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Introdução: A dormência é caracterizada pela incapacidade de sementes viáveis germinarem em condições favoráveis, sendo causada por fatores genéticos e/ou ambientais. A mamona (Ricinus communis Linné), cuja dormência não apresenta causa definida, é uma planta da família Euphorbiaceae. O óleo extraído de suas sementes apresenta diversas caraterísticas interessantes para aplicação industrial, sendo comercializado com alto valor agregado. Objetivo: Este estudo tem como objetivo investigar as causas da dormência em sementes de mamona e identificar genes relacionados ao fenômeno. Materiais e métodos: Inicialmente realizamos uma revisão de literatura para identificar genes relacionados a dormência em Arabidopsis thaliana, e então uma busca pelas sequências proteicas no banco de dados do TAIR. Após a seleção dos genes, utilizamos o banco de dados Phytozome para realizar um BLASTP no genoma de mamona, selecionando as sequências homólogas. Com a seleção das sequências, realizamos uma análise filogenética para montagem de árvore NJ e análise de motifs, identificando genes relacionados à dormência em mamona. Em seguida, realizamos uma curva de embebição com sementes escarificadas e não-escarificadas da Cultivar BRS Energia, que apresenta dormência, para verificar se o tegumento representa uma barreira à absorção de água. Paralelamente, preparamos extrato aquoso do tegumento, que foi utilizado em ensaio para verificar seu efeito sobre a germinação de sementes de alface. Resultados: Na análise filogenética, foram identificados 17 genes homólogos aos genes relacionados à dormência em Arabidopsis, indicando a possibilidade de dormência fisiológica na mamona. A curva de embebição demonstrou que as sementes dormentes atingem a fase II de embebição, descartando-se a possibilidade de dormência física. O extrato possuiu efeito inibitório na germinação de sementes de alface, que é atribuída a substâncias presentes no tegumento. Conclusão: Os resultados obtidos indicam a possibilidade de dormência fisiológica/química nas sementes de Ricinus communis. Com esse trabalho, esperamos contribuir em futuros estudos voltados à dormência na espécie.

Osmopriming of seeds can increase seed vigour, allowing faster germination and field emergence, especially under adverse field conditions. The restricted imbibition during priming treatments reactivates metabolism although germination will be prevented. In order to identify and characterize genes that are involved in the improvement of vigour of Poincianella pyramidalis (Catingueira) seeds upon priming, we produced two different Suppression Subtractive Hybridization (SSH) cDNA libraries. These were a Forward and Reverse subtraction of cDNA samples isolated from untreated dry seeds (unprimed) and osmoprimed seeds. The two different libraries were sequenced by Illumina GAII next generation sequencing, resulting in almost 20 million reads that could be assigned to 5298 different contigs. Of these, 999 were only found in the unprimed seeds library and 2711 were specific for osmoprimed seeds. The contigs were annotated and subjected to gene set enrichment analysis and differential expression of several genes was confirmed by qRT-PCR analysis. The identified differentially expressed genes might play an important role in vigour improvement upon priming and may be potential markers for tolerance to water stress in P. pyramidalis seeds.

Ricinus communis L. is an important oilseed crop species which is mostly cultivated in Brazil by family farmers in the northeastern semiarid region. This species is renowned by the quality and properties of the oil extracted from its seeds, which can be used for a diverse range of industrial applications. Furthermore, this species is considered to be tolerant to abiotic stresses such as drought stress. The use of post-harvest and pre-germination treatments on mature seeds can enhance their viability and vigor. In this context, the objective of this study was to investigate the effect of osmoconditioning on R. communis L. seed germinability parameters and events related to the reactivation of cell cycle, i.e . tubulin accumulation and microtubular cytoskeleton configurations. Osmoconditioning was applied to both fresh-harvested (FH) and dry seeds (DS), originating fresh-harvested osmoconditioned (FHO) and dry osmoconditioned (DO) seeds. In general, osmoconditioning promoted seed priming by enhancing seed germinability of FHO and DO seeds by means of better germination speed and uniformity, except for a decrease of uniformity in FHO seeds. Immunocytochemical and immunohistochemical analysis revealed an increasing accumulation of tubulin in FHO and DO seeds, followed by the accumulation of tubulin granules in DO seeds. Mitotic microtubules were only visible in FHO and DO seeds after radicle protrusion and, therefore, seed germination occurred solely by cell elongation, whereas FH and DS seeds required cell cycle reactivation and cell proliferation during imbibition, i.e . prior to radicle protrusion or germination per se . The prompt activation of microtubular cytoskeleton synthesis in seeds that were osmoconditioned might be a possible strategy of the species for faster and more uniform and vigorous seed germination and seedling establishment and, therefore, better survival during periods of drought in harsh environments. These findings provide important insights into the mechanism underlying plant adaptation to harsh environmental conditions, which would be very helpful to develop germination and seedling production protocols to obtain faster and more vigorous and uniform germination seedling stand for the sustainable exploitation of R. communis by small family farmers under the typical harsh conditions of the semiarid regions worldwide.

Metabolomics provides vital information for the understanding of biological processes and has been vastly applied in plant studies. Several metabolite-profiling studies have correlated physiological events, such as germination or seedling establishment, with metabolic and molecular changes under different environmental conditions. Castor bean displays high plasticity during initial vegetative growth, which is reflected in the metabolome of the seeds and seedlings. In general, several metabolite-profiling techniques are required to obtain a complete response in terms of metabolism plasticity of the studied biological system. Carbohydrates, amino acids, and organic acids have been measured in castor bean seeds and seedlings by nuclear magnetic resonance, gas chromatography coupled to a quadrupole time of flight mass spectrometry (GC-TOF-MS), as well as by high-performance liquid chromatography (HPLC). Fatty acids and some secondary metabolites have been quantified in castor bean seeds and seedlings by gas chromatography coupled to a triple-axis detector (GC-MS). In this chapter, we initially discuss how metabolomics studies suggested a possible role of gamma-aminobutyric acid (GABA) accumulation during early imbibitions and seedling establishment. Later, we consider a specific metabolic signature of castor bean: a shift in carbon–nitrogen metabolism as its main biochemical response to high temperatures. This metabolic shift is usually associated with adjusted growth, and it is likely involved in maintaining cellular homeostasis under heat stress. The castor bean metabolome has been vastly investigated, especially with regard to its ability to respond to external stimuli. These results might help us understand the molecular requirements for vigorous castor bean seed germination and seedling growth under different environmental conditions.

Abiotic stresses are the most important environmental factors that negatively affects crop production worldwide, whereas germination is the most critical phase in the plants life cycle. Water availability is essential for proper seed imbibition and vigorous germination and often scarce for Ricinus communis (castor bean) cultivated in semiarid regions of Brazil. Water restriction generally leads to the generation of reactive oxygen species (ROS) and one of the major families of enzymes that counteract ROS are superoxide dismutases (SODs). The objective of this study was to identify and characterize the SOD gene family of R. communis by phylogenetic, biochemical and gene expression analysis and link it to the effect of osmoconditioning on seed vigor. We characterized the effect of osmotic conditioning treatments of seeds of two contrasting R. communis cultivars, EBDA MPA34 and PARAGUACU, on germination performance. We also searched the R. communis genome for SOD genes and analyzed them phylogenetically and evaluated their activity and expression in embryos of seeds imbibed under different PEG concentrations (water restriction stress). We could show that some of these treatments improved germinability and vigor of seeds and seedlings in a cultivar-dependent manner. Additionally, R. communis seeds turned out to be very sensitive to the inhibition of germination by osmotic stress even at relative low concentrations. Eight SOD genes were found in the R. communis genome and classified based on phylogenetic analysis, protein domains and prediction of subcellular localization. SOD gene expression was mostly induced during seed imbibition, although their expression was generally inhibited in osmotic potentials below −0.6 MPa and showed partly different gene expression profiles between contrasting cultivars. Cv. PARAGUACU showed increased seed vigor after osmoconditioning as compared to cv. EBDA MPA34 which could be related to higher responses of some cv. PARAGUACU SOD genes to stress. Our results indicate that SOD genes might be of special interest for R. communis genetic engineering aimed at the increase of production in semiarid regions.

The newly described Streptococcus pneumoniae serotype 6C accounted for 2.3% (16/709) of meningitis cases and 3.2% (3/95) of nasopharyngeal isolates from healthy individuals in Brazil. The strains were multidrug resistant (18.8%) and genetically diverse. Despite low serotype 6C prevalence, continuous surveillance is necessary to guide vaccine strategies.

SummaryBackgroundOver recent decades, a resurgence of invasive group A streptococcal (GAS) infections has been observed; GAS remains a rare cause of pyogenic meningitis. We report herein population-based findings of long-term surveillance for GAS meningitis in Salvador, Brazil, and estimate the overall burden of invasive GAS infections.MethodsFrom February 1996 to February 2007 we conducted active surveillance for GAS meningitis in the state reference hospital for infectious diseases in Salvador, Brazil. Data on clinical presentation, laboratory records, and outcome were collected through interviews and chart review. GAS isolates were evaluated for antimicrobial susceptibility and emm type.ResultsWe identified 20 cases of GAS meningitis, which accounted for 0.9% of all culture-proven bacterial meningitis in the study period. The mean annual incidence of GAS meningitis was 0.03 cases per 100000 population in metropolitan Salvador and peaked in children

Even though the major benefit of arbuscular mycorrhizae is the increased uptake of phosphate from the soil solution and translocation to the plant, changes in the activity of enzymes involved in nitrogen (N) metabolism have been detected in mycorrhizal roots. Using differential display of reverse-transcripts of tobacco roots not-inoculated or inoculated with Glomus intraradices (Gi), we have cloned two partial cDNAs (NtGi2 and NtGi3). The presence of a conserved CobW/HypB/UreG domain and phylogenetic analyses suggest that NtGi2 and NtGi3 encode isoforms of urease accessory protein G (ureG) highly similar to ureG from fungi. The steady state levels of the putative ureG transcripts were shown to be higher in roots colonized by Gi, as compared to non-mycorrhizal controls. Urease activities were also determined in tobacco roots inoculated with Glomus clarum (Gc) or Gi and grown in substrate containing 50, 100 or 150 mg N kg-1 in the form of ammonium sulfate (N-AMS) or urea (N-URE). Urease activities were shown to be induced in mycorrhizal roots fertilized with 100 mg N-AMS kg-1. In Gc-colonized roots fertilized with N-URE, induction of urease activities was observed at the lowest N concentration. In contrast, at the highest N-URE concentration, suppression of urease activities was observed in Gc and Gi-colonized roots, as compared to non-mycorrhizal controls. Urease activities in roots were modulated by soil N availability and source, and arbuscular mycorrhizal fungal inoculation.Muito embora o maior benefício de micorrizas arbusculares seja o incremento na absorção de fosfato da solução do solo e translocação para a planta, alterações nas atividades de enzimas envolvidas no metabolismo de nitrogênio (N) têm sido detectadas em raízes micorrizadas. Usando "differential display of reverse-transcripts" de raízes de tabaco não-inoculadas ou inoculadas com Glomus intraradices (Gi), dois cDNAs parciais (NtGi2 e NtGi3) foram clonados. A presença de um domínio CobW/HypB/UreG conservado e a análise filogenética sugerem que NtGi2 e NtGi3 codificam isoformas de proteínas acessórias da urease G (ureG) altamente similares a ureG de fungos. Os níveis de transcritos dos genes ureG putativos foram mais elevados em raízes colonizadas por Gi, em relação ao controle não-micorrizado. As atividades de urease foram determinadas em raízes de tabaco inoculadas com Glomus clarum (Gc) ou Gi e cultivadas em substrato contendo 50, 100 ou 150 mg N kg-¹, na forma de sulfato de amônio (N-AMS) ou uréia (N-URE). As atividades de ureases foram induzidas em raízes micorrizadas cultivadas com 100 mg N-AMS kg-¹. Em raízes colonizadas por Gc e cultivadas com N-URE, a indução das atividades de urease foi observada na concentração mais baixa de N. Em contraste, na concentração mais elevada de N-URE, supressão das atividades de urease em raízes colonizadas por Gc e Gi, em relação aos controles não-micorrizados, foi observada. As atividades de urease nas raízes foram moduladas pela disponibilidade e fonte de N no solo, e pela inoculação com fungos micorrízicos arbusculares.

Background Few studies of dengue have shown group-level associations between demographic, socioeconomic, or geographic characteristics and the spatial distribution of dengue within small urban areas. This study aimed to examine whether specific characteristics of an urban slum community were associated with the risk of dengue disease. Methodology/Principal Findings From 01/2009 to 12/2010, we conducted enhanced, community-based surveillance in the only public emergency unit in a slum in Salvador, Brazil to identify acute febrile illness (AFI) patients with laboratory evidence of dengue infection. Patient households were geocoded within census tracts (CTs). Demographic, socioeconomic, and geographical data were obtained from the 2010 national census. Associations between CTs characteristics and the spatial risk of both dengue and non-dengue AFI were assessed by Poisson log-normal and conditional auto-regressive models (CAR). We identified 651 (22.0%) dengue cases among 2,962 AFI patients. Estimated risk of symptomatic dengue was 21.3 and 70.2 cases per 10,000 inhabitants in 2009 and 2010, respectively. All the four dengue serotypes were identified, but DENV2 predominated (DENV1: 8.1%; DENV2: 90.7%; DENV3: 0.4%; DENV4: 0.8%). Multivariable CAR regression analysis showed increased dengue risk in CTs with poorer inhabitants (RR: 1.02 for each percent increase in the frequency of families earning ≤1 times the minimum wage; 95% CI: 1.01-1.04), and decreased risk in CTs located farther from the health unit (RR: 0.87 for each 100 meter increase; 95% CI: 0.80-0.94). The same CTs characteristics were also associated with non-dengue AFI risk. Conclusions/Significance This study highlights the large burden of symptomatic dengue on individuals living in urban slums in Brazil. Lower neighborhood socioeconomic status was independently associated with increased risk of dengue, indicating that within slum communities with high levels of absolute poverty, factors associated with the social gradient influence dengue transmission. In addition, poor geographic access to health services may be a barrier to identifying both dengue and non-dengue AFI cases. Therefore, further spatial studies should account for this potential source of bias., Author Summary Dengue is influenced by the environment; however, few studies have investigated the relationship between neighborhood characteristics and the spatial distribution of dengue within small urban areas. We examined whether specific characteristics of an urban slum community were associated with dengue risk. From January 2009 to December 2010, we conducted community-based surveillance in a slum in Salvador, Brazil to identify patients with acute febrile illness (AFI) and to test them for dengue. We identified 651 (22.0%) patients with laboratory evidence of dengue infection among 2,962 AFI patients. All the four dengue serotypes were detected, but DENV2 predominated (DENV1 8.1%; DENV2 90.7%; DENV3 0.4%; DENV4 0.8%). Estimated risk of symptomatic dengue was 21.3 and 70.2 cases per 10,000 inhabitants in 2009 and 2010, respectively. We found that neighborhood poverty level and proximity to the health center were associated with higher risk of detection of dengue and other AFI. This study highlights the large burden of dengue in poor urban slums of Brazil and indicates that socioeconomic development could potentially mitigate risk factors for both dengue and non-dengue AFI cases. In addition, we found that residential proximity to a health care facility was associated with improved case detection. Therefore, further studies on disease distribution should consider household proximity to health care facilities when assessing risk.

Fetal hemoglobin (HbF), encoded by the HBG2 and HBG1 genes, is the best-known genetic modulator of sickle cell anemia, varying dramatically in concentration in the blood of these patients. This variation is partially associated with polymorphisms located in the promoter region of the HBG2 and HBG1 genes. In order to explore known and unknown polymorphisms in these genes, the sequences of their promoter regions were screened in sickle cell anemia patients and correlated with both their HbF levels and their betaS-globin haplotypes. Additionally, the sequences were compared with genes from 2 healthy groups, a reference one (N = 104) and an Afro-descendant one (N = 98), to identify polymorphisms linked to the ethnic background.The reference group was composed by healthy individuals from the general population. Four polymorphisms were identified in the promoter region of HBG2 and 8 in the promoter region of HBG1 among the studied groups. Four novel single nucleotide polymorphisms (SNP) located at positions -324, -317, -309 and -307 were identified in the reference group. A deletion located between -396 and -391 in the HBG2 promoter region and the SNP -271 C--T in the HBG1 promoter region were associated with the Central African Republic betaS-globin haplotype. In contrast, the -369 C--G and 309 A--G SNPs in the HBG2 promoter region were correlated to the Benin haplotype. The polymorphisms -396_-391 del HBG2, -369 SNP HBG2 and -271 SNP HBG1 correlated with HbF levels. Hence, we suggest an important role of HBG2 and HBG1 gene polymorphisms on the HbF synthesis.

Even though the major benefit of arbuscular mycorrhizae is the increased uptake of phosphate from the soil solution and translocation to the plant, changes in the activity of enzymes involved in nitrogen (N) metabolism have been detected in mycorrhizal roots. Using differential display of reverse-transcripts of tobacco roots not-inoculated or inoculated with Glomus intraradices (Gi), we have cloned two partial cDNAs (NtGi2 and NtGi3). The presence of a conserved CobW/HypB/UreG domain and phylogenetic analyses suggest that NtGi2 and NtGi3 encode isoforms of urease accessory protein G (ureG) highly similar to ureG from fungi. The steady state levels of the putative ureG transcripts were shown to be higher in roots colonized by Gi, as compared to non-mycorrhizal controls. Urease activities were also determined in tobacco roots inoculated with Glomus clarum (Gc) or Gi and grown in substrate containing 50, 100 or 150 mg N kg-1 in the form of ammonium sulfate (N-AMS) or urea (N-URE). Urease activities were shown to be induced in mycorrhizal roots fertilized with 100 mg N-AMS kg-1. In Gc-colonized roots fertilized with N-URE, induction of urease activities was observed at the lowest N concentration. In contrast, at the highest N-URE concentration, suppression of urease activities was observed in Gc and Gi-colonized roots, as compared to non-mycorrhizal controls. Urease activities in roots were modulated by soil N availability and source, and arbuscular mycorrhizal fungal inoculation. Muito embora o maior benefício de micorrizas arbusculares seja o incremento na absorção de fosfato da solução do solo e translocação para a planta, alterações nas atividades de enzimas envolvidas no metabolismo de nitrogênio (N) têm sido detectadas em raízes micorrizadas. Usando "differential display of reverse-transcripts" de raízes de tabaco não-inoculadas ou inoculadas com Glomus intraradices (Gi), dois cDNAs parciais (NtGi2 e NtGi3) foram clonados. A presença de um domínio CobW/HypB/UreG conservado e a análise filogenética sugerem que NtGi2 e NtGi3 codificam isoformas de proteínas acessórias da urease G (ureG) altamente similares a ureG de fungos. Os níveis de transcritos dos genes ureG putativos foram mais elevados em raízes colonizadas por Gi, em relação ao controle não-micorrizado. As atividades de urease foram determinadas em raízes de tabaco inoculadas com Glomus clarum (Gc) ou Gi e cultivadas em substrato contendo 50, 100 ou 150 mg N kg-¹, na forma de sulfato de amônio (N-AMS) ou uréia (N-URE). As atividades de ureases foram induzidas em raízes micorrizadas cultivadas com 100 mg N-AMS kg-¹. Em raízes colonizadas por Gc e cultivadas com N-URE, a indução das atividades de urease foi observada na concentração mais baixa de N. Em contraste, na concentração mais elevada de N-URE, supressão das atividades de urease em raízes colonizadas por Gc e Gi, em relação aos controles não-micorrizados, foi observada. As atividades de urease nas raízes foram moduladas pela disponibilidade e fonte de N no solo, e pela inoculação com fungos micorrízicos arbusculares.

A formação de micorrizas arbusculares é um processo regulado geneticamente e envolve alterações da expressão gênica da planta e do fungo. Poucos genes essenciais para o desenvolvimento da simbiose foram identificados até o presente. Para identificar genes possivelmente envolvidos no controle da colonização fúngica intrarradicular, o perfil de seqüências expressas em raízes de cana-de-açúcar colonizadas por Glomus clarum, em condições de baixo e alto fósforo (P), foi avaliado através do seqüenciamento em larga escala de ESTs e hibridização em macroarranjos de cDNAs. Plântulas micropropagadas de cana-de-açúcar foram plantadas em vasos contendo substrato composto por areia:vermiculita (3:1, vol/vol) e adubado com 20 ou 200 mg P kg-1 de substrato e inoculadas com Glomus clarum. A biomassa seca da parte aérea e a taxa de colonização micorrízica das raízes foram avaliadas no momento da colheita que foi feita oito ou doze semanas após o plantio. O RNA poliA foi extraído das raízes e utilizado para a síntese de cDNAs. Neste trabalho, quatro bibliotecas de cDNA e uma biblioteca subtrativa supressiva foram preparadas a partir de cDNAs de raízes de cana-de-açúcar colonizadas por G. clarum. Esta abordagem resultou na identificação de 1,925 ESTs as quais foram agrupadas em 1615 genes supostamente regulados pela micorrização. Estes clones foram analisados por Northern blots eletrônico, e aqueles que foram diferencialmente expressos foram submetidos a hibridização em macroarranjos de cDNAs. Um total de 386 genes foram arranjados em membranas de náilon e sua expressão avaliada por sondas sintetizadas a partir de cDNA extraído de raízes em diferentes condições. Dentre os genes com expressão diferencial significativa estatística em raízes colonizadas por G. clarum, em condições de baixo e alto P, foram detectados genes codificando proteínas putativamente envolvidas na percepção de moléculas sinais (receptor tipo quinase de proteína), transporte de íons (canais de íons), transdução de sinais (quinases de proteína e calmodulina), regulação da transcrição (fatores de transcrição), alterações de parede celular e citoesqueleto (extensina, arabinogalactanas, tubulinas), respostas de defesa e estresse (síntese de fitoalexinas e metalotioneínas), síntese de fitohormônios (nitrilase). Os dados sugerem que a transdução de sinais em MAs se dá através da fosforilação de proteínas e que a indução de respostas anti-oxidantes pode ser importante para o desenvolvimento da simbiose. Da mesma forma, os dados sugerem que a atividade de aspartatoproteases podem ser essenciais para o controle do crescimento fúngico intrarradicular. A caracterização desses genes e seus padrões de expressão em MAs poderá elucidar os mecanismos genéticos envolvidos no controle da simbiose. The formation of arbuscular mycorrhizae (AM) is a genetically regulated process and involves alteration in both plant and fungus gene expression. Few genes essential for the development of the symbioses have been identified so far. In order to identify genes possible involved in the intraradical fungal colonization control, the profiles of expressed sequences in sugarcane roots colonized roots with Glomus clarum at low or high phosphate (P) conditions were evaluated using large scale EST sequencing and cDNA macroarray hybridization. Sugarcane micropropagated seedlings were planted in pots containing sand:vermiculite (3:1, vol:vol), and fertilized with 20 or 200 mg P kg-1 substrate. Seedlings were inoculated with G. clarum and cultivated under greenhouse conditions for 8 or 12 weeks. After harvesting shoot dry weight and root colonization rates were evaluated. PolyA RNA was extracted from the root tissues and used for cDNA synthesis. In this work, one suppressive subtractive library and four cDNA libraries were synthesized using cDNA from sugarcane roots not-colonized or colonized by G. clarum, at low and high P conditions. This approach resulted in the identification of 1,925 ESTs which were clustered in 1,615 genes. The expression of these genes was evaluated using electronic Northern blots. A total of 386 genes with putative differential expression were spotted into nylon membranes in macroarrays and their expression profiles in roots under different conditions were evaluated using macroarray hybridization. Among the genes with statistically significant differential expression, in roots colonized by G. clarum grown at low or high P conditions, it was detected genes encoding proteins putatively involved in the perception of signal molecules (receptor-like protein kinase), ion transporte (channel-like protein precursor), signal transduction (protein kinases and calmodulin), transcriptional regulation (transcription factor), cell wall and cytoskeleton alterations (extensin, arabinogalactans, and tubulins), defense and stress responses (phythoalexins synthesis and metalothioneins), and phytohormone biosynthesis (nitrilases). Our data suggest that the signal transduction in AM occurs through protein phosphorylation, and that the induction of anti-oxidant responses may be important for the development of the symbioses. Additionally, the data suggest that aspartic-proteases might be essential for the control of intraradical fungal growth. Further characterization of these genes and their expression patterns in AM would contribute to elucidate the genetic mechanisms controlling the symbioses.

O papel das micorrizas arbusculares (MAs) no metabolismo de nitrogênio nos vegetais é pouco conhecido. Pouco se sabe também sobre os mecanismos que regulam a absorção, assimilação de nitrogênio em MAs. Recentemente, um gene altamente induzido em raízes de tabaco micorrizadas foi clonado e caracterizado. Esse gene codifica uma proteína acessória da urease (UreG). Sua regulação diferencial em MAs sugere que fungos micorrízicos arbusculares podem induzir alterações no metabolismo de nitrogênio nos vegetais, e/ou que ureases podem ser importantes no controle do desenvolvimento da simbiose. O objetivo deste trabalho foi verificar se ocorrem ou não alterações nas atividades de ureases em raízes de tabaco micorrizadas, e se essas alterações estão relacionadas com alterações do perfil de aminoácidos solúveis e com o crescimento fingico intrarradicular. Para isso, plântulas de tabaco foram inoculadas com Glomus clarum ou Glomus intraradices, e cultivadas em substrato esterilizado contendo 50, 100 ou 150 mg kg-1 de Nitrogênio, na forma de uréia ou sulfato de amônio. Oito semanas após a inoculação, foram avaliados: matéria seca da parte aérea e sistema radicular, colonização micorrízica intraradicular, atividades de ureases nas raízes, e concentração e perfil de aminoácidos solúveis na parte aérea. A colonização intraradicular por G. clarum teve aumento induzido pelo aumento das doses de nitrogênio. As atividades de ureases de plantas não-inoculadas aumentaram com o aumento da dose de N-uréia aplicada ao substrato. Quando cultivadas com 50 mg kg-1 de N-sulfato de amônio, as raízes de plantas inoculadas com G. clarum apresentaram atividades de ureases mais elevadas do que nos controles não-inoculados. Já, as raízes de plantas inoculadas com G. intraradices apresentaram maiores atividades de ureases, em relação aos controles não-inoculados, quando cultivadas com 100 mg kg-1 de N-sulfato de amônio. Com 150 mg kg-1 N-uréia, as atividades de ureases foram 3 vezes menores nas raízes micorrizadas do que nas não-micorrizadas. As atividades de ureases apresentaram correlação positiva e significativa com a colonização intrarradicular nos tratamentos com N-sulfato de amônio. A inoculação de plântulas com G. intraradices ou G. clarum resultou em diminuição da concentração de aminoácidos livres, em substrato com 50 mg kg-1 N-sulfato de amônio, em relação aos controles não inoculados. Em substrato 150 mg kg-1 N-uréia a concentração de aminoácidos solúveis aumentou aproximadamente 10 vezes, em relação aos controles não-inoculados. O aminoácido encontrado em maior quantidade na parte aérea de tabaco foi a glutamina. Alterações nos perfis de aminoácidos na parte aérea de plantas micorrizadas, em relação aos controles não micorrizados, foram observadas. Correlação significativa entre atividades de ureases nas raízes e concentração de aminoácidos solúveis na parte aérea não foi observada. Os dados obtidos neste trabalho sugerem que as atividades de ureases nas raízes, em condições de baixo N, podem estar relacionadas com a capacidade de colonização intrarradicular dos fungos, e que fungos micorrízicos arbusculares podem alterar o metabolismo de N nas plantas The role of arbuscular mycorrhizae (AM) in the nitrogen (N) metabolism of host plants is not understood, and the mechanisms regulating N uptake and assimilation in AM are not known. Recently, a gene highly induced in mycorrhizal tobacco roots was cloned and characterized. Its deduced amino acid sequence showed a high degree of homology to urease accessory proteins (UreG). The differential regulation of ureG in AM suggests that arbuscular mycorrhizal fungi may alter N metabolism in the host plants, and/or that ureases may be important in controlling symbiosis development. The aim of this work was to determine whether urease activities are altered in mycorrhizal roots, and whether these alterations are related to changes in the relative abundance of free amino acids in shoots, and/or intraradical fungaI growth. Tobacco seedlings were inoculated with Glomus clarum or Glomus intraradices, and grown in sterilized substrate containing 50, 100 or 150 mg N kg-1, as urea or ammonium sulphate. Eight weeks after inoculation, the following parameters were evaIuated: shoot and root dry weight, intraradical fungaI growth, urease activities in roots, and free amino acid concentration in shoots. IntraradicaI colonization by G. clarum was higher than by G. intraradices, at the different experimental conditions. Root coIonization by G. clarum and G. intraradices was induced in substrate with 50 and 100 mg N-urea kg-1. Urease activities in non-mycorrhizaI roots were higher at the highest N-urea concentration. When grown in 50 mg N-urea kg-1, urease activities in roots coIonized by G. clarum were higher than in non-mycorrhizaI controIs. In contrast, roots coIonized by G. intraradices showed higher urease activities than non-mycorrhizaI controIs when grown in 100 mg N-ammonium sulphate kg-1. Urease activities in mycorrhiza I roots grown in substrate containing 150 mg N-urea kg-1 were 3-fold Iower than in non-mycorrhizal controIs. A significant positive correlation between urease activities and intraradical fungaI growth was observed in roots grown with N-ammonium suIphate. InocuIation of tobacco seedlings with G. intraradices or G. clarum resuIted in decreased free amino acid concentration in the shoots, when grown with 50 mg N-ammonium suIphate kg-1, as compared to the not- inoculated controls. In mycorrhizaI roots grown in substrate with 150 mg N-urea kg

Submitted by MARCOS VINICIUS SILVA DE ANDRADE (mvandrade57@gmail.com) on 2022-07-02T12:59:32Z No. of bitstreams: 1 TESE Completa - Marcos Vinicius__VERSÃO FINAL.pdf: 4112661 bytes, checksum: b81289f7bc98803927d3e07e49341f73 (MD5) Approved for entry into archive by Delba Rosa (delba@ufba.br) on 2022-07-05T14:26:35Z (GMT) No. of bitstreams: 1 TESE Completa - Marcos Vinicius__VERSÃO FINAL.pdf: 4112661 bytes, checksum: b81289f7bc98803927d3e07e49341f73 (MD5) Made available in DSpace on 2022-07-05T14:26:35Z (GMT). No. of bitstreams: 1 TESE Completa - Marcos Vinicius__VERSÃO FINAL.pdf: 4112661 bytes, checksum: b81289f7bc98803927d3e07e49341f73 (MD5) Previous issue date: 2022-05-06 CAPES A Stevia rebaudiana (Bert.) Bertoni tem se destacado nos últimos anos devido a características adoçantes proporcionadas pela presença de glicosídeos de esteviol (StGly – Steviol Glycosides), principalmente em suas folhas. Em razão do seu rico perfil nutricional e fitoquímico, a estévia também fornece efeitos benéficos contra uma infinidade de condições de saúde, sendo assim, um material de interesse no campo das plantas com propriedades medicinais. Entretanto, o cultivo em larga escala da estévia enfrenta alguns obstáculos, uma vez que esta espécie é altamente responsiva a estímulos ambientais, entre os quais o fotoperíodo, é um dos mais relevantes, afetando em suas características morfofisiológicas e bioquímicas. Portanto, torna-se de fundamental importância compreender os mecanismos subjacentes às respostas da estévia a diferentes condições ambientais. Neste sentido, o presente estudo teve como objetivo (i) caracterizar o perfil morfoanatômico da germinação de sementes e enzimático de plântulas, além de (ii) realizar uma caracterização bioquímica e metabolômica do desenvolvimento de plantas de Stevia rebaudiana (Bert.) Bertoni cultivadas sob diferentes fotoperíodos. No CAPÍTULO 1, reportamos inicialmente sobre a análise do perfil morfofisiológico da germinação dos diferentes aquênios comumente produzidos pelas plantas, onde definimos um padrão de viabilidade de sementes de estévia baseado no teste de tetrazólio. Posteriormente buscamos investigar o perfil das enzimas antioxidantes em plântulas submetidas a diferentes condições de fotoperíodo. Os resultados demonstraram que as enzimas antioxidantes podem ser uma ferramenta importante para a avaliação do estado fisiológico de plântulas/plantas de estévia sob as diferentes condições testadas, fornecendo informações muito úteis para o cultivo baseado em sementes. No CAPÍTULO 2 reportamos sobre o crescimento de plantas de estévia sob diferentes regimes de fotoperíodo, onde avaliamos o conteúdo de StGly e a composição química geral por Ressonância Magnética Nuclear, atividade antioxidante (SOD / DPPH) e atividade antimicrobiana de extratos fitoquímicos, bem como o teor de fenólicos totais.de extratos obtidos de amostras de folha e do caule. Percebemos que, os fotoperíodos com menos de 12h de luz induzem modo reprodutivo precoce indesejado reduzindo o conteúdo de StGlys em suas folhas. Em contrapartida, dias longos (modo vegetativo) aumentaram a produção de StGly e outros compostos bioativos produzidos pelas plantas. Assim, fornecendo informações valiosas potencialmente servirão para o desenvolvimento de um protocolo de cultivo de estévia em larga escala. No CAPÍTULO 3, reportamos sobre o perfil fitoquímico com enfoque mais detalhado nos glicosídeos de esteviol (Esteviosídeo e Rebaudiosídeo A), em que promovemos a extração e quantificação destes metabólitos via RP–HPLC–UV (Reversed-Phase High-Performance Liquid Chromatography) para amostras de folha e caule de estévia. Como previsto, a folha sendo a matéria-prima de maior interesse na cultura da estévia, apresentou maiores teores dos StGly, sendo a síntese dos mesmos dependente das condições de luz testadas, onde plantas cultivadas sob 16/8h de luz apresentam um maior potencial para a síntese dos principais glicosídeos. Dentre tudo, vale destacar a presença de quantidades relevantes dos metabólitos de interesse também nas mostras de caule, podendo também apresentar propriedades potencialmente úteis para exploração futura. Stevia rebaudiana (Bert.) Bertoni has stood out in recent years due to the sweetening characteristics provided by the presence of steviol glycosides (StGly - Steviol Glycosides), mainly in its leaves. Due to its rich nutritional and phytochemical profile, stevia also provides beneficial effects against a multitude of health conditions, making it a biomaterial of interest in the field of plants with medicinal properties. However, the large-scale cultivation of stevia faces some obstacles, since this species is highly responsive to environmental stimuli, among which the photoperiod is one of the most relevant, affecting its morphophysiological and biochemical characteristics. Therefore, it is of fundamental importance to understand the mechanisms underlying stevia's responses to different environmental conditions. In this sense, the present thesis aimed to (i) characterize the morphoanatomical profile of seed germination and seedling enzymatic, in addition to (ii) perform a biochemical and metabolomic characterization of the development of Stevia rebaudiana (Bert.) Bertoni plants cultivated under different photoperiods. In CHAPTER 1, we report on the characterization of the morphophysiological profile of germination of the different achenes normally produced by the plants, where we could define a viability pattern for Stevia seeds based on tetrazolium test analysis. Further on, we report on the profile of antioxidant enzymes in seedlings submitted to different photoperiod conditions. The results showed that antioxidant enzymes can be an important tool for evaluating the physiological state of stevia seedlings/plants under the different conditions tested, providing very useful information for seed-based cultivation. In CHAPTER 2 we reported on the growth of stevia plants under different photoperiod regimes, where we evaluated the StGly content and the general chemical composition by Nuclear Magnetic Resonance, antioxidant activity (SOD / DPPH) and antimicrobial activity of phytochemical extracts, as well as the total phenolic content. of extracts obtained from leaf and stem samples. We noticed that photoperiods with less than 12h of light induce unwanted early reproductive mode by reducing the StGlys content in their leaves. In contrast, long days (vegetative mode) increased the production of StGly and other bioactive compounds produced by plants. Thus, providing valuable information will potentially serve for the development of a large-scale stevia cultivation protocol. In CHAPTER 3, we report on the phytochemical profile with a more detailed focus on steviol glycosides (Stevioside and Rebaudioside A), in which we promote the extraction and quantification of these metabolites via RP–HPLC–UV (Reversed-Phase High-Performance Liquid Chromatographic) for stevia leaf and stem samples. As expected, the leaf being the raw material of greatest interest in the stevia culture, showed higher levels of StGly, and their synthesis depends on the light conditions tested, where plants grown under 16/8h of light have a greater potential for the synthesis of the main glycosides. Among all, it is worth mentioning the presence of relevant amounts of the metabolites of interest also in the stem samples, which may also present potentially useful properties for future exploration.

Submitted by Programa de Pós-graduação em Biotecnologia (mebiotec.ufba@gmail.com) on 2017-09-14T14:16:25Z No. of bitstreams: 1 Dissertação final - Luciana Lima.pdf: 2423067 bytes, checksum: a4f37f089f064b722852c1a6565ea7de (MD5) Approved for entry into archive by Edvaldo Souza (edvaldosouza@ufba.br) on 2017-10-02T20:39:39Z (GMT) No. of bitstreams: 1 Dissertação final - Luciana Lima.pdf: 2423067 bytes, checksum: a4f37f089f064b722852c1a6565ea7de (MD5) Made available in DSpace on 2017-10-02T20:39:39Z (GMT). No. of bitstreams: 1 Dissertação final - Luciana Lima.pdf: 2423067 bytes, checksum: a4f37f089f064b722852c1a6565ea7de (MD5) Fapesb A acidificação e a Corrosão Microbiologicamente influenciada (CMI) são os principais problemas associados às Bactérias Redutoras de Sulfato (BRS) dentro da indústria de petróleo. Como forma minimizar os problemas citados, a indústria recorre a métodos de controles diversos, dentre os quais se destaca a utilização de biocidas, que por apresentar baixa eficiência e um custo elevado nem sempre são suficientes para conter a ação corrosiva das BRS. Compreender a dinâmica populacional das BRS em campos de petróleo é fundamental, tendo em vista a possibilidade de esclarecer os processos de acidificação e corrosão que acometem os reservatórios e os sistemas de transporte. Deste modo, métodos de cultivo tradicionais são utilizados e apesar da sua importância por vezes subestimam populações complexas. Por outro lado, métodos independentes de cultivo já promoveram uma compreensão destas comunidades em ambientes extremos, quando empregados em paralelo aos métodos tradicionais. O objetivo deste estudo foi testar e validar técnicas moleculares para monitorar em tempo real BRS em amostras de água produzida (AP), a partir de quatro poços (A, B, C e D) em diferentes pontos na região do Recôncavo Baiano, utilizando os métodos FISH, DAPI e NMP (número mais provável). Os dados obtidos evidenciaram a reunião de baixas populações de BRS nos poços pesquisados, e por meio de análise de variância (ANOVA) utilizando o teste de Scott Knott, foi observada a significância estatística (p≤0,05), entre as médias avaliadas para o poço de petróleo C, quando os três métodos (DAPI, FISH e NMP) foram comparados. Os métodos DAPI e FISH também apresentaram um p≤0,05 para a avaliação do poço D, sugerindo que as limitações encontradas pelo método NMP, para o respectivo poço, foram supridas pelos métodos independentes de cultivo. O qPCR foi empregado no referido estudo para quantificar o número de cópias do gene dsrA, responsável pela redução do sulfato, através da quantificação absoluta. Entanto, foi necessário previamente bioestimular uma amostra de AP em três concentrações diferentes (20%, 40% e 60%). Os resultados evidenciaram que a amostra de AP na concentração de 60% permitiu a amplificação do DNA extraído e a validação da metodologia, uma vez que a análise de qPCR mostrou um slope -3,32, uma eficiência de 100,081% e um R2 de 0,999.