Submitted by Geandra Rodrigues (geandrar@gmail.com) on 2018-03-27T12:00:11Z No. of bitstreams: 0 / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2018-03-27T13:49:39Z (GMT) No. of bitstreams: 0 / Made available in DSpace on 2018-03-27T13:49:39Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-03-02 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O queijo Minas Frescal (QMF) representa um dos queijos mais consumidos no Brasil. Diversos fatores do seu processamento influenciam suas características microbiológicas, e consequentemente, sua qualidade e propriedades organolépticas. Seu alto teor de umidade e os riscos de contaminação durante a cadeia produtiva favorecem a ocorrência de microrganismos contaminantes, muitas vezes apresentando resistência aos antimicrobianos. Dessa forma, do ponto de vista da segurança alimentar e frente ao crescente fenômeno da resistência bacteriana às drogas, torna-se importante a investigação sobre a estrutura da comunidade bacteriana em QMF, bem como a avaliação da ocorrência de marcadores genéticos microbianos relacionados à resistência a drogas e seu potencial de mobilização. Neste estudo foram obtidas 5 amostras de um mesmo lote de 7 marcas de QMF identificadas de A a G, totalizando 35 amostras. Após a extração de DNA total microbiano das amostras, foram utilizadas abordagens de DNA fingerprint, pela amplificação de sequências palindrômicas extragênicas repetitivas (rep-PCR) para avaliação comparativa da similaridade da estrutura global da comunidade bacteriana. Posteriormente, PCR-DGGE foi utilizada para avaliar o perfil e a riqueza das amostras com relação a grupos de bactérias láticas. Matrizes de similaridade foram obtidas utilizando o método de agrupamento UPGMA. Os resultados obtidos pela técnica de rep-PCR revelaram que as amostras de queijos foram claramente agrupadas de acordo com as suas respectivas marcas. Além disso, perfis semelhantes entre amostras de marcas diferentes foram observados, indicando a presença de um núcleo microbiano comum. As amostras avaliadas também foram agrupadas de acordo com suas respectivas marcas de fabricação de acordo com os padrões de DGGE obtidos para bactérias láticas. A elevada similaridade entre a maioria das amostras do mesmo lote obtida nas técnicas de fingerprint sugere a reprodutibilidade e aplicabilidade das técnicas, e controle no processamento dos queijos ao longo da cadeia produtiva. Para a avaliação do resistoma clínico, a presença de 40 marcadores de resistência a diferentes classes de antibióticos foi avaliada por reação de PCR. Um núcleo comum de marcadores genéticos em todas as marcas foi detectado, associado à resistência aos beta-lactâmicos, tetraciclinas, quinolonas e sulfonamidas. Outros marcadores, incluindo aqueles relacionados a bombas de efluxo e resistência aos aminoglicosídeos, também foram observados. Integrons de classes 1 e 2 foram detectados, respectivamente, em 77% e 97% das amostras. As diferentes amostras de QMF puderam ser agrupadas de acordo com seu perfil de marcadores genéticos de resistência aos antimicrobianos, o que sugere epidemiologia peculiar que pode estar relacionada a qualidade e aos níveis de contaminação dos queijos ao longo da cadeia produtiva. Em conjunto, os dados sugerem que embora a cadeia produtiva do QMF seja controlada na indústria, riscos sanitários são inerentes pela contaminação dos queijos por bactérias putativas resistentes a antimicrobianos. Como um todo, os dados apontam para a necessidade de discussão dos parâmetros de qualidade microbiológica na produção, armazenamento e distribuição de QMF. Além disso, a detecção de integrons de classe 1 e 2 levanta questões a respeito do potencial de transferência horizontal de genes de resistência para a microbiota humana através do consumo destes alimentos. / Minas Frescal cheese (QMF) represents one of the most consumed cheeses in the country. Several factors of its processing influence its microbiological characteristics, and, consequently, its quality and properties. Its high moisture content and the risks of contamination during the production chain favor the occurrence of contaminating microorganisms, often presenting antimicrobial resistance. Thus, from the point of view of food safety and the growing phenomenon of bacterial resistance to drugs, it is important to investigate the structure of the bacterial community in Minas Frescal cheese, as well as the evaluation of the occurrence of microbial genetic markers related to drug resistance and its potential for mobilization. In this study 5 samples from the same batch of 7 brands of Minas Frescal cheeses were identified from A to G, totaling 35 samples. After the extraction of total DNA from the samples, DNA fingerprint approaches were used, by the amplification of repetitive extragenic palindromic sequences (rep-PCR) to evaluate the similarity of the global structure of the bacterial community. Afterwards, PCR-DGGE was used to evaluate the profile and richness of the samples in relation to groups of lactic bacteria. Similarity matrices were obtained using the UPGMA clustering method. The results obtained by the rep-PCR technique revealed that the cheese samples were clearly brand-clustered. In addition, similar profiles among samples of different brands were observed, indicating the presence of a common microbial nucleus. The evaluated samples were also separated according to their respective manufacturing brands by the DGGE for lactic acid bacteria. The high similarity among the majority of the samples from the same batch obtained in the fingeprint techniques suggests the reproducibility and applicability of the techniques, and control in the cheese processing along the production chain. For the evaluation of clinical resistance, the presence of resistance markers to different classes of antibiotics was evaluated by PCR reaction. A common core of genetic markers was detected, associated with resistance to beta-lactams, tetracyclines, quinolones and sulfonamides. Other markers, including those related to efflux pumps and aminoglycoside resistance, have also been observed, but not in all brands. Integrons of classes 1 and 2 were detected, respectively, in 77% and 97% of the samples. The different QMF samples could be grouped according to their profile of genetic markers of antimicrobial resistance, which suggests peculiar epidemiology that may be related to the quality and levels of contamination of the cheeses along the production chain. Taken together, the data suggest that although the productive chain of QMF is controlled in the industry, health risks are inherent in the contamination of cheeses by putative antimicrobial resistant bacteria. As a whole, the data point to the need to discuss the parameters of microbiological quality in the production, storage and distribution of QMF. In addition, the detection of class 1 and 2 integrons raises questions about the potential for horizontal transfer of resistance genes to the human microbiota through the consumption of these foods.

Adoption of no-till (NT) and reduced tillage management is widespread on the Canadian prairies and together form the basic platform of soil management upon which most crop production is based. Elimination of tillage in cropping systems changes the physical and chemical characteristics of the soil profile and can affect crop growth and ultimately yield. As such, understanding how soil biota, as drivers of nutrient turnover, adapt to NT is important for maximizing crop productivity and mitigating environmental damage in agroecosystems. This work aims to achieve a greater understanding of microbial community structure and function in long-term NT versus conventionally tilled (CT) soils. Community phospholipid and DNA fingerprinting did not reveal any consistent tillage-induced shifts in microbial community structure, but demonstrated a clear influence of depth within the soil profile. While tillage did not result in broad changes in the community structure, total, bacterial and fungal biomass was consistently greater near the surface of NT soils. Further examination at one site near Swift Current, SK revealed differences in microbial biomass and community structure in NT and CT in field-formed aggregate size fractions. Measurement of mineralization and nitrification at the same site indicated that differences in the early-season turnover of N may be related to physical rather than microbial differences in NT and CT soils. Potential nitrification was higher prior to seeding than mid-season, was not affected by tillage and was correlated with ammonia oxidizer population size of archaea, but not bacteria. This work indicates that edaphic soil properties and spatial distribution of resources in the soil profile, rather than tillage management, are the primary factors driving microbial community structure in these soils.

Adoption of no-till (NT) and reduced tillage management is widespread on the Canadian prairies and together form the basic platform of soil management upon which most crop production is based. Elimination of tillage in cropping systems changes the physical and chemical characteristics of the soil profile and can affect crop growth and ultimately yield. As such, understanding how soil biota, as drivers of nutrient turnover, adapt to NT is important for maximizing crop productivity and mitigating environmental damage in agroecosystems. This work aims to achieve a greater understanding of microbial community structure and function in long-term NT versus conventionally tilled (CT) soils. Community phospholipid and DNA fingerprinting did not reveal any consistent tillage-induced shifts in microbial community structure, but demonstrated a clear influence of depth within the soil profile. While tillage did not result in broad changes in the community structure, total, bacterial and fungal biomass was consistently greater near the surface of NT soils. Further examination at one site near Swift Current, SK revealed differences in microbial biomass and community structure in NT and CT in field-formed aggregate size fractions. Measurement of mineralization and nitrification at the same site indicated that differences in the early-season turnover of N may be related to physical rather than microbial differences in NT and CT soils. Potential nitrification was higher prior to seeding than mid-season, was not affected by tillage and was correlated with ammonia oxidizer population size of archaea, but not bacteria. This work indicates that edaphic soil properties and spatial distribution of resources in the soil profile, rather than tillage management, are the primary factors driving microbial community structure in these soils.

Beluga whales in Canadian waters are subdivided into at least six genetically distinct stocks maintained by geographic separation and philopatry to estuaries in summer. Belugas in eastern and western Hudson Bay have previously been shown to be compose genetically distinct populations using mitochondrial restriction analysis. It is not known whether these stocks are further subdivided on the basis of specific estuarine use. Mitochondrial DNA control region sequences were used to investigate variation among belugas sampled at several sites along eastern Hudson Bay, Hudson Strait and Ungava Bay. 320 bp were sequenced, including the highly variable 5' region of control region, in 126 belugas. 17 variable sites and 17 haplotypes, which clustered into 2 related groups, were detected among the whales sequenced. Haplotypes of group A were found mostly in eastern Hudson Bay sites, while B group haplotypes were predominant in northern populations. Significant differences in frequencies of haplotype groups were found between eastern Hudson Bay and Southern Hudson Strait/Ungava Bay populations, indicating they are genetically distinct populations. Haplotype distribution patterns also suggested possible differences between belugas using different estuaries along eastern Hudson Bay. The presence of both groups in each population indicated some exchange of individuals between populations, and/or between eastern and western Hudson Bay. Multilocus DNA fingerprinting was used to investigate the extent of gene flow between eastern and western Hudson Bay belugas via interbreeding on common wintering grounds in Hudson Strait. Belugas from St. Lawrence estuary and the Mackenzie Delta were also analyzed to measure their genetic relatedness to Hudson Bay whales as well as for purposes of comparison to earlier fingerprinting analyses. While results supported lower genetic diversity within the St. Lawrence population, the range of bandsharing within and between populations was otherwise low (0.09 -0.17 for Jeffreys 33.15 and 0.12-0.22 for Jeffreys 33.6). Mantel tests showed differences among St. Lawrence, Hudson Bay, and Mackenzie Delta populations, but not within Hudson Bay. The conflicting nature of the data did not allow conclusions regarding gene flow. Therefore, DNA fingerprinting was not considered to have provided sufficient resolution in addressing this issue. / Thesis / Master of Science (MS)

In der vorliegenden Arbeit wurden 2460 Viertelgemelksproben aus 16 hessischen Milcherzeugerbetrieben untersucht. 115 S. canis-Isolate konnten gefunden und auf ihre morphologischen, biochemischen und bei molekularbiologischen Eigenschaften untersucht werden. Die Isolate stammten von Viertelgemelksproben bzw. Tankproben, die zu einem oder mehreren Zeitpunkten in den Betrieben genommen wurden. Die Untersuchung der biochemischen Eigenschaften erbrachte 24 verschiedene Reaktionsmuster. Der Vergleich dieser 24 Biotypen mit einem S. canis-Referenzstamm mittels tDNA-PCR und 16S-RNA-PCR ergab eine völlige Übereinstimmung (100%) und damit eine sichere Spezies-Identifizierung. Zur Aufklärung epidemiologischer Zusammenhänge und zur Intra-Spezies-Identifizierung wurde von allen 115 Isolaten mittels PFGE nach Makrorestriktionsverdau mit SmaI ein DNA-Fingerprint erstellt. Dabei ergaben sich 21 verschiedene Restriktionsmuster. Von den 21 nach Makrorestriktion mit Sma I und anschließender PFGE unterscheidbaren Restriktionsmustern wurde je ein Isolat zur Bestimmung der Differenzierungsfähigkeit der Restriktionsenzyme Cla I und Apa I sowie der RAPD-PCR weitergehend untersucht. Für die Beurteilung epidemiologischer Zusammenhänge bei S. canis erwies sich die PFGE nach Makrorestriktion mittels Sma I als die differenzierteste Variante. Die mittels PFGE nach Makrorestriktionsverdau mit Sma I durchgeführten Untersuchungen der 115 Isolate zeigten, dass zu einem Probennahme-Termin gewonnene Isolate identisch waren; vom gleichen Betrieb zu unterschiedlichen Zeiten entnommene Proben zeigten z.T. deutliche Unterschiede, und bei Isolaten von verschiedenen Betrieben konnten keine Verwandtschaftsbeziehungen nachgewiesen werden. Aufgrund dieser genotypischen Eigenschaften der Kulturen konnte gezeigt werden, dass es sich bei durch S. canis verursachte Mastitiden um ein infektiöses Bestandsproblem handelt, bei dem der Erreger von Viertel zu Viertel und von Kuh zu Kuh übertragen wird.

In der vorliegenden Arbeit wurden 2460 Viertelgemelksproben aus 16 hessischen Milcherzeugerbetrieben untersucht. 115 S. canis-Isolate konnten gefunden und auf ihre morphologischen, biochemischen und bei molekularbiologischen Eigenschaften untersucht werden. Die Isolate stammten von Viertelgemelksproben bzw. Tankproben, die zu einem oder mehreren Zeitpunkten in den Betrieben genommen wurden. Die Untersuchung der biochemischen Eigenschaften erbrachte 24 verschiedene Reaktionsmuster. Der Vergleich dieser 24 Biotypen mit einem S. canis-Referenzstamm mittels tDNA-PCR und 16S-RNA-PCR ergab eine völlige Übereinstimmung (100%) und damit eine sichere Spezies-Identifizierung. Zur Aufklärung epidemiologischer Zusammenhänge und zur Intra-Spezies-Identifizierung wurde von allen 115 Isolaten mittels PFGE nach Makrorestriktionsverdau mit SmaI ein DNA-Fingerprint erstellt. Dabei ergaben sich 21 verschiedene Restriktionsmuster. Von den 21 nach Makrorestriktion mit Sma I und anschließender PFGE unterscheidbaren Restriktionsmustern wurde je ein Isolat zur Bestimmung der Differenzierungsfähigkeit der Restriktionsenzyme Cla I und Apa I sowie der RAPD-PCR weitergehend untersucht. Für die Beurteilung epidemiologischer Zusammenhänge bei S. canis erwies sich die PFGE nach Makrorestriktion mittels Sma I als die differenzierteste Variante. Die mittels PFGE nach Makrorestriktionsverdau mit Sma I durchgeführten Untersuchungen der 115 Isolate zeigten, dass zu einem Probennahme-Termin gewonnene Isolate identisch waren; vom gleichen Betrieb zu unterschiedlichen Zeiten entnommene Proben zeigten z.T. deutliche Unterschiede, und bei Isolaten von verschiedenen Betrieben konnten keine Verwandtschaftsbeziehungen nachgewiesen werden. Aufgrund dieser genotypischen Eigenschaften der Kulturen konnte gezeigt werden, dass es sich bei durch S. canis verursachte Mastitiden um ein infektiöses Bestandsproblem handelt, bei dem der Erreger von Viertel zu Viertel und von Kuh zu Kuh übertragen wird.

Cette thèse précise les conditions pour introduire une sélection sur apparentés de caractères de qualité assistée par empreintes génétiques dans un programme de sélection massale sur la croissance chez la truite arc-enciel. Ce travail confirme l’intérêt à maîtriser l’effet maternel avec une doublement de l’héritabilité du poids à 70 g (0,36 vs 0,16), des estimations d’héritabilités des caractères mesurés de valeurs intermédiaires (0,37-0,54), l’intérêt d’utiliser la résiduelle des parties mesurées régressées linéairement au poids vif pour une sélection indépendante du poids pour les rendements, l’intérêt à remplacer la mesure du rendement au filetage par celle du rendement en carcasse éviscérée-étêtée plus héritable et très corrélé au rendement au filetage (0,97),la corrélation génétique élevée entre les rendements avec le ratio échographique e8/e23 (0,72-0,85) permettant une sélection sur candidats plus efficace qu’une sélection sur apparenté sur le rendement lui-même, des précisions élevées des valeurs génétiques (0.63–0.82) malgré très peu d’individus par famille (3,5-4) et une efficacité surestimée d’une sélection sur apparentés pré-sélectionnés (de 14 % à 62 %). Les conclusions sont qu’il est possible d’introduire une sélection sur apparentés dans un programme de sélection massale avec des gains génétiques au moins équivalents à ceux attendus en sélection familiale classique avec familles élevées initialement de façon séparées / The thesis aims at describing conditions to improve mass selection on growth by sib-selection on quality traits assisted by DNA parentage assignment in rainbow trout. Main results are that the control and management of differences in egg size between dams doubles heritability of body weight at 70 g (0.36 vs 0.16), heritabilities of quality traits are intermediates (0.37-0.54), the opportunity to use the residual of the body part (e.g. fillet weight) regressed to the whole body weight to select independently of body weight for ratios, the higher efficiency to replace fillet yield by deheaded and gutted carcass weight more heritable and highly genetically correlated to fillet yield (0.97), the possibility to use e8/e23ultrasound measures to predict genetically yields (0.72-0.85) allowing a direct selection on breeding candidates more efficient than in using sib information, a surprising intermediate to high accuracy of EBV even with a very low mean number of sib per full-sib family (3.5-4) and the medium to high overestimation of EBV when using pre- selected sibs (14 % to 62 %). The general conclusion are that sib-selection on quality traits can be introduced in mass selection with at least similar expected genetic gains than in traditional breeding design based on families reared initially separated