<i>Rhizobium leguminosarum</i> are well known for their ability to fix nitrogen (N). In addition, their capacity to solubilize phosphate has been receiving attention in recent years. The work presented in this thesis examined two aspects of screening and evaluating dicalcium phosphate (Pi) (CaHPO4) solubilizing <i> R. leguminosarum</i>. The objectives of this study were to: 1) identify a medium that is sensitive and effective as a screening tool for phospahte solubilizing <i>R. leguminosarum</i>; 2) determine the effect of N and carbon (C) on growth and P solubilization of <i>R. leguminosarum</i> isolates; 3) determine the relationship between the ability to solubilize CaHPO4 by <i>R. leguminosarum</i> isolates on solid medium and in liquid broth of same composition; and 4) assess and compare the ability of <i>R. leguminosarum</i> isolates to solubilize different P sources in soil under growth chamber conditions.<p>In this study, 30 <i>R. leguminosarum</i> isolates were evaluated for phosphate solubilization in broth and solid formulations of three different media, Yeast Mannitol Extract (YEM), Botanical Research Institute Phosphate Nutrient medium (MNBRI) and Pikovskaya Phosphate medium (PVK). All media contain CaHPO4 as the only phosphorus (P) source. The <i>R. leguminosarum</i> isolates were selected on the basis of their different plasmid profiles, indicative of genetically distinct isolates. <p>All 30 isolates increased the Pi concentration in solution to varying degrees in liquid cultures but performance varied from one medium to another. The highest average solution Pi concentration achieved by the 30 <i>R. leguminosarum</i> isolates was obtained from PVK cultured broth. CaHPO4 solubilization by <i>R. leguminosarum</i> isolates in liquid was associated with a decrease in pH. Among the three tested media, the lowest pH by the thirty <i>R. leguminosarum</i> isolates was obtained in PVK. Ability of the isolates to solubilize CaHPO4 on the solid media was not comparable to the performance of the isolates grown in liquid because only fewer <i>R. leguminosarum</i> isolates showed visible P solubilization on the solid media.<p>The composition and formulation of medium influence the ability of the <i>R. leguminosarum</i> isolates to solubilize CaHPO4. Effects of N and C concentrations on the growth and CaHPO4 solubilization by nine <i>R. leguminosarum</i> isolates were examined in liquid formulation. Ammonium N had a greater influence on the growth and CaHPO4 solubilization by <i>R. leguminosarum</i> isolates than C at the tested levels. The growth of isolates was inhibited by ammonium N at 0.5 g L-1 as (NH4)2SO4 meaning there were less viable cells in this N concentration than were of ammonium N at 0.1 g L-1. The ability of isolates to solubilize Pi however was not affected by ammonium N at 0.5 g L-1 as (NH4)2SO4. The media containing low N (0.1 g (NH4)2S04 L-1) both Pi solubilization and growth of <i>R. leguminosarum</i> isolates were not affected.<p><i>R. leguminosarum</i> isolates were tested for their effects on growth and P uptake of canola plants in P-deficient soils amended with different P sources. <i>R. leguminosarum</i> isolates were selected separately based on their ability to solubilize CaHPO4 from the three screening media. A quadrant model was used based on the ability of the 30 <i>R. leguminosarum</i> isolates to solubilize CaHPO4 on both solid and liquid formulations within a medium. The effect of <i>R. leguminosarum</i> on canola dry mass, tissue Pi content and total Pi uptake varied from one isolate to another, but was not different from the controls. The quadrant model failed to correlate isolates able to solubilize CaHPO4 in laboratory screening to isolates able to solubilize P in the growth chamber. Despite the influence of the medium composition and formulation, none of tested media predicted Pi solubilization ability by the <i>R. leguminosarum</i> isolates in soils under growth chamber conditions, from their Pi solubilization of laboratory screenings. <p>The work of this thesis demonstrates that phosphate solubilization is a complex process that depends on both organism and soil. Growth condition is an important factor for a <i>R. leguminosarum</i> isolate to express its ability to solubilize CaHPO4. Liquid media screenings illustrate an isolates ability to solubilize CaHPO4 under nonstressful conditions, but solid media screenings demonstrate the P solubilization result of an isolate under more stressful conditions. The lack of relationship in P solubilization ability by <i>R. leguminosarum</i> isolates, between laboratory methods to soil test, means neither liquid or solid media can provide a definitive selection process. Additional parameters should be investigated to modify the soil bioassay protocols and ultimate selection procedures. These include pH conditions, isolate colonization, growth, and survival on plants and rhizosphere.

<i>Rhizobium leguminosarum</i> are well known for their ability to fix nitrogen (N). In addition, their capacity to solubilize phosphate has been receiving attention in recent years. The work presented in this thesis examined two aspects of screening and evaluating dicalcium phosphate (Pi) (CaHPO4) solubilizing <i> R. leguminosarum</i>. The objectives of this study were to: 1) identify a medium that is sensitive and effective as a screening tool for phospahte solubilizing <i>R. leguminosarum</i>; 2) determine the effect of N and carbon (C) on growth and P solubilization of <i>R. leguminosarum</i> isolates; 3) determine the relationship between the ability to solubilize CaHPO4 by <i>R. leguminosarum</i> isolates on solid medium and in liquid broth of same composition; and 4) assess and compare the ability of <i>R. leguminosarum</i> isolates to solubilize different P sources in soil under growth chamber conditions.<p>In this study, 30 <i>R. leguminosarum</i> isolates were evaluated for phosphate solubilization in broth and solid formulations of three different media, Yeast Mannitol Extract (YEM), Botanical Research Institute Phosphate Nutrient medium (MNBRI) and Pikovskaya Phosphate medium (PVK). All media contain CaHPO4 as the only phosphorus (P) source. The <i>R. leguminosarum</i> isolates were selected on the basis of their different plasmid profiles, indicative of genetically distinct isolates. <p>All 30 isolates increased the Pi concentration in solution to varying degrees in liquid cultures but performance varied from one medium to another. The highest average solution Pi concentration achieved by the 30 <i>R. leguminosarum</i> isolates was obtained from PVK cultured broth. CaHPO4 solubilization by <i>R. leguminosarum</i> isolates in liquid was associated with a decrease in pH. Among the three tested media, the lowest pH by the thirty <i>R. leguminosarum</i> isolates was obtained in PVK. Ability of the isolates to solubilize CaHPO4 on the solid media was not comparable to the performance of the isolates grown in liquid because only fewer <i>R. leguminosarum</i> isolates showed visible P solubilization on the solid media.<p>The composition and formulation of medium influence the ability of the <i>R. leguminosarum</i> isolates to solubilize CaHPO4. Effects of N and C concentrations on the growth and CaHPO4 solubilization by nine <i>R. leguminosarum</i> isolates were examined in liquid formulation. Ammonium N had a greater influence on the growth and CaHPO4 solubilization by <i>R. leguminosarum</i> isolates than C at the tested levels. The growth of isolates was inhibited by ammonium N at 0.5 g L-1 as (NH4)2SO4 meaning there were less viable cells in this N concentration than were of ammonium N at 0.1 g L-1. The ability of isolates to solubilize Pi however was not affected by ammonium N at 0.5 g L-1 as (NH4)2SO4. The media containing low N (0.1 g (NH4)2S04 L-1) both Pi solubilization and growth of <i>R. leguminosarum</i> isolates were not affected.<p><i>R. leguminosarum</i> isolates were tested for their effects on growth and P uptake of canola plants in P-deficient soils amended with different P sources. <i>R. leguminosarum</i> isolates were selected separately based on their ability to solubilize CaHPO4 from the three screening media. A quadrant model was used based on the ability of the 30 <i>R. leguminosarum</i> isolates to solubilize CaHPO4 on both solid and liquid formulations within a medium. The effect of <i>R. leguminosarum</i> on canola dry mass, tissue Pi content and total Pi uptake varied from one isolate to another, but was not different from the controls. The quadrant model failed to correlate isolates able to solubilize CaHPO4 in laboratory screening to isolates able to solubilize P in the growth chamber. Despite the influence of the medium composition and formulation, none of tested media predicted Pi solubilization ability by the <i>R. leguminosarum</i> isolates in soils under growth chamber conditions, from their Pi solubilization of laboratory screenings. <p>The work of this thesis demonstrates that phosphate solubilization is a complex process that depends on both organism and soil. Growth condition is an important factor for a <i>R. leguminosarum</i> isolate to express its ability to solubilize CaHPO4. Liquid media screenings illustrate an isolates ability to solubilize CaHPO4 under nonstressful conditions, but solid media screenings demonstrate the P solubilization result of an isolate under more stressful conditions. The lack of relationship in P solubilization ability by <i>R. leguminosarum</i> isolates, between laboratory methods to soil test, means neither liquid or solid media can provide a definitive selection process. Additional parameters should be investigated to modify the soil bioassay protocols and ultimate selection procedures. These include pH conditions, isolate colonization, growth, and survival on plants and rhizosphere.

To understand the soil microbial community structures of different locations and altitudes, as well as the fluctuation of microbial populations foward seasonal change, this study investigated the biodiversity of soil microbiota of eight sampling sites along the Lao-Nong River basin during 4/11/2003~3/21/2004. The results showed that the soil properties were different among these sampling sites. The soil temperatures were ranging in between 24~32¢J, except Tianchr, the highest altitude sampling site (7~19¢J) ; pH values were 4.00 (Tianchr) ~8.20 (Tauyuan) ; moisture contents were 2.31% (Liouguei) ~70.23% (Tianchr) ; total organic carbon contents were 0.38% (Linkou) ~36.21% (Tianchr) ; total organic nitrogen contents were 0.067% (Tauyuan) ~0.672% (Tianchr); the C/N ratio were 7.0~25.0, except Tianchr (42 in average) . In microbial community structures analysis, bacteria were the most abundant organisms among all soil samples (106~108 CFU per gram of soil) . The amount of other microbes were as follow: nitrogen-fixing microbes 105~107 CFU, actinomycetes 104~107 CFU, fungi 104~106 CFU, cellulolytic microbes 104~106 CFU, and the least one was phosphate- solubilizing microbes 103~105 CFU. For molecular biological microbiota analysis, the bead-beating DNA extraction method resulted the richest bacterial 16S rDNA DGGE profile. The16S rDNA DGGE profiles of various samples taken from different seasons showed somewhat seasonal variations that might relate to the specific climates of Taiwan, such as the plum rains and typhoons. Moreover, we also successfully cultured and preserved many microbes with specific biological functions. These microorganisms will provide a useful resource for further research and development in the biotechnology industries.

Made available in DSpace on 2016-12-08T15:50:04Z (GMT). No. of bitstreams: 1 PGMS13DA016.pdf: 762204 bytes, checksum: 51d4b0b6a684c027977c08cf9ff670a1 (MD5) Previous issue date: 2013-05-21 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / The use of microorganisms that help plant nutrition and growth is an important alternative to the use of chemical fertilizers. This study was divided into four distinct works, the first is related to the multiplication of mycorrhizal inoculant using the on farm method with pre inoculated plants, the second study involves the production of mycorrhizal inoculum using the on farm methodology and its application incorn,, the third evaluated the process of single or dual inoculation of phosphate solubilizing bacteria and arbuscular mycorrhizal fungi (AMF) in two forest species, and the fourth assessed the mycorrhizal dependency of the two forest species. The overall goal was to improve existing technologies for the use of soil microorganisms capable of providing better P uptake by plants. For the first experiment, pre inoculated sorghum plants were transplanted into bags with a capacity of 20L. After 3 months of growth under field conditions, the mycorrhizal inoculum potential, root mycorrhizal colonization and number of spores were evaluated. For the second experiment, we used field soil mixed with vermiculite (1:1) in 20 L bags.. Each bag received 10% of AMF inoculum mixed with the soil : vermiculite mix and seeded with oats as the host plant. After 3 months,this inoculums was used in corn to evaluate its effect on plant productivity. The third work was the isolation of phosphate solubilizing bacteria from soils near spring waters and forests in the municipality of Laurentino SC and subsequent inoculation (single or dual) with the AMF Rhizophagus clarus, in Acca sellowiana and Citharexilum myrianthum, under greenhouse and field conditions. The fourth experiment was conducted in a greenhouse where the Acca sellowiana and Citharexylum myrianthum were grown in 3 different levels of soil P and 4 mycorrhizal treatments (Rhizophagus clarus, Claroideoglomus etunicatus, Dentiscutata heterogama and control). After 3 months the experiment was collected and the mcyorrhizal dependency calculated. Results demonstrate that the process of dual inoculating of phosphate solubilizing bacteria and AMF was benefit to both forest species tested. Considering the production of mycorrhizal inoculant using the on farm method, it was demonstrated that: i)both method of inoculation (pre colonized plants and direct inoculation)increased the mycorrhizal inoculum potential, ii) the inoculant is feasible to apply, and iii) using on farm inoculant produced by direct inoculaton increased corn productivity.. Both forest species varied in their mycorrhizal dependency and the highest values were obtained when inoculated with Rhizophagus clarus compared to other two isolates.. The technologies studied here have shown the benefit although further studies are still nee de in order to be applied commercially / O uso de microrganismos que auxiliam na nutrição e no crescimento de plantas é uma importante alternativa ao uso de fertilizantes químicos. O presente estudo foi dividido em quatro trabalhos distintos, sendo o primeiro relacionado a multiplicação de inoculante micorrízico pelo método on farm com plantas pré inoculadas, o segundo relacionada a multiplicação de inóculo pelo método on farm e sua aplicação na cultura do milho, o terceiro tratou de processos de inoculação conjunta e isolada de bactérias solubilizadoras de fosfato e fungos micorrízicos arbusculares (FMAs) em duas espécies florestais e o quarto avaliou a dependência micorrízica das duas espécies florestais. Sendo assim, o objetivo geral foi aprimorar tecnologias já existentes na utilização de microrganismos do solo capazes de propiciar melhor absorção de P pelas plantas. Para o primeiro experimento foram utilizadas plantas de sorgo pré-inoculadas para multiplicação de inoculante em sacos de mudas com capacidade de 20L. Após 3 meses de crescimento em condições de campo foi avaliado o potencial de inóculo micorrízico, porcentagem de colinização radicular e número de esporos produzidos. Para o segundo experimento, foi utilizado solo do campo misturado com vermiculita (1:1) acondicionados em sacos de 20L. Cada saco recebeu 10% de inoculo contendo FMA adicionado de forma direta e com planta hospedeira utilizou-se aveia. Após 3 meses o inoculante produzido via inoculação direta foi utilizado na cultura do milho para posterior avaliação de produtividade. O terceiro trabalho contou com o isolamento de bactérias solubilizadoras de fosfato de solos de áreas de nascente e de florestas do município de Laurentino-SC e posterior inoculação de 5 bactérias solubilizadoras de fosfato, de forma conjunta ou não, ao isolado de FMA Rhizophagus clarus, nas espécies florestais Acca sellowiana e Citharexilum myrianthum em condições de casa de vegetação e a campo. O quarto experimento foi realizado em condições de casa de vegetação onde as duas espécies florestais utilizadas no experimento 1 foram submetidas a crescimento em 3 níveis distintos de P no solo e 4 tratamentos de FMAs (Rhizophagus clarus, Claroideoglomus etunicatus, Dentiscutata heterogama e controle). Após 3 meses, o experimento foi coletado e analisada a dependência micorrízica. Os resultados obtidos demonstram que o processo de inoculação conjunta de bactérias solubilizadoras de fosfato e FMAs beneficiaram o crescimento das duas espécies florestais testadas. Em relação a produção on farm de inoculante micorrízico, as duas metodologias apresentaram resultados positivos no aumento do potencial de inoculo micirrízico dos inoculantes produzidos, foram de fácil aplicabilidade e a utilização de inoculante multiplicado via inoculação direta proporcionou incremento significativo na produtividade do milho. As duas espécies florestais apresentaram variação na dependência micorrízica e o isolado R. clarus apresentou maiores valores absolutos em relação aos outros dois isolados testados. As tecnologias aqui estudadas, embora tenham demonstrado beneficio, ainda necessitam de estudos para poderem ser aplicadas de forma comercial

Made available in DSpace on 2017-07-10T19:24:14Z (GMT). No. of bitstreams: 1 Juliana _Matter.pdf: 980208 bytes, checksum: 39b0f232a5a4474d2605637e0dc10ea5 (MD5) Previous issue date: 2015-04-28 / Chemical fertilizers management in agriculture is the main source of nutrients for crops development aiming at increasing yield. Thus, phosphorus, among other nutrients, is required by plants since it is the most limiting mineral on yield crops of commercial interest, because of its low availability and immobility in soil, as it also gets unavailable to absorption by the plants. Studies have shown that phosphate solubilization up from bacteria inoculation can improve agricultural production. Many bacteria can take part on the dissolution process through different mechanisms and promote plant growth either by dissolving phosphates or determining nitrogen (N) and indole acetic acid production (IAA). Due to the isolation of these microorganisms, they can be inoculated and make the fixed phosphorus available, reducing the use of fertilizers or readjusting excessive percentages of this element in their formulas to improve a development of sustainable agricultural yield. According to this theme, it was possible to apply a methodology for the isolation of subtropical soil bacteria with maize cropping from different soil fertilization managements. After isolating the micro-organisms, they were submitted to endurance tests after pricking, phosphatase production ability, IAA production with and without L-tryptophan and identification method by rDNA extraction technique. A greater diversity of mineralizing bacteria was found in treatment 1 soil (no fertilization). The results related to the largest percentage increase in Calcium phytate can be justified by the fact that bacterial communities can produce enzymes. The stability of the solubilizing function was greater in calcium phytate when compared to the other culture media that have been used. The study has indicated that soil microbiota keeps a similarity in which Ochrobactrum gender has prevailed and under different management situations, its diversity slightly varies, but it is able to produce metabolites promoters of plant growth; while the isolates of Treatment 2 (mineral fertilization) have kept the best acid phosphatase production. The IAA production, without L-tryptophan addition in T1, suggests that the microbiota uses metabolic pathways for different purposes, since the studied soil is poor in minerals, thus it affects the production of plant growth promoters. It differs from Treatment 3 that received fertilization with swine wastewater, and diversified IAA production for its isolates with stronger values. With this study it was possible to identify genera of bacteria with high phytate mineralization capacity. At last, according to the study results, it was possible to identify the bacteria genera with high capacity to mineralize phytate and produce IAA, indicating the promotion of vegetal growth capacity. / A utilização de fertilizantes químicos na agricultura constitui-se como a principal fonte de nutrientes para o desenvolvimento das culturas visando ao aumento da produtividade. O fósforo, dentre outros nutrientes, é o mais requerido pelas plantas e o que mais limita a produtividade nas culturas de interesse comercial em virtude da sua baixa disponibilidade e imobilização no solo, haja vista ficar indisponível para absorção pelo vegetal. Estudos comprovam que a solubilização de fosfatos a partir da inoculação de bactérias pode ajudar na melhoria da produção agrícola. Muitas bactérias realizam processos de dissolução a partir de diferentes mecanismos e promovem crescimento vegetal tanto pela dissolução de fosfatos como na fixação de nitrogênio (N) e produção de ácido indol acético (AIA). Com o isolamento desses microrganismos, é possível inoculá-los e disponibilizar o fósforo fixado, reduzir o uso de fertilizantes ou readequar as porcentagens excessivas desse elemento em suas fórmulas, visando ao desenvolvimento da produção agrícola sustentável. A partir desta temática, foi utilizada metodologia para isolamento de bactérias de solo subtropical sob cultivo de milho com diferentes históricos de adubação. Além do isolamento, os microrganismos foram submetidos a testes de resistência após repicagem, produção de fosfatases, produção de AIA com e sem L- triptofano e identificação por técnica de extração de rDNA. Uma maior diversidade de bactérias mineralizadores foi encontrada no solo Tratamento 1 (sem adubação). Os resultados relacionados à maior porcentagem de crescimento em fitato de cálcio podem ser justificados pelo fato de as comunidades bacterianas serem capazes de produzir enzimas. A estabilidade da função solubilizadora foi maior em fitato de cálcio do que nos outros meios usados. O estudo indicou que a microbiota do solo mantém similaridade onde prevalece o gênero Ochrobactrum e que, em diferentes situações de manejo, pouco varia sua diversidade, mas apresenta capacidade para produzir metabólitos promotores de crescimento de plantas. Os isolados do Tratamento 2 (adubação mineral) mantiveram a produção de fosfatase ácida mais acentuada. A produção de AIA sem a adição de L-triptofano no tratamento T1 sugere que a microbiota utiliza vias metabólicas distintas para tal, visto que o solo estudado é pobre em minerais. Isso afeta a produção de promotores de crescimento de plantas, contrapondo-se ao Tratamento 3, o qual recebeu adubação com água residuária de suinocultura e diversificou a produção de AIA por seus isolados com valores mais robustos. Finalmente, de acordo com os resultados do presente estudo, foi possível a identificação de gêneros de bactéria com elevada capacidade de solubilização de fitatos e produção de AIA, indicando capacidade de promoção de crescimento vegetal.

Made available in DSpace on 2017-05-12T14:47:31Z (GMT). No. of bitstreams: 1 Juliana _Matter.pdf: 980208 bytes, checksum: 39b0f232a5a4474d2605637e0dc10ea5 (MD5) Previous issue date: 2015-04-28 / Chemical fertilizers management in agriculture is the main source of nutrients for crops development aiming at increasing yield. Thus, phosphorus, among other nutrients, is required by plants since it is the most limiting mineral on yield crops of commercial interest, because of its low availability and immobility in soil, as it also gets unavailable to absorption by the plants. Studies have shown that phosphate solubilization up from bacteria inoculation can improve agricultural production. Many bacteria can take part on the dissolution process through different mechanisms and promote plant growth either by dissolving phosphates or determining nitrogen (N) and indole acetic acid production (IAA). Due to the isolation of these microorganisms, they can be inoculated and make the fixed phosphorus available, reducing the use of fertilizers or readjusting excessive percentages of this element in their formulas to improve a development of sustainable agricultural yield. According to this theme, it was possible to apply a methodology for the isolation of subtropical soil bacteria with maize cropping from different soil fertilization managements. After isolating the micro-organisms, they were submitted to endurance tests after pricking, phosphatase production ability, IAA production with and without L-tryptophan and identification method by rDNA extraction technique. A greater diversity of mineralizing bacteria was found in treatment 1 soil (no fertilization). The results related to the largest percentage increase in Calcium phytate can be justified by the fact that bacterial communities can produce enzymes. The stability of the solubilizing function was greater in calcium phytate when compared to the other culture media that have been used. The study has indicated that soil microbiota keeps a similarity in which Ochrobactrum gender has prevailed and under different management situations, its diversity slightly varies, but it is able to produce metabolites promoters of plant growth; while the isolates of Treatment 2 (mineral fertilization) have kept the best acid phosphatase production. The IAA production, without L-tryptophan addition in T1, suggests that the microbiota uses metabolic pathways for different purposes, since the studied soil is poor in minerals, thus it affects the production of plant growth promoters. It differs from Treatment 3 that received fertilization with swine wastewater, and diversified IAA production for its isolates with stronger values. With this study it was possible to identify genera of bacteria with high phytate mineralization capacity. At last, according to the study results, it was possible to identify the bacteria genera with high capacity to mineralize phytate and produce IAA, indicating the promotion of vegetal growth capacity. / A utilização de fertilizantes químicos na agricultura constitui-se como a principal fonte de nutrientes para o desenvolvimento das culturas visando ao aumento da produtividade. O fósforo, dentre outros nutrientes, é o mais requerido pelas plantas e o que mais limita a produtividade nas culturas de interesse comercial em virtude da sua baixa disponibilidade e imobilização no solo, haja vista ficar indisponível para absorção pelo vegetal. Estudos comprovam que a solubilização de fosfatos a partir da inoculação de bactérias pode ajudar na melhoria da produção agrícola. Muitas bactérias realizam processos de dissolução a partir de diferentes mecanismos e promovem crescimento vegetal tanto pela dissolução de fosfatos como na fixação de nitrogênio (N) e produção de ácido indol acético (AIA). Com o isolamento desses microrganismos, é possível inoculá-los e disponibilizar o fósforo fixado, reduzir o uso de fertilizantes ou readequar as porcentagens excessivas desse elemento em suas fórmulas, visando ao desenvolvimento da produção agrícola sustentável. A partir desta temática, foi utilizada metodologia para isolamento de bactérias de solo subtropical sob cultivo de milho com diferentes históricos de adubação. Além do isolamento, os microrganismos foram submetidos a testes de resistência após repicagem, produção de fosfatases, produção de AIA com e sem L- triptofano e identificação por técnica de extração de rDNA. Uma maior diversidade de bactérias mineralizadores foi encontrada no solo Tratamento 1 (sem adubação). Os resultados relacionados à maior porcentagem de crescimento em fitato de cálcio podem ser justificados pelo fato de as comunidades bacterianas serem capazes de produzir enzimas. A estabilidade da função solubilizadora foi maior em fitato de cálcio do que nos outros meios usados. O estudo indicou que a microbiota do solo mantém similaridade onde prevalece o gênero Ochrobactrum e que, em diferentes situações de manejo, pouco varia sua diversidade, mas apresenta capacidade para produzir metabólitos promotores de crescimento de plantas. Os isolados do Tratamento 2 (adubação mineral) mantiveram a produção de fosfatase ácida mais acentuada. A produção de AIA sem a adição de L-triptofano no tratamento T1 sugere que a microbiota utiliza vias metabólicas distintas para tal, visto que o solo estudado é pobre em minerais. Isso afeta a produção de promotores de crescimento de plantas, contrapondo-se ao Tratamento 3, o qual recebeu adubação com água residuária de suinocultura e diversificou a produção de AIA por seus isolados com valores mais robustos. Finalmente, de acordo com os resultados do presente estudo, foi possível a identificação de gêneros de bactéria com elevada capacidade de solubilização de fitatos e produção de AIA, indicando capacidade de promoção de crescimento vegetal.

Made available in DSpace on 2014-06-11T19:32:54Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-10-19Bitstream added on 2014-06-13T19:03:38Z : No. of bitstreams: 1 barroso_cb_dr_jabo.pdf: 424873 bytes, checksum: c11fcf246d6ada6dcb329194e9814468 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Devido a baixa disponibilidade de P no solo e a alta capacidade do fungo Aspergillus niger F111 em solubilizar fosfatos inorgânicos, este trabalho teve por objetivo geral avaliar a possibilidade de inocular no solo esporos ou pellets imobilizados com vista a prolongar sua habilidade de solubilização e averiguar o mecanismo de solubilização de fosfatos inorgânicos de Ca, Al e Fe por este fungo. Os pellets inoculados em meio de cultura agitado proporcionaram maior solubilização dos fosfatos, principalmente o fosfato de Fe por ser de baixa solubilidade. No solo, os pellets livres e imobilizados promoveram as maiores solubilizações de fosfato de Fe e maior produção de CO2. Avaliando-se o efeito da fonte de N, as seguintes proporções foram obtidas na solubilização dos fosfatos de Ca, glicina > Al, nitrato de amônio > Fe, ácido l-glutâmico. Os açúcares que mais solubilizaram os fosfatos foram manitol, maltose e d-galactose. Dentre os metais somente o FeCl3.6H2O promoveu maior solubilização do fosfato de Fe e os metais FeSO4.7H2O e FeCl3.6H2O promoveram maiores solubilizações do fosfato de Ca. As concentrações de álcoois que mais favoreceram a solubilização do fosfato de Fe foram 3 e 4% de etanol e metanol, para o fosfato de Ca foi 3% de etanol. A combinação dos metais com o metanol, indicou que o metanol foi o principal responsável pela solubilização. Fatores como queda do pH, a maior produção de ácidos e o menor crescimento do fungo influíram neste trabalho, principalmente em relação a solubilização do fosfato de Fe. No solo, os pellets solubilizaram quantidades semelhantes de fosfato de Fe que os esporos imobilizados de A. niger, podendo ser utilizados com vantagem devido a sua facilidade de obtenção. / Considering the low P availability in the soil and the high capability of Aspergillus niger F111 in solubilizing inorganic phosphates, this work aimed to evaluate the possibility of inoculating spores or immobilized pellets in the soil to prolong the solubilization capability and study the solubilization mechanism of inorganic calcium phosphate, aluminum phosphate and iron phosphate by this fungus. Pellets inoculated in culture medium under agitation allowed higher phosphate solubilization, especially iron phosphate, which is low soluble. In the soil, free and immobilized pellets allowed the highest solubilization of iron phosphate and CO2 production. Evaluating the effect of N sources, the following proportions were obtained in the solubilization of calcium phosphates, glycine > Al, ammonium nitrate > Fe, l-glutamic acid. The sugars that most solubilized phosphates were mannitol, maltose and d-galactose. Among the metals, only FeCl3.6H2O promoted higher iron phosphate solubilization, and FeSO4.7H2O and FeCl3.6H2O promoted higher solubilization of calcium phosphate. The alcohol concentrations that most favored iron phosphate solubilization were 3 and 4% of ethanol and methanol, while the highest solubilization of calcium phosphate was reached with 3% ethanol. The combination of metals with methanol indicated this alcohol was mainly responsible for solubilization. Factors as pH decrease, higher acid production and lower A. niger growth influenced the results, especially in the solubilization of iron phosphate. In the soil, pellets and immobilized spores solubilized similar amounts of iron phosphate. Pellets are thus preferable because they are more easily obtained.

Submitted by MILENA NOVA CASTELBLANCO null (milenan80@gmail.com) on 2016-05-18T23:09:22Z No. of bitstreams: 1 Dissertação_Biotecnologia_Milena_Nova.pdf: 2319209 bytes, checksum: 5f8a2cf09d53469b118673ce803f4c05 (MD5) / Approved for entry into archive by Ana Paula Grisoto (grisotoana@reitoria.unesp.br) on 2016-05-20T19:50:02Z (GMT) No. of bitstreams: 1 castelblanco_mn_me_araiq_par.pdf: 1007182 bytes, checksum: a277306214e78520e7169816799bf058 (MD5) / Made available in DSpace on 2016-05-20T19:50:02Z (GMT). No. of bitstreams: 1 castelblanco_mn_me_araiq_par.pdf: 1007182 bytes, checksum: a277306214e78520e7169816799bf058 (MD5) Previous issue date: 2016-05-06 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A demanda crescente de metais como ferro, cobre, entre outros ocasionou o esgotamento progressivo dos depósitos minerais; de tal modo que as companhias mineradoras desenvolveram tecnologias alternativas aos métodos convencionalmente aplicados para a recuperação e extração de metais valiosos partir de minerais de baixo teor e outras fontes de metais polimetálicos como minerais de ferro que contem metais de base associados. Uma dessas alternativas é a biomineração que utiliza principalmente microrganismos procariontes e eucariontes para acelerar a dissolução oxidativa de minerais sulfetados presentes em minérios de baixo teor produzindo a solubilização de metais associados; esta biotecnologia é atualmente usada apenas para processar minérios reduzidos e rejeitos minerais uma vez que a maioria destes minerais são sulfetados. No entanto, muitos metais de valor econômico também são encontrados em minerais que são parcial ou totalmente oxidados como as lateritas de cobre ou níquel, minerais que não são susceptíveis à dissolução oxidativa; portanto, os minerais de ferro férrico contidos nesses minérios não podem ser processados oxidativamente, pois estes compostos são susceptíveis de serem reduzidos por processos biológicos, derivando na solubilização de metais associados. Microrganismos acidófilos tais como Acidithiobacillus ferrooxidans podem catalisar a redução dissimilatória de íons férricos na ausência de oxigênio para acelerar a solubilização destes metais. Estudos recentes têm mostrado que essa nova abordagem pode ser utilizada para extrair metais tais como níquel e cobre a partir de minérios oxidados a uma velocidade mais elevada do que pode ser conseguido por processos oxidativos. Neste trabalho, foram realizados ensaios de redução biológica de íons férricos acoplado a oxidação anaeróbia de enxofre elementar em um biorreator automatizado de 2 L com temperatura, agitação e pH constantes, usando um minério de ferro e uma cultura pura da bactéria anaeróbia facultativa At. ferrooxidans. A amostra foi fornecida pelo Instituto Tecnológico Vale - Desenvolvimento Sustentável (ITV-DS) e analisada por difração de raios X (DRX) evidenciando fases dominantes de monazita (Ce, La, Nd, Th) PO4, coesita (SiO2) e goethita (FeO (OH)). Os resultados do processo mostraram que metais como Cr, Al, Ca, Co Cu, K, Mg Mn, Ni, Pb e Zn foram solubilizados no processo bioredutivo com porcentagens de remoção de 93,3% para Mn e 34,7 para Zn, Nb um metal raro e as terras raras como lantânio (La) e cério (Ce) foram também solubilizadas no processo com 61%,17,9 % e 3,2% respetivamente, as outras terras raras que não solubilizaram como Sm e Nd foram expostas no resíduo mineral. Também foram solubilizados fosfatos nos primeiros dez dias atingindo 1% de solubilização. O resíduo solido foi avaliado por DRX e apresentou formação de novas fases como sodalita (K7,7Na0,3(AlSiO4)6(Cl)4)2), berlinita (AlPO4) e hematolita ((Mn2+, Mg, Al)15(AsO3) (AlO4)2(OH)2) e uma aparente diminuição da presença das fases de goethita e monazita. A análise do resíduo por microscopia eletrônica de varredura com emissão de campo (MEV-FEG) mostrou uma diferença na diminuição da presença das partículas menores presentes na amostra original e claramente uma maior presença de partículas maiores no sólido biolixiviado. Os resultados obtidos neste trabalho mostraram a recuperação de diversos metais, além de terras raras num minério de ferro, a recuperação de fosfatos que não estava prevista como objetivo inicial do trabalho, também foi observada. Estes resultados evidenciam que a dissolução redutiva catalisada por bactérias é uma alternativa promissória para a utilização de diversos tipos de minério de baixo teor que não poderiam ser processados por processos oxidativos e que seriam uma fonte para a recuperação de metais e outros compostos de interesse econômico, além de ser. ambientalmente amigável comparada com os processos convencionalmente aplicados para a recuperação de metais. / The growing demand for metals such as iron, copper, and others has caused the gradual exhaustion of mineral deposits; such a way that the mining companies have developed technologies alternative to the methods conventionally applied for the recovery and extraction of valuable metals from low grade minerals and other sources of polymetallic metals such as iron minerals which contains base metals associated. One such alternative is the biomining which uses mainly microorganisms prokaryotes and eukaryotes to accelerate the oxidative dissolution of sulphide minerals present in low grade ores producing solubilization of associated metals; This biotechnology is currently used only to process reduced ores and minerals tailings since most of these minerals are sulphides. However, many metals of economic value are also found in minerals that are partially or fully oxidized like copper or nickel laterites, minerals that are not susceptible to oxidative dissolution; therefore, the mineral ferric iron contained in these ores can not be oxidatively processed, since these compounds are capable of being reduced by biological processes, deriving the associated solubilizing metals. Acidophilic microorganisms such as Acidithiobacillus ferrooxidans can catalyze the dissimilatory reduction of ferric iron in the absence of oxygen to accelerate the solubilization of these metals. Recent studies have shown that this new approach can be used to extract metals such as copper and nickel from oxide ores at a higher speed than can be achieved by oxidative processes. In this work were carried out biological reduction tests of ferric iron coupled anaerobic oxidation of elemental sulfur in an automated bioreactor 2 L with temperature, agitation and pH constant in anaerobic conditions using an iron ore and a pure culture of facultative anaerobic bacterium At. ferrooxidans. The sample was provided by the Vale Technology Institute - Sustainable Development (ITV-DS) and analyzed by X-ray diffraction (XRD) showing dominant phases of monazite ((Ce, La, Nd, Th) PO4), coesita (SiO2) and goethite (FeO (OH)). The process results showed that metals such as Cr, Al, Ca, Co Cu, K, Mg, Mn, Ni, Pb and Zn were solubilized in the bioreductive process with removal percentages 93.3% by Mn and 34.7, for Zn, Nb, a rare metal and rare earths elements such as lanthanum (La) and cerium (Ce) were also solubilized in the process with 61%, 17.9% and 3,2% respectively, other rare earths not solubilized as Sm and Nd were exposed in the mineral residue. Also were solubilized phosphates in the first ten days to reach 1% solubilization. The solid residue was evaluated by XRD and showed formation of new phases such as sodalite (K7,7Na0,3(AlSiO4)6(Cl)4)2), berlinita (AlPO4) and hematolita ((Mn2+, Mg, Al)15(AsO3) (AlO4)2(OH)2) and an apparent reduction in the presence of goethite and monazite phases Analysis of the residue by scanning electron microscopy with field emission (SEM-FEG) showed a difference in reducing the presence of smaller particles present in the original sample and clearly a greater presence of larger particles in the solid biolixiviado. The results of this study showed the recovery of various metals, besides rare earths in an iron ore, phosphates recovery which was not expected as initial objective of this study was also observed. These results show that the reductive dissolution catalyzed by bacteria is a promising alternative to the use of different types of low grade ores that could be processed by oxidative processes and it would be a source for recovery of metals and other compounds of economic interest, besides being environmentally friendly compared to the processes conventionally applied to the recovery of metals.

The nitrogen fixing bacteria (NFB) and phosphate solubilizing bacteria (PSB) are used widely for producing of microbiological fertilizers. This study aims to seek nitrogen-fixing and phosphate-solubilizing bacteria strains to add to the collection of candidate strains for producing single and multi-function microbiological fertilizers. From 40 soil samples of 8 fields for cultivating rice, medicinal plants and vegetables, 15 NFB strains and 12 PSB strains were isolated and determined the ability of fixing nitrogen and solubilizing inorganic phosphate compound through creation of NH₄⁺ and PO₄⁻ in culture medium. Among 15 NFB strains, the fixing nitrogen activities of 7 strains were much higher than the remaining strains, including NFBR3, NFBV2, NFBM5, NFBM3, NFBM1, NFBV5 and NFBR2 with NH₄⁺ concentration 18.85 mg/l, 18.41 mg/l, 17.32 mg/l, 16.19 mg/l, 15.49 mg/l, 12.83 mg/l and 12.57 mg/l, respectively Among 12 PSB strains, The ability of solubilizing phosphate of 5 strains were higher than the others, including PSBM2, PSBR1, PSBV1, PSBR5 and PSBR3 with PO₄⁻ concentration 14.49 mg/l, 11.83 mg/l, 11.33 mg/l, 10.65 mg/l, 10.37 mg/l, respectively. 3 NFB strains (NFBR3, NFBV2, NFBM5) and 3 PSB strains (PSBM2, PSBR1, PSBV1) with higher activity were identified by 16S-rDNA sequence analysis and comparing to some homologous sequences in genbank. The results showed that NFBR3 was identified as Azotobacter vinelandii, NFBV2 as Azopirillum brasilense, NFBM5 as Azotobacter chroococum, PSBM2 and PSBV1 as Pseudomonas fluorescens and PSBR1 as Bacillus subtilis. / Vi khuẩn cố định nitơ (NFB) và vi khuẩn phân giải phosphate (PSB) được sử dụng rộng rãi trong sản xuất phân bón vi sinh. Nghiên cứu này nhằm mục đích tìm kiếm các chủng vi khuẩn cố định nitơ và hòa tan phosphate, bổ sung vào bộ sưu tập các chủng dự tuyển cho sản xuất phân bón vi sinh đơn và đa chức năng. Từ 40 mẫu đất của 8 ruộng trồng lúa, cây dược liệu và rau màu, 15 chủng NFB và 12 chủng PSB đã được phân lập và xác định khả năng cố định nitơ và phân giải phosphate vô cơ thông qua sự tạo thành NH₄⁺ và PO₄⁻ trong môi trường nuôi cấy. Trong số 15 chủng NFB, có 7 chủng có hoạt tính cố định nitơ cao hơn những chủng còn lại, bao gồm các chủng NFBR3, NFBV2, NFBM5, NFBM3, NFBM1, NFBV5 và NFBR2 với nồng độ NH₄⁺ lần lượt là 18.85mg/l, 18.41 mg/l, 17.32 mg/l, 16.19 mg/l, 15.49 mg/l, 12.83 mg/l và 12.5 7mg/l. Trong số 12, có 5 chủng có khả năng phân giải phosphate cao hơn những chủng khác, bao gồm chủng PSBM2, PSBR1, PSBV1, PSBR5 và PSBR3 với nồng độ PO₄⁻ lần lượt là 14.49 mg/l, 11.83 mg/l, 11.33 mg/l, 10.65 mg/l và 10.37 mg/l. Các chủng NFB và PSB này đều xuất hiện ơ các mẫu đất trồng lúa, đất trồng cây dược liệu và đất trồng rau màu. 3 chủng NFB và 3 chủng PSB với hoạt tính cố định nitơ và phân giải phosphate cao hơn được định loại bằng phân tích trình tự gen 16S-rDNA và so sánh với một số trình tự tương đồng trong genebank. Kết quả chỉ ra rằng chủng NFBR3 được định danh là Azotobacter vinelandii, chủng NFBV2 là Azopirillum brasilense, chủng NFBM5 là Azotobacter chroococum, chủng PSBM2 và chủng PSBV1 là Pseudomonas fluorescens và chủng PSBR1 là Bacillus subtilis.

O fósforo (P) que é um nutriente pouco disponível em solos tropicais devido a sua deficiência natural e também por causa da imobilização do mesmo em constituintes do próprio solo, formando fosfatos insolúveis e assim indisponíveis as plantas. A solubilização destes fosfatos inorgânicos é uma estratégia para aumentar a disponibilidade deste nutriente no solo. Uma forma seria a utilização de microrganismos solubilizadores de P que deixam este nutriente disponível às plantas. Desta forma, o presente trabalho teve como objetivos: (i) avaliar o crescimento do cafeeiro em função das fontes e doses de P2O5; (ii) determinar o fósforo disponível e (iii) avaliar o estado nutricional do cafeeiro, em função da fonte e dose de P2O5. O desenvolvimento das mudas de cafeeiro é superior com o uso de fonte de P solúvel, em relação à fonte de solubilidade gradual, independentemente da dose. A determinação da capacidade de substituição da fonte solúvel pela fonte de solubilidade gradual depende do tempo de reação no solo, cuja máxima equivalência é igual a 84 %, aos 180 dias após a aplicação do fertilizante (MSF). Em solos com teor de P disponível suficiente para a cultura do cafeeiro é viável a substituição de uma fonte solúvel (SFS) por outra de baixa solubilidade associada à micro-organismos (MSF). / Phosphorus (P) which is a little available nutrient in tropical soils due to their disability and also because natural impoundment of the same constituents in the soil itself, forming insoluble phosphates and thus unavailable to plants. The solubilization of inorganic phosphate is a strategy to increase the availability of this nutrient in the soil. One way would be to use P-solubilizing microorganisms that make nutrients available to plants that. Thus, this study aimed to: (i) evaluate the growth of coffee as a function of the sources and levels of P2O5, (ii) determine the available phosphorus and (iii) assess the nutritional status of coffee, depending on the source and dose of P2O5. The development of coffee seedlings is higher with the use of soluble P source, in relation to the source of gradual solubility regardless of dose. The determination of the replacement capacity of the water soluble source of gradual solubility depends on the reaction time in the soil, whose maximum equivalence is equal to 84% at 180 days after fertilizer application (MSF). In soils with a sufficient level of available P for the cultivation of coffee is feasible to replace a water soluble (SFS) for another low solubility associated with micro-organisms (MSF).