esses períodos quanto à presença desses sintomas, nos dois grupos. Diferença entre os grupos não foram constatadas em relação à presença de sonolência diurna excessiva, ronco, apneia, sono/fadiga, histórico de obesidade ou hipertensão arterial e risco potencial para SAOS. No grupo estudo houve semelhante distribuição entre a realização de veloplastia intravelar e retalho faríngeo, diferentemente do grupo controle em que prevaleceu a veloplastia intravelar, não havendo relação entre a frequência de sintomas respiratórios e o tipo de cirurgia, para ambos os grupos. Conclusão: Nos pacientes com sinais clínicos da SVCF, sintomas de respiração oral e ronco estão presentes antes e após a correção cirúrgica da DVF; não há diferença quanto aos sintomas respiratórios entre indivíduos com sinais clínicos da SVCF e indivíduos com fissura isolada de palato sem sinais da SVCF; a condição respiratória antes da cirurgia não interferiu na escolha do tipo de procedimento cirúrgico para correção da DVF. / Objectives: To investigate the occurrence of respiratory complaint in individuals with clinical signs of Velocardiofacial Syndrome (VCFS) submitted to surgery for correction of Velopharyngeal Dysfunction (VPD), compared to individuals without signs of VCFS, and analyzed if the preoperative respiratory condition interferes with surgical treatment planning for correction of VPD. Material and method: Retrospective and prospective study of 30 individuals of both genders, who were submitted to surgery for correction of VPD (intravelar veloplasty or pharyngeal flap), being 15 with clinical signs of VCFS (study group) and 15 with isolated cleft palate without clinical signs of VCFS, matched for gender and age (control group). A survey of respiratory complaints was performed using three questionnaires, one applied before and after surgery (Caouette-Laberge et al 1992) and two applied only after surgery (Petry et al 2008, Berlin, proposed by Netzer et al 1999). Comparisons were performed by the exact Fisher test, at a significance level of 5%. Results: Symptoms as mouth breathing and snoring were present in pre- and postoperative periods in both groups, without difference between periods concerning the presence of these symptoms, in the two groups. No differences were observed between groups as to the presence of excessive somnolence during the day, snoring, apnea, sleep/fatigue, history of obesity or arterial hypertension and potential risk to OSA. The study group presented similar distribution of intravelar veloplasty and pharyngeal flap, different from the control group that presented predominance of intravelar veloplasty, without relationship between the frequency of respiratory symptons and type of surgery, for both groups. Conclusion: In patients with clinical signs of VCFS, complaints of mouth breathing and snoring are present before and after surgical correction of VPD; there is no difference in the complaints of respiratory symptoms between individuals with clinical signs of VCFS and individuals with isolated cleft palate without signs of VCFS; the respiratory condition before surgery did not interfere with selection of the type of surgical procedure for correction of VPD.

INTRODUÇÃO: Os defeitos cardíacos congênitos (DCC) são um grupo de anormalidades estruturais mais prevalentes ao nascimento e uma das principais causas de morbidade e mortalidade infantil. Os fatores genéticos são importantes na etiologia dos DCC. Estudos têm mostrado a contribuição da variação no número de cópias (CNV) na gênese das malformações cardíacas. A deleção 22q11.2 é a causa mais comum de microdeleção humana e está relacionada com defeito cardíaco (DCC) conotruncal. O MLPA (Multiplex Ligation-dependent Probe Amplification) é um método eficaz para detectar microdeleções/microduplicações em pacientes com DCC. OBJETIVO: Detectar a presença da variação no número de cópias gênicas em pacientes portadores de cardiopatia conotruncal pela técnica de MLPA e associar ao fenótipo do paciente. MÉTODOS: Foram avaliados 39 pacientes (23 do sexo masculino, 16 do sexo feminino) com idade entre 2 dias e 19 anos (mediana de 6 anos), todos com cardiopatia conotruncal, a maioria dos pacientes (56%) apresentavam tetralogia de Fallot. Avaliação clínica e laboratorial foi realizada em todos os pacientes. O cariótipo foi normal em todos pacientes. MLPA foi realizada com os kits P064, P036/P070 e P250. RESULTADOS: Foram detectadas CNVs em sete pacientes: deleção 22q11.2, duplicação 22q11.2, duplicação 15q11.2, duplicação 20p12.2, deleção 19p, duplicação 15q e duplicação 8p23.2 com duplicação 10p12.31. As cardiopatias encontradas nestes pacientes foram: dupla via de saída de ventrículo direito (2), coartação da aorta, tetralogia de Fallot (3) e transposição de grandes artérias. Os achados clínicos extracardíacos encontrados nestes pacientes foram dismorfismo facial, dente neonatal, atrofia e displasia cerebral, atresia duodenal, dificuldade de aprendizado, insuficiência velofaríngea, aplasia de timo, refluxo gastroesofágico, hérnia umbilical, asma, infecções de vias aéreas frequente, déficit de crescimento e somente três apresentavam retardo no desenvolvimento neuropsicomotor (dup 15q11.2, dup 15q, del 22q11.2). As características clínicas foram compatíveis com o relatado na literatura associado com a microdeleção/microduplicação encontrada. Nenhuma destas alterações foram herdadas de seus pais testados em seis casos. CONCLUSÃO: O uso do MLPA possibilitou a detecção de CNVs em pacientes com DCC. O diagnóstico precoce das CNVs em pacientes com DCC auxilia na prevenção de morbidade e diminuição da mortalidade nestes pacientes, contudo em um país com regiões com poucos recursos laboratoriais genéticos uma avaliação clínica minuciosa em todo paciente com DCC é imprescindível para direcionar qual melhor exame deve ser realizado / INTRODUCTION: Congenital heart defects (CHD) are a group of structural abnormalities most prevalent birth and a major cause of infant morbidity and mortality. Genetic factors are important in the etiology of CHD. Studies have shown the contribution of copy number variation (CNV) in the genesis of cardiac malformations. The deleletion 22q11.2 is the most common cause of human microdeletion and is related conotruncal cardiac defect (DCC). The MLPA (Multiplex Ligation-dependent Probe Amplification) is an effective method to detect microdeletions/micoduplications in patients with CHD. PURPOSE: Detect the presence of gene copies number variation in the patients with conotruncal heart defect by MLPA technique and associate the phenotype of the patient. METHODS: 39 patients (23 males, 16 females) aged 2 days old - 19 years old (median= 6 years old) with conotruncal cardiac defect were evaluated. Tetralogy of Fallot was more prevalent heart defect (56%). All patients were evaluated clinical and laboratory. Karyotypes were normal in all pacients. MLPA was performed with the P064, P036/P070 and P250 kits. RESULTS: CNVs were detected in seven patients: 22q11.2 deletion, 22q11.2 duplication, 15q11.2 duplication, 20p12.2 duplication, 19p deletion, 15q duplication and 8p23.2 duplication with 10p12.31 duplication. The congenital heart defect found in these patients were: double outlet right ventricle (2), coarctation of the aorta, tetralogy of Fallot (3) and transposition of the great arteries. Clinical findings in these patients were facial dysmorphism, neonatal tooth, brain atrophy and dysplasia, duodenal atresia, learning disabilities, velopharyngeal insufficiency, thymic aplasia, gastroesophageal reflux, umbilical hernia, asthma, frequent infections of the airways , failure to thrive, and only three had delayed psychomotor development (dup 15q 11.2, dup 15q, del 22q11.2) The clinical features were consistent with those reported in the literature associated with the microdeletion /microduplication found. None of these alterations were inherited from six parents tested. CONCLUSIONS: MLPA was effective to detect CNVs in patients with CHD. Early diagnosis of CNVs in patients with CHD assists in preventing morbidity and decreased mortality in these patients, however, in a country with regions with few genetic laboratory resources a thorough clinical evaluation in all patients with CHD is essential to direct which should be further analyzed performed

L'hévéa (Hevea brasiliensis) est la seule source de caoutchouc naturel commercialisé à travers le monde. En Afrique et en Asie, la maladie 'Corynespora Leaf Fall' (CLF), causée par le champignon nécrotrophe Corynespora cassiicola, affecte les plantations hévéicoles en provoquant des défoliations massives sur les clones les plus sensibles. L’évaluation précoce de la sensibilité des clones dans les programmes de sélection est un enjeu majeur pour écarter les individus les plus sensibles des programmes de développement et ainsi réduire la pression de la maladie. Une des méthodes d’évaluation envisagée consiste à tester indirectement la sensibilité des clones aux effecteurs fongiques responsables de la virulence (test toxinique). Parmi tous les effecteurs potentiels du champignon identifiés in silico, seule la cassiicoline Cas1 a été purifiée et caractérisée à ce jour. Il s’agit d’une petite glycoprotéine sécrétée qui jouerait un rôle dans les phases précoces de l’infection en induisant la nécrose des tissus. Les souches porteuses du gène Cas1 sont parmi les plus agressives sur les clones d'hévéa testés. Néanmoins, certaines souches de C. cassiicola ne produisant pas de cassiicoline présentent tout de même une agressivité modérée, suggérant l'implication d'autres effecteurs dans l'établissement de la maladie CLF. Les objectifs de cette thèse sont (i) de déterminer si la sensibilité à la cassiicoline Cas1 est un critère de sélection pertinent pour identifier les clones d’hévéa les plus sensibles à la maladie CLF, et (ii) d’identifier chez l’hévéa des facteurs de sensibilité à la cassiicoline Cas1. Nous avons d'abord analysé l’inoculum naturel et montré que les souches porteuses du gène Cas1 représentent un quart de la population de C. cassiicola dans les plantations d’hévéa d’Afrique de l’Ouest, le reste étant majoritairement constitué de souches dépourvues du gène codant la cassiicoline (Type A/Cas0). Nous avons ensuite créé un mutant de délétion du gène Cas1 pour la souche de référence CCP et comparé sa virulence à celle de la souche sauvage. Nous avons ainsi pu montrer que la cassiicoline Cas1 est bien un effecteur de nécrotrophie déterminant pour la virulence de C. cassiicola chez l’hévéa puisque la souche délétée perd toute virulence sur les clones testés. Enfin, nous avons recherché chez l'hévéa des facteurs de sensibilité à la cassiicoline Cas1 à travers deux approches. La technique de "double hybride en levures" nous a permis d'identifier une trentaine de protéines candidates qui pourraient interagir physiquement avec la toxine. Une approche transcriptomique, nous a permis d’identifier les gènes d’hévéa dont l’expression est modifiée suite à l’application de la cassiicoline purifiée, en comparant un clone sensible (PB260) et un clone tolérant (RRIM600). En conclusion, ces travaux ont permis de mieux comprendre les mécanismes impliqués dans l'interaction compatible entre C. cassiicola et l'hévéa et ouvrent la voie de la sélection assistée par effecteur. / The rubber tree (Hevea brasiliensis) is the primary commercial source of natural rubber worldwide. In Asia and Africa, H. brasiliensis is affected by the Corynespora leaf fall (CLF) disease, caused by the broad-spectrum necrotrophic fungus Corynespora cassiicola. During severe attacks, massive fall of young leaves can occur in susceptible cultivars. Early evaluation of the susceptibility of rubber clones in breeding programs is required to avoid developing highly susceptible clones that would amplify the disease. An indirect phenotyping procedure envisaged consists in testing the sensitivity to the fungal toxins (or effectors) rather than the susceptibility to the fungus itself (toxin test). Among all putative effectors identified in silico, only cassiicolin Cas1 has been purified and characterized to date. This small secreted glycoprotein was for long suspected to play a role in the early phase of infection by inducing tissue necrosis. Strains carrying the Cas1 gene are the most aggressive on tested rubber clones. However, strains without cassiicolin gene (called Cas0) still show moderate aggressiveness, suggesting the existence of effectors other than cassiicolin. The objectives of this study are (i) to determine if susceptibility to cassiicolin Cas1 is a relevant selection criterion to eliminate the rubber clones most susceptible to CLF disease, and (ii) identify molecular factors involved in the sensitivity to Cas1, in rubber tree. We have thus analyzed the typology of a large set of C. cassiicola isolates collected from various rubber plantations in West Africa. Our results show that isolates carrying the cassiicolin isoform Cas1 are widely represented, but that the most represented type (A/Cas0) are isolates without cassiicolin gene. Here we show that deletion of the cassiicolin gene in the isolate CCP resulted in a total loss of virulence. This clearly demonstrated that cassiicolin is indeed a necrotrophic effector required for the virulence of isolate CCP in rubber tree. Finally, we have investigated susceptibility factors to cassiicolin Cas1 on rubber tree with two different approaches. We identified about thirty candidate proteins that could physically interact with the toxin, through the two-hybrid assay. A transcriptomic approach allowed us to identify the rubber genes differentially expressed in response to the purified cassiicolin, comparing a susceptible clone (PB260) and a tolerant clone (RRIM600). In conclusion, we think that the necrotrophic effector Cas1 can be an interesting tool for effector-based selection of tolerant clones for African plantations; however, efforts should also be placed on A/Cas0 isolates, in order to identify potential necrotrophic effector(s) responsible for their virulence. This would enlarge the potential of effector-based selection.

Protein-coding genes evolve together with their genome and acquire changes, some of which affect the length of their protein products. This explains why equivalent proteins from different species can exhibit length differences. Variation in length of proteins during evolution arguably presents a large number of possibilities for improvement and innovation of protein structure and function. In order to contribute to an increased understanding of this process, we have studied variation caused by tandem domain duplications and insertions or deletions of intrinsically disordered residues. The study of two proteins, Nebulin and Filamin, together with a broader study of long repeat proteins (&gt;10 domain repeats), began by confirming that tandem domains evolve by internal duplications. Next, we show that vertebrate Nebulins evolved by duplications of a seven-domain unit, yet the most recent duplications utilized different gene parts as duplication units. However, Filamin exhibits a checkered duplication pattern, indicating that duplications were followed by similarity erosions that were hindered at particular domains due to the presence of equivalent binding motifs. For long repeat proteins, we found that human segmental duplications are over-represented in long repeat genes. Additionally, domains that have formed long repeats achieved this primarily by duplications of two or more domains at a time. The study of homologous protein pairs from the well-characterized eukaryotes nematode, fruit fly and several fungi, demonstrated a link between variation in length and variation in the number of intrinsically disordered residues. Next, insertions and deletions (indels) estimated from HMM-HMM pairwise alignments showed that disordered residues are clearly more frequent among indel than non-indel residues. Additionally, a study of raw length differences showed that more than half of the variation in fungi proteins is composed of disordered residues. Finally, a model of indels and their immediate surroundings suggested that disordered indels occur in already disordered regions rather than in ordered regions. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: In press. Paper 4: Manuscript.</p>

Introduction: Amyotrophic lateral sclerosis (ALS) is a deadly, progressive neuromuscular disease that affects individuals all over the world. About 10% of the patients have a familial predisposition (FALS) while the remainder of cases are isolated or sporadic (SALS) and of unknown cause. To date, the principal recognized risk factors for ALS are higher age, male gender, slim figure (BMI&lt;23) and a family history of ALS. In 1993, Rosen et al. observed that some FALS cases were associated with mutations in the gene encoding the CuZn superoxide dismutase enzyme (SOD1). Since then, several mutations in the SOD1 gene have been discovered, and mutations in more than 18 other genes have been associated with causing ALS. The aim of this thesis was to identify new mutations associated with ALS pathogenesis, and by comparing patients from different countries, were we also able to identify population-specific genetic variations. The studies are referred to as I–V. Methods: With written informed consent and adhering to the tenets of the Declaration of Helsinki, through a national network of ALS clinicians´, venous blood samples were collected from ALS patients and healthy subjects in Europe and the USA. The patients were diagnosed according to the El Escorial criteria, and as having FALS according to the criteria of Byrne et al. (2011). The DNA variations were amplified by various PCR techniques. (I, III and IV) The amplicons of ataxin 2 (ATXN2), profilin 1 (PFN1), and vesicle-associated membrane protein type B (VAPB) were characterised by direct sequencing. (II) After quantitative PCR, a genotype-phenotype correlation was performed to assess whether the survival motor neuron gene (SMN) modulates the phenotype of ALS. (V) The amplicons of the 50 base pair deletion in the SOD1 promotor (50 bp) were separated by electrophoresis on agarose. Results: (I) We observed a significant association between CAG expansions in the ATXN2 gene and ALS in a European cohort. (II) Abnormal copy number of the SMN1 gene was identified as a risk factor in France, but not in Sweden. Homozygosity of the SMN2 deletion prolonged survival among Swedish ALS patients, compared to French patients. (III) We identified two mutations in the PFN1 gene, the novel p.Thr109Met mutation and the p.Gln117Gly mutation, in two unrelated FALS patients. (IV) In our cohort, we identified five VAPB mutations p.Asp130Glu, p.Ser160del, p.Asp162Glu, p.Met170Ile, and p.Arg184Trp, two of which are novel. (V) The 50 bp deletion upstream of the SOD1 gene was found in equal frequencies in both the patient and control cohorts. The 50 bp deletion did not affect SOD1 enzymatic activity. Furthermore, we found no differences in age of onset or disease duration in relation to the 50 bp deletion genotype.VI Conclusions: (I) Our findings indicate that ATXN2 plays an important role in the pathogenesis of ALS, and that CAG expansions in ATXN2 are a significant risk factor for the disease. (II) We suggest that abnormal SMN1 gene copynumber cannot be considered a universal genetic susceptibility factor for ALS. We also propose that the effect of abnormal SMN2 gene copy number on ALS phenotype may differ between populations. (III) This work provides evidence that PFN1 mutations can cause ALS as a Mendelian dominant trait. The novel p.Thr109Met mutation also shows that disturbance of actin dynamics can cause motor neuron degeneration. (IV) We find it unlikely that the VAPB mutations cause ALS in our cohorts. (V) We find it unlikely that the 50 bp region contains important regulatory elements for SOD1 expression. This thesis supports the theory that ALS is a multigenetic disease, but there appears to be great genetic variation among apparently identical populations. These studies emphasise the importance of continuous genetic screening, to identify further mutations and genes involved in ALS disease, but it also highlights the importance of cooperation and comparison between countries. / On the aetiology of ALS: A comprehensive genetic study

This thesis investigates the impact of fragmentation in the ownership of complementary patents or patent thickets on firms' market value. This question is motivated by the increase in the patent ownership fragmentation following the pro-patent shifts in the US since 1982. The first chapter uses panel data on patenting US manufacturing firms from 1979 to 1996, and estimates the impact of patent thickets on firms' market value. I find that patent thickets lower firms' market value, and firms with a large patent portfolio size experience a smaller negative effect from their thickets. Moreover, no systematic difference exists in the impact of patent thickets on firms' market value over time. The second chapter extends this analysis to account for the indirect impacts of patent thickets on firms' market value. These indirect effects arise through the effects of patent thickets on firms' R\&D and patenting activities. Using panel data on US manufacturing firms from 1979 to 1996, I estimate the impact of patent thickets on market value, R\&D, and patenting as well as the impacts of R\&D and patenting on market value. Employing these estimates, I determine the direct, indirect, and total impacts of patent thickets on market value. I find that patent thickets decrease firms' market value, while I hold the firms’ R\&D and patenting activities constant. I find no evidence of a change in R\&D due to patent thickets. However, there is evidence of defensive patenting (an increase in patenting attributed to thickets), which helps to reduce the direct negative impact of patent thickets on market value. The data sets used in Chapters 1 and 2 have a number of missing observations on regressors. The commonly used methods to manage missing observations are the listwise deletion (complete case) and the indicator methods. Studies on the statistical properties of these methods suggest a smaller bias using the listwise deletion method. Employing Monte Carlo simulations, Chapter 3 examines the properties of these methods, and finds that in some cases the listwise deletion estimates have larger biases than indicator estimates. This finding suggests that interpreting estimates arrived at with either approach requires caution.

ZUSAMMENFASSUNG Haukur Lindberg Sigmarsson PATHOGENITÄTSVERGLEICH VON SALMONELLA TYPHIMURIUM DT104 - WILDTYP UND SALMONELLA TYPHIMURIUM - DELETIONSMUTANTEN (sseD::aphT & invC::aphT) IN PERSISTENT INFIZIERTEN SCHWEINEN Salmonella (S.) Typhimurium DT104 ist ein gram-negatives Bakterium. Es weist keine Wirtsspezifität auf und gilt als Zoonoseerreger. Jährlich erkranken daran allein in Deutschland mehrere Tausend Menschen unter dem Bild einer schwerwiegenden Diarrhö mit zum Teil tödlichem Ausgang. Das Schwein gilt als eines der Reservoire für S. Typhimurium DT104 des Menschen. S. Typhimurium DT104 gelangt über vom Schwein stammende Produkte in den menschlichen Verzehr. Die Kontrolle von S. Typhimurium DT104 einschließlich effektiver Eradikationsmassnahmen in unseren Schweinebeständen ist deshalb von entscheidender Bedeutung, um den Eintrag dieses Bakteriums in die menschliche Nahrungskette wenn möglich zu eliminieren. Dafür ist das Verständnis über S. Typhimurium DT104 einschließlich der Kenntnis seine Pathogenitätseigenschaften notwendig. Ziel dieser Arbeit waren Untersuchungen zur Pathogenität von S. Typhimurium DT104. Dabei wurden der Wildstamm mit zwei seiner Deletionsmutanten (sseD::aphT und invC::aphT) verglichen. Die Untersuchungen erfolgten im Infektionsversuch an insgesamt 25 sechs Wochen alten männlichen Schweinen, die in einem vollklimatisierten Versuchsstall gehalten wurden. Den Tieren wurde im Anschluss an eine einwöchige Akklimatisierungsphase eines der nachfolgenden Stämme von S. Typhimurium DT104 oral in einer Konzentration von 1 x 1011 KBE verabreicht: Wildtyp (n = 8 Schweine), Deletionsmutante seeD::aphT (n = 8) und Deletionsmutante invC::aphT (n = 9). Bei den Mutanten handelt es sich um Varianten von S. Typhimurium DT104, die an den entsprechenden Abschnitten des Bakteriumgenoms (d.h. sseD-Gen bzw. invC-Gen) deletiert wurden. SseD regelt die Überlebensfähigkeit von S. Typhimurium in Makrophagen, invC dessen Invasionsvermögen. Im Mäusemodel war die Pathogenität beider Mutanten deutlich vermindert. Nach der Infektion schloss sich ein 20 tägiger Beobachtungszeitraum an, während dessen nachfolgend genannte Parameter erfasst bzw. Proben genommen wurden: klinische Symptome (Allgemeinbefinden, Erbrechen, Durchfall, Futteraufnahme, Atmung, Temperatur); Blutentnahme für Erstellung des weißen Blutbildes; Kotentnahme zum Nachweis der Ausscheidung von S. Typhimurium. Einen Tag nach Ende der Beobachtung wurden die Tiere getötet und Proben von insgesamt 15 Organen (unter anderem Tonsille; Colon und Caecum sowie dazugehörige Lymphknoten; Leber; Milz; Muskulatur) genommen. Kot sowie Gewebeproben wurden kulturell und wenn positiv auch mittels PCR untersucht. Alle mit dem Wildtyp infizierten Schweine wurden mehr oder weniger stark krank. Häufig zeigten erkrankte Schweine zeitgleich mehrere Krankheitssymptome (z. B. Erbrechen und Durchfall). Die Erkrankung hielt über mehrere Tage an. Im Vergleich dazu waren die Krankheitssymptome der Tiere, die mit Mutanten infiziert wurden, mild. Nur wenige Tiere erkrankten und dann auch nur kurzzeitig. Gewöhnlich war nur einer der erfassten Parameter verändert. Typische Veränderungen im weißen Blutbild waren nur bei Wildtyp-infizierten Tieren zu beobachten, während Tiere beider Mutanten kaum auf die Infektion reagierten. Alle 25 infizierten Tiere schieden S. Typhimurium mit dem Kot während der ersten Woche post inocculationem aus. Danach wurden in allen drei Gruppen etwa gleichviel intermittierende Ausscheider beobachtet. Zwischen 65 und 67 % der Gewebeproben der mit dem Wildtyp und mit der sseD::aphT-Mutante infizierten Tiere waren sowohl in der Kultur als auch mittels PCR S. Typhimurium positiv, während dieser Anteil nach Infektion mit invC::aphT nur 49 % betrug. Alle Tiere waren in Mandibularlymphknoten und im Colon positiv, während S. Typhimurium nur selten in Muskulatur und Leber nachzuweisen war. Die Ergebnisse dieser Arbeit bestätigen, dass Infektionen mit dem Wildtyp von S. Typhimurium zu einer schweren Erkrankung führen können. Gleichzeitig konnte gezeigt werden, dass beide in dieser Arbeit verwendeten Mutanten weniger krankmachend sind. Es muss davon ausgegangen werden, dass die Deletionen in den sseD bzw. invC-Bereichen tatsächlich zu Veränderungen bestimmter Eigenschaften geführt haben, die Teil der Pathogenitätsmechanismen für das Schwein sind. Im Unterschied zur Maus war sseD beim Schwein allerdings invasiv. Es kann vermutet werden, dass die durch sseD kodierten Pathogenitätseigenschaften von S. Typhimurium bei der Maus anders als beim Schwein wirken und somit unterschiedliche Bedeutung haben. Da die invC::aphT-Mutante jedoch und wie erwartet wesentlich schwächer als Wildtyp und sseD::aphT invadierte ist davon auszugehen, dass die Deletion im invC Bereich das Invasionsvermögen der Mutante beim Schwein ähnlich wie bei der Maus verringerte.

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This thesis investigates the impact of fragmentation in the ownership of complementary patents or patent thickets on firms' market value. This question is motivated by the increase in the patent ownership fragmentation following the pro-patent shifts in the US since 1982. The first chapter uses panel data on patenting US manufacturing firms from 1979 to 1996, and estimates the impact of patent thickets on firms' market value. I find that patent thickets lower firms' market value, and firms with a large patent portfolio size experience a smaller negative effect from their thickets. Moreover, no systematic difference exists in the impact of patent thickets on firms' market value over time. The second chapter extends this analysis to account for the indirect impacts of patent thickets on firms' market value. These indirect effects arise through the effects of patent thickets on firms' R\&D and patenting activities. Using panel data on US manufacturing firms from 1979 to 1996, I estimate the impact of patent thickets on market value, R\&D, and patenting as well as the impacts of R\&D and patenting on market value. Employing these estimates, I determine the direct, indirect, and total impacts of patent thickets on market value. I find that patent thickets decrease firms' market value, while I hold the firms’ R\&D and patenting activities constant. I find no evidence of a change in R\&D due to patent thickets. However, there is evidence of defensive patenting (an increase in patenting attributed to thickets), which helps to reduce the direct negative impact of patent thickets on market value. The data sets used in Chapters 1 and 2 have a number of missing observations on regressors. The commonly used methods to manage missing observations are the listwise deletion (complete case) and the indicator methods. Studies on the statistical properties of these methods suggest a smaller bias using the listwise deletion method. Employing Monte Carlo simulations, Chapter 3 examines the properties of these methods, and finds that in some cases the listwise deletion estimates have larger biases than indicator estimates. This finding suggests that interpreting estimates arrived at with either approach requires caution.