Submitted by Ana Damasceno (ana.damasceno@unipampa.edu.br) on 2016-09-12T20:02:35Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Relação Da Nutrição Apícola Com A Microbiota Do Pólen E Do Sistema Digestório De Abelhas Melíferas Verificada Por Sequenciamento De Nova Geração.pdf: 2887795 bytes, checksum: 12218b912a445c8eb21706a94911e90f (MD5) / Made available in DSpace on 2016-09-12T20:02:35Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Relação Da Nutrição Apícola Com A Microbiota Do Pólen E Do Sistema Digestório De Abelhas Melíferas Verificada Por Sequenciamento De Nova Geração.pdf: 2887795 bytes, checksum: 12218b912a445c8eb21706a94911e90f (MD5) Previous issue date: 2015-03-16 / A microbiota e os genes funcionais ativamente envolvidos no processo de decomposição e utilização de grãos de pólen em pão de mel e no trato digestório de abelha ainda não são completamente compreendidos. O objetivo deste trabalho foi avaliar a estrutura e diversidade da comunidade de bactérias e Archaeas em amostrasde pão de mel e sistema digestório de abelhas africanizadas, bem como para prever os genes envolvido na bioprocessamento microbiano do pólen, usando a tecnologia de seqüenciamento de nova geração. Um total de 11 filos bacterianos foram encontrados dentro do sistema de digestório de abelhas e 10 filos bacterianos foram encontrado dentro pão de mel. Embora a comparação a nível de filo mostre mais filos em comum, a análise filogenética mais profunda mostrou maior variação de composição taxonômica. A família Enterobacteriaceae, Ricketsiaceae, Spiroplasmataceae e Bacillaceae, foram os principais grupos responsáveis por a especificidade do intestino de abelhas, enquanto as principais famílias responsáveis pela especificidade do pão de mel foram Neisseriaceae, Flavobacteriaceae, Acetobacteraceae e Lactobacillaceae. Em termos da estrutura da comunidade microbiana, a análise mostrou que as comunidades dos dois ambientes foram bastante diferentes umas das outras, com apenas 7% dos táxons a nível de espécies compartilhados entre o sitema digestório de abelhas e o pão de mel. Os resultados indicaram a presença de um elevado nível de especialização e uma microbiota intestinal bem adaptada dentro de cada abelha e do pão de mel.A comunidade associada ao pão de mel, apresentou maior abundância relativa de genes relacionados com a degradação de aminoácidos, carboidratos, e o metabolismo lipídico, sugerindo que biodegradação do pólen ocorre predominantemente pela microbiota associada ao pão de mel. Estes resultados sugerem uma complexa e importante relação entre nutrição de abelhas e suas comunidades microbianas. / The microbiota and the functional genes actively involved in the process of breakdown and utilization of pollen grains in beebread and beeguts are not yet understood. The aim of this work was to assess the diversity and community structure of bacteria and archaea in Africanized honeybee guts and beebread as well as to predict the genes involved in the microbial bioprocessing of pollen using state of the art ‘post-light’ based sequencing technology. A total of 11 bacterial phyla were found within bee guts and 10 bacterial phyla were found within beebread. Although the phylum level comparison shows most phyla in common, a deeper phylogenetic analysis showed greater variation of taxonomic composition. The families Enterobacteriaceae, Ricketsiaceae, Spiroplasmataceae and Bacillaceae, were the main groups responsible for the specificity of the bee gut while the main families responsible for the specificity of the beebread were Neisseriaceae, Flavobacteriaceae, Acetobacteraceae and Lactobacillaceae. In terms of microbial community structure, the analysis showed that the communities from the two environments were quite different from each other with only 7 % of species-level taxa shared between beegut and beebread. The results indicated the presence of a highly specialized and well-adapted microbiota within each bee gut and beebread. The beebread community included a greater relative abundance of genes related to amino acid, carbohydrate, and lipid metabolism, suggesting that pollen biodegradation predominantly occurs in the beebread. These results suggests a complex and important relationship between honeybee nutrition and their microbial communities.

Submitted by Ana Damasceno (ana.damasceno@unipampa.edu.br) on 2016-10-31T16:19:59Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Sequenciamento por Ion Torrent revela padrões de interação e distribuição de comunidades microbianas em um perfil de solo ornitogênico da Ilha Seymour, Península Antártica.pdf: 907768 bytes, checksum: 05c430f88f8f32e041e763c1d453d54a (MD5) / Made available in DSpace on 2016-10-31T16:19:59Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Sequenciamento por Ion Torrent revela padrões de interação e distribuição de comunidades microbianas em um perfil de solo ornitogênico da Ilha Seymour, Península Antártica.pdf: 907768 bytes, checksum: 05c430f88f8f32e041e763c1d453d54a (MD5) Previous issue date: 2014 / Neste estudo, foram analisadas e comparadas comunidades bacterianas do solo de uma pinguineira da Ilha Seymour (Península Antártica) em termos de abundância, estrutura, diversidade e rede de interações, a fim de se identificar padrões de interação entre os vários grupos de bactérias presentes em solos ornitogênicos em diferentes profundidades (camadas). A análise das sequências revelou a presença de oito filos distribuídos em diferentes proporções entre as Camadas 1 (0-8 cm), 2 (20-25 cm) e 3 (35-40 cm). De acordo com os índices de diversidade, a Camada 3 apresentou os maiores valores de riqueza, diversidade e uniformidade quando comparado com as Camadas 1 e 2. Em termos de estrutura da comunidade microbiana, a análise UniFrac mostrou que as comunidades microbianas das três camadas foram muito diferentes umas das outras. A análise de redes revelou a existência de um padrão único de interações no qual a rede microbiana formou uma topologia de agrupamento, mas não estruturado em módulos, como de costume em comunidades biológicas. Da mesma forma, através da utilização de análise de redes, foi possível identificar táxons específicos como sendo potencialmente importantes para a estruturação e funcionamento da comunidade microbiana. Além disso, as análises de simulação indicaram que a perda de grupos importantes de microorganismos pode alterar significativamente os padrões de interação dentro da comunidade microbiana. Estes resultados fornecem novos insights sobre as interações bacterianas e ecologia microbiana desse importante, mas ameaçado ambiente. / In this study, we analyzed and compared the soil bacterial communities from a penguin rookery site at Seymour Island (Antarctic Peninsula) in terms of abundance, structure, diversity and interaction network in order to identify interaction patterns among the various groups of bacteria presented in an ornithogenic site at three depths (layers). The analysis of the sequences revealed the presence of 8 phyla distributed in different proportions among the Layers 1 (0-8 cm), 2 (20-25 cm) and 3 (35-40 cm). According to the diversity indexes, Layer 3 presented the highest values of richness, diversity and evenness when compared to Layers 1 and 2. In terms of bacterial community structure, the unweighted and weighted UniFrac analysis showed that the soil bacterial communities from the three layers were quite different from each other. Network analysis revealed the existence of a unique pattern of interactions in which the soil microbial network formed a clustered topology, but not structured in modules, as usual in biological communities. In addition, through the use of network analysis, it was possible to identify specific taxa as potentially important for the structuring and functioning of the microbial community. Furthermore, simulation analyzes indicated that the loss of potential keystone groups of microorganisms may significantly alter the patterns of interactions within the microbial communityThese findings provide new insights into the bacterial interactions and microbial ecology of this important, but threatened environment.

Submitted by Ana Damasceno (ana.damasceno@unipampa.edu.br) on 2016-07-21T18:21:38Z No. of bitstreams: 1 Relação Da Nutrição Apícola Com A Microbiota Do Pólen E Do Sistema Digestório De Abelhas Melíferas Verificada Por Sequenciamento De Nova Geração.pdf: 2887795 bytes, checksum: 12218b912a445c8eb21706a94911e90f (MD5) / Approved for entry into archive by Ana Damasceno (ana.damasceno@unipampa.edu.br) on 2016-08-23T16:34:05Z (GMT) No. of bitstreams: 1 Relação Da Nutrição Apícola Com A Microbiota Do Pólen E Do Sistema Digestório De Abelhas Melíferas Verificada Por Sequenciamento De Nova Geração.pdf: 2887795 bytes, checksum: 12218b912a445c8eb21706a94911e90f (MD5) / Made available in DSpace on 2016-08-23T16:34:05Z (GMT). No. of bitstreams: 1 Relação Da Nutrição Apícola Com A Microbiota Do Pólen E Do Sistema Digestório De Abelhas Melíferas Verificada Por Sequenciamento De Nova Geração.pdf: 2887795 bytes, checksum: 12218b912a445c8eb21706a94911e90f (MD5) Previous issue date: 2015-03-16 / A microbiota e os genes funcionais ativamente envolvidos no processo de decomposição e utilização de grãos de pólen em pão de mel e no trato digestório de abelha ainda não são completamente compreendidos. O objetivo deste trabalho foi avaliar a estrutura e diversidade da comunidade de bactérias e Archaeas em amostrasde pão de mel e sistema digestório de abelhas africanizadas, bem como para prever os genes envolvido na bioprocessamento microbiano do pólen, usando a tecnologia de seqüenciamento de nova geração. Um total de 11 filos bacterianos foram encontrados dentro do sistema de digestório de abelhas e 10 filos bacterianos foram encontrado dentro pão de mel. Embora a comparação a nível de filo mostre mais filos em comum, a análise filogenética mais profunda mostrou maior variação de composição taxonômica. A família Enterobacteriaceae, Ricketsiaceae, Spiroplasmataceae e Bacillaceae, foram os principais grupos responsáveis por a especificidade do intestino de abelhas, enquanto as principais famílias responsáveis pela especificidade do pão de mel foram Neisseriaceae, Flavobacteriaceae, Acetobacteraceae e Lactobacillaceae. Em termos da estrutura da comunidade microbiana, a análise mostrou que as comunidades dos dois ambientes foram bastante diferentes umas das outras, com apenas 7% dos táxons a nível de espécies compartilhados entre o sitema digestório de abelhas e o pão de mel. Os resultados indicaram a presença de um elevado nível de especialização e uma microbiota intestinal bem adaptada dentro de cada abelha e do pão de mel.A comunidade associada ao pão de mel, apresentou maior abundância relativa de genes relacionados com a degradação de aminoácidos, carboidratos, e o metabolismo lipídico, sugerindo que biodegradação do pólen ocorre predominantemente pela microbiota associada ao pão de mel. Estes resultados sugerem uma complexa e importante relação entre nutrição de abelhas e suas comunidades microbianas. / The microbiota and the functional genes actively involved in the process of breakdown and utilization of pollen grains in beebread and beeguts are not yet understood. The aim of this work was to assess the diversity and community structure of bacteria and archaea in Africanized honeybee guts and beebread as well as to predict the genes involved in the microbial bioprocessing of pollen using state of the art ‘post-light’ based sequencing technology. A total of 11 bacterial phyla were found within bee guts and 10 bacterial phyla were found within beebread. Although the phylum level comparison shows most phyla in common, a deeper phylogenetic analysis showed greater variation of taxonomic composition. The families Enterobacteriaceae, Ricketsiaceae, Spiroplasmataceae and Bacillaceae, were the main groups responsible for the specificity of the bee gut while the main families responsible for the specificity of the beebread were Neisseriaceae, Flavobacteriaceae, Acetobacteraceae and Lactobacillaceae. In terms of microbial community structure, the analysis showed that the communities from the two environments were quite different from each other with only 7 % of species-level taxa shared between beegut and beebread. The results indicated the presence of a highly specialized and well-adapted microbiota within each bee gut and beebread. The beebread community included a greater relative abundance of genes related to amino acid, carbohydrate, and lipid metabolism, suggesting that pollen biodegradation predominantly occurs in the beebread. These results suggests a complex and important relationship between honeybee nutrition and their microbial communities.

ObjectifLe diagnostic prénatal (DPN) a pour but de détecter des pathologies foetales in utero. L’objectif de ce travail était de mettre au point et d’appliquer les techniques d’exploration chromosomique en prénatal. Nous avons, tout d’abord, validé et évalué une plateforme de séquençage basée sur la technologie des semi-conducteurs, Ion Proton®, pour le dépistage prénatal non-invasif (DPNI) des principales aneuploïdies en routine clinique, puis évalué l’intérêt de l’Analyse Chromosomique sur Puces à ADN (ACPA) dans le diagnostic prénatal des retards de croissance intra-utérin (RCIU) foetaux. Matériel et Méthodes Nous avons inclus prospectivement 2505 patientes enceintes analysées par huit laboratoires hospitalo-universitaires de génétique : 695 grossesses à haut risque pour la trisomie 21 (risque ≥1/250 ou avec anomalie échographique) dans une étude de validation de la technique du test ADN libre circulant (ADNlc), et 1810 grossesses à risque, sans anomalie échographique, en routine clinique. Les issues de grossesses étaient toutes disponibles dans l’étude de validation et pour 521 grossesses dans l’étude en routine clinique. L’ADNlc extrait d’échantillons plasmatiques était séquencé, puis les données étaient analysées à l’aide du logiciel WISECONDOR. Les résultats des tests ADNlc étaient comparés aux caryotypes foetaux ou 7 aux données à la naissance. Nous avons aussi évalué le taux d’échec et comparé trois méthodes d’évaluation de la fraction foetale (FF) (RASSF1A, DEFRAG et SANEFALCON). Nous avons rétrospectivement inclus tous les foetus référés pour un prélèvement invasif pour RCIU et étudié les résultats de technique d’hybridation in situ en fluorescence (FISH), caryotypes et ACPA. Résultats Les résultats des deux cohortes de l’étude sur l’ADNlc étaient cohérents et les âges gestationnels n’étaient pas significativement différents ; les données ont été combinées afin d’étoffer la cohorte à analyser. Respectivement, la sensibilité et la spécificité étaient de : 98.3% (95% IC, 93.5–99.7%) et 99.9% (95% IC, 99.4–100%) pour la trisomie 21; 96.7% (95% IC, 80.9–99.8%) et 100% (95% IC, 99.6–100%) pour la trisomie 18 ; et 94.1% (95% IC, 69.2–99.7%) et 100% (95% IC, 99.6–100%) pour la trisomie 13. Le taux de non rendus était de 1.2% initialement puis après réanalyse de 0.6%. L’estimation de la FF avec les méthodes RASSF1A et DEFRAG étaient comparables, toutes deux compatibles avec une utilisation en routine clinique. Parmi les 162 foetus RCIU (78 associés et 84 isolés) inclus dans l’étude ACPA, 15 avaient une FISH pathologique : 10 RCIU associés et cinq RCIU isolés. Parmi 143 foetus étudiés par ACPA, 10 (7%) présentaient un variant du nombre de copies (CNV), tous étaient des RCIUs associés (10/65 soit 15.4%; 95 IC: 8.4%‐26.2%), versus 0/78 dans le groupe RCIUs isolés (95% IC: 0%‐5.6%). Six foetus (4.2%) ont présenté des variants de signification inconnue (VSI) (trois RCIU associés et trois RCIU isolés). Conclusion : Notre étude évaluant le test ADNlc utilisant la technologie des semi-conducteurs est la première étude clinique à rapporter les issues de grossesses dans une population aussi large. La plateforme est performante pour le DPNI des principales aneuploïdies. Notre protocole robuste est facilement applicable en routine clinique. Notre étude souligne une augmentation de rendement diagnostique de l’ACPA de 6.1% (4/65) par rapport au caryotype pour le DPN des foetus présentant un RCIU associé. Aucun CNV pathogène n’a été mis en évidence dans le groupe RCIU isolé. L’ADNlc pourrait-il supplanter l’ACPA dans cette population de RCIU isolé ? Le développement du test ADNlc a permis de limiter le nombre de prélèvements invasifs et donc leurs complications [...]. / ObjectivePrenatal diagnosis allows to detect fetal pathologies in-utero. The goal of this work was both technical development and application of the chromosomal exploration technics in prenatal diagnosis. First, we aimed to validate and evaluate the performance metrics of the highthroughput semiconductor sequencing platform, Ion Proton®, in non-invasive prenatal genetic screening (NIPS) for common fetal aneuploidies in a clinical setting and, then to evaluate the diagnostic utility of prenatal diagnosis using the chromosomal microarray analysis (CMA) for fetuses presenting with isolated or associated intrauterine growth restriction (IUGR). Methods : First, regarding NIPS, a prospective cohort study including 2505 pregnant women from eight academic genetics laboratories (695 high risk pregnancies for trisomy 21 (risk ≥1/250 or with ultrasound anomalies) in a validation study, and 1810 such pregnancies, without ultrasound anomalies, in a real-life NIPS clinical setting) was conducted. An outcome was available for all cases in the validation cohort and for 521 in the clinical cohort. Cell-free DNA from plasma samples was sequenced using the Ion Proton sequencer, and sequencing data were analyzed using the open-access software, WISECONDOR. Performance metrics for detection 10 of trisomies 21, 18 and 13 were calculated based on either fetal karyotype result or clinical data collected at birth. We also evaluated the failure rate and compared three methods of fetal fraction quantification (RASSF1A assay, and DEFRAG and SANEFALCON software). Then, regarding the CMA study, we retrospectively included all fetuses with IUGR referred for prenatal testing and studied by rapid fluorescence in situ hybridization (FISH), karyotype, and CMA. Results :In the NIPS study, results from both cohorts were consistent and their gestational age was not significantly different, so their data were combined to increase the sample size for analysis. Sensitivities and specificities, respectively, were as follows: for trisomy 21, 98.3% (95% CI, 93.5–99.7%) and 99.9% (95% CI, 99.4–100%); for trisomy 18, 96.7% (95% CI, 80.9–99.8%) and 100% (95% CI, 99.6–100%); and for trisomy 13, 94.1% (95% CI, 69.2–99.7%) and 100% (95% CI, 99.6–100%). Our failure rate was 1.2% initially and as low as 0.6% after retesting some of the failed samples. Fetal fraction estimation by the RASSF1A assay was consistent with DEFRAG results, and both were adequate for routine diagnosis. Among the 162 IUGR fetuses (78 associated and 84 isolated IUGR) included in the CMA study, 15 had an abnormal FISH result: 10 associated and five isolated fetal IUGRs. Among the 143 fetuses studied by CMA, 10 (7%) presented pathogenic copy number variations (CNVs). All 10 were in the associated fetal IUGR group (10/65 or 15.4%; 95% confidence interval [CI]: 8.4%‐26.2%) versus 0/78 in the isolated fetal IUGR group (95% CI: 0%‐5.6%). Six fetuses (4.2%) carried variants of unknown significance (VOUS) (three associated and three isolated fetal IUGRs). Conclusion: We described one of the largest studies evaluating Ion Proton-based NIPS and the first clinical study reporting pregnancy outcome in a large series of patients. This platform is highly efficient in detecting the three most common trisomies. Our protocol is robust and can be implemented easily in any medical genetics’ laboratory. Our second study highlighted the added value of CMA in the case of associated fetal IUGR with an incremental yield of 6.1% (4/65) over karyotyping. No pathogenic CNVs were reported in the isolated fetal IUGR group. Could NIPS supplant CMA in isolated fetal IUGR? The development of the NIPS test has reduced prenatal invasive testing and therefore its complications [...].