Hybridization and Population Structure of the Culex Pipiens Complex in the Islands of Macaronesia

The Culex pipiens complex includes two widespread mosquito vector species, Cx. pipiens and Cx. quinquefasciatus. The distribution of these species varies in latitude, with the former being present in temperate regions and the latter in tropical and subtropical regions. However, their distribution range overlaps in certain areas and interspecific hybridization has been documented. Genetic introgression between these species may have epidemiological repercussions for West Nile virus (WNV) transmission. Bayesian clustering analysis based on multilocus genotypes of 12 microsatellites was used to determine levels of hybridization between these two species in Macaronesian islands, the only contact zone described in West Africa. The distribution of the two species reflects both the islands’ biogeography and historical aspects of human colonization. Madeira Island displayed a homogenous population of Cx. pipiens, whereas Cape Verde showed a more intriguing scenario with extensive hybridization. In the islands of Brava and Santiago, only Cx. quinquefasciatus was found, while in Fogo and Maio high hybrid rates (~40%) between the two species were detected. Within the admixed populations, second-generation hybrids (~50%) were identified suggesting a lack of isolation mechanisms. The observed levels of hybridization may locally potentiate the transmission to humans of zoonotic arboviruses such as WNV.

Identification of naturally processed hepatitis C virus-derived major histocompatibility complex class I ligands.

Fine mapping of human cytotoxic T lymphocyte (CTL) responses against hepatitis C virus (HCV) is based on external loading of target cells with synthetic peptides which are either derived from prediction algorithms or from overlapping peptide libraries. These strategies do not address putative host and viral mechanisms which may alter processing as well as presentation of CTL epitopes. Therefore, the aim of this proof-of-concept study was to identify naturally processed HCV-derived major histocompatibility complex (MHC) class I ligands. To this end, continuous human cell lines were engineered to inducibly express HCV proteins and to constitutively express high levels of functional HLA-A2. These cell lines were recognized in an HLA-A2-restricted manner by HCV-specific CTLs. Ligands eluted from HLA-A2 molecules isolated from large-scale cultures of these cell lines were separated by high performance liquid chromatography and further analyzed by electrospray ionization quadrupole time of flight mass spectrometry (MS)/tandem MS. These analyses allowed the identification of two HLA-A2-restricted epitopes derived from HCV nonstructural proteins (NS) 3 and 5B (NS3₁₄₀₆₋₁₄₁₅ and NS5B₂₅₉₄₋₂₆₀₂). In conclusion, we describe a general strategy that may be useful to investigate HCV pathogenesis and may contribute to the development of preventive and therapeutic vaccines in the future.

Purification and ligand binding of a soluble class I major histocompatibility complex molecule consisting of the first three domains of H-2Kd fused to beta 2-microglobulin expressed in the baculovirus-insect cell system.

A recombinant baculovirus encoding a single-chain murine major histocompatibility complex class I molecule in which the first three domains of H-2Kd are fused to beta 2-microglobulin (beta 2-m) via a 15-amino acid linker has been isolated and used to infect lepidopteran cells. A soluble, 391-amino acid single-chain H-2Kd (SC-Kd) molecule of 48 kDa was synthesized and glycosylated in insect cells and could be purified in the absence of detergents by affinity chromatography using the anti-H-2Kd monoclonal antibody SF1.1.1.1. We tested the ability of SC-Kd to bind antigenic peptides using a direct binding assay based on photoaffinity labeling. The photoreactive derivative was prepared from the H-2Kd-restricted Plasmodium berghei circumsporozoite protein (P.b. CS) peptide 253-260 (YIPSAEKI), a probe that we had previously shown to be unable to bind to the H-2Kd heavy chain in infected cells in the absence of co-expressed beta 2-microglobulin. SC-Kd expressed in insect cells did not require additional mouse beta 2-m to bind the photoprobe, indicating that the covalently attached beta 2-m could substitute for the free molecule. Similarly, binding of the P.b. CS photoaffinity probe to the purified SC-Kd molecule was unaffected by the addition of exogenous beta 2-m. This is in contrast to H-2KdQ10, a soluble H-2Kd molecule in which beta 2-m is noncovalently bound to the soluble heavy chain, whose ability to bind the photoaffinity probe is greatly enhanced in the presence of an excess of exogenous beta 2-m. The binding of the probe to SC-Kd was allele-specific, since labeling was selectively inhibited only by antigenic peptides known to be presented by the H-2Kd molecule.

Increased mobility of major histocompatibility complex I-peptide complexes decreases the sensitivity of antigen recognition.

CD8(+) cytotoxic T lymphocytes (CTL) can recognize and kill target cells expressing only a few cognate major histocompatibility complex (MHC) I-peptide complexes. This high sensitivity requires efficient scanning of a vast number of highly diverse MHC I-peptide complexes by the T cell receptor in the contact site of transient conjugates formed mainly by nonspecific interactions of ICAM-1 and LFA-1. Tracking of single H-2K(d) molecules loaded with fluorescent peptides on target cells and nascent conjugates with CTL showed dynamic transitions between states of free diffusion and immobility. The immobilizations were explained by association of MHC I-peptide complexes with ICAM-1 and strongly increased their local concentration in cell adhesion sites and hence their scanning by T cell receptor. In nascent immunological synapses cognate complexes became immobile, whereas noncognate ones diffused out again. Interfering with this mobility modulation-based concentration and sorting of MHC I-peptide complexes strongly impaired the sensitivity of antigen recognition by CTL, demonstrating that it constitutes a new basic aspect of antigen presentation by MHC I molecules.

Impact and cost of a 2-week community-based screening and awareness program for diabetes and cardiovascular risk factors in a Swiss canton.

BACKGROUND: Community-based diabetes screening programs can help sensitize the population and identify new cases. However, the impact of such programs is rarely assessed in high-income countries, where concurrent health information and screening opportunities are common place. INTERVENTION AND METHODS: A 2-week screening and awareness campaign was organized as part of a new diabetes program in the canton of Vaud (population of 697,000) in Switzerland. Screening was performed without appointment in 190 out of 244 pharmacies in the canton at the subsidized cost of 10 Swiss Francs per participant. Screening included questions on risk behaviors, measurement of body mass index, blood pressure, blood cholesterol, random blood glucose (RBG), and A1c if RBG was >/=7.0 mmol/L. A mass media campaign promoting physical activity and a healthy diet was channeled through several media, eg, 165 spots on radio, billboards in 250 public places, flyers in 360 public transport vehicles, and a dozen articles in several newspapers. A telephone survey in a representative sample of the population of the canton was performed after the campaign to evaluate the program. RESULTS: A total of 4222 participants (0.76% of all persons aged >/=18 years) underwent the screening program (median age: 53 years, 63% females). Among participants not treated for diabetes, 3.7% had RBG >/= 7.8 mmol/L and 1.8% had both RBG >/= 7.0 mmol/L and A1c >/= 6.5. Untreated blood pressure >/=140/90 mmHg and/or untreated cholesterol >/=5.2 mmol/L were found in 50.5% of participants. One or several treated or untreated modifiable risk factors were found in 78% of participants. The telephone survey showed that 53% of all adults in the canton were sensitized by the campaign. Excluding fees paid by the participants, the program incurred a cost of CHF 330,600. CONCLUSION: A community-based screening program had low efficiency for detecting new cases of diabetes, but it identified large numbers of persons with elevated other cardiovascular risk factors. Our findings suggest the convenience of A1c for mass screening of diabetes, the usefulness of extending diabetes screening to other cardiovascular risk factors, and the importance of a robust background communication campaign.

Molecular modeling study of the binding and signaling properties of the TCRpMHC complex

The activation of the specific immune response against tumor cells is based on the recognition by the CD8+ Cytotoxic Τ Lymphocytes (CTL), of antigenic peptides (p) presented at the surface of the cell by the class I major histocompatibility complex (MHC). The ability of the so-called T-Cell Receptors (TCR) to discriminate between self and non-self peptides constitutes the most important specific control mechanism against infected cells. The TCR/pMHC interaction has been the subject of much attention in cancer therapy since the design of the adoptive transfer approach, in which Τ lymphocytes presenting an interesting response against tumor cells are extracted from the patient, expanded in vitro, and reinfused after immunodepletion, possibly leading to cancer regression. In the last decade, major progress has been achieved by the introduction of engineered lypmhocytes. In the meantime, the understanding of the molecular aspects of the TCRpMHC interaction has become essential to guide in vitro and in vivo studies. In 1996, the determination of the first structure of a TCRpMHC complex by X-ray crystallography revealed the molecular basis of the interaction. Since then, molecular modeling techniques have taken advantage of crystal structures to study the conformational space of the complex, and understand the specificity of the recognition of the pMHC by the TCR. In the meantime, experimental techniques used to determine the sequences of TCR that bind to a pMHC complex have been used intensively, leading to the collection of large repertoires of TCR sequences that are specific for a given pMHC. There is a growing need for computational approaches capable of predicting the molecular interactions that occur upon TCR/pMHC binding without relying on the time consuming resolution of a crystal structure. This work presents new approaches to analyze the molecular principles that govern the recognition of the pMHC by the TCR and the subsequent activation of the T-cell. We first introduce TCRep 3D, a new method to model and study the structural properties of TCR repertoires, based on homology and ab initio modeling. We discuss the methodology in details, and demonstrate that it outperforms state of the art modeling methods in predicting relevant TCR conformations. Two successful applications of TCRep 3D that supported experimental studies on TCR repertoires are presented. Second, we present a rigid body study of TCRpMHC complexes that gives a fair insight on the TCR approach towards pMHC. We show that the binding mode of the TCR is correctly described by long-distance interactions. Finally, the last section is dedicated to a detailed analysis of an experimental hydrogen exchange study, which suggests that some regions of the constant domain of the TCR are subject to conformational changes upon binding to the pMHC. We propose a hypothesis of the structural signaling of TCR molecules leading to the activation of the T-cell. It is based on the analysis of correlated motions in the TCRpMHC structure. - L'activation de la réponse immunitaire spécifique dirigée contre les cellules tumorales est basée sur la reconnaissance par les Lymphocytes Τ Cytotoxiques (CTL), d'un peptide antigénique (p) présenté à la suface de la cellule par le complexe majeur d'histocompatibilité de classe I (MHC). La capacité des récepteurs des lymphocytes (TCR) à distinguer les peptides endogènes des peptides étrangers constitue le mécanisme de contrôle le plus important dirigé contre les cellules infectées. L'interaction entre le TCR et le pMHC est le sujet de beaucoup d'attention dans la thérapie du cancer, depuis la conception de la méthode de transfer adoptif: les lymphocytes capables d'une réponse importante contre les cellules tumorales sont extraits du patient, amplifiés in vitro, et réintroduits après immunosuppression. Il peut en résulter une régression du cancer. Ces dix dernières années, d'importants progrès ont été réalisés grâce à l'introduction de lymphocytes modifiés par génie génétique. En parallèle, la compréhension du TCRpMHC au niveau moléculaire est donc devenue essentielle pour soutenir les études in vitro et in vivo. En 1996, l'obtention de la première structure du complexe TCRpMHC à l'aide de la cristallographie par rayons X a révélé les bases moléculaires de l'interaction. Depuis lors, les techniques de modélisation moléculaire ont exploité les structures expérimentales pour comprendre la spécificité de la reconnaissance du pMHC par le TCR. Dans le même temps, de nouvelles techniques expérimentales permettant de déterminer la séquence de TCR spécifiques envers un pMHC donné, ont été largement exploitées. Ainsi, d'importants répertoires de TCR sont devenus disponibles, et il est plus que jamais nécessaire de développer des approches informatiques capables de prédire les interactions moléculaires qui ont lieu lors de la liaison du TCR au pMHC, et ce sans dépendre systématiquement de la résolution d'une structure cristalline. Ce mémoire présente une nouvelle approche pour analyser les principes moléculaires régissant la reconnaissance du pMHC par le TCR, et l'activation du lymphocyte qui en résulte. Dans un premier temps, nous présentons TCRep 3D, une nouvelle méthode basée sur les modélisations par homologie et ab initio, pour l'étude de propriétés structurales des répertoires de TCR. Le procédé est discuté en détails et comparé à des approches standard. Nous démontrons ainsi que TCRep 3D est le plus performant pour prédire des conformations pertinentes du TCR. Deux applications à des études expérimentales des répertoires TCR sont ensuite présentées. Dans la seconde partie de ce travail nous présentons une étude de complexes TCRpMHC qui donne un aperçu intéressant du mécanisme d'approche du pMHC par le TCR. Finalement, la dernière section se concentre sur l'analyse détaillée d'une étude expérimentale basée sur les échanges deuterium/hydrogène, dont les résultats révèlent que certaines régions clés du domaine constant du TCR sont sujettes à un changement conformationnel lors de la liaison au pMHC. Nous proposons une hypothèse pour la signalisation structurelle des TCR, menant à l'activation du lymphocyte. Celle-ci est basée sur l'analyse des mouvements corrélés observés dans la structure du TCRpMHC.

Molecular analysis and mating behaviour of the Trichophyton mentagrophytes species complex.

Isolates of the Trichophyton mentagrophytes complex vary phenotypically. Whether the closely related zoophilic and anthropophilic anamorphs currently associated with Arthroderma vanbreuseghemii have to be considered as members of the same biological species remains an open question. In order to better delineate species in the T. mentagrophytes complex, we performed a mating analysis of freshly collected isolates from humans and animals with A. benhamiae and A. vanbreuseghemii reference strains, in comparison to internal transcribed spacer (ITS) and 28S rDNA sequencing. Mating experiments as well as ITS and 28S sequencing unambiguously allowed the distinction of A. benhamiae and A. vanbreuseghemii. We have also shown that all the isolates from tinea pedis and tinea unguium identified as T. interdigitale based on ITS sequences mated with A. vanbreuseghemii tester strains, but had lost their ability to give fertile cleistothecia. Therefore, T. interdigitale has to be considered as a humanized species derived from the sexual relative A. vanbreuseghemii.

Impactos Decorrentes da Expansão da Cidade de Assomada, Município de Santa Catarina – Cabo Verde

Esta dissertação tem por finalidade evidenciar os “Impactos Decorrentes da Expansão da Cidade de Assomada”. Propõe-se ainda, analisar a relação urbanorural no município de Santa Catarina, Cabo Verde. Essa análise incidiu sobre os dados da sua ocupação, que remonta à 2ª metade do século XIX, no decurso da crise e decadência da primeira cidade de Cabo Verde (Ribeira Grande, atual Cidade- Velha) e às conjunturas regionais e nacionais que condicionaram sua trajetória, bem como, os principais agentes intervenientes na transformação do espaço urbano de Assomada (Estado, proprietários de terra e comerciantes), sobretudo, no período que decorre de 1961 a 2003. A pesquisa empreendida demonstra que a “ruralidade” é um dos aspectos fundamentais de Santa Catarina, abrangendo 21,3% da população rural do território nacional e 37,9% da ilha de Santiago. Do ponto de vista urbanístico, o mapeamento da evolução da cidade de Assomada, referente à área ocupada pelas edificações nesse lapso de tempo, permite-nos perceber um crescimento bastante acelerado desse espaço urbano, sobretudo, a partir dos anos 90. Dois aspectos importantes se encontram na base dessa transformação: a Independência Nacional ocorrida em 1975 e a Emigração. Com a Independência, o país passou a beneficiar de amplos programas de cooperação internacional o que veio a reforçar a sua importância econômica e política, visualizada através de projetos industrializantes, infra-estruturas e equipamentos de diversas naturezas. Esses investimentos foram por sua vez reforçados por aportes dos emigrantes, que passaram a ser um dos parceiros fundamentais no desenvolvimento de Santa Catarina, através de iniciativas privadas e empresariais, como a construção de residências e constituição de empresas. A análise do indicador uso e ocupação do solo urbano permitiu-nos identificar alguns impactos ambientais causados, sobretudo, pela falta de instrumentos de gestão urbanística, ausência de cadastro municipal e uma deficiente fiscalização dos órgãos administrativos.

Anopheles goeldii Rozeboom & Gabaldón (Diptera, Culicidae): a species of the Nuneztovari Complex of Anopheles Meigen

Anopheles (Nyssorhynchus) goeldiiRozeboom & Gabaldón, 1941, a species of the Nuneztovari Complex, was described based on morphological characteristics of the male, female, larva, pupa, and eggs. The type locality is Boa Vista (= Fordlândia), a district in the vicinity of Rio Tapajós, in the municipality of Aveiro, in the state of Pará, Brazil. Anopheles goeldii is redescribed based on morphological traits of the fourth instar larva, pupa, egg, and male and female. DNA sequences from the cytochrome oxidase subunit I (COI barcode region) of the mitochondrial genome were utilized for species characterization. Specimens of An. goeldii from the Pará, Amapá, and Amazonas states were employed to redescribe the species and to compare with morphologically similar taxa.

Pre-Mesozoic Alpine basements - Their place in the European Paleozoic framework

Prior to their Alpine overprinting, most of the pre-Mesozoic basement areas in Alpine orogenic structures shared a complex evolution, starting with Neoproterozoic sediments that are thought to have received detrital input from both West and East Gondwanan cratonic sources. A subsequent Neoproterozoic-Cambrian active margin setting at the Gondwana margin was followed by a Cambrian-Ordovician rifting period, including an Ordovician cordillera-like active margin setting. During the Late Ordovician and Silurian periods, the future Alpine domains recorded crustal extension along the Gondwana margin, announcing the future opening of the Paleotethys oceanic domain. Most areas then underwent Variscan orogenic events, including continental subduction and collisions with Avalonian-type basement areas along Laurussia and the juxtaposition and the duplication of terrane assemblages during strike slip, accompanied by contemporaneous crustal shortening and the subduction of Paleotethys under Laurussia. Thereafter, the final Pangea assemblage underwent Triassic and Jurassic extension, followed by Tertiary shortening, and leading to the buildup of the Alpine mountain chain. Recent plate-tectonic reconstructions place the Alpine domains in their supposed initial Cambrian-Ordovician positions in the eastern part of the Gondwana margin, where a stronger interference with the Chinese blocks is proposed, at least from the Ordovician onward. For the Visean time of the Variscan continental collision, the distinction of the former tectonic lower-plate situation is traceable but becomes blurred through the subsequent oblique subduction of Paleotethys under Laurussia accompanied by large-scale strike slip. Since the Pennsylvanian, this global collisional scenario has been replaced by subsequent and ongoing shortening and strike slip under rising geothermal conditions, and all of this occurred before all these puzzle elements underwent the complex Alpine reorganization.

Evaluation of an online, case-based interactive approach to teaching pathophysiology

Rapport de synthèse : L'article qui fait l'objet de ma thèse évalue une nouvelle approche pédagogique pour l'apprentissage de certains chapitres de physiopathologie. Le dispositif pédagogique se base sur l'alternance d'apprentissage ex-cathedra et de l'utilisation d'un site web comprenant des vignettes cliniques. Lors de la consultation de ces-dernières, l'étudiant est invité à demander des examens de laboratoire dont il pourrait justifier la pertinence selon le cas clinique étudié. La nouveauté du procédé réside dans le fait que, préalablement à son cours ex-cathedra, l'enseignant peut consulter les statistiques de demandes de laboratoire et ainsi orienter son cours selon les éléments mal compris par les étudiants. A la suite du cours ex-cathedra, les étudiants peuvent consulter sur internet la vignette clinique complète avec des explications. A l'issue de tout le cours, une évaluation auprès des étudiants a été conduite. Le procédé a été mis en place durant deux années consécutives et l'article en discute notamment les résultats. Nous avons pu conclure que cette méthode innovatrice d'enseignement amène les étudiants à mieux se préparer pour les cours ex-cathedra tout en permettant à l'enseignant d'identifier plus précisément quelles thématiques étaient difficiles pour les étudiants et donc d'ajuster au mieux son cours. Mon travail de thèse a consisté à créer ce dispositif d'apprentissage, à créer l'application web des vignettes cliniques et à l'implanter durant deux années consécutives. J'ai ensuite analysé les données des évaluations et écrit l'article que j'ai présenté à la revue 'Medical Teacher'. Après quelques corrections et précisions demandées par le comité de lecture, l'article a été accepté et publié. Ce travail a débouché sur une seconde version de l'application web qui est actuellement utilisée lors du module 3.1 de 3è année à l'Ecole de Médecine à Lausanne. Summary : Since the early days of sexual selection, our understanding of the selective forces acting on males and females during reproduction has increased remarkably. However, despite a long tradition of experimental and theoretical work in this field and relentless effort, numerous questions remain unanswered and many results are conflicting. Moreover, the interface between sexual selection and conservation biology has to date received little attention, despite existing evidence for its importance. In the present thesis, I first used an empirical approach to test various sexual selection hypotheses in a population of whitefish of central Switzerland. This precise population is characterized by a high prevalence of gonadal alterations in males. In particular, I challenged the hypothesis that whitefish males displaying peculiar gonadal features are of lower genetic quality than other seemingly normal males. Additionally, I also worked on identifying important determinant of sperm behavior. During a second theoretical part of my work, which is part of a larger project on the evolution of female mate preferences in harvested fish populations, I developed an individual-based simulation model to estimate how different mate discrimination costs affect the demographical behavior of fish populations and the evolutionary trajectories of female mate preferences. This latter work provided me with some insight on a recently published article addressing the importance of sexual selection for harvesting-induced evolution. I built upon this insight in a short perspective paper. In parallel, I let some methodological questions drive my thoughts, and wrote an essay about possible synergies between the biological, the philosophical and the statistical approach to biological questions.

Photodamage to Oxygen Evolving Complex - An Initial Event in Photoinhibition of Photosystem II.

Photosystem II (PSII) of oxygenic photosynthesis is susceptible to photoinhibition. Photoinhibition is defined as light induced damage resulting in turnover of the D1 protein subunit of the reaction center of PSII. Both visible and ultraviolet (UV) light cause photoinhibition. Photoinhibition induced by UV light damages the oxygen evolving complex (OEC) via absorption of UV photons by the Mn ion(s) of OEC. Under visible light, most of the earlier hypotheses assume that photoinhibition occurs when the rate of photon absorption by PSII antenna exceeds the use of the absorbed energy in photosynthesis. However, photoinhibition occurs at all light intensities with the same efficiency per photon. The aim of my thesis work was to build a model of photoinhibition that fits the experimental features of photoinhibition. I studied the role of electron transfer reactions of PSII in photoinhibition and found that changing the electron transfer rate had only minor influence on photoinhibition if light intensity was kept constant. Furthermore, quenching of antenna excitations protected less efficiently than it would protect if antenna chlorophylls were the only photoreceptors of photoinhibition. To identify photoreceptors of photoinhibition, I measured the action spectrum of photoinhibition. The action spectrum showed resemblance to the absorption spectra of Mn model compounds suggesting that the Mn cluster of OEC acts as a photoreceptor of photoinhibition under visible light, too. The role of Mn in photoinhibition was further supported by experiments showing that during photoinhibition OEC is damaged before electron transfer activity at the acceptor side of PSII is lost. Mn enzymes were found to be photosensitive under visible and UV light indicating that Mn-containing compounds, including OEC, are capable of functioning as photosensitizers both in visible and UV light. The experimental results above led to the Mn hypothesis of the mechanism of continuous-light-induced photoinhibition. According to the Mn hypothesis, excitation of Mn of OEC results in inhibition of electron donation from OEC to the oxidized primary donor P680+ both under UV and visible light. P680 is oxidized by photons absorbed by chlorophyll, and if not reduced by OEC, P680+ may cause harmful oxidation of other PSII components. Photoinhibition was also induced with intense laser pulses and it was found that the photoinhibitory efficiency increased in proportion to the square of pulse intensity suggesting that laser-pulse-induced photoinhibition is a two-photon reaction. I further developed the Mn hypothesis suggesting that the initial event in photoinhibition under both continuous and pulsed light is the same: Mn excitation that leads to the inhibition of electron donation from OEC to P680+. Under laser-pulse-illumination, another Mn-mediated inhibitory photoreaction occurs within the duration of the same pulse, whereas under continuous light, secondary damage is chlorophyll mediated. A mathematical model based on the Mn hypothesis was found to explain photoinhibition under continuous light, under flash illumination and under the combination of these two.

Atrioventricular septal defects among infants in Europe: a population-based study of prevalence, associated anomalies, and survival.

OBJECTIVE: To describe the epidemiology of chromosomal and non-chromosomal cases of atrioventricular septal defects in Europe. METHODS: Data were obtained from EUROCAT, a European network of population-based registries collecting data on congenital anomalies. Data from 13 registries for the period 2000-2008 were included. RESULTS: There was a total of 993 cases of atrioventricular septal defects, with a total prevalence of 5.3 per 10,000 births (95% confidence interval 4.1 to 6.5). Of the total cases, 250 were isolated cardiac lesions, 583 were chromosomal cases, 79 had multiple anomalies, 58 had heterotaxia sequence, and 23 had a monogenic syndrome. The total prevalence of chromosomal cases was 3.1 per 10,000 (95% confidence interval 1.9 to 4.3), with a large variation between registers. Of the 993 cases, 639 cases were live births, 45 were stillbirths, and 309 were terminations of pregnancy owing to foetal anomaly. Among the groups, additional associated cardiac anomalies were most frequent in heterotaxia cases (38%) and least frequent in chromosomal cases (8%). Coarctation of the aorta was the most common associated cardiac defect. The 1-week survival rate for live births was 94%. CONCLUSION: Of all cases, three-quarters were associated with other anomalies, both chromosomal and non-chromosomal. For infants with atrioventricular septal defects and no chromosomal anomalies, cardiac defects were often more complex compared with infants with atrioventricular septal defects and a chromosomal anomaly. Clinical outcomes for atrioventricular septal defects varied between regions. The proportion of termination of pregnancy for foetal anomaly was higher for cases with multiple anomalies, chromosomal anomalies, and heterotaxia sequence.

Genetic and maternal contributions to offspring viability under different environmental conditions in various salmonids

Summary : Due to anthropogenic impacts and natural fluctuations, fish usually have to cope with constantly changing and often hostile environments. Whereas adult fish have various possibilities to counteract unfavourable environmental conditions, embryos have much fewer options. Besides by their developing immune system, they are protected by the egg envelopes and several immune substances provided by their mothers. In addition to this, they may also adjust their hatching timing in reaction to various risks. However, individuals may vary in their defensive potential. This variation may be either based on their genetics and/or on differential maternal investments and may be dependent on the experienced stress. Nevertheless, in fish, the impact of such parental contributions on embryo and/or juvenile viability is still poorly investigated. The main objective of this thesis was to investigate the importance of paternal (i.e. genetic) and maternal (i.e. genetic + egg investment) contributions to offspring viability under different environmental conditions and at different life stages. In order to investigate this, we used gametes of various salmonids for in vitro fertilisation experiments based on full-factorial breeding designs. The individual studies are summarised in the following chapters: In the first chapter, we tested the effectiveness of the embryonic immune system in Lake whitefish (Coregonus palaea). Namely, we investigated paternal and maternal contributions to the embryos' tolerance to different kinds of pathogen exposure. Additionally, we tested whether an early sub-léthal exposure has a positive or a negative effect on an embryo's susceptibility to later pathogen exposures with the same pathogen. We found that pre-challenged embryos were more susceptible to future challenges. Moreover, pathogen susceptibility was dependent on maternal investments and/or the embryos' own genetics, depending on the challenge level. Chapter 2 summarises a similar study with brown trout (Salmo trutta). In addition to the previously described investigations, we analysed if genetic effects on offspring viability are mediated either by parental MHC genotypes or relatedness based on neutral microsatellite markers, and we tested if males signal their genetic quality either by their body size or their melanin-based skin colouration. We found that embryo survival was lower at higher stress levels and dependent on the embryos' genetics. Addirionally, parents with similar and/or, very common MHC genotypes had higher offspring viabilities. Finally, darker males produced more viable offspring. In the first two chapters we investigated the embryos' defensive potential based on their immune system, i.e. their pathogen tolerance. In chapter 3 we investigate whether hatching timing of Lake whitefìsh (C. palaea) is dependent on parental contributions and/or on pathogen pressure, and whether there are parental-environmental interactions. We found that whitefish embryos hatch earlier under increasing pathogen pressure. Moreover, hatching timing was affected by embryo genetics and/or maternally provided resources, but the magnitude of the effect was dependent on the pathogen. pressure. We also found a significant paternal-environmental interaction, indicating that the hatching efficiency of a certain sib group is dependent on the pathogen environment. Chapter 4 describes an analogous study with brown trout (S. trutta), with similar findings. In the former chapters, we only looked at offspring performance during the embryonic period, and only under semi-natural conditions. In chapter 5 we now test the performance and viability of embryonic and juvenile brown trout (S. trutta) under natural conditions. To measure embryo viability, we put them in brood boxes, buried them in the gravel of a natural river, and analysed survival after several months. To investigate juvenile survival and performance, wé reared embryos under different stress levels in the laboratory and subsequently released the resulting hatchlings in to a closed river section. Juvenile size and survival was then determined one year later. Additionally, we investigated if sires differ in their genetic quality, determined by embryo and juvenile survival as well as juvenile size, and if they signal their quality by either body size or melanin-based body darkness. We found hat juvenile size was dependent on genetic effects and on maternal investment, whereas this was neither the case for embryo nor for juvenile survival. Additionally, we found that offspring of darker males grew larger, and larger juveniles had also an increased survival. Finally, we found acarry-over effect of the early non-lethal challenge: exposing embryos to higher stress levels resulted in smaller juveniles. To evaluate the long-term performance of differently treated groups, mark-recapture studies are inevitable. For this purpose, effective mass-marking techniques are essential. In chapter 6 we tested the suitability of the fluorescent pigment spray marking method for the mass marking of European graylings (Thymallus thymallus), with very promising results. Our in vitro fertilisation studies on whitefish may reveal new insights on potential genetic benefits of mate choice, but the mating system of whitefish under natural conditions is still poorly investigated. In order to study this, we installed underwater cameras at the spawning place of a Coregonus suidteri population, recorded the whole mating period and subsequently analysed the recordings. Confirmations of previous findings as well as exciting new observations are listed and discussed in chapter 7. Dus aux impacts anthropogéniques et aux fluctuations naturelles, les poissons doivent faire face à des environnements en perpétuel changement. Ces changements font que les poissons doivent s'adapter à de nouvelles situations, souvent hostiles pour eux. Les adultes ont différentes possibilités d'échapper à un environnement peu favorable, ce n'est par contre pas le cas des embryons. Les embryons sont protégés d'une part par leur système immunitaire en développement, d'autre part, par la coquille de l'eeuf et différentes substances immunitaires fournies par leur mère. De plus, ils sont capables d'influencer leur propre date d'éclosion en réponse à différents facteurs de stress. Malgré tout, les individus varient dans leur capacité à se défendre. Cette variation peut être basé sur des facteurs génétiques et/ou sur des facteurs maternels, et est dépendante du stress subi. Néanmoins, chez les poissons, l'impact de telles contributions parentales sur la survie d'embryons et/ou juvéniles est peu étudié. L'objectif principal de cette thèse a été d'approfondir les connaissances sur l'importance de la contribution paternelle (c.a.d. génétique) et maternelle (c.a.d. génétique + investissement dans l'oeuf) sur la survie des jeunes dans différentes conditions expérimentales et stades de vie. Pour faire ces analyses, nous avons utilisé des gamètes de divers salmonidés issus de croisements 'full-factorial'. Les différentes expériences sont résumées dans les chapitres suivants: Dans le premier chapitre, nous avons testé l'efficacité du système immunitaire des embryons chez les corégones (Coregonus palea). Plus précisément nous avons étudié la contribution paternelle et maternelle à la tolérance des embryons à différents niveaux de stress pathogène. Nous avons aussi testé, si une première exposition non létale à un pathogène avait un effet positif ou négatif sur la susceptibilité d'un embryon a une deuxième exposition au même pathogène. Nous avons trouvé que des embryons qui avaient été exposés une première fois étaient plus sensibles au pathogène par la suite. Mais aussi que la sensibilité au pathogène était dépendante de l'investissement de la mère et/ou des gènes de l'embryon, dépendamment du niveau de stress. Le deuxième chapitre résume une étude similaire avec des truites (Salmo truffa). Nous avons examiné, si la survie des jeunes variait sous différentes intensités de stress, et si la variance observée était due aux gènes des parents. Nous avons aussi analysé si les effets génétiques sur la survie des juvéniles étaient dus au MHC (Major Histocompatibility Complex) ou au degré de parenté des parents. De plus, nous avons analysé si les mâles signalaient leur qualité génétique par la taille du corps ou par leur coloration noire, due à la mélanine. On a trouvé que la survie des embryons était plus basse quand le niveau de stress était plus haut mais que la variation restait dépendante de la génétique des embryons. De plus, les parents avec des MHC similaires et/ou communs avaient des embryons avec une meilleure survie. Par contre, des parents avec un degré de parenté plus haut produisent des embryons avec une survie plus mauvaise. Finalement nous avons montré que les mâles plus foncés ont des embryons qui survivent mieux, mais que la taille des mâles n'a pas d'influence sur la survie de ces mêmes embryons. Dans les deux premiers chapitres, nous avons étudié le potentiel de défense des embryons basé sur leur système immunitaire, c.a.d. leur tolérance aux pathogènes. Dans le troisième chapitre, nous nous intéressons à la date d'éclosion des corégones (C. palea), pour voir si elle est influencée par les parents ou par la pression des pathogènes, et si il y a une interaction entre ces deux facteurs. Nous avons trouvé que les jeunes naissent plus rapidement lorsque la pression en pathogènes augmente. La date d'éclosion est influencée par la génétique des embryons et/ou l'investissement des parents, mais c'est la magnitude des effets qui est dépendante de la pression du pathogène. Nous avons aussi trouvé une interaction entre l'effet paternel et l'environnement, ce qui indique que la rapidité d'éclosion de certains croisements est dépendante des pathogènes dans l'environnement. Le chapitre 4 décrit une étude analogue avec de truites (S. truffa), avec des résultats sitzimilaires. Dans les précédents chapitres nous nous sommes uniquement concentrés sur les performances des jeunes durant leur stade embryonnaire, et seulement dans des conditions semi naturelles. Dans le chapitre 5 nous testons la performance et la viabilité des embryons et de juvéniles de truites (S. truffa) dans des conditions naturelles. Nous avons trouvé que la taille des juvéniles était dépendante d'effets génétiques et de l'investissement maternel, mais ceci n'était ni les cas pour les survie des embryons et des juvéniles. De plus, nous avons trouvé que les jeunes des mâles plus foncés devenaient plus grands et que les grands ont un meilleur taux de survie. Finalement nous avons trouvé un 'carry-over effect' d'une première exposition non létale à un pathogène: exposer des embryons à des plus hauts niveaux de stress donnait des juvéniles plus petits. Pour évaluer la performance à long terme de groupes traités dé manières différentes, une méthode de marquage-recapture est inévitable. Pour cette raison, des techniques de marquage en masse sont nécessaires. Dans le chapitre 6, nous avons testé l'efficacité de la technique `fluorescent pigment spray marking' pour le marquage en masse de l'Ombre commun (Thymallus thymallus), avec des résultats très prometteurs. Les études de fertilisations in vitro avec les corégones nous donnent une idée du potentiel bénéfice génétique que représente la sélection d'un bon partenaire, même si le système d'accouplement des corégones en milieu naturel reste peu connu. Pour combler cette lacune, nous avons installé des caméras sous-marines autour de la frayère d'une population de corégones (C. suidteri), nous avons enregistré toute la période de reproduction et nous avons analysé les données par la suite. Ainsi, nous avons été capables de confirmer bien des résultats trouvés précédemment, mais aussi de faire de nouvelles observations. Ces résultats sont reportés dans le septième chapitre, où elles sont comparées avec des observations antérieures.

Prediction of 18-month survival in patients with primary myelodysplastic syndrome. A regression model and scoring system based on the combination of chromosome findings and the Bournemouth score.

The predictive potential of six selected factors was assessed in 72 patients with primary myelodysplastic syndrome using univariate and multivariate logistic regression analysis of survival at 18 months. Factors were age (above median of 69 years), dysplastic features in the three myeloid bone marrow cell lineages, presence of chromosome defects, all metaphases abnormal, double or complex chromosome defects (C23), and a Bournemouth score of 2, 3, or 4 (B234). In the multivariate approach, B234 and C23 proved to be significantly associated with a reduction in the survival probability. The similarity of the regression coefficients associated with these two factors means that they have about the same weight. Consequently, the model was simplified by counting the number of factors (0, 1, or 2) present in each patient, thus generating a scoring system called the Lausanne-Bournemouth score (LB score). The LB score combines the well-recognized and easy-to-use Bournemouth score (B score) with the chromosome defect complexity, C23 constituting an additional indicator of patient outcome. The predicted risk of death within 18 months calculated from the model is as follows: 7.1% (confidence interval: 1.7-24.8) for patients with an LB score of 0, 60.1% (44.7-73.8) for an LB score of 1, and 96.8% (84.5-99.4) for an LB score of 2. The scoring system presented here has several interesting features. The LB score may improve the predictive value of the B score, as it is able to recognize two prognostic groups in the intermediate risk category of patients with B scores of 2 or 3. It has also the ability to identify two distinct prognostic subclasses among RAEB and possibly CMML patients. In addition to its above-described usefulness in the prognostic evaluation, the LB score may bring new insights into the understanding of evolution patterns in MDS. We used the combination of the B score and chromosome complexity to define four classes which may be considered four possible states of myelodysplasia and which describe two distinct evolutional pathways.