The archaeal branch of life represents some of the oldest life forms on Earth. Archaea are believed to have diverged from Prokaryotes roughly 3.5 billion years ago and it’s theorized that biological methane production started around this time as well. This would make methanogenesis one of the oldest metabolisms on our planet. Methanogenesis is a process that, so far, is known to be unique to archaea. Since their evolution, methanogens have had massive impacts on Earth’s climate and biology. Methane is an important part of the global carbon cycle, but is also a major greenhouse gas, making it a vital area of research. The methanogen studied in this thesis is referred to as the wild type (WT) and was isolated from an active serpentinization system at Chimaera seep in Çıralı, Antalya Gulf, Turkey. The Chimaera seep is a geological formation analogous to mid-ocean ridges, but exposed and above land. The research in this thesis focuses on the description of the WT, which is believed to be a new species. Describing the WT consisted of the characterization of extremes as well as optimal growth conditions. The WT was found to grow at initial pH levels of 9.0 and 9.5. It grew from 15 °C to 45 °C, but not at 47 °C and not at 12 °C, and had an optimum at 39 °C. The WT had measurable growth up to 40 g/L NaCl, and had its optimum at 0 g/L. The WT grew best with H2/CO2 substrate, but also grew well on formate.

Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2018-07-27T22:02:00Z No. of bitstreams: 1 Dissertação THAYLI RAMIRES ARAUJO 2018.pdf: 1826233 bytes, checksum: bb42c5caf9e8edf5af4dce72d1cca93f (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2018-07-31T19:05:52Z (GMT) No. of bitstreams: 1 Dissertação THAYLI RAMIRES ARAUJO 2018.pdf: 1826233 bytes, checksum: bb42c5caf9e8edf5af4dce72d1cca93f (MD5) / Made available in DSpace on 2018-07-31T19:05:52Z (GMT). No. of bitstreams: 1 Dissertação THAYLI RAMIRES ARAUJO 2018.pdf: 1826233 bytes, checksum: bb42c5caf9e8edf5af4dce72d1cca93f (MD5) Previous issue date: 2018-05-21 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / A atividade das agroindústrias tem causado impactos ao meio ambiente, dentre eles a contaminação de solo, águas subterrâneas e superficiais, atmosfera. Na indústria de alimentos o sal é utilizado em diversas etapas de processamento de alimentos, e também é considerado o ingrediente mais consumido pelos humanos. Na indústria pesqueira marítima o sal está presente desde a descarga de material e o efluente gerado quando tratado origina o chamado lodo de estação de efluentes de pescado salgado, esse resíduo lançado ao meio ambiente sem tratamento acarreta sérios problemas ambientais, tais como a inibição da microbiota natural do meio afetando o ciclo biogeoquímico. Uma das ferramentas para o tratamento desse resíduo salino é o processo de compostagem, que pode ser aplicada no tratamento de diversos resíduos sólidos agroindustriais. Uma questão a ser discutida é a presença de sal nesse resíduo. A pressão osmótica provoca a inibição do crescimento (atividade) da microbiota natural podendo levá-la a morte por extravasamento do conteúdo celular. Para solucionar esses problemas apresentados, a adição de cepas halofílicas e/ou halotolerantes seria uma alternativa para que ocorra a compostagem desse resíduo salino. Portanto, um dos objetivos desse trabalho foi isolar e identificar por PCR bactérias halofílicas de uma matriz alimentar salgada como o bacalhau seco salgado. Os resultados demostraram que as cepas Bacillus cereus, Enterococcus faecalis e Lactobacillus spp foram as mais promissoras quando submetidas a multiplicação bacteriana em caldo suplementado com 7,5% de NaCl. Outro objetivo deste trabalho constituiu em avaliar através de parâmetros físico-químicos a microcompostagem da mistura de lodo de pescado salino, casca-de-arroz e serragem com a adição de B. cereus, E. faecalis e consórcio delas nos dias de processo de compostagem. Os resultados demonstraram eficácia na queda da condutividade elétrica no período estudado pelas cepas utilizadas nos tratamentos. O pH apesar de ter apresentado queda durante a compostagem se manteve acima de 6, padrão estabelecido pela legislação brasileira para comercialização do composto. Durante todo o período de compostagem, o teor de nitrogênio encontrado permaneceu em porcentagens baixas, devido ao uso de bactérias halotolenrantes, apesar de serem utilizadas como probióticas causam a desnitrificação, assim elevando o percentual de nitrogênio no decorrer do experimento. Em relação ao carbono a porcentagem de carbono teve uma queda o insignificativa durante o processo, a porcentagem de carbono encontrada durante o experimento é respondido pela alta atividade microbiana obtido em porcentagens de carbono abaixo dos 45% ao final dos 26 dias de incubação. A umidade para os tratamentos com adição de probióticos estiveram dentro do padrão estipulado pelo MAPA, entre 40 e 60%, contrariamente dos tratamentos sem os inóculos que apresentaram valores acima. E a matéria mineral, apresentou valores maiores ao final da compostagem devido a ação microbiana e a redução da matéria orgânica e formação de minerais. O lodo de pescado salino de estação de tratamento de efluentes obteve melhores resultados com adição de inóculo, como adição de outro resíduo rico em nitrogênio, modificação de volume/volume e material estruturante, e maior tempo de compostagem. Apesar dessas observações os resultados apresentados foram dentro dos padrões da legislação brasileira. / The activity of the agroindustries has caused impacts to the environment, among them the contamination of soil, groundwater and surface, atmosphere. In the food industry, salt is used in several stages of food processing, and is also considered the most consumed ingredient by humans. In the marine fishing industry salt is present since the discharge of material and the effluent generated when treated gives rise to so-called salty fish effluent seasoning, this waste released to the environment without treatment causes serious environmental problems such as the inhibition of the microbiota natural environment affecting the biogeochemical cycle. One of the tools for the treatment of this saline residue is the composting process, which can be applied in the treatment of various agroindustrial solid wastes. One issue to be discussed is the presence of salt in this residue. The osmotic pressure causes the inhibition of the growth (activity) of the natural microbiota and can lead to death by extravasation of the cellular content. To solve these problems, the addition of halophilic and / or halotolerant strains would be an alternative for the composting of this saline residue. Therefore, one of the objectives of this work was to isolate and identify, by PCR, halophilic bacteria from a salt food matrix such as salted dry cod. The results showed that the Bacillus cereus, Enterococcus faecalis and Lactobacillus spp strains were the most promising when submitted to bacterial multiplication in broth supplemented with 7.5% NaCl. Another objective of this work was to evaluate the microcompaction of the sludge mixture of saline fish, rice husk and sawdust with the addition of B. cereus, E. faecalis and their consortium in the days of the composting process. The results demonstrated efficacy in the reduction of the electrical conductivity in the period studied by the strains used in the treatments. The pH, even though it showed a drop during composting, remained above 6, a standard established by the Brazilian legislation for the commercialization of the compost. During the whole composting period, the nitrogen content remained low, due to the use of halotolenating bacteria, although they are used as probiotics, causing denitrification, thus raising the percentage of nitrogen during the experiment. In relation to carbon the percentage of carbon had an insignificant drop during the process, the percentage of carbon found during the experiment is answered by the high microbial activity obtained in carbon percentages below 45% at the end of the 26 days of incubation. The humidity for the treatments with addition of probiotics were within the standard stipulated by MAPA, between 40 and 60%, contrary to the treatments without the inocula that presented values above. And mineral matter presented higher values at the end of composting due to microbial action and reduction of organic matter and mineral formation. The saline fish sludge from the effluent treatment station obtained better results with addition of inoculum, as addition of another residue rich in nitrogen, modification of volume / volume and structuring material, and greater time of composting. Despite these observations, the results presented were within the brazilian legislation.

Soil salinity is an increasing problem facing agriculture in many parts of the world. Climate change and irrigation practices have led to decreased yields of large areas of farmland due to increased salt levels in the soil. Irrigation introduces salts to the soil that with time accumulate and threaten crop yield. In arid climates like Utah, the practice of irrigation is especially threatening to salt-sensitive crops including alfalfa (Medicago sativa). Plants that have tolerance to salt are needed to feed livestock and the world's population. One approach to address this problem is to introduce genes encoding salt tolerance into the genomes of salt-sensitive plants through genetic engineering, but this approach has limitations. These limitations include the misinformed public perception of genetically modified organisms (GMOs). Even if the GMO salt-tolerant plants could produce palatable foods, in regions of the world with saline soils incongruous with farming, if consumers refuse to purchase the food then the engineering and upfront costs of production are negated. Another fairly new approach involves the isolation and development of salt-tolerant (halophilic) plant-associated bacteria. Several reports are now available demonstrating how the use of halophilic inoculants enhance plant growth in salty soil. This enhanced plant growth is most likely associated with changes in plant gene expression; however, the mechanisms behind this growth stimulation are not yet clear. Halomonas elongata 1H9, a rhizobacteria native to Goshen UT, has been identified as a plant growth-promoting rhizobacteria (PGPR) when used as an inoculant added to alfalfa seedlings grown in salty soils. Plants grown in the presence of this Halomonas sp. and 1% salt demonstrated an average increase of 2.4x the biomass of alfalfa plants grown without inoculum in salty soils. This suggests that this Halomonas sp. positively influences plant salt tolerance, which raises the question as to how the bacteria stimulate plant growth under these conditions. To identify and characterize plant genes induced by Halomonas elongata, transcriptional analysis was performed using RNA-sequencing (RNA-seq). This analysis identified a variety of differentially expressed genes (DEGs) including transcription factors (e.g. MYB14, GATA transcription factor 9, Ethylene-responsive transcription factors ER017 and ER109) and plant enzymes involved in growth and development (e.g. xyloglucan endotransglucosylase and phosphodiesterase). This was followed by gene validation via real-time quantitative PCR (RT-qPCR), the gold standard for RNA-seq validation, however this process was never successfully completed. Suggestions for next steps are included in the discussion section of chapter 3.

No description available.

Os organismos extremófilos são considerados atualmente reservatórios de novas biomoléculas de interesse biotecnológico. Dentro deste grupo encontram-se os microrganismos que requerem altas concentrações de sal para crescer, denominados halófilos. Também existem os halotolerantes que são aqueles microrganismos que não precisam de sal para proliferar, mas toleram altas concentrações de NaCl. Os ambientes salinos provaram ser uma fonte rica de microrganismos halotolerantes produtores de novos compostos naturais e, portanto, a pesquisa nestes ambientes torna-se de grande importância. No Brasil, na região salina de Areia Branca no bioma caatinga, foram isoladas bactérias halotolerantes que foram estudadas com o objetivo de avaliar a produção de novos biopolímeros de interesse biotecnológico. Acredita-se que os polímeros naturais desses microrganismos extremos podem ter aplicações inovadoras ou características diferentes às tradicionais. Neste estudo, os isolados foram identificados em nível de gênero com base na análise da sequência do gene 16s rRNA. Os isolados foram principalmente bactérias Gram-positivas atribuídas às famílias Bacillaceae, Staphylococcaceae, Microbacteriaceae e uma bactéria Incertae Sedis do filo firmicutes, afiliadas aos gêneros Bacillus, Staphylococcus, Curtobacterium e Exiguobacterium, respectivamente. Apenas um isolado Gram-negativo foi identificado e atribuído como membro da família Pseudomonadaceae, incluso no gênero Pseudomonas. Avaliou-se a tolerância ao sal dos isolados em meio TSB suplementado com 5, 35, 60 e 120 g/L de NaCl. Todos os isolados apresentaram a capacidade de crescer nas quatro concentrações de NaCl avaliadas, com exceção do isolado Exiguobacterium sp. sac36 que não cresceu na concentração de 120 g/L de NaCl no meio. Realizaram-se ensaios de acúmulo de polihidroxialcanoatos (PHA) e evidenciou-se que quatro isolados do gênero Bacillus são capazes de acumular 3-hidroxibutirato (3HB) a partir de glicose, xilose, e alguns destes em glicerol. Adicionalmente, confirmou-se que quando há altas concentrações de NaCl no meio, o acúmulo de 3HB dos isolados produtores diminui. Observou-se também que doze isolados halotolerantes são produtores de exopolissacarídeos (EPS). Testes realizados indicaram que os mesmos podem ter efeitos imunoestimulantes em macrófagos. Finalmente, avaliou-se a produção de ácido hialurônico (AH) pelos isolados halotolerantes. Segundo sugere o método de Alcian blue, todos os isolados foram capazes de produzir AH, mostrando que a maior parte deles acumulou o biopolímero em concentrações maiores ou semelhantes ao controle. Os resultados obtidos evidenciam que os isolados halotolerantes avaliados são uma fonte rica em compostos com atividades promissoras para as diferentes indústrias. O presente trabalho contribui no estudo do potencial biotecnológico de microrganismos isolados no bioma caatinga, destacando sua biodiversidade, versatilidade e a necessidade de continuar explorando esses ambientes extremos pouco estudados. / Extremophile organisms are considered reservoirs of new biomolecules of biotechnological interest. In this group there are microorganisms that require high salt concentration to grow, called halophiles, and halotolerant microorganisms, that do not need salt to proliferate but can tolerate high concentrations of NaCl. Saline environments proved to be a rich source of new natural compounds by halotolerant producers and therefore, research in these environments becomes of great importance. In Brazil, in the saline region of Areia Branca in the caatinga biome, halotolerant bacteria were isolated and studied in order to evaluate the production of new biopolymers of biotechnological interest. It is believed that the natural polymers of those extreme microorganisms could have innovative applications or different characteristics from the traditional biopolymers. In this study, the isolates were identified at the genus level based on 16S rRNA gene sequence analysis. Isolates were mainly Gram-positive bacteria from Bacillaceae, Staphylococcaceae and Microbacteriaceae families, and Bacillus, Exiguobacterium, Staphylococcus and Curtobacterium genera. One of the Gram-negative isolate was identified as member of the Pseudomonadaceae family, genus Pseudomonas. The evaluation of salt tolerance of the bacterial isolates on TSB medium supplemented with 5, 35, 60 and 120 g / L NaCl was performed. All the isolates showed the ability to grow in the four concentrations evaluated, except for Exiguobacterium sp. sac36, that did not grow at 120 g / L NaCl. Polyhydroxyalkanoate (PHA) accumulation assays were performed using glucose, xylose and glycerol as carbon source. The results showed that four strains of the genus Bacillus were able to accumulate 3-hydroxybutyrate (3HB) in all conditions. Additionally, it was confirmed that the presence of high concentrations of NaCl in the medium causes a decrease in 3HB accumulation in the cells. It was observed that twelve halotolerant bacteria produced exopolysaccharides (EPS). Tests performed indicated that those EPS could have immunostimulatory effects on macrophages. Finally, hyaluronic acid (HA) production was evaluated. According to Alcian blue method, all strains were able to produce HA, showing that most of the isolates accumulated the biopolymer in higher or similar concentrations to the control. The results showed that the halotolerant isolates are a rich source of compounds with promising activities for different industries. This study contributes to the knowledge of microorganisms from the caatinga biome and their biotechnological potential, highlighting their biodiversity, versatility and the need to continue exploring these poorly studied extreme environments.