The role played by the protein UnkG from the plant growth-promoting bacterium Pseudomonas putida UW4 in the ability of the bacterium to facilitate plant growth was studied. Previous work showed that over-expressing UnkG decreased the ability of P. putida UW4 to facilitate plant growth. In contrast, an unkG knock-out mutant of P. putida UW4 displayed an increased ability to promote plant growth. Various biological activities of P. putida UW4, P. putida UW4/pETP and P. putida UW4/pETP-unkG have been compared. Thus, the growth curves were measured; the Biolog™ system was used to test the ability of these strains to utilize various carbon sources; the strains were observed by scanning electron microscopy to assess their relative cell sizes; biochemical assays were conducted to quantify 3-indoleacetic acid production and to measure the enzymatic activity of 1-aminocyclopropane-1-carboxylate (ACC) deaminase; proteome-level changes of P. putida UW4/pETP and P. putida UW4/pETP-unkG were profiled using two-dimensional difference in-gel electrophoresis (DIGE), followed by mass spectrometry identification of the altered proteins. After running DIGE, sixteen altered proteins were identified and their possible roles in the interactions between the bacterium and plants were discussed. Based on the preliminary results, we hypothesize that 1) UnkG may be detrimental to plant growth; 2) UnkG may negatively regulate a number of key cellular functions in a general way related to the energy balance of the bacterium.

The role played by the protein UnkG from the plant growth-promoting bacterium Pseudomonas putida UW4 in the ability of the bacterium to facilitate plant growth was studied. Previous work showed that over-expressing UnkG decreased the ability of P. putida UW4 to facilitate plant growth. In contrast, an unkG knock-out mutant of P. putida UW4 displayed an increased ability to promote plant growth. Various biological activities of P. putida UW4, P. putida UW4/pETP and P. putida UW4/pETP-unkG have been compared. Thus, the growth curves were measured; the Biolog™ system was used to test the ability of these strains to utilize various carbon sources; the strains were observed by scanning electron microscopy to assess their relative cell sizes; biochemical assays were conducted to quantify 3-indoleacetic acid production and to measure the enzymatic activity of 1-aminocyclopropane-1-carboxylate (ACC) deaminase; proteome-level changes of P. putida UW4/pETP and P. putida UW4/pETP-unkG were profiled using two-dimensional difference in-gel electrophoresis (DIGE), followed by mass spectrometry identification of the altered proteins. After running DIGE, sixteen altered proteins were identified and their possible roles in the interactions between the bacterium and plants were discussed. Based on the preliminary results, we hypothesize that 1) UnkG may be detrimental to plant growth; 2) UnkG may negatively regulate a number of key cellular functions in a general way related to the energy balance of the bacterium.

The presented diploma thesis focuses on the use of plant growth promoting bacteria as an ecological alternative to conventional fertilizers. The incorporation of bacterial cells into hydrogel carriers is already a well-studied topic, but due to its disadvantages it has not yet found wider application in agriculture. This work offers a novel concept of encapsulating bacteria by gelation directly from the culture. This is achieved by crosslinking the bacterial alginate produced by the model microorganism Azotobacter vinelandii. Since this process was not described before, first its optimization was needed. Alginate production was determined gravimetrically, and its parameters were further characterized using available analytical methods – infrared spectroscopy to monitor structural parameters (monomer composition and the extent of acetylation), dynamic light scattering to characterize the size distribution and AF4-MALS-dRI to obtain the molecular weight. Bacterial PHB production was also investigated using gas chromatography and infrared spectroscopy. The second part of the work is focused on the optimization of the gelling process using bacterial alginate from the culture and CaCl2 as a crosslinking agent. Rheological experiments were used as a tool in understanding the viscoelastic properties of the prepared gels. Gelation was demonstrated within the first day after inoculation. Maximum production of alginate (1,9 ± 0,3) g/l was reached on the fourth day after inoculation. It was found that the addition of 5 g/l of calcium carbonate promotes the production of alginate. Nevertheless, further addition of CaCO3 (30 g/l) showed adverse effects on the molecular weight and is therefore not recommended. Production of PHB was confirmed by both FTIR and GC measurements, with a maximum yield of (23 ± 3) % CDW. Rheological testing confirmed that the product of the crosslinking was a gel. It was found that the crosslinker concentration plays an important role at time 0 min of the gelation, forming a denser network in the structure and causing higher rigidity. Using the highest studied concentration of CaCl2, the critical strain reached values of (5,0 ± 0,7) %. Finally, the incorporation of bacterial cells into the hydrogel was confirmed using fluorescence microscope.

Here, we embarked a bioprospecting project that focuses on the isolation and characterization of plant root endophytes, collected from the Thar Desert. A total of 381 endophytes were isolated and based on their 16S rRNA gene sequences, genus Bacillus (58 strains) was identified as the major taxon and only endophytes from this genus were isolated from all plant types. Of the 58 Bacillus strains, only 16 strains were selected for screening of plant growth promotion traits such as P and Zn solubilization, indole-3-acetic acid and siderophore production, and antimicrobial activity. Based on the presence of specific plant growth promotion traits 10 strains were shortlisted for further in vitro screening with A. thaliana; to confirm that these bacteria can confer resilience to plants under salt stress conditions. B. circulans (PK3-15 and PK3-109), B. cereus (PK6-15) B. subtilis (PK3-9) and B. licheniformis (PK5-26) displayed the ability to increased the fresh weight of A. thaliana under salt stress conditions by more than 50 % compared to the uninoculated control. An interesting observation was that B. circulans (PK3-109) (shown to produce IAA exopolysaccharide) and B. circulans (PK3-138) (shown to produce IAA) in vitro results were substantially different as B. circulans (PK3-138) decreased the total fresh weight of A. thaliana by 47 %, whilst B. circulans (PK3-109) was one of the best performing strains. Thus, the genomes of these two strains were sequences to unravel the molecular versatility of B. circulans strains, specifically with respect to their interaction with plants. Most of the genome of these strains is identical but the most interesting feature was the presence of 1/ the DegS–DegU two-component system that is known to mediate the salt stress response and DegU also represses toxin wapA similar to antitoxin wapI, and 2/ YxiG, a gene in the unique orthogroup of PK3-109 was found to be linked to WapI. Thus, PK3-138 substantially decreasing the total fresh weight of A. thaliana under salt stress conditions suggests that the toxic activity of a toxin such as WapA is not effectively ameliorated by the antitoxin such as WapI in the absence of a functional YxiG gene.

With the increase of agricultural production, there is an improvement in the use of mineral fertilizers, which may cause different environmental problems, besides the soil salinization. A possible alternative for reducing the application of these products is the use of plant growth-promoting bacteria (PGPB), that can be used alone or in co-inoculation, resulting in an alternative environmentally and economically feasible. Better results can be obtained if the interaction among bacteria-bacteria and bacteria-plant be elucidated, and strategy developed to optimize these interactions. Thus, the plant growth-promoting Bacillus sp. RZ2MS9, previous described as a potential PGPB in maize and soybean, was GFP-tagged and monitored alone and co-inoculated with Azospirillum brasilense (Ab-v5::pWM1013) during maize colonization. The interaction of tagged strains in maize were monitored by fluorescent microscopy (FM) and quantitative PCR (qPCR), demonstrating an endophytic behavior of Bacillus sp. RZ2MS9. Although the non-detection of Ab-v5::pWM1013, the co-inoculation resulted in the best increase in root and shoot dried weight, root volume and in root diameter, showing that inoculation with more than one strain can be a good choice to development of bio-fertilizers. One important system to bacterial interaction is the quorum sensing (QS). The QS is an important cell-cell communication system that allows bacterial cells to recognize their own population and modulate their gene expression. This, system is also involved in the interspecific communication, including other bacterial species and plants. In the other hand, enzymes able to detect and degrade these molecules evolved, the called quorum quenching (QQ) system, that has been evolved in some bacteria as competitive advantage for niches colonization. The aiiA gene, was one of the first gene related with the QQ in Bacillus. The aiiA was found in Bacillus sp. RZ2MS9 genome. Through construction of a new QQ biosensor, Agrobacterium tumefaciens At11006, and validated by A. tumefaciens NTL4, the ability of RZ2MS9 to degrade QS molecules was confirmed. The knockout of aiiA gene was performed using the CRISPR-Cas9 system, confirming this gene function. By these results, the influence of QQ system of Bacillus sp. RZ2MS9 during maize colonization and RZ2MS9 - A. brasilense - maize can be better investigated, opens the possibility to better understand the role of QQ system in the interaction among PGPB and plants. / Concomitantemente ao aumento da produção agrícola, há o aumento do uso de fertilizantes minerais, que pode acarretar no desenvolvimento de diferentes problemas ambientais, além de causar a salinização dos solos. Uma possível alternativa para tentar reduzir a aplicação desses produtos é o uso de bactérias promotoras de crescimento de plantas (BPCPs), que podem ser usadas isoladamente ou em co-inoculação com outras bactérias, tornando-as uma alternativa ambientalmente e economicamente viável. Melhores resultados podem ser obtidos se a interação bactéria-bactéria e bactéria-planta for elucidada, permitindo que estratégias sejam desenvolvidas para otimizar essas interações. Em vista disso, a bactéria Bacillus sp. RZ2MS9, previamente descrita como uma potencial BPCP em milho e soja, foi marcada com GFP e monitorada durante a colonização de milho inoculada sozinha, bem como em co-inoculação com Azospirillum brasilense (Ab-v5::pWM1013). A interação dessas linhagens marcadas em milho, foi monitorada por microscopia de fluorescência (FM) e PCR quantitativo (qPCR), revelando um comportamento endofítico de Bacillus sp. RZ2MS9. Em plantas co-inoculadas, apesar da linhagem Ab-v5::pWM1013 não ter sido detectada por qPCR, a co-inoculação resultou no aumento do peso seco das raízes e da parte aérea, no volume e no diâmetro do sistema radicular, demonstrando que a inoculação com mais de uma linhagem bacteriana pode ser uma boa alternativa para o desenvolvimento de bio-fertilizantes. O quorum sensing (QS) é um importante sistema de comunicação célula-célula que permite que as bactérias reconheçam sua própria população e modulem sua expressão gênica. Este sistema também está envolvido na comunicação interespecífica, incluindo outras espécies bacterianas e plantas. Co-evolutivamente, enzimas capazes de detectar e degradar essas moléculas evoluíram, dando origem ao chamado quorum quenching (QQ), sistema que evoluiu em algumas bactérias como uma vantagem competitiva para a colonização de nichos. O gene aiiA, foi um dos primeiros genes relacionados ao sistema QQ descrito no gênero Bacillus, gene este que foi anotado no genoma de RZ2MS9. Através da construção de uma nova linhagem biossensora de QQ, Agrobacterium tumefaciens At11006, e validada através da linhagem A. tumefaciens NTL4, a capacidade de RZ2MS9 de degradar moléculas de QS foi confirmada. O knockout do gene aiiA foi realizado utilizando o sistema CRISPR-Cas9, confirmando a função desse gene. Através dos resultados obtidos neste trabalho, a influência do sistema QQ de Bacillus sp. RZ2MS9 durante a colonização do milho, bem como a interação RZ2MS9 - A. brasilense - milho pode ser melhor investigada, abrindo a possibilidade de uma melhor compreensão do papel do sistema QQ na interação entre bactérias promotoras de crescimento e plantas.

Soils have been contaminated with cadmium (Cd) by the use of fertilizers, calcareous, pesticides and industrial and/or domestic effluents. It can be leached to groundwater, as well as be taken up by plants potentially leading to reduce growth and yield. It causes different damages to the cell, generating oxidative stress which is responsible for its toxicity, affecting all living organism. A balance in the redox state of the cell to maintain cellular integrity and metabolism is essential for organism tolerance. Thus, the antioxidant response of bacteria exposed to Cd was studied to understand the tolerance mechanism, and be able to develop a methodology to bioremediate contaminated soils. MDA and hydrogen peroxide contents and different enzymes activity of antioxidant system (SOD, CAT, GR and GST) of two strains from Burkholderia genus, one from a soil contaminated with Cd in high concentrations (strain SCMS54) and the other from soil without Cd (strain SNMS32) in two exposure time (5 and 12 h), were analyzed. Stress measurement (MDA and hydrogen peroxide content) and antioxidant enzyme activities (SOD, CAT, GR and GST) increased in almost all treatments in the presence of Cd. These results also indicate that strain SCMS54 (isolated from Cd contaminated soil) presents a higher metabolic diversity and plasticity due the expression of more isoforms of the enzymes SOD, CAT and GR. The strain also accumulates 50% more Cd. We also analyzed the response to Ni of these two strain, observing a similar response to Cd, except for GST enzyme expression, which in strain SCMS54 this enzyme was induced in the presence of Ni, indicating that this enzyme can be essential on Ni tolerance. After that, the strain isolated from Cd contaminated soil (SCMS54) was selected to proceed the studies to evaluate the benefits of tolerant microorganism-tomato plant interaction. The use of plants to remove heavy metals from contaminated soilhas less impact and a lower cost. Soil microorganisms can be able to solubilize or mobilize soil metals acting also as bioremediator. Besides the high tolerance to Cd, the strain SCMS54 can produce indole-acetic acid (IAA), solubilize inorganic phosphate and produce siderophore, revealing its potential in plantmicroorganism mutual and beneficial interaction. When interacting with tomato plants exposed to Cd, this bacterium led to decrease in plant peroxide concentration and chlorosis levels, promoted relative plant growth and reduced the root absorption of Cd resulting in an increase in plant tolerance to this highly toxic heavy metal. Indicating that inoculation of tomato plants with Burkholderia sp. SCMS54 promotes better growth when cultivated in the presence of Cd by a mechanism that appears to decrease Cd concentration in roots as a result of a bacterial-plant root beneficial interaction. / O cádmio (Cd) tem contaminado solos pelo uso de fertilizantes, calcário, agrotóxicos e resíduos industriais e/ou domésticos. Podendo ser lixiviado ao lençol freático ou absorvido pelas plantas,resultando na redução do crescimento e da produtividade. Esse metal afeta todos os organismos vivos e causa diferentes danos às células. A tolerância a esse metal se deve principalmente ao balanço do estado redox da célula para manter a integridade celular e metabolismo.Assim, foram isoladas bactérias de solo contaminado e não contaminado com Cd, selecionando isolados tolerantes a altas concentrações de diferentes metais (Cd, Ni e Zn), em seguida, foi observado a resposta do sistema antioxidante da bactéria na presença do Cd, a fim de auxiliar no desenvolvimento de metodologias para biorremediar solos contaminados. Foi quantificado MDA e peróxido de hidrogênio e a atividade de diferentes enzimas do sistema antioxidante (SOD, CAT, GR e GST) de duas estirpes do gênero Burkholderia tolerantes a todos os metais testados, uma isolada do solo contaminado com altas concentrações de Cd (estirpe SCMS54) e a outra do solo sem Cd (estirpe SNMS32) em dois tempos de exposição (5 e 12 h). Na estirpe SCMS54, as medidas de estresse (peroxidação lipídica e peróxido de hidrogênio) e a atividade das enzimas antioxidantes (SOD, CAT, GR e GST) da maioria dos tratamento com cádmio aumentaram, esta estirpe também expressa mais isoformas de SOD, CAT e GR, além de acumular 50% mais Cd. Esses resultados mostram que a estirpe SCMS54 (isolada do solo contaminado com Cd) apresenta uma maior diversidade metabólica e plasticidade. Foram analisadas também a resposta dessas duas estirpes ao Ni, observando uma resposta semelhante ao Cd, exceto na expressão da enzima GST, que no estirpe SCMS54 foi induzida na presença do Ni, indicando que essa enzima pode ser essencial na tolerância ao Ni. Portanto, a estirpe isoladado solo contaminado com Cd (SCMS54) foi selecionada para prosseguir os estudos e avaliar os benefícios da interação entre microrganismos tolerantes-plantas de tomate na fitorremediação. Essa técnica é usada remover para metais pesados do solo com um menor impacto e baixos custos. Os microrganismos do solo podem solubilizar e mobilizar metais do solo, atuando como biorremediador. Além da alta tolerância ao Cd, a estirpe SCMS54 produz ácido indol acético (AIA), solubiliza fosfato inorgânico e produz sideroforo, mostrando seu potencial na interação benéfica planta-microorganismo. Quando interagindo com as plantas de tomate expostas ao Cd, essa bactéria diminui a concentração de peróxido da planta e a clorose ocasionado pelo Cd,e reduz a absorção de Cd pela raiz resultando em um aumento da tolerância da planta ao metal pesado altamente tóxico. Assim, a inoculação de plantas de tomate com Burkholderia sp. SCMS54 promove crescimento da planta na presença de Cd, desencadeando um mecanismo que diminui a concentração de Cd nas raízes devido a essa interação benéfica bactéria-raiz da planta.

Soils have been contaminated with cadmium (Cd) by the use of fertilizers, calcareous, pesticides and industrial and/or domestic effluents. It can be leached to groundwater, as well as be taken up by plants potentially leading to reduce growth and yield. It causes different damages to the cell, generating oxidative stress which is responsible for its toxicity, affecting all living organism. A balance in the redox state of the cell to maintain cellular integrity and metabolism is essential for organism tolerance. Thus, the antioxidant response of bacteria exposed to Cd was studied to understand the tolerance mechanism, and be able to develop a methodology to bioremediate contaminated soils. MDA and hydrogen peroxide contents and different enzymes activity of antioxidant system (SOD, CAT, GR and GST) of two strains from Burkholderia genus, one from a soil contaminated with Cd in high concentrations (strain SCMS54) and the other from soil without Cd (strain SNMS32) in two exposure time (5 and 12 h), were analyzed. Stress measurement (MDA and hydrogen peroxide content) and antioxidant enzyme activities (SOD, CAT, GR and GST) increased in almost all treatments in the presence of Cd. These results also indicate that strain SCMS54 (isolated from Cd contaminated soil) presents a higher metabolic diversity and plasticity due the expression of more isoforms of the enzymes SOD, CAT and GR. The strain also accumulates 50% more Cd. We also analyzed the response to Ni of these two strain, observing a similar response to Cd, except for GST enzyme expression, which in strain SCMS54 this enzyme was induced in the presence of Ni, indicating that this enzyme can be essential on Ni tolerance. After that, the strain isolated from Cd contaminated soil (SCMS54) was selected to proceed the studies to evaluate the benefits of tolerant microorganism-tomato plant interaction. The use of plants to remove heavy metals from contaminated soilhas less impact and a lower cost. Soil microorganisms can be able to solubilize or mobilize soil metals acting also as bioremediator. Besides the high tolerance to Cd, the strain SCMS54 can produce indole-acetic acid (IAA), solubilize inorganic phosphate and produce siderophore, revealing its potential in plantmicroorganism mutual and beneficial interaction. When interacting with tomato plants exposed to Cd, this bacterium led to decrease in plant peroxide concentration and chlorosis levels, promoted relative plant growth and reduced the root absorption of Cd resulting in an increase in plant tolerance to this highly toxic heavy metal. Indicating that inoculation of tomato plants with Burkholderia sp. SCMS54 promotes better growth when cultivated in the presence of Cd by a mechanism that appears to decrease Cd concentration in roots as a result of a bacterial-plant root beneficial interaction. / O cádmio (Cd) tem contaminado solos pelo uso de fertilizantes, calcário, agrotóxicos e resíduos industriais e/ou domésticos. Podendo ser lixiviado ao lençol freático ou absorvido pelas plantas,resultando na redução do crescimento e da produtividade. Esse metal afeta todos os organismos vivos e causa diferentes danos às células. A tolerância a esse metal se deve principalmente ao balanço do estado redox da célula para manter a integridade celular e metabolismo.Assim, foram isoladas bactérias de solo contaminado e não contaminado com Cd, selecionando isolados tolerantes a altas concentrações de diferentes metais (Cd, Ni e Zn), em seguida, foi observado a resposta do sistema antioxidante da bactéria na presença do Cd, a fim de auxiliar no desenvolvimento de metodologias para biorremediar solos contaminados. Foi quantificado MDA e peróxido de hidrogênio e a atividade de diferentes enzimas do sistema antioxidante (SOD, CAT, GR e GST) de duas estirpes do gênero Burkholderia tolerantes a todos os metais testados, uma isolada do solo contaminado com altas concentrações de Cd (estirpe SCMS54) e a outra do solo sem Cd (estirpe SNMS32) em dois tempos de exposição (5 e 12 h). Na estirpe SCMS54, as medidas de estresse (peroxidação lipídica e peróxido de hidrogênio) e a atividade das enzimas antioxidantes (SOD, CAT, GR e GST) da maioria dos tratamento com cádmio aumentaram, esta estirpe também expressa mais isoformas de SOD, CAT e GR, além de acumular 50% mais Cd. Esses resultados mostram que a estirpe SCMS54 (isolada do solo contaminado com Cd) apresenta uma maior diversidade metabólica e plasticidade. Foram analisadas também a resposta dessas duas estirpes ao Ni, observando uma resposta semelhante ao Cd, exceto na expressão da enzima GST, que no estirpe SCMS54 foi induzida na presença do Ni, indicando que essa enzima pode ser essencial na tolerância ao Ni. Portanto, a estirpe isoladado solo contaminado com Cd (SCMS54) foi selecionada para prosseguir os estudos e avaliar os benefícios da interação entre microrganismos tolerantes-plantas de tomate na fitorremediação. Essa técnica é usada remover para metais pesados do solo com um menor impacto e baixos custos. Os microrganismos do solo podem solubilizar e mobilizar metais do solo, atuando como biorremediador. Além da alta tolerância ao Cd, a estirpe SCMS54 produz ácido indol acético (AIA), solubiliza fosfato inorgânico e produz sideroforo, mostrando seu potencial na interação benéfica planta-microorganismo. Quando interagindo com as plantas de tomate expostas ao Cd, essa bactéria diminui a concentração de peróxido da planta e a clorose ocasionado pelo Cd,e reduz a absorção de Cd pela raiz resultando em um aumento da tolerância da planta ao metal pesado altamente tóxico. Assim, a inoculação de plantas de tomate com Burkholderia sp. SCMS54 promove crescimento da planta na presença de Cd, desencadeando um mecanismo que diminui a concentração de Cd nas raízes devido a essa interação benéfica bactéria-raiz da planta.

>Magister Scientiae - MSc / As the global human population grows, so does the demand for faster food production rates. Owing to this, agricultural practices have had to expand and move into semi-arid and arid regions, too, where frequent irrigation is essential. However, irrigated ground water contains many salt ions (mainly Na+ and Cl-) which contribute to soil salinization on croplands. Soil salinity negatively impacts crop growth and yield and thus, strategies for the alleviation of salt stress on crop plants have had to be developed. This study assessed the use of plant growth promoting bacteria (PGPB). The aim of this study was to isolate, identify and characterize bacterial endophytes isolated from the halophyte, Arctotheca calendula. Endophytes were identified using 16S rDNA and were screened for plant growth promoting properties including nitrogen fixation, phosphate and zinc solubilization, siderophore, ammonia and indole-3-acetic acid (IAA) when exposed to 0 mM, 300 mM and 600 mM NaCl. The endophytes had been identified as Erwinia persicina NBRC 102418T, Bacillus marisflavi JCM 11544T, Ochrobactrum rhizosphaerae PR17T, Microbacterium gubbeenense DSM 15944T and Bacillus zhangzhouensis DW5-4T and all of which had demonstrated some plant growth promoting characteristics. Thereafter, we aimed to demonstrate plant growth promotion of P. vulgaris cv. Star 2000 inoculated with PGPB under salinity stress. P. vulgaris cv. Star 2000 seeds were inoculated with the PGPB and exposed to 0 mM and 100 mM NaCl. Post-harvest, plants were assessed for their dry mass, cell death, superoxide concentration and nutrient content. It was discovered that salinity negatively impacted P. vulgaris cv. Star 2000’s dry mass, NaCl-induced cell death, and differentially influenced superoxide concentration, nutrient uptake and content of the leaf and root material in the inoculated and control treatments. However, the isolated PGPB had been able to mitigate the negative effects of soil salinity on P. vulgaris cv. Star 2000.

Made available in DSpace on 2017-07-10T17:37:57Z (GMT). No. of bitstreams: 1 2012_Walkyria_Neiverth.pdf: 2694760 bytes, checksum: 7b3db9a364fcea380b3a53433bdbf6aa (MD5) Previous issue date: 2012-06-26 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Nitrogen is one of the limiting factors of crop production, but its continuous availability by applying chemical fertilizer in agriculture, will lead to high costs, both economic and environmental. To reduce these costs, several studies have been conducted regarding the biological nitrogen fixation (BNF) and use of plant growth-promoting bacteria (PGPB). Considering that the great diversity of soil microorganisms has not been yet catalogued, the goal of this work was to study the morphological and genetic diversity of endophytic rhizobacteria obtained from soils of different classes and crop managements in order to isolate individuals with potential for PGPB. Bacteria were isolated from diluted soils and inoculated in bait-plants of wheat grown in the nutrient solution. After 30 days, the plant tissue was superficially sterilized and macerated and an aliquot of serial dilutions was inoculated in selective medium Dygs with successive plating to obtain pure colonies. Bacterial growth was quantified by counting colony forming units (c.f.u.) and 400 morphologically distinct colonies were characterized: size, shape, color, edge, homogeneity, transparency, elevation, and mucus. Based on this information, the isolates were analyzed and grouped by using the program Past for morphological typing. From these 269 strains were genetically characterized by rep-PCR with specific primer. Polymorphic profiles obtained were analyzed and grouped with BioNumerics software. Calculations regarding the rates of morpho-genetic diversity and richness were performed by SPADE program. The managements with succession of corn and soybean in organic system (M10) and corn - soybean RR (M8) were the ones showing higher and lower c.f.u, respectively. The great biodiversity found in the various types of soil under different culture managements from evaluated regions have resulted in a total of 35 morphological and 25 genetics groups. As for genetic groups, there was, in addition to the 25 major groups, 16 profiles when used 60% of similarity. In morphological groups, it was observed greater number of profiles relating to riparian forest area (M4), whereas the smallest number was observed in succession corn and organic soybean on zero tillage (M10). It was found higher morphological diversity index, richness and groups expected in future collection management with monoculture of sugar cane, high levels of P in soil and application of vinasse (M1), being the lowest indexes for the monoculture management with sugar cane with low P in soil and mineral fertilization (M2). As for diversity indexes and genetic richness the M1 management stands out as showing the higher diversity indexes, but the highest rates of richness and groups expected in future collection were observed in the succession corn - soybean (M9) management. By contrast, the lowest rates of genetic diversity and richness were appointed by management with organic soybean on zero tillage. The results showed a high diversity in all areas assessed, both for the different cultivation managements, soil types, as to characteristic vegetation of the different areas, pointing the concomitant influence of these agri-environmental characteristics on the dominance of specific morpho-genetic groups within the communities / O nitrogênio é um dos fatores mais limitantes da produção vegetal, porém sua disponibilização contínua pela agricultura através da aplicação de fertilizantes químicos acarreta em altos custos financeiros, energéticos e ambientais. Com intuito de reduzir estes custos, vários estudos vêm sendo realizados quanto à fixação biológica de nitrogênio (FBN) e utilização de bactérias promotoras de crescimento vegetal (BPCV). Considerando-se a grande diversidade destes micro-organismos de solo ainda não catalogados, o objetivo deste trabalho foi estudar a diversidade morfológica e genética de rizobactérias endofíticas obtidas a partir de solos de diferentes classes e manejos de cultivo, a fim de isolar indivíduos com potenciais biotecnológicos para a FBN e para a promoção de crescimento vegetal (PCV). Bactérias foram isoladas a partir de solos diluídos e inoculados em plantas-iscas de trigo cultivadas em solução nutritiva. Após 30 dias, o tecido vegetal foi superficialmente esterilizado, macerado e uma alíquota plaqueada em meio de cultura generalizado, Dygs. O crescimento bacteriano foi quantificado pela contagem de unidades formadoras de colônias (u. f. c.) e as colônias aparentemente distintas foram caracterizadas morfologicamente. Com base nestes dados, foram escolhidas 400 estirpes, analisadas e agrupadas pela utilização do programa Past. A caracterização genética de 269 estirpes foi realizada pela reação de rep-PCR com primer específico BOX e os perfis polimórficos apresentados foram analisados e agrupados com auxílio do programa BioNumerics. Cálculos quanto aos índices de diversidade e riqueza morfo-genética foram realizados a partir do programa SPADE. Os manejos com sucessão milho - soja em sistema orgânico e milho - soja RR foram os que apresentaram maiores e menores médias de u. f. c., respectivamente. A grande biodiversidade encontrada nas diferentes classes de solos sob os manejos de cultivo das regiões avaliadas estabeleceram um total de 35 agrupamentos morfológicos e 25 agrupamentos genéticos. Quanto aos agrupamentos genéticos, verificou-se, além dos 25 GG formados, a obtenção de 16 perfis isolados quando comparados a 60% de similaridade. Nos agrupamentos morfológicos, observou-se maior número de perfis referentes à área de mata ciliar (M4), enquanto que o menor número foi observado no manejo de sucessão milho - soja orgânico (M10). Observou-se ainda, maior índice de diversidade, riqueza e grupos esperados em coleta futura no manejo com monocultivo de cana-de-açúcar, alto teor de P no solo e aplicação de vinhaça (M1), sendo os menores índices referentes ao manejo com monocultivo de cana-de-açúcar com baixo teor de P e adubação mineral (M2). Quanto aos índices de diversidade e riqueza genética destaca-se o manejo M1 como sendo o de maior diversidade, porém os maiores índices de riqueza e grupos esperados em coleta futura foram observados no manejo de sucessão milho - soja (M9). Em contrapartida, os menores índices de diversidade e riqueza são apontados ao manejo M10, sendo a área com pousio (M5) a menos favorável ao desenvolvimento dos micro-organismos de acordo com o índice de grupos esperados em coleta futura. Os resultados demonstram uma alta diversidade em todas as regiões avaliadas, tanto para os diferentes manejos de cultivo, classes de solo quanto à vegetação característica das diferentes áreas, além da influência concomitante destes sobre a dominância de determinados grupos morfo-genéticos dentro da comunidade

Plant growth-promoting bacteria (PGPB) have been studied in many agriculturally interesting plants, but never in pitcher plants. Sarracenia oreophila (the green pitcher plant) is an endangered species in Georgia, Alabama, and North Carolina (Rice 2010). With the help of Dr. Jim Spain's lab, a previous student in Dr. Gerald Pullman's lab discovered evidence that nitrogen-fixing bacteria (Burkholderia spp.) live within these pitcher plants. This study aims to determine whether these nitrogen-fixing bacteria confer a benefit to their host plants by providing fixed nitrogen. To do this, pitcher plants were inoculated with the Burkholderia and grown on a control medium, a medium without sugar (as the sugar causes the bacteria to grow until they hinder the plants), various media that are missing nitrogen-containing compounds usually provided in growth media, and a medium completely lacking nitrogen. These plants were compared to control plants on the same media that had not been inoculated with Burkholderia. The plants' biomass and root growth were measured. The data suggest that Burkholderia may stimulate plant biomass growth when sufficient nitrogen is present and there may be a nitrogen-threshold that needs to be met in order to sustain the Burkholderia-Sarracenia symbiosis. Also, the Burkholderia has a negative effect on roots grown in high-nitrogen media, possibly due to competition for nutrients.