Rozprawy doktorskie: „Soil DNA”

1

Mueller, Sabrina R. "Chromium, DNA, and Soil Microbial Communities". Cincinnati, Ohio : University of Cincinnati, 2006. http://rave.ohiolink.edu/etdc/view.cgi?acc_num=ucin1141334651.

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Thesis (Ph. D.)--University of Cincinnati, 2008.
Advisor: Brian K. Kinkle. Title from electronic thesis title page (viewed Apr. 23, 2009). Keywords: SEC-ICP-MS; Fungal community; bacterial community; DGGE. Includes abstract. Includes bibliographical references.

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2

Kapadia, Jaimin Maheshbhai. "DNA transfer in the soil bacterium Rhodococcus". Digital Commons @ East Tennessee State University, 2020. https://dc.etsu.edu/honors/565.

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Gene transfer plays an important role in bacterial evolution. Especially in an under explored species like Rhodococcus, a type of bacteria found in the soil. Rhodococcus has several applications in the pharmaceutical industry and in the production of antibiotics. Rhodococcus possess several unique sets of properties which makes it beneficial to have a reliable method of producing mutants of Rhodococcus. The goal of the experiment was to find an efficient way of forming Rhodococcus colonies with kanamycin resistant genes. The project began from an unexpected observation from an earlier experiment with Rhodococcus strain MTM3W5.2. where I attempted to transform this strain with a transposon via electro-transformation. The colonies that grew/ appeared transformants were screened to confirm the presence of kanamycin gene, however there was no amplified DNA seen on the PCR gel (i.e. absence of the kanamycin gene). The electro-transformant colonies were selected on LB plates containing different higher concentrations of kanamycin. Then the appeared transformants were again screened via disk diffusion assay and were classified into 3 different kanamycin resistant phenotypes. Majority of the “C” phenotypic colonies (i.e., high level resistance to kanamycin) appear to contain the kanamycin gene, but these colonies were less in numbers. This led us to try another method of gene transfer which is conjugation. Conjugation was carried on a double selection antibiotic plate containing both chloramphenicol (30 µg) and kanamycin (100 µg). The transconjugate colonies that appeared on the double selection plates were also screened by PCR, but none of the colonies had amplified DNA suggesting absence of the kanamycin gene. The colonies seen on the double selection plate were possibly due to spontaneous mutation or some type of unknown phenotypic variation. However, in the future, double selection plates with higher concentrations of antibiotics can possibly give us transconjugants with kanamycin genes.

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3

au, N. Williams@murdoch edu, i Nari Michelle Anderson. "DNA methods for the detection of Phytophthora cinnamomi from soil". Murdoch University, 2006. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20070820.130155.

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This project assesses two aspects of DNA detection of Phytophthora species from soil samples. Firstly, a nested PCR protocol was established with both primary and nested PCR specific for P. cinnamomi detection. PCR amplification of P. cinnamomi DNA isolated from soil was optimised with the addition of bovine serum albumin and formamide. This was found to improve both the specificity and sensitivity of PCR amplification of DNA in the presence of inhibitors co-extracted along with the target DNA from soil samples. The application of diagnostic nested PCR with the addition of BSA and formamide was verified by comparison with routine culture based detection methods. In all cases, nested PCR detection incorporating BSA and formamide was found to be considerably more sensitive than the culture based detection methods. The second component of this thesis investigates the simultaneous detection of multiple species of Phytophthora using microarray analysis. Microarray based detection has been previously limited by variable and inconsistent hybridisation intensities across the diversity of probes used in each array. In this study a novel concept for the differentiation of detection targets using duplex melting kinetics is introduced. A microarray assay was developed on a PamChip ¥ microarray enabling the differentiation of target Phytophthora species using the melting kinetics of probe-target duplexes. In the majority of cases the hybridization kinetics of target and non-target duplexes differed significantly. Analysis of the melting kinetics of duplexes formed by probes with target and non-target DNA was found to be an effective method for determining specific hybridization and was independent of fluctuations in hybridization signal intensity. This form of analysis was more robust than the traditional approach based on hybridisation intensity, and allowed the detection of individual Phytophthora species and mixtures there of.

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4

Anderson, Nari Michelle. "DNA methods for the detection of Phytophthora cinnamomi from soil". Thesis, Anderson, Nari Michelle (2006) DNA methods for the detection of Phytophthora cinnamomi from soil. PhD thesis, Murdoch University, 2006. https://researchrepository.murdoch.edu.au/id/eprint/42/.

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This project assesses two aspects of DNA detection of Phytophthora species from soil samples. Firstly, a nested PCR protocol was established with both primary and nested PCR specific for P. cinnamomi detection. PCR amplification of P. cinnamomi DNA isolated from soil was optimised with the addition of bovine serum albumin and formamide. This was found to improve both the specificity and sensitivity of PCR amplification of DNA in the presence of inhibitors co-extracted along with the target DNA from soil samples. The application of diagnostic nested PCR with the addition of BSA and formamide was verified by comparison with routine culture based detection methods. In all cases, nested PCR detection incorporating BSA and formamide was found to be considerably more sensitive than the culture based detection methods. The second component of this thesis investigates the simultaneous detection of multiple species of Phytophthora using microarray analysis. Microarray based detection has been previously limited by variable and inconsistent hybridisation intensities across the diversity of probes used in each array. In this study a novel concept for the differentiation of detection targets using duplex melting kinetics is introduced. A microarray assay was developed on a PamChip microarray enabling the differentiation of target Phytophthora species using the melting kinetics of probe-target duplexes. In the majority of cases the hybridization kinetics of target and non-target duplexes differed significantly. Analysis of the melting kinetics of duplexes formed by probes with target and non-target DNA was found to be an effective method for determining specific hybridization and was independent of fluctuations in hybridization signal intensity. This form of analysis was more robust than the traditional approach based on hybridisation intensity, and allowed the detection of individual Phytophthora species and mixtures there of.

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5

Anderson, Nari Michelle. "DNA methods for the detection of Phytophthora cinnamomi from soil". Anderson, Nari Michelle (2006) DNA methods for the detection of Phytophthora cinnamomi from soil. PhD thesis, Murdoch University, 2006. http://researchrepository.murdoch.edu.au/42/.

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This project assesses two aspects of DNA detection of Phytophthora species from soil samples. Firstly, a nested PCR protocol was established with both primary and nested PCR specific for P. cinnamomi detection. PCR amplification of P. cinnamomi DNA isolated from soil was optimised with the addition of bovine serum albumin and formamide. This was found to improve both the specificity and sensitivity of PCR amplification of DNA in the presence of inhibitors co-extracted along with the target DNA from soil samples. The application of diagnostic nested PCR with the addition of BSA and formamide was verified by comparison with routine culture based detection methods. In all cases, nested PCR detection incorporating BSA and formamide was found to be considerably more sensitive than the culture based detection methods. The second component of this thesis investigates the simultaneous detection of multiple species of Phytophthora using microarray analysis. Microarray based detection has been previously limited by variable and inconsistent hybridisation intensities across the diversity of probes used in each array. In this study a novel concept for the differentiation of detection targets using duplex melting kinetics is introduced. A microarray assay was developed on a PamChip microarray enabling the differentiation of target Phytophthora species using the melting kinetics of probe-target duplexes. In the majority of cases the hybridization kinetics of target and non-target duplexes differed significantly. Analysis of the melting kinetics of duplexes formed by probes with target and non-target DNA was found to be an effective method for determining specific hybridization and was independent of fluctuations in hybridization signal intensity. This form of analysis was more robust than the traditional approach based on hybridisation intensity, and allowed the detection of individual Phytophthora species and mixtures there of.

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6

Kasu, Mohaimin. "The validation of forensic DNA extraction systems to utilize soil contaminated biological evidence". Master's thesis, University of Cape Town, 2013. http://hdl.handle.net/11427/5921.

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7

Smart, Trevor Blake. "Microbial Community Response to Fumigation in Potato Soils". BYU ScholarsArchive, 2018. https://scholarsarchive.byu.edu/etd/7355.

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Soil microorganisms have a variety of beneficial and deleterious effects on plants, impacting such processes as plant growth, soil nutrient cycling, crop yield, disease resistance and tolerance to an array of biotic and abiotic stressors. The disruption of soil microbial community structures, particularly when beneficial soil biota are altered, has been shown to reduce crop yield and leave plants susceptible to disease. Long-term disruption of microbial communities may occur with repeated fumigation, being the application of gaseous pesticides, in agricultural soils. For this reason, we characterized bacterial, fungal, oomycete and nematode populations in paired fumigated and nonfumigated potato fields located in Idaho, Oregon, Washington and Minnesota. Samples were taken at three distinct timepoints: one before a fall fumigation event and two others at important stages in potato production, row closure and vine death. Soil biota populations were assessed by targeting the 16S, 18S and ITS1 gene regions. FunGuild, a database capable of guild and trophic assignment of fungal lineages, was used to sort fungal OTUs in different trophic modes. Fungal analyses indicated an increase in relative abundances of saprotrophic fungal populations and a decrease in pathotrophic fungal populations, both during row closure. Principally, the fungal genera of Humicola and Mortierella were responsible for the increase of saprotrophs while Alternaria decreased the most for pathotrophs. Other fungi occupying multiple trophic modes, such as Fusarium, also decreased during row closure. We found that fumigation treatments, in combination with various pesticide and fertilizer applications, alter both alpha- and beta- bacterial soil diversity although certain treatments, i.e. chloropicrin, may alter bacterial populations more than other treatment types such as metam-sodium. Nematode populations were likewise distinct at each location with soils from Boardman, OR, Minidoka, ID and Pine Point, MN with these having higher levels of nematodes associated with better soil health, i.e. Dorylaimidae. Conversely, nematodes associated with plant pathogenesis were found in higher relative abundances at Minidoka, ID and Quincy, WA. In this study, we characterize the populations of bacteria, fungi, oomycetes and nematodes with an emphasis on fungal taxa. We found that relative abundances of fungal trophic modes vary temporally. Additionally, we catalogue several other high abundance taxa with seasonal differential abundances whose functional capacity in potatoes remain uncharacterized.

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8

He, Jizheng, i n/a. "Molecular Biological Studies of Soil Microbial Communities Under Different Management Practices in Forest Ecosystems of Queensland". Griffith University. Australian School of Environmental Studies, 2005. http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20060309.095702.

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Soil microorganisms play important roles in maintaining soil quality and ecosystem health. Development of effective methods for studying the composition, diversity, and behavior of microorganisms in soil habitats is essential for a broader understanding of soil quality. Forest management strategies and practices are of vital significance for sustainable forest production. How the different forest management measures will influence soil microbial communities is a widespread concern of forest industry and scientific communities. Only a small proportion (~0.1%) of the bacteria from natural habitats can be cultured on laboratory growth media. Direct extraction of whole-community DNA from soil, followed by polymerase chain reaction (PCR) and other analysis circumvents the problems of the culture-dependent methods and may shed light on a broader range of microbial communities in the soil. DNA-based molecular methods rely on high quality soil microbial DNA as template, and thus extraction of good quality DNA from soil samples has been a challenge because of the complex and heterogeneous nature of the soil matrix. The objectives of this research were to establish a set of DNA-based molecular methods and to apply them to investigate forest soil microbial composition and diversity. Soil samples were collected from different forest ecosystems, i.e., the natural forest (YNF) and the first rotation (~ 50 years) (Y1R) and the second rotation (~ 1 year) (Y2R) of hoop pine plantations at Yarraman, and from different forest residue management practices (the experiments had established 6.4 years before the samples were collected) at Gympie, two long-term experimental sites of the Queensland Department of Primary Industry-Forestry in subtropical Queensland, Australia. Some DNA-based molecular techniques, including DNA extraction and purification, PCR amplification, DNA screening, cloning, sequencing and phylogenetic analyses, were explored using Yarraman soil samples, which were high in organic matter, clay and iron oxide contents. A set of methods was assembled based on the recommendations of the method development experiments and applied to the investigations of the microbial composition and diversity of the Yarraman and Gympie soil samples. Four soil DNA extraction methods, including the Zhou method (Zhou et al., 1996), the Holben method (Holben, 1994), the UltraClean (Mo Bio) and FastDNA (Bio 101) soil DNA extraction kits, were explored. It was necessary to modify these methods for Yarraman soil. I designed and introduced a pre-lysis buffer washing step, to partially remove soil humic substances and promote soil dispersion. This modification greatly improved the quality of the extracted DNA, decreasing co-extracted humic substances by 31% and increasing DNA yield by 24%. The improved Holben method was recommended for fungal community studies, and the improved Zhou method for bacterial community studies. The extracted DNA was good in quality, with a consistent size of ~20 kb and a yield of 48-87 g g-1 soil, and could be successfully used for 16S (Zhou method) and 18S (Holben method) rDNA amplifications. For less difficult environmental samples, UltraClean kits could be a good option, because they are simple and fast and the extracted DNA are also of good quality. Screening of the DNA PCR products using TGGE, Heteroduplex-TGGE and SSCP was also explored. These methods were not so effective for the screening of the soil DNA PCR products, owing to the difficulty in interpretation of the results. Cloning was a necessary step to obtain a single sequence at species level in soil microbial community studies. The screening of the clone library by TGGE, Heteroduplex-TGGE and SSCP could only separate the clones into several major bands, although SSCP gave better separation. Sequencing of selected clones directly from the clone library obtained ultimate results of microbial taxonomic composition and diversity through well-established sequence analysis software packages and the databases. It was recommended that, in this project with the target of microbial community composition and diversity, soil DNA PCR products were directly cloned to construct clone libraries and a sample of clones were sequenced to achieve an estimate of the taxonomic composition of the soil. Fungal communities of the Yarraman soil samples under the natural forest (YNF) and the hoop pine plantations (YHP) were investigated using 18S rDNA based cloning and sequencing approaches. Twenty-eight clone sequences were obtained and analysed. Three fungal orders, i.e., Zygomycota, Ascomycota and Basidiomycota were detected from the YNF and YHP samples. By contrast, culture-based analyses of fungi in the literature were mostly Ascomycetes. YNF appeared to have more Ascomycota but less Zygomycota than YHP, and within the Zygomycota order, YHP had more unidentified species than YNF. Bacterial communities of Yarraman soil samples of YNF, Y1R and Y2R were investigated using 16S rDNA-based cloning and sequencing approaches. 305 16S rDNA clone sequences were analysed and showed an overall bacterial community composition of Unclassified bacteria (34.4%), Proteobacteria (22.0%), Verrucomicrobia (15.7%), Acidobacteria (10.2%), Chloroflexi (6.9%), Gemmatimonadetes (5.6%), and Actinobacteria (5.2%). There was a significant difference among YNF, Y1R and Y2R in the taxonomic group composition. YNF had a greater proportion of Acidobacteria (18.0%), Verrucomicrobia (23.0%) and Chloroflexi (9.0%) than Y1R and Y2R (corresponding to 6.3%, 12.1% and 5.9%, respectively), while Y1R and Y2R had a higher percentage of the Unclassified group (38.5% for Y1R and 46.5% for Y2R) than YNF (18.0%). For the Proteobacteria group, YNF had more Alpha-subdivision but Y1R and Y2R had more Delta-subdivision. From YNF to Y1R to Y2R, the clone sequence variable site ratios, 5% and 10% OTU numbers and Shannon's diversity index H' values tended to decrease, indicating the soil bacterial diversity decreased from the natural forest to the first and the second rotation hoop pine plantations. The large amount of unclassified clone sequences could imply a novel group of bacteria in the soil, particularly in the hoop pine soil samples. Alternatively they may result from artefacts during the PCR process. Bacterial communities of the Gympie soil under different residue management practices, i.e., residue (litter plus logging residue) removed (G0R), residue retained (G1R), and residue doubled (G2R), were also investigated using the 16S rDNA-based cloning and sequencing approaches. Acidobacteria (37.6%) and Proteobacteria (35.6%, including Alpha-subdivision of 29.9% and Gamma-subdivision of 5.7%) were dominant components of the communities, followed by Actinobacteria (14.7%), Verrucomicrobia (7.3%) and Unclassified bacteria. There was no significant difference among G0R, G1R and G2R in the bacterial community compositions and diversity. These findings provided an in-depth vision of the soil microbial communities under different forest management practices. Their combination with other soil analysis results, such as physical and chemical properties, and forest production data, could provide an improved understanding of sustainable forest management strategies.

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9

He, Jizheng. "Molecular Biological Studies of Soil Microbial Communities Under Different Management Practices in Forest Ecosystems of Queensland". Thesis, Griffith University, 2005. http://hdl.handle.net/10072/367075.

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Soil microorganisms play important roles in maintaining soil quality and ecosystem health. Development of effective methods for studying the composition, diversity, and behavior of microorganisms in soil habitats is essential for a broader understanding of soil quality. Forest management strategies and practices are of vital significance for sustainable forest production. How the different forest management measures will influence soil microbial communities is a widespread concern of forest industry and scientific communities. Only a small proportion (~0.1%) of the bacteria from natural habitats can be cultured on laboratory growth media. Direct extraction of whole-community DNA from soil, followed by polymerase chain reaction (PCR) and other analysis circumvents the problems of the culture-dependent methods and may shed light on a broader range of microbial communities in the soil. DNA-based molecular methods rely on high quality soil microbial DNA as template, and thus extraction of good quality DNA from soil samples has been a challenge because of the complex and heterogeneous nature of the soil matrix. The objectives of this research were to establish a set of DNA-based molecular methods and to apply them to investigate forest soil microbial composition and diversity. Soil samples were collected from different forest ecosystems, i.e., the natural forest (YNF) and the first rotation (~ 50 years) (Y1R) and the second rotation (~ 1 year) (Y2R) of hoop pine plantations at Yarraman, and from different forest residue management practices (the experiments had established 6.4 years before the samples were collected) at Gympie, two long-term experimental sites of the Queensland Department of Primary Industry-Forestry in subtropical Queensland, Australia. Some DNA-based molecular techniques, including DNA extraction and purification, PCR amplification, DNA screening, cloning, sequencing and phylogenetic analyses, were explored using Yarraman soil samples, which were high in organic matter, clay and iron oxide contents. A set of methods was assembled based on the recommendations of the method development experiments and applied to the investigations of the microbial composition and diversity of the Yarraman and Gympie soil samples. Four soil DNA extraction methods, including the Zhou method (Zhou et al., 1996), the Holben method (Holben, 1994), the UltraClean (Mo Bio) and FastDNA (Bio 101) soil DNA extraction kits, were explored. It was necessary to modify these methods for Yarraman soil. I designed and introduced a pre-lysis buffer washing step, to partially remove soil humic substances and promote soil dispersion. This modification greatly improved the quality of the extracted DNA, decreasing co-extracted humic substances by 31% and increasing DNA yield by 24%. The improved Holben method was recommended for fungal community studies, and the improved Zhou method for bacterial community studies. The extracted DNA was good in quality, with a consistent size of ~20 kb and a yield of 48-87 g g-1 soil, and could be successfully used for 16S (Zhou method) and 18S (Holben method) rDNA amplifications. For less difficult environmental samples, UltraClean kits could be a good option, because they are simple and fast and the extracted DNA are also of good quality. Screening of the DNA PCR products using TGGE, Heteroduplex-TGGE and SSCP was also explored. These methods were not so effective for the screening of the soil DNA PCR products, owing to the difficulty in interpretation of the results. Cloning was a necessary step to obtain a single sequence at species level in soil microbial community studies. The screening of the clone library by TGGE, Heteroduplex-TGGE and SSCP could only separate the clones into several major bands, although SSCP gave better separation. Sequencing of selected clones directly from the clone library obtained ultimate results of microbial taxonomic composition and diversity through well-established sequence analysis software packages and the databases. It was recommended that, in this project with the target of microbial community composition and diversity, soil DNA PCR products were directly cloned to construct clone libraries and a sample of clones were sequenced to achieve an estimate of the taxonomic composition of the soil. Fungal communities of the Yarraman soil samples under the natural forest (YNF) and the hoop pine plantations (YHP) were investigated using 18S rDNA based cloning and sequencing approaches. Twenty-eight clone sequences were obtained and analysed. Three fungal orders, i.e., Zygomycota, Ascomycota and Basidiomycota were detected from the YNF and YHP samples. By contrast, culture-based analyses of fungi in the literature were mostly Ascomycetes. YNF appeared to have more Ascomycota but less Zygomycota than YHP, and within the Zygomycota order, YHP had more unidentified species than YNF. Bacterial communities of Yarraman soil samples of YNF, Y1R and Y2R were investigated using 16S rDNA-based cloning and sequencing approaches. 305 16S rDNA clone sequences were analysed and showed an overall bacterial community composition of Unclassified bacteria (34.4%), Proteobacteria (22.0%), Verrucomicrobia (15.7%), Acidobacteria (10.2%), Chloroflexi (6.9%), Gemmatimonadetes (5.6%), and Actinobacteria (5.2%). There was a significant difference among YNF, Y1R and Y2R in the taxonomic group composition. YNF had a greater proportion of Acidobacteria (18.0%), Verrucomicrobia (23.0%) and Chloroflexi (9.0%) than Y1R and Y2R (corresponding to 6.3%, 12.1% and 5.9%, respectively), while Y1R and Y2R had a higher percentage of the Unclassified group (38.5% for Y1R and 46.5% for Y2R) than YNF (18.0%). For the Proteobacteria group, YNF had more Alpha-subdivision but Y1R and Y2R had more Delta-subdivision. From YNF to Y1R to Y2R, the clone sequence variable site ratios, 5% and 10% OTU numbers and Shannon's diversity index H' values tended to decrease, indicating the soil bacterial diversity decreased from the natural forest to the first and the second rotation hoop pine plantations. The large amount of unclassified clone sequences could imply a novel group of bacteria in the soil, particularly in the hoop pine soil samples. Alternatively they may result from artefacts during the PCR process. Bacterial communities of the Gympie soil under different residue management practices, i.e., residue (litter plus logging residue) removed (G0R), residue retained (G1R), and residue doubled (G2R), were also investigated using the 16S rDNA-based cloning and sequencing approaches. Acidobacteria (37.6%) and Proteobacteria (35.6%, including Alpha-subdivision of 29.9% and Gamma-subdivision of 5.7%) were dominant components of the communities, followed by Actinobacteria (14.7%), Verrucomicrobia (7.3%) and Unclassified bacteria. There was no significant difference among G0R, G1R and G2R in the bacterial community compositions and diversity. These findings provided an in-depth vision of the soil microbial communities under different forest management practices. Their combination with other soil analysis results, such as physical and chemical properties, and forest production data, could provide an improved understanding of sustainable forest management strategies.
Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
Australian School of Environmental Studies
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10

Spiegelman, Dan. "Exploring the fusion of metagenomic library and DNA microarray technologies". Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=98805.

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We explored the combination of metagenomic library and DNA microarray technologies into a single platform as a novel way to rapidly screen metagenomic libraries for genetic targets. In the "metagenomic microarray" system, metagenomic library clone DNA is printed on a microarray surface, and clones of interest are detected by hybridization to single-gene probes. This study represents the initial steps in the development of this technology. We constructed two 5,000-clone large-insert metagenomic libraries from two diesel-contaminated Arctic soil samples. We developed and optimized an automated fosmid purification protocol to rapidly-extract clone DNA in a high-throughput 96-well format. We then created a series of small prototype arrays to optimize various parameters of microarray printing and hybridization, to identify and resolve technical challenges, and to provide proof-of-principle of this novel application. Our results suggest that this method shows promise, but more experimentation must be done to establish the feasibility of this approach.

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11

Cappelini, Luciana Teresa Dias. "Influência das substâncias húmicas na degradação do pesticida fipronil pela bactéria Burkholderia sp". Universidade de São Paulo, 2013. http://www.teses.usp.br/teses/disponiveis/75/75135/tde-30072013-093208/.

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Devido à expansão da cultura canavieira, e aos altos custos dos insumos agrícolas, buscam-se novas formas de manejo do solo dentre elas o uso de substancias húmicas. Apesar disso os produtores aliam as técnicas alternativas as tradicionais como o uso do fipronil, um inseticida fenil - pirazólico muito utilizado na cultura de cana-de-açúcar. Apesar desta prática, pouco se sabe como ocorre à degradação biótica do fipronil, fipronil-sulfeto e fipronil sulfona, quando se utiliza as substâncias húmicas como forma de adubação. Por esse motivo o objetivo deste trabalho foi avaliar a degradação biológica do fipronil quando se utiliza substâncias húmicas. Para este estudo, empregou-se um método indireto de extração de DNA do solo, onde as bactérias foram cultivadas em meio contendo apenas água estéril e fipronil e posteriormente seu DNA foi extraído, fez-se a PCR utilizando o primer 27F/1100R e o produto da PCR foi clonado e sequenciado; as bactérias identificadas estão afiliadas aos gêneros: Clostridium sp, Bdellovibrio sp, Flavisolibacter sp, Burkholderia sp e Herbaspirillum sp. O gênero selecionado para o estudo foi afiliada a Burkholderia sp, devido a seu potencial de degradação citado na literatura. Adquiriu-se uma cepa pura de Burkholderia thailandensis Para determinação do fipronil e seus metabólitos, fipronil sulfeto e fipronil sulfona validou-se o método QuEChERS GC- MS que resultou nas seguintes condições: limites de detecção e quantificação foram de 0,06 mgL-1 0,25 mgL-1 respectivamente, a linearidade foi de 0,99, a precisão foi em torno de 10,9 a 7,3% para o nível baixo e 1,84 a 1,24% para o nível mais alto. Calculou-se a recuperação do método de extração que variou entre 78 e 98%, Em seguida montou-se oito tratamentos os quais foram medidos a degradação do fipronil frente a utilização de SH que obteve-se o seguintes resultados: A Burkholderia thailandensis degradou cerca de 0,75 mg L-1 de fipronil com ou sem a presença das SH adicionada no solo de estudo; não foi possível quantificar os produtos de degradação pois ambos (fipronil sulfeto e fipronil sulfona) ficaram abaixo do LOQ do método.
Due to the expansion of sugar cane, and the high costs of agricultural inputs, currently looking up forms new of soil management among them the use of humic substances. Nevertheless producers combine traditional and alternative techniques, for example addition of fipronil to humic substances. Fipronil is an insecticide phenyl - pyrazole widely used in cane sugar cultivation . Despite this practice, few is known about a biotic degradation of the fipronil, fipronil sulfide and fipronil sulfone, when using humic substances as fertilization. Therefore the objective of this study was to evaluate the biological degradation of fipronil when it used in humic substances. For this study, we used an indirect method of DNA extraction from the soil, where bacteria were cultured in medium containing only sterile water and fipronil. After, the DNA was extracted,a PCR was performing using primer 27F/1100Rr and the PCR product was cloned and sequenced. The bacteria identified were affiliated to the genus Clostridium sp., Bdellovibrio sp., Flavisolibacter sp., Burkholderia sp. and Herbaspirillum sp. The gene selected for the study was affiliated with Burkholderia sp due to its potential degradation reported in the literature. The study strain was acquired in public banks, and was selected to Burkholderia thailandensis. For the determination of fipronil and its metabolites, the method QuEChERS follow by analysis with GC-MS was validated to fipronil, fipronil sulfide, and fipronil sulfone. To this method was found the follow conditions: detection and quantitation limits were 0.06 mg L-1 0.25 mgL-1 respectively, the linearity was 0.99, the precision was about 10.9 to 7.3% for the lowest level and 1.84 to 1.24% for the highest level. We calculated the recovery of the extraction method which ranged between 78 and 98%. Then were assembled eight treatments, to evaluate the degradation of the fipronil and the formation of its two degradation products biotic, fipronil sulfide and fipronil sulfone, when using humic substances in fertilization. The Burkholderia thailandensis degraded approximately 0.75 mg L-1 fipronil with or without the presence of SH in the study soil, but it was not possible to quantify the degradation products of the fipronil (fipronil sulfide and fipronil sulfone) because both were detected but cannot be quantified because their values are below the limit of quantification of the method.

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12

Rosa, Márcia Maria. "Avaliação de diferentes metodologias para extração de DNA de solo sob cultivo de cana-de-açúcar /". Rio Claro : [s.n.], 2006. http://hdl.handle.net/11449/95004.

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Resumo: O solo é um ecossistema caracterizado pela grande complexidade de difícil estudo, devido à sua heterogeneidade, especialmente os microrganismos do solo. Atualmente, ferramentas da biologia molecular têm sido usadas para mostrar o potencial biotecnológico do solo, através dos genes microbianos. A extração direta do DNA é uma etapa importante nesse tipo de estudo, porém, continua sendo um obstáculo para a avaliação da diversidade microbiana do solo. Esses estudos no Brasil apresentam algumas dificuldades, uma vez que a maioria das técnicas foi desenvolvida para solos de clima temperado. Desta forma, o objetivo deste estudo foi avaliar dez diferentes técnicas para extração direta de DNA de solos em áreas de cultura de cana-de-açúcar sob manejo orgânico e convencional. A eletroforese se apresentou como a técnica mais adequada para se conhecer a eficiência da extração do DNA, através da intensidade e tamanho de suas bandas. A técnica de Selbach (SEL) apresentou melhores resultados, com bandas de DNA mais intensas e sem arraste, indicando a obtenção de uma solução com menores teores de contaminantes. Com a técnica de Direito (DIR) também se verificou bandas de DNA, porém menos fortes e sem arraste. A técnica proposta neste estudo (PRO) resultou em bandas de DNA fortes, porém com arraste, indicando a necessidade de uma etapa para purificação do DNA extraído. Esta mostrou ser a metodologia de mais fácil e rápida execução, sendo que, após processo de purificação a região 16S do DNA ribossomal, utilizando-se primers universais, foi amplificada satisfatoriamente a partir do DNA obtido do solo. Bandas de DNA apresentadas na eletroforese a partir das amostras de solo sob viii manejo orgânico foram mais intensas do que aquelas de manejo convencional. Estes resultados estão relacionados com a maior quantidade de biomassa microbiana presente no solo orgânico... (Resumo completo, clicar acesso eletrônico abaixo).
Abstract: Soil is an ecosystem characterized by a great complexity and hard to study due to its heterogeneity, especially the soil microorganisms. Nowadays, molecular biology tools have been used to show the biotechnological potential of the soil mainly through microbial genes. Direct extraction of DNA from soil is an important step in this kind of study, however it is still an obstacle for microbial diversity evaluation. Besides, the majority of techniques proposed was developed for soils from temperate climate. This work aimed to evaluate ten different techniques for soil direct DNA extraction from sugar cane crop areas under organic and conventional managements, which presented distinct results. Gel electrophoresis was the most appropriate technique to evaluate the efficiency of DNA extraction, through the intensity and size of the bands. The Selbach technique (SEL) showed the best results, with more intense DNA bands and without smearing , indicating that a DNA solution with low concentration of contaminants was obtained. With the Direito technique (DIR) DNA bands were also verified, but less intensive and also without smearing. The technique proposed in this study (PRO) resulted in intense DNA however with smearing, indicating that a DNA purification step is necessary. This technique was easy, cheap and rapid to execute, enabling the amplification of 16S rDNA (using universal primers) after DNA solution purification. The intensity of DNA bands, as revealed by electrophoresis, was higher when using DNA solution extracted from soil under organic management, which also presented higher microbial biomass. This study is a contribuition for the selection and improvement of molecular tools to the study of microbial diversity applied to Brazilian soils.
Orientador: Sâmia Maria Tauk-Tornisielo
Coorientador: Sandra Regina Ceccato-Antonini
Banca: Marli de Fatima Fiore
Banca: Welington Luiz de Araujo
Mestre

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Rosa, Márcia Maria [UNESP]. "Avaliação de diferentes metodologias para extração de DNA de solo sob cultivo de cana-de-açúcar". Universidade Estadual Paulista (UNESP), 2006. http://hdl.handle.net/11449/95004.

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O solo é um ecossistema caracterizado pela grande complexidade de difícil estudo, devido à sua heterogeneidade, especialmente os microrganismos do solo. Atualmente, ferramentas da biologia molecular têm sido usadas para mostrar o potencial biotecnológico do solo, através dos genes microbianos. A extração direta do DNA é uma etapa importante nesse tipo de estudo, porém, continua sendo um obstáculo para a avaliação da diversidade microbiana do solo. Esses estudos no Brasil apresentam algumas dificuldades, uma vez que a maioria das técnicas foi desenvolvida para solos de clima temperado. Desta forma, o objetivo deste estudo foi avaliar dez diferentes técnicas para extração direta de DNA de solos em áreas de cultura de cana-de-açúcar sob manejo orgânico e convencional. A eletroforese se apresentou como a técnica mais adequada para se conhecer a eficiência da extração do DNA, através da intensidade e tamanho de suas bandas. A técnica de Selbach (SEL) apresentou melhores resultados, com bandas de DNA mais intensas e sem arraste, indicando a obtenção de uma solução com menores teores de contaminantes. Com a técnica de Direito (DIR) também se verificou bandas de DNA, porém menos fortes e sem arraste. A técnica proposta neste estudo (PRO) resultou em bandas de DNA fortes, porém com arraste, indicando a necessidade de uma etapa para purificação do DNA extraído. Esta mostrou ser a metodologia de mais fácil e rápida execução, sendo que, após processo de purificação a região 16S do DNA ribossomal, utilizando-se primers universais, foi amplificada satisfatoriamente a partir do DNA obtido do solo. Bandas de DNA apresentadas na eletroforese a partir das amostras de solo sob viii manejo orgânico foram mais intensas do que aquelas de manejo convencional. Estes resultados estão relacionados com a maior quantidade de biomassa microbiana presente no solo orgânico... .
Soil is an ecosystem characterized by a great complexity and hard to study due to its heterogeneity, especially the soil microorganisms. Nowadays, molecular biology tools have been used to show the biotechnological potential of the soil mainly through microbial genes. Direct extraction of DNA from soil is an important step in this kind of study, however it is still an obstacle for microbial diversity evaluation. Besides, the majority of techniques proposed was developed for soils from temperate climate. This work aimed to evaluate ten different techniques for soil direct DNA extraction from sugar cane crop areas under organic and conventional managements, which presented distinct results. Gel electrophoresis was the most appropriate technique to evaluate the efficiency of DNA extraction, through the intensity and size of the bands. The Selbach technique (SEL) showed the best results, with more intense DNA bands and without smearing , indicating that a DNA solution with low concentration of contaminants was obtained. With the Direito technique (DIR) DNA bands were also verified, but less intensive and also without smearing. The technique proposed in this study (PRO) resulted in intense DNA however with smearing, indicating that a DNA purification step is necessary. This technique was easy, cheap and rapid to execute, enabling the amplification of 16S rDNA (using universal primers) after DNA solution purification. The intensity of DNA bands, as revealed by electrophoresis, was higher when using DNA solution extracted from soil under organic management, which also presented higher microbial biomass. This study is a contribuition for the selection and improvement of molecular tools to the study of microbial diversity applied to Brazilian soils.

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Kumpula, Kimmo. "Systematic comparison of the relative accuracy of vegetation surveys and soil DNA metabarcoding : Assessing plant biodiversity at different spatial scales". Thesis, Umeå universitet, Institutionen för ekologi, miljö och geovetenskap, 2020. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-172130.

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Analysis of soil-derived DNA has been shown to minimize problems seen during traditional vegetation surveys, e.g. by matching the eDNA to a reference database for taxonomic identification rather than relying solely on taxonomic expertise. However, it has been debated to what extent and how accurately eDNA acts as a proxy for biodiversity. The reliability on eDNA and the awareness on influencing factors is also important for palaeoenvironmental reconstructions where above-ground vegetation cannot be used. This study aimed to investigate how well modern soil-derived eDNA reflects the contemporary vascular vegetation communities in a subarctic environment, and if the efficiency of the taxonomic identification differed between spatial scales. Near-surface soil samples at altitudinal gradients along numerous transects were collected in combination with vegetation surveys. The eDNA was amplified through metabarcoding using the P6 loop region of the chloroplast trnL intron, followed by a high-throughput sequencing. No difference in the number of identified taxa between eDNA and vegetation survey was seen at landscape scale. In contrast, the number of identified taxa was consistently higher in the vegetation survey at smaller spatial scales. The efficiency of identified taxa per scale remained stable for the vegetation survey, whereas for eDNA, a decreasing trend was seen. This study highlights the variations on taxa identification between both methods and which factors might cause it. Combining the methods allows for a more precise modern biodiversity estimation, as well as to minimize wrongful conclusions. This allows for a more accurate palaeoenvironmental reconstructions, which in turn can improve future species conservation decisions.

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Igun, Onotasamiderhi Tarric. "Employing the power of DNA-based microbial community structure analysis for the rational design of hydrocarbon contaminated soil remediation". Thesis, University of Newcastle upon Tyne, 2017. http://hdl.handle.net/10443/3796.

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The impact of activated carbon (AC) amendments on the biodegradation of crude oil in soil was studied in batch microcosms. AC amendment slowed down biodegradation and removal of hydrocarbon pollutants and was more evident when AC when added from the beginning compared to after 5 months. The microbial sequencing analysis revealed that the overall bacterial community shifted more due to crude oil addition compared to AC amendment at the start and after 5 months. AC amendments reduced and slightly reduced the abundance of hydrocarbon degraders belonging to Actinobacteria and classes Gammaproteobacteria and Alphaproteobacteria (Such as Rhodococcus, Marinobacter, and Parvibaculum) in crude oil batches with AC from start and AC after 5 months, respectively. The effect of biofuels on the natural attenuation of toluene was also investigated. 13C/12C-CO2 batch production showed that biofuel components were preferentially degraded in blended fuels. Ethanol had a more negative effect on toluene degradation compared to biodiesel, as it was preferentially degraded even with nutrient bio-stimulation. The microbial analysis revealed nutrient effect on the microbial communities with nitrifying bacteria Nitrospira seen to make significant gains in OTU ranking and relative abundance due to the nutrient amendment. The microbial community analysis also showed a distinction in microbial communities that degrade biodiesel, ethanol and toluene in the presence or absence of nutrients. For instance the results showed that Rhodococcus can degrade toluene in the presence of biodiesel when nutrients are surplus or scarce and can degrade toluene in the presence of ethanol only when nutrients are scarce. Indicating a lower effect of biodiesel on toluene degraders compared to ethanol. Pseudomonas was identified as a key ethanol degrader and thrives in presence of both ethanol and toluene when nutrient availability is high but has preference for ethanol as a carbon source. Nocardia is main biodiesel degrader when nutrient availability is high or low. This study has shown that the use of DNA microbial community analysis gives a broader insight into microbes involved in the physiological activities and how they are affected by certain treatments and the findings in this study would possibly aid the understanding of the impact of adsorbents on hydrocarbon pollutants and the effect of blended fuels have on the microbiology within soil. It is thereby recommended that during hydrocarbon soil remediation studies/interventions, DNA microbial community analysis should be carried out in conjunction with chemical analysis such as the one carried out in this study as this would inform the proper utilization of the remediation strategies. The work was carried out in batch studies and it is important to repeat it in column, mesocosms or field studies to see if there are any significant changes in the results.

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Diawara, Aïssatou. "Development of DNA assays for the detection of single nucleotide polymorphism associated with benzimidazole resistance, in human soil-transmitted helminths". Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=19242.

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Soil-transmitted helminths are parasitic worms of humans, causing many disabilities in tropical parts of the developing world. Control programs such as "The Focussing Resources on Effective School Health” (FRESH) Partnership have been implemented to remove human soil transmitted nematodes through large-scale use of benzimidazole anthelmintic drugs for school-aged children in developing countries. The benzimidazole drugs, albendazole and mebendazole are used as a single annual dose in areas where the burden is high. Unfortunately, there is concern that increased use of anthelmintics in children could select for resistant populations of these human parasites. In filarial nematodes, a single amino acid substitution from phenylalanine (Phe) to tyrosine (Tyr), known to be associated with benzimidazole resistance in other nematodes, has been found in parasite ß-tubulin at position 200. We have developed pyrosequencer assays for the codon 200 in Trichuris trichiura, Ascaris lumbricoides, and the hookworm Necator americanus to screen for this single nucleotide polymorphism (SNP). Assays for this resistance-associated SNP could be useful for monitoring for anthelmintic resistance in control programs. These assays have been tested on adult worms from a benzimidazle-naïve population in Kenya. Following this, these assays have been applied on individual worms, pooled eggs and pooled larvae from people in East Africa, the Caribbean and Central America where mass drug anthelmintic programs have been implemented. The 200Tyr SNP was detected in T. trichiura from non-treated people and in T. trichiura and N. americanus from benzimidazole-treated people.
Les géo-helminthes sont des vers parasitant l'Homme et causant de nombreux handicaps dans les régions tropicales des pays en voie de développement. Des programmes de contrôles tels que le partenariat FRESH : ''Focussing Resources on Effective School Health'' ont été mis en place afin d'éliminer les géo-helminthes en administrant massivement en milieu scolaire des pays en voie de développement des medicaments anthelmintiques. L'albendazole et le mébendazole appartiennent au groupe des benzimidazoles et sont distribués dans les régions grandement infestées. Cependant, cette attribution massive de médicaments, aux enfants, pourrait entraîner une sélection de parasites résistants aux anthelmintiques. La substitution de l'acide aminé phénylalanine (Phe) par la tyrosine (Tyr) connue pour être associée à la résistance aux benzimidazoles chez les nématodes a été identifiée chez les filaires à la position 200 du gène de la ß-tubuline. Nous avons développés des tests pour les parasites Trichuris trichiura, Ascaris lumbricoides et Necator americanus en utilisant la méthode du pyroséquençage afin de détecter le polymorphisme d'un unique nucleotide (SNP) au niveau du codon 200 du gène de la ß-tubuline. Ce test a été appliqué sur des vers adultes provenant d'individus du Kenya n'ayant jamais été traités par des anthelmintiques. Puis ce même test a été appliqué à des vers adultes individuels, à des pools d'œufs et de larves provenant d'individus d'Afrique de l'est, des Caraïbes et d'Amérique centrale, où les programmes de contrôles de masse sont implantés. Le SNP fût détecté chez T. trichiura provenant d'individus non-traités aux benzimidazoles ainsi que chez T. trichiura et N. ameri

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Pillai, Suresh Divakaran. "Ecology and genetic stability of Tn5 mutants of bean rhizobia in Sonoran desert soils". Diss., The University of Arizona, 1989. http://hdl.handle.net/10150/184823.

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Five transposon Tn5 mutants of bean rhizobia (Rhizobium leguminosarum b.v. phaseoli) and the wild type strain were used in ecological studies to evaluate the efficacy of transposon Tn5 as a phenotypic marker in rhizobia for ecological studies in two Sonoran desert soils. All mutants possessed chromosomal insertions of the transposable element. Survival of each mutant strain was compared to that of the wild type strain under non stress, moisture stress and temperature stress conditions in Pima silty clay loam and Brazil to sandy loam. The genetic stability of Tn5 in terms of transposition of the element within the chromosome and the Tn5 coded antibiotic resistant phenotype was determined in cells recovered throughout the survival period. Under non stress conditions, the viable Tn5 mutant population decreased in size. Two mutants showed significantly (p < 0.01) lower populations than the wild type at the end of 30 days in the silty clay loam. In the sandy loam, four of the five mutant populations were significantly lower than the wild type. Tn5 was genetically stable in both soils. Under moisture stress conditions, the decline of the Tn5 mutant and wild type populations corresponded to a decline in soil moisture content. The finer textured soil afforded more protection to the cells than the coarse textured soil. There were no indications of Tn5 instability under moisture stress. In both soils under temperature stress, sizes of all populations declined rapidly and after 12 days, the mutant cells when screened using the Tn5 coded markers were significantly less in numbers than the wild type indicating a loss of Tn5 coded antibiotic resistance phenotype. There were no significant differences in numbers between wild type and mutant cells when screened using only the intrinsic markers. DNA:DNA hybridizations confirmed that the lack of Tn5 coded antibiotic resistance phenotype was probably not due to a deletion or transposition of the element. Under non stress conditions Tn5 is a useful ecological marker, but each Tn5 mutant has to be evaluated independently under specific environmental conditions to determine the efficacy of Tn5 as an ecological marker.

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Horton, Dean J. "Using molecular techniques to investigate soil invertebrate communities in temperate forests". Kent State University / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=kent1448799316.

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Moreno, Lilliana I. "The Effect of Sample and Sample Matrix on DNA Processing: Mechanisms for the Detection and Management of Inhibition in Forensic Samples". FIU Digital Commons, 2015. http://digitalcommons.fiu.edu/etd/1764.

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The presence of inhibitory substances in biological forensic samples has, and continues to affect the quality of the data generated following DNA typing processes. Although the chemistries used during the procedures have been enhanced to mitigate the effects of these deleterious compounds, some challenges remain. Inhibitors can be components of the samples, the substrate where samples were deposited or chemical(s) associated to the DNA purification step. Therefore, a thorough understanding of the extraction processes and their ability to handle the various types of inhibitory substances can help define the best analytical processing for any given sample. A series of experiments were conducted to establish the inhibition tolerance of quantification and amplification kits using common inhibitory substances in order to determine if current laboratory practices are optimal for identifying potential problems associated with inhibition. DART mass spectrometry was used to determine the amount of inhibitor carryover after sample purification, its correlation to the initial inhibitor input in the sample and the overall effect in the results. Finally, a novel alternative at gathering investigative leads from samples that would otherwise be ineffective for DNA typing due to the large amounts of inhibitory substances and/or environmental degradation was tested. This included generating data associated with microbial peak signatures to identify locations of clandestine human graves. Results demonstrate that the current methods for assessing inhibition are not necessarily accurate, as samples that appear inhibited in the quantification process can yield full DNA profiles, while those that do not indicate inhibition may suffer from lowered amplification efficiency or PCR artifacts. The extraction methods tested were able to remove >90% of the inhibitors from all samples with the exception of phenol, which was present in variable amounts whenever the organic extraction approach was utilized. Although the results attained suggested that most inhibitors produce minimal effect on downstream applications, analysts should practice caution when selecting the best extraction method for particular samples, as casework DNA samples are often present in small quantities and can contain an overwhelming amount of inhibitory substances.

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Buckley, Elan. "Change in the Structure of Soil Microbial Communities in Response to Waste Amendments". Thesis, Virginia Tech, 2020. http://hdl.handle.net/10919/101499.

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Soil microbial communities are affected extensively by addition of amendments to their environment. Of particular concern is the addition of poultry litter, which contains a substantial C, energy, and nutrient supply, but also antibiotic resistance genes (ARG), antimicrobials, and a multitude of microbial species. This project seeks to primarily assess if there is a change in bacterial community structure in response to poultry litter amendments to pasture land across geographically independent land across northern Georgia. It may be that changes in the relative abundance of bacterial communities also result in alteration in ARGs, and the community resistance to antibiotics (“resistome”) which in turn increases the potential threat of antibiotic resistance genes. While another part of this study will determine changes in integrons and specific ARGs, this project will focus on changes in bacterial communities and the potential functional changes in the community, which in turn have consequences for ARG levels and its horizontal transfer to various members of the soil community. Addition of waste from livestock is a historical method for increasing nutrients needed in the soil for the cultivation of crops, and in turn causes pronounced shifts in soil microbial communities due to the addition of large amounts of carbon, nutrients, foreign microbes, and other material. This study is unique because it utilizes a novel and relatively large landscape-scale to determine if there are discernable and repeatable patterns of bacterial community structure change in response to amendment regardless of exact soil type or source of chicken litter amendment. In the future, these data can also provide insight into the changes in the relative abundance antibiotic related genes associated with community change.
M.S.
Soil is complicated, both in terms of its physical makeup and the organisms that live inside of it. Predicting changes in soil based on the addition of foreign material such as chemicals or biological waste is not an easy process, and whether or not it is even possible to reliably predict those changes is a matter of some dispute. This study is designed to illustrate that such changes can in fact be reliably and consistently predicted even with regard to the addition of complicated materials to the soil. In this study, specifically, the material in question is chicken litter. A mix of the bedding and waste produced by chickens, litter is commonly handled by composting and is added to soil in farms as a fertilizer rich in organic matter. It is possible to point at specific elements of the soil such as the chemistry and bacteria and see how it is changed with the addition of chicken litter, which allows us to determine the nature and extent of the change that chicken litter has on soil. This study is conducted on a larger scale than similar experiments conducted in the past, making it apparent that these relationships exist on a repeated basis. It is the object of this study to pave the way and make it easier for scientists in the future to determine these relationships in other unique contexts.

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Mattiello, Lucia. "Fisiologia e transcriptoma de milho cultivado em solo ácido". [s.n.], 2010. http://repositorio.unicamp.br/jspui/handle/REPOSIP/317449.

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Orientadores: Marcelo Menossi Teixeira, Renato Atílio Jorge
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A presença do alumínio (Al) em solos ácidos é o principal fator limitante da produtividade agrícola no Brasil e no mundo. A resposta desenvolvida pelas plantas contra o Al é complexa e a identificação de genes responsivos após a exposição ao íon através de técnicas de análise em larga escala, como microarrays, pode facilitar a sua compreensão. Este projeto possui como objetivo ampliar o conhecimento sobre a fisiologia e a regulação gênica de raízes e folhas utilizando genótipos contrastantes de milho (Cat100-6 (Al-tolerante) e S1587-17 (Al-sensível)) cultivadas em solo ácido com concentração fitotóxica de Al. As linhagens de milho Cat100-6 e S1587-17 foram cultivadas por um ou três dias em solo ácido (pH 4,1) ou solo corrigido com Ca(OH)2 (pH 5,5). O genótipo S1587-17 apresentou uma maior inibição do crescimento radicular, resultado este altamente correlacionado com a acumulação de Al nos ápices radiculares e deposição de calose. Os dados fisiológicos confirmam a discriminação entre as duas linhagens em solo, abrindo perspectivas para entender pela primeira vez a base molecular das alterações das plantas em condições próximas à realidade de campo. O transcriptoma de raízes possibilitou a identificação possíveis candidatos a tolerância ao Al. Adicionalmente, com um experimento de hidroponia separamos as variáveis pH e presença de Al, ambas condições diferenciais do tratamento com solo. Identificamos, entre os candidatos, genes responsivos pela presença do Al e não pela acidez delimitando assim os genes com possíveis papéis na tolerância ao Al presente no solo ácido a apenas três: retinol desidrogenase, um fator de transcrição WRKY e uma proteína desconhecida. Esses resultados permitem concluir que o cultivo em solo é diferencial em relação à hidroponia, e outros fatores que apenas presentes no substrato solo podem provocar a indução de alguns genes. Diversas vias metabólicas são afetadas na linhagem sensível pelo tratamento em solo ácido e podem estar envolvidas na inibição radicular como a produção de lignina, celulose e calose e a síntese de etileno e auxina. O mapeamento nos cromossomos dos genes identificados pelo experimento de microarray das raízes de milho permitiu a identificação de genes localizados dentro de QTLs de milho previamente descritos na literatura como responsáveis pelo fenótipo tolerante. Diante esse resultado, podemos especular o papel de genes como uma proteína ligadora de RNA, uma inibidora de proteases e ciclinas na tolerância ao Al contido no solo ácido. Pela primeira vez na literatura, o transcriptoma de folhas coletadas após três dias de cultivo em solo ácido ou solo corrigido foi obtido com o uso de microarrays da Affymetrix. Essa análise indicou profundas alterações na Cat100-6, em contraposição à ausência de alteração significativa nas folhas na S1587-17. Genes referentes à fotossíntese e a fotorrespiração foram regulados negativamente pelo tratamento em solo ácido no genótipo tolerante. Contudo, o ciclo do ácido cítrico está ativado indicando uma putativa participação da produção de ácidos orgânicos nas folhas na resposta ao Al
Abstract: The presence of aluminum (Al) is the main factor limiting crops yield in Brazil and worldwide. The plant responses developed against this ion are complex and the identification of responsive genes after exposure to the ion with the use of a large scale technique, such as microarrays, can facilitate its comprehension. This project aimed to amplify the knowledge about physiology and gene expression regulation of roots and leaves associated towards Al resistance using contrasting maize genotypes (Cat100-6 (Al-tolerant) and S1587-17 (Al-sensitive) cultivated in acid soil containing phytotoxic concentrations of Al. Maize lines Cat100-6 and S1587-17 were cultivated for one or three days in acid soil (pH 4,1) or limed soil with Ca(OH)2 (pH 5,5). The genotype S1587-17 presented a higher root growth inhibition, which is highly correlated with Al accumulation in the root apexes and callose deposition. The physiological data confirms the discrimination of the two maize lines cultivated in soil, opening perspective to understand for the first time the molecular bases of alterations in plants on a closer condition to the field. Transcriptome from roots made possible the identification of possible tolerance candidates and genes constitutively expressed genes in the tolerant line. Additionally, throw a hydroponic experiment we splited the variables pH and Al presence, both differential conditions between soil treatments. It was possible to identify, among the candidates, genes responsive in the presence of Al in acid soil rather than acidity limiting genes with a possible roles in Al present in the acid soil tolerance to only three: retinol dehydrogenase, the transcription factor WRKY and an unknown protein. These results allow the conclusion that the soil culture is different in relation to hydropony, and other factors present only in soil substrate could provoke the induction of some genes. Several metabolic pathways were affected in the sensitive line after acid soil growth and could be involved on root growth inhibition such as lignin, cellulose and callose production and ethylene and auxin synthesis. The mapping of the identified genes through the microarray experiments into the chromosomes allowed the identification of genes localized into maize QTLs previously reported in the literature as responsible for the tolerant phenotype. Facing these results, we can speculate the role of these genes such as a RNA binding protein, a protease inhibitor, and cyclines in the Al present in the acid soil tolerance. For the first time in literature, the transcriptome of leaves collected after three days in culture with acid soil or limed soil with the Affymetrix microarrays. This analysis indicated great alterations in Cat100-6, meanwhile S1587-17 showed no significative alteration. Genes related to photosynthesis and photorespiration were down-regulated due acid soil treatment in the tolerant genotype. However, citric acid cycle was activated indicating the putative partitipation of organic acids produced in the leaves in thr Al response
Doutorado
Genetica Vegetal e Melhoramento
Doutor em Genetica e Biologia Molecular

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Sommeria-Klein, Guilhem. "From models to data : understanding biodiversity patterns from environmental DNA data". Thesis, Toulouse 3, 2017. http://www.theses.fr/2017TOU30390/document.

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La distribution de l'abondance des espèces en un site, et la similarité de la composition taxonomique d'un site à l'autre, sont deux mesures de la biodiversité ayant servi de longue date de base empirique aux écologues pour tenter d'établir les règles générales gouvernant l'assemblage des communautés d'organismes. Pour ce type de mesures intégratives, le séquençage haut-débit d'ADN prélevé dans l'environnement (" ADN environnemental ") représente une alternative récente et prometteuse aux observations naturalistes traditionnelles. Cette approche présente l'avantage d'être rapide et standardisée, et donne accès à un large éventail de taxons microbiens jusqu'alors indétectables. Toutefois, ces jeux de données de grande taille à la structure complexe sont difficiles à analyser, et le caractère indirect des observations complique leur interprétation. Le premier objectif de cette thèse est d'identifier les modèles statistiques permettant d'exploiter ce nouveau type de données afin de mieux comprendre l'assemblage des communautés. Le deuxième objectif est de tester les approches retenues sur des données de biodiversité du sol en forêt amazonienne, collectées en Guyane française. Deux grands types de processus sont invoqués pour expliquer l'assemblage des communautés d'organismes : les processus "neutres", indépendants de l'espèce considérée, que sont la naissance, la mort et la dispersion des organismes, et les processus liés à la niche écologique occupée par les organismes, c'est-à-dire les interactions avec l'environnement et entre organismes. Démêler l'importance relative de ces deux types de processus dans l'assemblage des communautés est une question fondamentale en écologie ayant de nombreuses implications, notamment pour l'estimation de la biodiversité et la conservation. Le premier chapitre aborde cette question à travers la comparaison d'échantillons d'ADN environnemental prélevés dans le sol de diverses parcelles forestières en Guyane française, via les outils classiques d'analyse statistique en écologie des communautés. Le deuxième chapitre se concentre sur les processus neutres d'assemblages des communautés.[...]
Integrative patterns of biodiversity, such as the distribution of taxa abundances and the spatial turnover of taxonomic composition, have been under scrutiny from ecologists for a long time, as they offer insight into the general rules governing the assembly of organisms into ecological communities. Thank to recent progress in high-throughput DNA sequencing, these patterns can now be measured in a fast and standardized fashion through the sequencing of DNA sampled from the environment (e.g. soil or water), instead of relying on tedious fieldwork and rare naturalist expertise. They can also be measured for the whole tree of life, including the vast and previously unexplored diversity of microorganisms. Taking full advantage of this new type of data is challenging however: DNA-based surveys are indirect, and suffer as such from many potential biases; they also produce large and complex datasets compared to classical censuses. The first goal of this thesis is to investigate how statistical tools and models classically used in ecology or coming from other fields can be adapted to DNA-based data so as to better understand the assembly of ecological communities. The second goal is to apply these approaches to soil DNA data from the Amazonian forest, the Earth's most diverse land ecosystem. Two broad types of mechanisms are classically invoked to explain the assembly of ecological communities: 'neutral' processes, i.e. the random birth, death and dispersal of organisms, and 'niche' processes, i.e. the interaction of the organisms with their environment and with each other according to their phenotype. Disentangling the relative importance of these two types of mechanisms in shaping taxonomic composition is a key ecological question, with many implications from estimating global diversity to conservation issues. In the first chapter, this question is addressed across the tree of life by applying the classical analytic tools of community ecology to soil DNA samples collected from various forest plots in French Guiana. The second chapter focuses on the neutral aspect of community assembly.[...]

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23

Whissell, Gavin. "Merging metagenomic and microarray technologies to explore bacterial catabolic potential of Arctic soils". Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=98518.

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A novel approach for screening metagenomic libraries by merging both metagenomic and microarray platforms was developed and optimized. This high-throughput screening strategy termed "metagenomic microarrays" involved the construction of two Arctic soil large-insert libraries and the high density arraying of the clone plasmid DNA (~50 kb) onto glass slides. A standard alkaline lysis technique used for the purification of plasmid DNA was adapted and optimized to function efficiently in a 96-well format, providing an economically viable means of producing sufficient high-quality plasmid DNA for direct printing onto microarrays. The amounts of printed material and probe, required for maximal clone detection, were optimized. To examine catabolic clone detection libraries were first screened by PCR for catabolic genes of interest. Two PCR-positive clones were printed onto microarrays, and detection of these specific clones in the printed libraries was achieved using labeled probes produced from PCR fragments of known sequence. Also, hybridizations were performed using labeled PCR fragments derived from the amplification of a catabolic gene from the total community DNA. The ability of selected probes to specifically target clones of interest was demonstrated. This merger of metagenomics and microarray technologies has shown great promise as a tool for screening the natural microbial community for catabolic potential and could also be used to profile microbial diversity in different environments.

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24

Paraskova, Julia V. "Organic phosphorus speciation in environmental samples : Method development and applications". Doctoral thesis, Uppsala universitet, Institutionen för kemi - BMC, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-228734.

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This thesis investigates the development of new methodology for the identification and quantification of organic phosphorus compounds in environmental samples. Phosphorus is a vital element for primary production and one of the factors contributing to eutrophication. Eutrophication of aquatic systems leads to algal blooms, changes in ecological balance and deteriorating water quality. Difficulties in studying organic phosphorus stem from the fact that organic phosphorus is present in the environment in a variety of forms and each form may have different degradation and turnover time, having very different effects on eutrophication. New methods for the quantification of phosphorus derived from three groups of organic phosphorus compounds were developed. For the determination of phosphorus derived from DNA and phospholipids selective extraction was combined with digestion and colorimetric determination of the extracted phosphate. For quantification of inositol phosphates high performance liquid chromatography was coupled with tandem mass spectrometry using electrospray ionization. The methods were applied to studying the distribution of these compounds in a small catchment and in the case of DNA-P and phospholipid-P, the degradation of the fractions in lake sediments. The studies showed that phosphorus bound to DNA, phospholipids and inositol phosphates constitute a sizeable part of the total phosphorus in different environmental samples. The phospholipid-P fraction was the smallest one, accounting for, on average, only a few percent of the total phosphorus in the sample. Inositol phosphates were most prevalent in the soils, with inositol hexakisphosphate accounting for over 10% of the total phosphorus content. The highest content of DNA-P was found in sediments and it was shown that DNA-P degrades more rapidly than phospholipid-P and therefore plays a more critical role in internal loading.

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25

Sullivan, Madsen Paul. "Effects of and Influences on Microbial Populations of Missouri Maize Fields". BYU ScholarsArchive, 2018. https://scholarsarchive.byu.edu/etd/7706.

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The role of individual soil microorganisms changes over the course of a plant's life - microorganisms that have no discernable role at one developmental stage may affect the plant later in its growth. Traditional analysis of the soil microbiome, which has focused principally on the relative abundances (RA) of individual organisms, may be incomplete, as underlying differences in population size cannot be addressed. We conducted a metagenomic analysis of soil microorganisms from various maize (Zea mays L.) fields at two depths, accompanied by crop yield components, to provide insight into influences of edaphic microbes on maize productivity under commercial maize production systems in Missouri. This study assesses the influence of fungi and bacteria, not only in terms of RA, but also in their estimated absolute abundances (EAA), derived by combining the results of Illumina HiSeq sequencing data and phospholipid fatty acid abundance data. Significant interactions were identified between maize yield components and soil microbes at critical developmental states. Most interactions between fungi and yield components were negative, with notable exceptions. Bacterial interactions were more complex, with most interactions during early ear development identified as positive, and most interactions during tasseling identified as negative. In addition to the effects that microbial populations have on yield, plant populations reciprocally changed the microbial community. Plant developmental state was the greatest predictor of bacteria, with the microbial communities present during the active growing season being most similar to each other, whereas the preplant microbiome and post-reproductive microbiome being most similar to each other. Fungal communities were primarily dependent on location.

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26

Sampietro, Bergua Mª Lourdes. "Genetic Analysis of the prehistoic peopling of Western Europe: Ancient DNA the role of contamination". Doctoral thesis, Universitat Pompeu Fabra, 2007. http://hdl.handle.net/10803/79128.

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In this thesis we have addressed three different although related topics. First, we studied the post-mortem mutation damage rate of contaminated DNA sequences in ancient human remains focusing on the development of strategies to avoid pre-laboratory derived contaminations. We proposed a guideline to control them consisting in typing every single person involved on the manipulation of the remains, especially when they have not been excavated and washed under controlled conditions. Second, we successfully develop a non-invasive technique to sequence ancient remains but preserving it from the destruction. And third, we sequenced ancient human remains from different evolutionary times (from Paleolithic to post-Neolithic) to make inferences about the peopling of Western Europe focusing mainly in the Iberia peninsula. We found that there is a long term genetic continuity at least since the Neolithic. The only clear genetic discontinuity found is that involving two different human species, H. sapiens and H. neanderthalensis.
En la presente tesis hemos tratado tres temas diferentes aunque muy relacionados. Primero, hemos estudiado la tasa de mutación post-mortem de secuencias de ADN contaminante en restos humanos antiguos centrándonos en el desarrollo de estrategias para evitar que las muestras se contaminen antes de llegar al laboratorio. Proponemos una guía que consiste en el tipado genético de cada persona implicada en la manipulación de los restos, especialmente cuando estos han sido excavados y lavados bajo condiciones no controladas. Segundo, hemos desarrollado una técnica no invasiva para secuenciar DNA de restos humanos antiguos pero sin destruirlos. Y por ultimo, hemos secuenciado restos humanos antiguos pertenecientes a diferentes periodos evolutivos (desde el Paleolitico hasta el post-Neolitico) que nos han permitido hacer inferencias sobre el poblamiento Europeo centrándonos básicamente en la Península Ibérica. Hemos encontrado que ha habido una continuidad genética desde el Neolítico. La única clara discontinuidad genética encontrada es entre dos especies distintas: H. Sapiens y H.neanderthalensis.

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27

Cury, Juliano de Carvalho. "Diversidade de Bacteria e Archaea em solos de mangue e marisma". Universidade de São Paulo, 2006. http://www.teses.usp.br/teses/disponiveis/11/11138/tde-11122006-144427/.

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Estudos sobre a diversidade de Bacteria em solos de mangue (Brasil) e marisma (Espanha) são escassos. A vegetação de mangue, composta por espécies como Spartina alterniflora, Rhizophora mangle, Avicennia schaueriana e Laguncularia racemosa, pode ser um dos fatores que determinam a estruturação das comunidades de procariotos. Determinações das estruturas das comunidades e de diversidade de Bacteria podem ocorrer em função das diferentes condições físico-químicas dos solos, refletindo na configuração dos processos biogeoquímicos. O objetivo deste trabalho foi avaliar a variação das estruturas das comunidades de Bacteria e Archaea, bem como a diversidade, em solos de mangue e marisma utilizando DGGE e sequenciamento parcial do rDNA 16S. As estruturas das comunidades de procariotos apresentaram variações em função de condições de vegetação. Proteobacteria e Bacteroidetes estão presentes em todos os solos estudados. A comunidade de Bacteria destes ambientes é dominada por Proteobacteria. Vários dos táxons detectados estão relacionados com ciclos biogeoquímicos importantes para os ambientes estudados. As estimativas não-paramétricas de riqueza de espécies (ACE e Chao1) mostram que solos de mangue e marisma podem conter milhares de espécies de bactérias. As comunidades de Bacteria dos solos de mangue e marisma são significativamene diferentes. Na camada mais superficial do sedimento de mangue predomina Euryarchaeota metanogênicas enquanto que na camada mais profunda predomina Crenarchaeota. Bactérias das ordens Desulfobacterales, Desulfovibrionales e Desulfuromonales podem estar relacionadas com a atividade de sulfato-redução e formação de pirita na camada anaeróbia do perfil de solo de marisma. De uma maneira geral, pode-se concluir que a diversidade e estrutura das comunidades de procariotos de ambientes estuarinos pode variar em função da vegetação estabelecida e do tipo de ambiente. Adicionalmente, solos de mangue e marisma possuem grande diversidade de procariotos, grande parte da qual é desconhecida, podendo representar elevado potencial genético para utilização biotecnológica.
The bacterial diversity in mangrove (Brazil) and marisma (Espanha) soils are largely unknown. Bacterial communities participate in biogeochemicals processes that occurs in soils of estuarine ecosystems. Determinations of the bacterial communities structures and diversity can occur in function of different physico-chemical conditions, reflecting in the biogeochemical processes. The aim of this work was to evaluate the variation of bacterial an archaeal communities structures utilizing DGGE and partial sequencing of 16S rDNA. Bacterial community structures showed more similarity between repetitions samples than the areas under different vegetation. Phylogenetic afiliation shows that several sequences were not clamped into known phyla. Proteobacteria prevails in bacterial communities of mangrove and marisma soils. Several taxa detected are associated to important biogeochemical cycles that occur in estuarine ecosystems. Analysis of species richness showed that mangrove and marisma soils can contain 200 to 6000 species of bacteria. Methanogenic Euryarchaeota was found specially in the upper sample of mangrove sediment analysed whereas the Crenarchaeota was found specially in the lower. Based on the data obtained, it can be concluded that the vegetation is one of the factors affecting the structure of bacterial and archaeal communities in mangrove soils. Additionaly, the effects of edafic factors and seasonal variations have to be considered as determining the prokaryotic community sctuctures, and bacterial and archaeal communities can respond independently to the factors that determine their community structures. Bacterial diversity can vary with the studied estuarine ecosystem. Studies are necessary concerning to diversity of Bacteria, it variation and correlation with biogeochemical process in the mangrove and marisma soils. These soils show a great diversity of bacteria, much of than unknown, which represent a great genetic potential to the biotechnology.

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Navarrete, Acácio Aparecido. "Estrutura e diversidade de comunidades microbianas em solos sob diferentes sistemas de uso da terra na Amazônia Ocidental". Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/91/91131/tde-13102009-135502/.

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O presente trabalho esteve inserido em um projeto mais amplo de cooperação internacional intitulado Conservation and Sustainable Management of Below-Ground Biodiversity CSMBGBD/ BiosBrasil, implementado pela United Nations Environment Programmer (UNEP) na bacia do Alto Solimões, Amazônia Ocidental, estado do Amazonas. Esta região é território remanescente de povos indígenas e permanece conservada, sendo um importante hotspot de biodiversidade. Ainda assim, as áreas de estudo foram caracterizadas por paisagens antropizadas com diferentes sistemas de usos da terra. Amostras de solos foram coletadas em um período de elevado índice pluviométrico, nos anos de 2008 e 2009, em áreas caracterizadas por floresta primária tropical, cultivos semiperenes de mandioca manejados por prática agrícola de corte-equeima, pastagens implantadas nos anos de 1970 e áreas florestais em estádios avançados de regeneração (>10 anos de abandono). As amostras foram analisadas pelas técnicas de DGGE, ARISA, clonagem e sequenciamento a fim de obter uma caracterização das estruturas de comunidades de Archaea, Bacteria e microfungos e da composição e diversidade de um grupo funcional de Archaea envolvido no processo de oxidação de amônia nos ambientes do solo. Os resultados permitiram concluir que o uso da terra tem um grande efeito sobre as estruturas de comunidades de Archaea, Bacteria e microfungos presentes no solo e sugerem que longo período de abandono das áreas é necessário para cumprir com a resiliência dos ecossistemas amazônicos no contexto de recomposição da paisagem. Adicionalmente, os dados revelaram que a riqueza e a diversidade de comunidades de Archaea oxidadoras de amônia foram capazes de refletir sensivelmente as alterações percebidas nos ambientes do solo em decorrência do desmatamento de áreas de floresta primária e uso subsequente com cultivo agrícola tradicional e pastagem na região do Alto Solimões, Amazônia Ocidental.
The present study was part of a wider project of international cooperation entitled Conservation and Sustainable Management of Below-Ground Biodiversity CSMBGBD/ BiosBrasil, established by the United Nations Environment Programmer (UNEP) at the basin of Alto Solimões, Western Amazon, Amazonas State. This region is a remanescent territory of indigenous people and remains conserved, being considered an important hotspot of biodiversity. Even though, the studied areas were characterized by mosaic landscapes under different land use systems. Soil samples were collected in a period of high pluviometric index in the years 2008 and 2009 in areas characterized by tropical rainforest, semi permanent manioc cultivation under agricultural management of slash-and-burn, pasture established in the 1970s and forested areas at a higher stage of regeneration (>10 years abandoned). The samples were analyzed by DGGE, ARISA techniques, cloning and sequencing in order to obtain a characterization of the community structure of Archaea, Bacteria and microfungi and the composition and diversity of an archaeal functional group involved in the process of ammonia oxidation in the soil environments. The results allowed to conclude that land use has a great effect on the community structure of Archaea, Bacteria and microfungi present in the soil and suggest that long period of abandon of the areas is necessary to accomplish with the resilience of Amazonian ecosystems in the context of landscape recomposition. Additionally, the data revealed that richness and community diversity of ammonia oxidizing Archaea were able to sensitively reflect the changes observed in the soil environment due to deforestation of rainforest areas and subsequent use with traditional agriculture cultivation and pasture in the region of Alto Solimões, Western Amazon.

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29

Lee, Sungeun. "Virus-host interactions across a soil pH gradient at the community and individual scale". Thesis, Lyon, 2020. http://www.theses.fr/2020LYSEC020.

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Les virus du sol sont capables d'influencer la structure de la communauté microbienne et le fonctionnement de l'écosystème en affectant l'abondance des cellules hôtes par lyse et par leurs caractéristiques à transférer des gènes entre les hôtes. Bien que notre compréhension sur la diversité et la fonction virales s’améliore, la connaissance des interactions hôte-virus dans le sol reste limitée. Pour mieux comprendre les interactions hôte-virus, un gradient du sol à long terme manipulé par le pH dans lequel la communauté microbienne change à travers, a été étudié. Les principaux objectifs de cette thèse ont consisté à (1) déterminer l'influence de la structure de la communauté microbienne et du pH du soil sur les virus par séquençage d’ADN haut-débit (Chapitre II), (2) déterminer l’infectivité des populations virales à partir de niches de sol co-localisées et non co-localisées avec son hôte grâce à une approche de l’essai de plaque combinée à un séquençage hybride (Chapitre III), (3) identifier les populations virales infectant des groupes fonctionnels microbiens spécifiques du sol, en particulier les méthanotrophes (Chapitre IV) et les nitrifiants (Chapitre V ), à l’aide d’une sonde isotopique stable à l'ADN. Nos premiers résultats ont montré que la structure de la communauté virale change selon le pH du sol, ce qui souligne que la communauté virale est étroitement liée aux populations hôtes. L’analyse de CRISPR systèmes a révélé des interactions virus-hôte dynamiques, avec le nombre et la taille des CRISPR systèmes distincts selon le soil à pH contrasté. L’analyse taxonomique de cette CRISPR systèmes suggère que les virus jouent un rôle essentiel dans la composition et de la fonction de la communauté procaryote du sol. Les processus co-évolutionnaires entre l'hôte (le système de restriction-modification et le CRISPR-Cas système) et les populations virales co-localisées (la mutation d’une séquence espaceur « spacer » et la méthyltransférase codée par le virus) fournissent des preuves de l'adaptation locale et que les interactions virus-hôte jouent un rôle important dans la susceptibilité d'un hôte à l'infection et par conséquent la régulation des populations bactériennes du sol. L'ADN-SIP-métagénomique ciblant des groupes fonctionnels microbiens spécifiques a permis l’analyse des populations hôte-virus individuelles. Le suivi du flux de carbone à travers les populations procaryotes et virales a révélé des interactions actives entre les virus et les hôtes méthanotrophes et nitrifiants, et les préférences de niches de pH du sol. Notre étude a montré une preuve de transfert horizontal de gènes et des gènes métaboliques auxiliaires codés par le virus, indiquant que les virus contribuent de manière significative aux cycles biogéochimiques dans le sol, tels que le carbone (les gènes qui codent pour les familles GH, peptidases et la sous-unité C de méthane monooxygénase particulaire), et l'azote (les gènes qui codent pour la nitrogénase et le cytochrome cd1-nitrite réductase). Dans l’ensemble, ces résultats ont montré que les virus du sol sont des régulateurs importants des communautés microbiennes par la lyse spécifique de l’hôte et des interactions dynamiques virus-hôte
Soil viruses have potential to influence microbial community structure and subsequent ecosystem functioning by directly affecting the abundance of host cells by lysis and through their ability to transfer genes between hosts. Although our understanding of soil viral diversity and functioning has increased, the role of viruses and their interactions with prokaryotes in soil is limited. To gain a better understanding of virus-host interactions in soil, a long-term pH-manipulated soil gradient, which microbial community structure changes across, was investigated. The main objectives of this thesis were to 1) determine the influence of microbial community structure and soil pH on viruses using metagenomics and viromics (Chapter II), 2) determine the infectivity of soil viral populations from co-localized and foreign pH soil niches using a plaque assay approach combined with hybrid metagenomics sequencing (Chapter III) and 3) identify virus populations infecting specific soil microbial functional groups, specifically methanotrophs (Chapter IV) and nitrifiers (Chapter V), using DNA stable isotope probing combined with metagenomic deep sequencing. Viral community structure was found to change with soil pH, demonstrating that viral communities are tightly linked to host populations, but also may have narrow host ranges. Analysis of clustered regularly interspaced short palindromic repeats (CRISPR) arrays revealed dynamic virus-host interactions, with the number and size of CRISPR arrays distinct across contrasting pH soil. Profiling of the host-virus linkages between soil pH, suggests that viruses play a critical role in shaping the composition and function of the soil prokaryotic community. Surprisingly, greater infectivity of a host bacterium by virus populations was found when viruses and host bacterium were not co-localized in the same pH soil. Coevolutionary processes between the host and virus populations, such as restriction modification/virus-encoded methyltransferase and CRISPR-Cas system/spacer mutation, provide evidence for local adaptation, and that virus-bacterial host interactions play an integral part in the susceptibility of a host to infection and consequently in the regulation of soil bacterial populations. Targeting specific microbial functional groups via stable isotope probing allowed analysis of individual host-virus populations. Tracking carbon flow through prokaryotic and viral populations revealed active interactions between viruses and methanotroph and nitrifier hosts, and soil pH niche preferences. Evidence of horizontal gene transfer and virus-encoded auxiliary metabolic genes, such as glycoside hydrolase families, peptidases, particulate methane monooxygenase subunit C (pmoC), nitrogenase (nifH) and cytochrome cd1-nitrite reductase, supports that viruses are significant contributors to host functioning and carbon and nitrogen cycling in soil. Overall, this work demonstrated that soil viruses are important regulators of microbial communities through specific host lysis and dynamic virus-host interactions

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30

Radomski, Nicolas. "Sources des mycobactéries non-tuberculeuses dans les bassins versants". Phd thesis, Université Paris-Est, 2011. http://pastel.archives-ouvertes.fr/pastel-00669399.

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L'eau et le sol sont considérés comme des sources potentielles de mycobactéries non-tuberculeuses (MNT). Parmi les infections humaines causées par les MNT d'origine environnementale, les infections pulmonaires et cutanées sont souvent décrites. Le manque de connaissances sur leur cycle de vie dans l'environnement requiert des outils analytiques, qui ne sont actuellement pas adaptés à ce type d'échantillons. Cette thèse vise donc premièrement à proposer des méthodes de quantification en bactériologie et en biologie moléculaire dans le but de déterminer les sources des MNT dans les bassins versants. Ainsi, la comparaison des méthodes d'isolement de MNT a montré que le traitement au chlorure de cetylpyridininium de l'eau suivi d'une culture en milieu riche supplémenté par un mélange d'antibiotiques (polymyxine B, amphotéricine, acide nalidixique, triméthoprime, carboxy-pénicilline) limitait la croissance des microorganismes interférents et éliminait moins de MNT que les autres méthodes comparées (Radomski et al. 2010, doi: 10.1128/AEM.00942-10). Bien que des espèces de MNT potentiellement pathogènes aient été isolées de l'eau de surface de la Seine en utilisant ces outils bactériologiques, la quantification des MNT ne s'est pas avérée reproductible. En conséquence, une méthode de quantification par polymérisation en chaîne en temps réel (qPCR) a été développée pour énumérer le genre Mycobacterium dans l'eau (Radomski et al. 2010, doi: 10.1128/AEM.02659-09). La nouvelle méthode développée, ciblant l'ARNr 16S, était plus spécifique que les autres méthodes qPCR publiées, ciblant un autre locus de l'ARNr 16S et le gène hsp65 (respectivement 100 % versus 44 % et 91 %). La comparaison des méthodes d'extraction d'ADN mycobactérien a montré que la lyse enzymatique combinée au bromure d'hexadécyltriméthylammonium était la procédure la plus efficace pour énumérer par qPCR les MNT dans des échantillons environnementaux. Ainsi, ces méthodes d'extraction d'ADN et de qPCR ont été utilisées pour étudier des sources de MNT dans des bassins versants. Dans un second temps, nous avons étudié trois sources potentielles de MNT : une ponctuelle et deux diffuses. Plus précisément, une station d'épuration (STEP) a été choisie comme source ponctuelle de MNT et a été étudiée en temps sec en fonction d'indicateurs de contamination fécale et des paramètres globaux habituellement contrôlés. Les MNT ont atteint 5,52×105±3,97×105 copies/L dans l'eau en entrée de STEP (84 % d'échantillons positifs), n'ont pas été détectées dans l'eau en sortie de STEP après décantation physico-chimique et biofiltration et ont été estimées à 1,04×106 ±1,75×106 copies/g dans les boues de STEP (50 % d'échantillons positifs). La plupart des MNT (98±2 %, correspondant à 2,45±0,78 log10) ont été éliminées par décantation physico-chimique et les MNT restantes (0,74×104 ±1,40×104 copies/L) ont été éliminées par biofiltration (53 % d'échantillons positifs). Ces résultats ont montré également que Mycobacterium, Escherichia coli et les entérocoques intestinaux possèdent des comportements significativement différents conduisant respectivement à trois modèles : hydrophobe, hydrophile et intermédiaire. Concernant les sources diffuses, la densité de MNT a été mesurée dans divers sols ruraux et urbains qui ont été caractérisés par différents paramètres physico-chimiques. Les densités de MNT les plus importantes ont été mesurées dans des sols de forêts tourbeuses (9,27×104±5,00×104 copies/g sec) et dans des sols faiblement urbanisés proches de marécages côtiers (1,71×106±2,85×106 copies/g sec) alors qu'aucune MNT n'a été détectée dans les autres types de sols étudiés. De plus, la densité de MNT a été significativement associée à des sols proches de zones acides et des teneurs fortes des sols en eau, matière organique et fer. Ces résultats suggéreraient que les MNT sont dépendantes de leur production intra et extracellulaire de chélateurs de fer et indiqueraient que les zones faiblement urbanisées pourraient être impactées par la proximité de marais acides. Afin d'étudier une autre source diffuse, les MNT et d'autres paramètres ont été mesurés lors d'événements pluvieux dans l'eau de surface de la Marne et de ses principaux affluents. Les densités de MNT ont été estimées à 2,16×105±2,36×105 copies/L dans environ 20 % des échantillons d'eau collectés, et elles ne différaient pas entre les zones péri-urbaines et rurales échantillonnées. Nos résultats ont montré que la pluviométrie et la durée de l'évènement expliquaient la diminution du nombre de MNT détectées dans l'eau de surface au cours de l'événement pluvieux de faible intensité (6,6 mm/h de pluviométrie cumulées en 5,5 h). Ces résultats ont souligné que certains affluents de la Marne pouvaient apporter des MNT en temps sec, mais qu'au cours de l'évènement pluvieux suivi les densités de MNT diminuaient.En guise d'amélioration à ces études appliquées, des réflexions sur les défis relatifs à la surveillance des microorganismes pathogènes dans l'environnement ont été explorées. En nous focalisant sur la MNT la plus pathogène, M. avium, nous avons discuté des défis de la détection et de l'énumération et proposé un guide d'adaptation des méthodes médicales aux échantillons environnementaux (Radomski et al. 2011, ed. A. Méndez-Vilas, Vol. 2). Ce guide se présente sous la forme d'un arbre de décision permettant de choisir les outils analytiques les plus appropriés pour surveiller les microorganismes pathogènes dans l'environnement. De plus, une stratégie in silico de comparaison de génomes bactériens totalement séquencés a été développée dans le but de décrire des nouvelles cibles de détection. L'analyse in silico des génomes totalement séquencés a permis de détecter 11 protéines présentant entre 80 % et 100 % de similarité dans les génomes mycobactériens et moins de 50 % de similarité dans les génomes non-mycobactériens des genres Corynebacterium, Nocardia et Rhodococcus. Sur la base d'alignements des séquences d'ADN de ces cibles potentielles, il a été possible de dessiner des amorces PCR et une sonde pour détecter le gène codant la sous-unité C de la synthase de l'adénosine triphosphate qui semble exclusivement conservée dans le génome mycobactérien. Le développement d'outils analytiques, en particulier la qPCR, a permis de montrer qu'une STEP éliminait efficacement les MNT et que le traitement des eaux usées est nécessaire pour préserver l'eau de surface de cette source ponctuelle de MNT. Il a été mis en évidence que les événements pluvieux diminuent la densité de MNT dans l'eau de surface et que les sols acides sont des sources naturelles majeures de MNT qui pourraient impacter des zones faiblement urbanisées en temps de pluie via le ruissellement. Concernant les réflexions sur la surveillance des microorganismes pathogènes dans l'environnement, l'arbre de décision des outils analytiques appropriés et la nouvelle stratégie in silico de détection de cibles moléculaires pourraient être appliqués pour l'étude d'autres microorganismes de l'environnement

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Arthur, Jennifer D., Noah W. Mark, Susan Taylor, J. Šimunek, M. L. Brusseau i Katerina M. Dontsova. "Batch soil adsorption and column transport studies of 2,4-dinitroanisole (DNAN) in soils". ELSEVIER SCIENCE BV, 2017. http://hdl.handle.net/10150/624067.

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The explosive 2,4,6-trinitrotoluene (TNT) is currently a main ingredient in munitions; however the compound has failed to meet the new sensitivity requirements. The replacement compound being tested is 2,4-dinitroanisole (DNAN). DNAN is less sensitive to shock, high temperatures, and has good detonation characteristics. However, DNAN is more soluble than TNT, which can influence transport and fate behavior and thus bio-availability and human exposure potential. The objective of this study was to investigate the environmental fate and transport of DNAN in soil, with specific focus on sorption processes. Batch and column experiments were conducted using soils collected from military installations located across the United States. The soils were characterized for pH, electrical conductivity, specific surface area, cation exchange capacity, and organic carbon content. In the batch rate studies, change in DNAN concentration with time was evaluated using the first order equation, while adsorption isotherms were fitted using linear and Freundlich equations. Solution mass-loss rate coefficients ranged between 0.0002 h(-1) and 0.0068 h(-1). DNAN was strongly adsorbed by soils with linear adsorption coefficients ranging between 0.6 and 6.3 L g(-1), and Freundlich coefficients between 1.3 and 34 mg(1-n) L-n kg(-1). Both linear and Freundlich adsorption coefficients were positively correlated with the amount of organic carbon and cation exchange capacity of the soil, indicating that similar to TNT, organic matter and clay minerals may influence adsorption of DNAN. The results of the miscible-displacement column experiments confirmed the impact of sorption on retardation of DNAN during transport. It was also shown that under flow conditions DNAN transforms readily with formation of amino transformation products, 2-ANAN and 4-ANAN. The magnitudes of retardation and transformation observed in this study result in significant attenuation potential for DNAN, which would be anticipated to contribute to a reduced risk for contamination of ground water from soil residues.

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Nunes, Gisele Lopes. "Diversidade e estrutura de comunidades de Bacteria e Archaea em solo de mangue contaminado com hidrocarbonetos de petróleo". Universidade de São Paulo, 2007. http://www.teses.usp.br/teses/disponiveis/11/11138/tde-23032007-162450/.

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Os impactos da poluição por hidrocarboneto de petróleo sobre a diversidade e funcionalidade das comunidades microbianas em manguezais não são totalmente conhecidos, principalmente devido às limitações metodológicas para acessar os microrganismos nãocultiváveis. No entanto, vários métodos moleculares independentes de cultivo têm sido utilizados para investigar a diversidade e a estrutura das comunidades microbianas em ecossistemas naturais. O objetivo deste trabalho foi avaliar as variações da estrutura das comunidades de Bacteria e Archaea e a diversidade de Bacteria em uma transeção de solo de mangue do rio Iriri (Bertioga, SP) com um gradiente de contaminação por hidrocarbonetos de petróleo. As análises por eletroforese em gel com gradiente desnaturante (DGGE) mostraram que as comunidades de Bacteria e Archaea nas diferentes posições geográficas foram mais similares entre si do que entre diferentes profundidades ao longo do perfil em uma mesma posição geográfica. A análise das seqüências de clones de rDNA 16S de Bacteria dos diferentes pontos amostrados em abril de 2000, mostrou que a diversidade genética, avaliada pelo índice de Shannon, das comunidades microbianas diferem estatisticamente somente entre ponto o P1 (ponto menos contaminado) e P3 (ponto mais contaminado). As estimativas não-paramétricas da riqueza de espécies mostraram que P1, P2 e P3 possuem mais de 3539, 2524 e 1421 espécies bacterianas, respectivamente. Já, para as amostras do ponto P2 coletadas nos anos 2000 e 2004, muito embora os valores dos índices de Shannon tenham sido semelhantes, houve uma provável dominância de grupos específicos nas amostras coletadas em 2004, verificada pelos altos valores da recíproca do índice de Simpson. Os dados mostraram também que o número estimado de espécies bacterianas no ponto P2 diminuiu com o tempo, sendo menor em amostras de 2004, se comparado com amostras de 2000. No geral, a afiliação filogenética dos clones de rDNA 16S mostrou a grande diversidade de espécies, a maioria não conhecidas. Os dados sugerem que a contaminação do solo de mangue do rio Iriri está selecionando microrganismos mais adaptados às fontes de carbono introduzidas no solo.
The impacts of petroleum hydrocarbon pollution on the diversity and functionality of the microbial communities in mangrove soils are not totally understood, mainly due to the methodological limitations to access unculturable microorganisms. However, several cultureindependent molecular methods have been used to investigate the diversity and structure of microbial communities in natural ecosystems. The aim of this work was to evaluate shifts in Bacteria and Archaea community structures and the diversity of Bacteria in a soil transection of the Iriri river mangrove (Bertioga, SP) showing a petroleum hydrocarbon contamination gradient. The analyses by denaturing gradient gel electrophoresis (DGGE) showed that the communities of Bacteria and Archaea in different geographical positions were more similar among them than the communities in different depths along the soil profile at the same geographical position. Sequence analyses of bacterial 16S rDNA clones from different points sampled in April 2000 showed that the genetic diversity of the bacterial communities, based on the Shannon index, differ statistically only between P1 (less polluted) and P3 (more polluted) locations. Nonparametric estimates of species richness showed that P1, P2 and P3 may have more than 3539, 2524 and 1421 bacterial species, respectively. For P2 sampled in years 2000 and 2004, even though the Shannon indices were similar, there was a probable dominance of specific bacterial groups in year 2004, based on the high values of the reciprocal of Simpson\'s index. The data also showed that the estimated number of bacterial species in P2 decreased with the time, being lower in samples collected in 2004, as compared to samples collected in 2000. In the general, the phylogenetic affiliation of the 16S rDNA clones showed high bacterial species diversity, and most of the bacteria were of unknown species. The data suggest that the contamination of Iriri river mangrove soil with petroleum hydrocarbon is selecting microorganisms more adapted to the introduced carbon sources into the soil.

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Gruber, Helga [Verfasser], Martin Akademischer Betreuer] Müller, Heinrich H. D. [Akademischer Betreuer] [Meyer i Karl [Akademischer Betreuer] Kramer. "Surveillance of Cry1Ab protein and cry1Ab DNA in liquid manure, soil and agricultural crops under Bt-maize cropping and slurry management of cows fed Bt-maize (MON810) / Helga Gruber. Gutachter: Karl Kramer. Betreuer: Martin Müller ; Heinrich H. D. Meyer". München : Universitätsbibliothek der TU München, 2011. http://d-nb.info/1019590017/34.

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Kapusuz, Derya. "Sol-gel Synthesis Of Dna Encapsulated Silica". Master's thesis, METU, 2009. http://etd.lib.metu.edu.tr/upload/2/12610627/index.pdf.

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Sol-gel processing routes for encapsulation of double stranded DNA in solid porous silica hosts have been established. The encapsulation was carried out in two steps: hydrolysis of a silica-forming alkoxide-based sol was followed by condensation/gelation to a solid form upon addition of a buffer solution containing DNA molecules. The effects of gelation chemistry and DNA amount on chemical and microstructural properties of resultant silica matrices and on DNA encapsulation efficiency were investigated. The analytical characterization was performed by UV-vis spectroscopy, 29Si nuclear magnetic resonance spectroscopy and by nitrogen adsorption studies. It was demonstrated that DNA incorporation had a pH-dependent catalytic effect on gelation kinetics and promoted silica network completion. In addition, the scale of porosity and the average pore size of the resultant silica increased with gelation pH and also with DNA-buffer solution in the starting sol-gel formulation. The chemistry-derived pore size variation controls the DNA encapsulation efficiency in the silica matrices and the DNA holding capacity strongly depends on the scale of the porosity attained. The selective adsorption of ethidium bromide- a DNA-intercalative reagent molecule- on DNA-silica gels confirmed that the DNA molecules remained entrapped within the silica host in their native state without any deterioration. Besides pure silica, amine-functionalized hybrid silica hosts were also formed by sol-gel. The hybrid gels were found not to be suitable for DNA encapsulation, as these matrices dissolve in aqueous environment due to incomplete silica network formation. The DNA-doped silica hosts may provide promising matrices for development of biosensors, bioreactors and bioassay platforms.

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Anderson, Dominique Elizabeth. "Gene Discovery in Antarctic Dry Valley Soils". Thesis, University of the Western Cape, 2008. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_6598_1265941858.

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The metagenomic approach to gene discovery circumvents conventional gene and gene product acquisition by exploiting the uncultured majority of microorganisms in the environment. It was demonstrated in this study that metagenomic methods are suitable for gene mining in extreme environments that harbor very high levels of unculturable microorganisms. DNA was extracted from Antarctic mineral soil samples taken from the Miers Valley, Antarctica. The metagenomic DNA was also used to construct a fosmid library comprising over 7900 clones with an average insert size of 29 kb. PCR amplification using bacterial and archaeal 16S rRNA gene specific primers and subsequent denaturing gradient gel electrophoresis (DGGE) of bacterial 16S rDNA amplicons showed that a small percentage of bacterial diversity (>
1%) was captured in the metagenomic fosmid library. Activity-based screening for lipase and esterase genes using a tributyrin plate assay yielded twelve positive clones. LD1, a putative, novel cold-active GDSL lipase/esterase was identified and sequenced. The C-terminal domain of the ORF was found to be an autotransporter similar to those associated with type V secretion systems in Gram negative bacteria. Sub-cloning of the gene resulted in lipolytic activity in E. coli. Preliminary enzyme assays have determined that LD1 hydrolyses p-nitrophenyl esters with chain lengths shorter than C10, an indication that the enzyme is an esterase. Complete purification and characterisation of this enzyme is subject to further study.

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van, Woerkom Anne. "Ancient DNA from soils and sediments from the Krigstjärn area, northern Sweden : Preservation and detection of Holocene mammal sedaDNA". Thesis, Umeå universitet, Institutionen för ekologi, miljö och geovetenskap, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-127680.

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Current knowledge of past vegetation and faunal diversity has been based on pollen and macrofossil analysis from lake sediments. The innovative method of sedimentary ancient DNA (sedaDNA) is a promising, complementary proxy to reconstruct information about past environments. However, to what extent animal DNA can be extracted from old sediments and soils has not been frequently studied. This study explored if ancient DNA of moose (Alces alces), reindeer (Rangifer tarangus), goat (Capra aegagrus) and plants could be extracted from millennia old lake sediments of Lake Krigstjärn and archaeological soil samples in northern Sweden. SedaDNA was successfully extracted and detected from both reindeer and plants DNA, while goats sedaDNA was absent in all sediments. Moose ancient DNA (aDNA) was only detected in the archaeological soils. Yet, there were signs that the applied moose primer set was not optimal for heavily degraded DNA and the validity of this primer needs further research. Earliest detections of reindeer DNA can be dated to ~6500 c. years ago. Oldest sediments contained DNA, indicating sufficient DNA preservation conditions in the sediments of Lake Krigstjärn. Finds of plants DNA in pre-deglaciational sediments may indicate the existence of >9500 year old glacial vegetation. Altogether is sedaDNA a highly promising tool to reconstruct diversity, origin and immigration routes of mammals, but technical issues such as primer set specificity and its purpose should be considered and tested carefully in advance.

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Cristea, Pop Casandra Ioana <1978&gt. "Minori rumeni tra percorsi migratori e rappresentazioni. Nè soli, nè accompagnati a Bologna". Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2008. http://amsdottorato.unibo.it/732/.

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The thesis deals with the heterogenous category of the “unaccompanied minors”, concentrating the scientific work on those who migrate from Romania to the Italian city of Bologna. Between different migratory routes that include Romanian minors, I chose to explore the ones linked with the underground and illegal contexts. In order to analyse the reasons and the morphology of their migratory career, I used the multisituated field research which allowed me to consider the social policies in both the Romanian and the Italian environment. The main debate on the situation of the “unaccompanied children” refers to the extent to which these minors leave their country of origin “accompanied” by different adult figures and it also involves the role played by these adults. The first chapter is dedicated to a brief theoretical and methodological introduction to the main arguments of the thesis such as Romanian migration to Italy, trafficking in human beings, transnationality of migrant’s migration and decentered cooperation as a means of contrasting illegal migration and trafficking. Each field of research is characterized by a specific methodological approach, but they are all linked by the anthropological perspective I adopted throughout the entire work. The Romanian context, analized from a diachronic and a synchronic perspective represents the object of the second chapter. Some aspects of the Regime policies and other characteristics of the Romanian poscomunist period of “transition” are useful frameworks that become a background of the migration flows outside the country. The third chapter focuses on the Romanian patterns of migration. The reconstruction of some past attitudes that Romanians adopted towards migration are relevant in order to reveal the continuity with the present migratory practices. A consistent part is dedicated to a concrete example based on a field research in Bologna on a group of Romanian roma migrating from the south of Romania. The contact with these persons opened a debate on the limits between legal and illegal migration practices among the Romanians. The conclusion is that minors’ migration to Italy follows the adult patterns and flows. The nucleus of the field researches is included in the fourth and the fifth chapter. Before presenting the settings and the itineraries of the field researches, some deconstructive reflections are made on the representations that common sense and social sciences create on concepts as “child”, “minor” and “childhood”. A first perspective on the Romanian migrant minors emerges from a research concentrated on a group of roma teenagers engaged in Bologna in activities like windscreen washing, pocket-picking, begging and street prostitution. The aim of the research was to gain access to their daily life, to observe their relationship with the adults who “accompany” them and the strategies they activate in order to take some material profit out of their migratory experience. A parallel field research focuses on the Romanian minors who are part of the roma group coming from the south of Romania. Most of them are reunited with their family in Bologna, but according to the Italian law, they are all living as illegal migrants. Others are only temporary sheltered by these families and they meanwhile dedicate to illegal survviving practices. An interesting point of my participant observation was to reveal the motivations that these minors give when asked about the refusal to start a legal career inside the local Centres dedicated to the “non accompanied minors”. Their autoreflexivity brings some light on the controversy regarding the adequacy of the local and national care system and the migratory projects the minors have. In this respect, a small part of the research is dedicated to the phenomena of minors’ street prostitution in Bologna, as a useful contribution to the fragmented vision researchers have on the “unaccompanied” or “separated” children. The last chapter focuses on a decentered cooperation project that emerged as an alternative response the local administration from Bologna had chosen for facing the presence of numerous migrants coming from the south of Romania. The group of Romanian roma who was also the object of my field research became the starting point for the cooperation proposals between the city of Bologna and the city of Craiova. Although there are three projects involving the two administrations, throughout a period of stage in the Romanian city of Craiova I chose to analyse, only the one dedicated to the “urgent measures” requested in order to contrast the illegal migration and the trafficking in minors. This final part of the thesis highlightens the possible contribution that such a project might bring to the study of a complex and in some parts contradictory phenomena as that of the “unaccompanied” migrant minors.

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Barroso, Haline. "Análise de metilação de DNA nos genes da citoqueratina 14 (KRT14) e 19 (KRT19) em amostras de pele exposta e não exposta ao sol". Universidade Federal da Paraíba, 2015. http://tede.biblioteca.ufpb.br:8080/handle/tede/9427.

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It is well established that solar UV radiation can cause mutations in DNA and increase the risk of developing skin cancer. However, little is known about the ability of UV radiation to cause epigenetic changes in the skin. DNA methylation, characterised by the addition of a methyl group in cytosines within CpG dinucleotides, can modify gene transcription, leading to decreased expression or even silencing of a gene. Epigenetic changes could represent an important pathway by which environmental factors influence aging and disease risks, with a tissue-specific manner. Epithelial keratins are called cytokeratins, the main function of cytokeratins is to maintain the integrity and mechanical stability through cell-cell contacts with epithelial tissue. The aim of this study was to investigate the sun exposure influence on DNA methylation status in the cytokeratin 14 (KRT14) and 19 (KRT19) genes of skin cells of subjects whithout history of skin disease. Skin biopsies were obtained by punch of sun-exposed (outer forearm) and sun-protected areas (inner arm) from 30 corpses of the Brazilian Services of Death Investigation. The KRT14 gene DNA methylation analysis was performed using Methylation-Specific PCR (MSP), and the KRT19 gene DNA methylation analysis was performed using Methylation-Sensitive Restriction Enzymes (MSRE) of sun-exposed and sun-protected skin areas. Statistical analysis showed no significant differences between sun-protected and sun-exposed areas and the most frequently methylated condition for CpG studied for KRT14 and KRT19 genes (p> 0.05; McNemar). We conclude that sun exposure does not induce changes in DNA methylation status in the KRT14 and KRT19 genes.
Pesquisas tem mostrado que a radiação UV do sol pode causar mutações no DNA e aumentar o risco para o desenvolvimento de câncer de pele. Entretanto, ainda pouco se sabe sobre a capacidade da radiação UV em causar alterações epigenéticas na pele. A metilação do DNA é caracterizada pela adição do grupo metil em uma citosina precedida por uma guanina (dinucleotídeo CpG), o que pode alterar a transcrição gênica, diminuindo a expressão ou silenciando um gene. Alterações epigenéticas podem representar um importante caminho de como os fatores ambientais influenciam no envelhecimento e no desenvolvimento de certas doenças de maneira tecido específica. Queratinas epiteliais são chamadas de citoqueratinas, e sua principal função é manter a integridade e estabilidade mecânica do tecido epitelial. Neste trabalho investigamos se há influência da exposição solar sobre o perfil de metilação de DNA nos genes das citoqueratinas 14 (KRT14) e (KRT19), em células da pele de indivíduos sem histórico de doenças de pele. Biopsias de pele foram obtidas através de um Punch circular da de área exposta e não exposta ao sol de 30 cadáveres do Serviço de Verificação de Óbito. A análise de metilação do gene KRT14 foi realizada pelo método de PCR Específica para Metilação (MSP), e para o gene KRT19 foi realizado o método de Restrição Enzimática Sensível à Metilação (MSRE) das áreas expostas e protegidas do sol. A análise estatística mostrou que não há diferenças significativas entre as regiões exposta e não exposta ao sol, sendo a condição metilada a mais frequente tanto para o gene KRT14 quanto para o gene KRT19 (p>0,05; McNemar). Assim, concluímos que não há influência da exposição solar no perfil de metilação de DNA nos genes KRT14 e KRT19.

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Arthur, Jennifer, i Jennifer Arthur. "The Investigation of the Environmental Fate and Transport of 2,4- dinitroanisole(DNAN) in Soils". Diss., The University of Arizona, 2017. http://hdl.handle.net/10150/623164.

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New explosive compounds that are less sensitive to shock and high temperatures are being tested on military ranges as replacements for 2, 4, 6-trinitrotoluene (TNT) and hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX). One of the two compounds being tested is 2, 4-dinitroanisole (DNAN), which has good detonation characteristics and is one of the main ingredients in a suite of explosive formulations being tested. Data on the fate and transport of DNAN is needed to determine its potential to reach groundwater and be transported off base, a result which could create future contamination problems on military training ranges and trigger regulatory action. In this study, I measured how DNAN in solution interacts with different types of soils from across the United States. I conducted kinetic and equilibrium batch soil adsorption experiments, saturated column experiments with DNAN and dissolution and transport studies of insensitive munitions (IMX-101, IMX -104), which include DNAN, 3-nitro-1,2,4-triazol-5-one (NTO), nitroguanidine (NQ) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), under steady state and transient conditions. In the rate studies, change in DNAN concentration with time was evaluated using the first order kinetic equation. Solution mass-loss rate coefficients ranged between 0.0002 h-1 and 0.0068 h-1. DNAN was strongly adsorbed by soils with linear adsorption coefficients ranging between 0.6 and 6.3 L kg-1, and Freundlich coefficients between 1.3 and 34 mg1-n Ln kg-1. Both linear and Freundlich adsorption coefficients were positively correlated with the amount of organic carbon and cation exchange capacity of the soil. In saturated miscible-displacement experiments, it was shown that under flow conditions DNAN transforms readily with formation of amino transformation products, 2-amino-4-nitroanisole (2-ANAN) and 4-amino-2-nitroanisole (4-ANAN). Dissolution miscible-displacement experiments demonstrated that insensitive munition compounds dissolved in order of aqueous solubility as indicated by earlier lab and outdoor dissolution studies. The sorption of NTO and NQ was low, while RDX, HMX, and DNAN all adsorbed to the soils. DNAN transformed in soils with formation of amino-reduction products, 2- ANAN and 4-ANAN. Adsorption parameters determined by HYDRUS-1D generally agreed with batch and column study adsorption coefficients for pure NTO and DNAN. The magnitudes of retardation and transformation observed in these studies result in significant attenuation potential for DNAN in soils, which would reduce risk of groundwater contamination.

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Delgado, de la flor Yvan A. "Spider and Beetle Communities across Urban Greenspaces in Cleveland, Ohio: Distributions, Patterns, and Processes". The Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1587656050129337.

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Olivares, Martinez Christopher Ignacio. "Environmental Fate, (Bio)transformation, and Toxicology of 2,4-dinitroanisole (DNAN) in Soils and Wastewater Sludge". Diss., The University of Arizona, 2016. http://hdl.handle.net/10150/596139.

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Insensitive munition compounds (IMC) are an emerging class of explosives that are less susceptible to accidental explosions compared to the conventional explosives they will be replacing. An IMC that has been incorporated in several explosives formulations is 2,4-dinitroanisole (DNAN). As the manufacture, storage, and use of these compounds increases, the expected releases in natural and engineered systems might pose an environmental hazard to public health and ecosystems. To date there is little information on the environmental fate and toxicology of DNAN. However, nitroaromatic compounds are known to be toxic, mutagenic and difficult to completely biodegrade. In order to study the fate and (bio)transformation of DNAN, microcosm studies with soils and anaerobic wastewater sludge were performed to determine (bio)transformation pathways and key factors influencing (bio)conversion. Transformation was enhanced in anaerobic conditions, in particular when exogenous electron donor was added. Abiotic transformation (in heat-killed soil) was also significant and dominated transformation reactions in soils that were not amended with exogenous electron donor. The organic carbon content of soils was a key factor that correlated to the anaerobic biotransformation rate. Having identified (bio)transformation products using liquid chromatography coupled to quadrupole time-of-flight mass spectrometry, an overall pathway of (bio)transformation was devised and consistent with nitro-group reduction to form aromatic amines. During the nitro-group reduction, reactive products (e.g. nitroso-intermediates) coupled with amines to form azo-dimers and oligomers. Subsequent transformation pathways included N-alkylation, N-acetylation, and stepwise demethoxylation of these oligomers. The assessment of the toxicity of DNAN and its (bio)transformation products was performed utilizing microbial toxicity assays and ecotoxicity evaluation with zebrafish (Danio rerio) embryos. Overall DNAN severely inhibited methanogens (IC₅₀ = 41 μM ), the bioluminescent marine bacterium Aliivibrio fischeri utilized in the Microtox test (IC₅₀ = 57 μM), and nitrifiers (IC₅₀ = 49 μM). Reduced aromatic amine products in general were less toxic than DNAN with the exception of 2-methoxy-5-nitroaniline and 3-nitro-4-methoxyaniline, which were similar in toxicity to some of the test organisms as DNAN. Azo-oligomer surrogates were as toxic or more toxic than DNAN, although at trace levels they significantly stimulated activity. N-acetylated amines were found to have by far the lowest toxicity to microorganisms. In zebrafish embryos, the (bio)transformation product or surrogates 3-nitro-4-methoxyaniline and 2,2'-dimethoxy-4,4'-azodianiline caused developmental abnormalities (each with lowest observable effect level of 6.4 μM). An integrated approach which monitored (bio)transformation product mixture profile in parallel with their toxicity to microbial and zebrafish toxicity was used to characterize toxicity during the time course of the anaerobic (bio)transformation of DNAN. Enhanced inhibition of methanogenic activity and zebrafish mortality were associated with the onset of dimer formation indicating they were being mostly impacted by reactive intermediates formed early in the biotransformation of DNAN. Further accumulation of oligomers was associated with a decrease toxicity. On the other hand, A. fischeri bioluminescence became more and more inhibited as the oligomers formed, indicating different responses depending on target organism. Taken globally, the results indicate that DNAN can be readily transformed in soils and wastewater sludge forming both highly toxic (e.g. azo-oligomers) and non-toxic intermediates (e.g. N-acetylated 2,4-diaminoanisole). Depending on target organism, the prolonged formation of oligomer mixtures either resulted in detoxification or recovery of activity.

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Gianfrancesco, Richard Umberto. "Phosphorus nutrition of mycorrhizal and non-mycorrhizal plants of upland soils with special reference to the utilization of the phosphodiester DNA under sterile conditions". Thesis, University of Sheffield, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.299580.

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Sivander, Malin. "Din sol, din himmel, dina ICA-butiker röda : om hur Sverige framställs i ICA:s reklamfilmer". Thesis, Stockholms universitet, Institutionen för mediestudier, 2015. http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-113107.

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Syftet med den här uppsatsen har varit att undersöka på vilket sätt ICA-reklamen gestaltar Sverige. Med nationalism, myter och ideologi som teoretiska utgångspunkter syftar uppsatsen till att komma närmare hur myten om Sverige rekonstrueras i ICA:s reklamfilmer, samt vilken roll ICA har i det Sverige som framställs. Efter en förstudie på de av ICA:s reklamfilmer som sänts i TV det senaste halvåret har ett strategiskt val gjorts. Utifrån ett semiotiskt angreppssätt har en kvalitativ innehållsanalys genomförts på tre av ICA:s reklamfilmer från 2014. Analysen visar att genom användningen av tecken som symboliserar Sverige i reklamfilmerna konstrueras myter om den svenska familjen, svensken och Sverige. ICA bidrar därmed genom sina reklamfilmer till mytifieringen av Sverige. Slutsatserna dras till att genom de här tre reklamfilmerna gestaltar ICA Sverige som ett land med hälsosamma och medvetna invånare som helst äter svenskproducerad mat och lever i kärnfamiljskonstellationer. Samtidigt har ICA själva blivit en symbol för det svenska och representerar därmed nationell identitet. I idyllen agerar ICA-butiken som den familjära mittpunkten för det svenska vardagslivet. Spåren av Sverige som återfinns i ICA:s reklamfilmer, till exempel debatten om föräldraledighet, firandet av svenska traditioner och (myten om) svenskens längtan efter det genuina och traditionella, bidrar till att ICA-reklamen kan ses som ett ”miniatyr-Sverige”.

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Rábová, Petra. "Využití testů s destruenty pro ekotoxikologické hodnocení kvality půd". Master's thesis, Vysoké učení technické v Brně. Fakulta chemická, 2020. http://www.nusl.cz/ntk/nusl-433143.

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The aim of this diploma thesis was to determine the dehydrogenase activity of matrix samples that can enter the environment and the use of ecotoxicological tests to assess the quality of the affected soil. The effects of sludge, biochar samples, two landfill samples and a sediment sample were monitored. Ecotoxicology included tests on decomposers - tests of avoidance behavior (Eisenia fetida and Folsomia candida), acute, chronic and reproductive toxicity tests (Eisenia fetida) and Lactuca sativa root growth inhibition tests. The results indicate that dehydrogenase activity assays serve as a suitable complement to ecotoxicological tests. They provide valuable additional information on soil quality and sample effect after application to soil. The sediment sample had the least favorable effect on the activity of the enzyme and the vitality of the organisms, both in the determination of dehydrogenase activity and in ecotoxicological tests. Furthermore, it was found that biochar as a product of sewage sludge treatment is less toxic to organisms than the original sludge.

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Marlin, Constanze von. "Public Art Space : zum Öffentlichkeitscharakter der Minimal Art : Carl Andre - Dan Flavin - Donald Judd - Sol LeWitt - Robert Morris /". Weimar : Verlag und Datenbank für Geisteswissenschaften, 2008. http://aleph.unisg.ch/hsgscan/hm00227183.pdf.

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Hasegawa, Aline Yuri. "Um dia de sol para encontrar os antepassados : o Shokonsai como estudo de caso do Obon no Brasil". reponame:Repositório Institucional da UFABC, 2018.

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Orientadora: Profª. Drª. Marilda Aparecida de Menezes
Coorientadora: Profª. Drª. Adriana Capuano de Oliveira
Tese (doutorado) - Universidade Federal do ABC, Programa de Pós-Graduação em Ciências Humanas e Sociais, Santo André, 2018.
Esta pesquisa trata do Shokonsai, um rito de culto aos ancestrais realizado no município de Álvares Machado, há 99 anos, em um cemitério étnico de nikkei. Além das peculiaridades do local e da longevidade do evento, um fator climático também é mobilizado no discurso do público: nos registros e nas narrativas orais, jamais choveu ao longo do dia e, durante o espetáculo de acendimento de velas, os ventos cessam de soprar e somente retornam após a última apagar-se naturalmente. Por meio de obervação participante, pretendo dar conta da descrição e análise das práticas dos interlocutores que articulam fenômenos climáticos à vontade dos mortos. Essa prática ritualística levanta questões identitário-étnicas, que se conectam a processos históricos nacionais japoneses e brasileiros, de modo que se faz necessário também compreender como os discursos de pertencimento étnico contemporâneos mobilizam elementos raciais e culturais de cada contexto.
This research approaches the Shokonsai, an ancestor worship rite performed in Álvares Machado since 1918, in an ethnic Nikkei cemetery. In addition to the peculiarities of the place and the longevity of the event, a climate factor is also mobilized in the public discourse: according to the records and oral narratives, it never rained during the day, and during the candlelight show the winds have ceased to blow and have only returned after the last candle is put out naturally. Through participant observation, I intend to describe and analyze the practices of the interlocutors who articulate climate phenomena to the will of the dead. This ritual practice raises some ethnic and identity issues, which necessarily connect to Japanese and Brazilian national historical processes. In that sense, it is also necessary to understand how contemporary discourses about ethnic belonging mobilize racial and cultural elements of each context.

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Marlin, Constanze von. "Public - art - space zum Öffentlichkeitscharakter der Minimal Art ; Carl Andre, Dan Flavin, Donald Judd, Sol LeWitt und Robert Morris". Weimar VDG, 2005. http://d-nb.info/985889950/04.

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Marlin, Constanze von Andre Carl Flavin Dan Judd Donald LeWitt Sol Morris Robert. "Public - Art - Space : zum Öffentlichkeitscharakter der Minimal Art ; Carl Andre, Dan Flavin, Donald Judd, Sol LeWitt und Robert Morris /". Weimar : VDG, Verl. und Datenbank für Geisteswiss, 2007. http://www.gbv.de/dms/weimar/toc/547381905_toc.pdf.

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Oliver, Fojkar. "Azotofiksirajuće cijanobakterije u zemljištima Vojvodine i njihova ultrastrukturna i genetička karakterizacija". Phd thesis, Univerzitet u Novom Sadu, Poljoprivredni fakultet u Novom Sadu, 2016. https://www.cris.uns.ac.rs/record.jsf?recordId=101459&source=NDLTD&language=en.

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U radu je ispitana zastupljenost azotofiksirajućih cijanobakterija, ukupnog broja algi i ukupnog broja bakterija u različitim tipovima zemljišta na jedanaest lokaliteta u Vojvodini, od čega se sedam nalaze u zaštićenim prirodnim dobrima. Ispitana je brojnost u zavisnosti od dubine pedološkog profila, kao i od godišnjeg doba. Izvršena je izolacija sojeva azotofiksirajućih cijanobakterija, određena njihova taksonomska pripadnost i osnovne citološke karakteristike. Ispitana je ultrastruktura vegetativnih ćelija, heterocista i spoljnih struktura na ćelijama fimbrije i pili, transmisionim elektronskim mikroskopom. Izvršena je genetička karakterizacija izolovanih sojeva azotofiksirajućih cijanobakterija PCR metodom analizom STRR fragmenata DNA.Brojnost azotofiksirajućih cijanobakterija i ukupna brojnost algi je bila znatno veća kod hidromorfnih i halomorfnih zemljišta, nego kod automorfnih zemljišta. Najveća prosečna godišnja brojnost azotofiksirajućih cijanobakterija, u površinskom sloju 0-5cm, je utvrđena kod zemljišta fluvisol u SRP “Koviljsko petrovaradinskom ritu”, 150864 jedinki po gramu apsolutno suvog zemljišta. Zemljište sa najnižom brojnošću azotofiksirajućih cijanobakterija, je gajnjača u NP Fruška gora, 1582 jed./gr zemljišta u površinskom sloju.Kod svih ispitivanih zemljišta brojnost azotofiksirajućih cijanobakterija je bila najveća u površinskom sloju zemljišta, 0-5 cm dubine, opadala je sa dubinom zemljišta i bila najmanja u najdubljem sloju, 30 - 60 cm. Kod najvećeg broja ispitivanih zemljišta brojnost azotofiksirajućih cijanobakterija je bila najveća tokom zimskog perioda. Azotofiksirajuće cijanobakterije su dominantne u našim zemljištima i zastupljene sa 56.27% u odnosu na druge grupe algi.Izolovano je 30 sojeva azotofiksirajućih cijanobakterija: 19 sojeva Nostoc-a, 4 soja Anabaena, 4 Cylindrospermum, i po jedan soj Calothrix, Tolypothrix i Phormidium. Prosečna zastupljenost heterocista, ćelija koje vrše azotofiksaciju, kod roda Nostoc je iznosila 8.28%, Anabaena 4.25%, Cylindrospermum-a 2.93%, Calotrix elenkinii 6.19% i Tolypothrix 7.76%.Ultrastrukturnim ispitivanjem, TEM mikroskopom, vegetativnih ćelija azotofiksirajućih cijanobakterija uočili smo inkluzije redovnog pojavljivanja: karboksizome (Cs), cijanoficinkse granule (CG), polifosfatne granule (PG), ribozome (R), lipidne granule (ß –granule) i tilakoide (T), kao i inkluzije neredovnog pojavljivanja: membranom ograničene kristalne inkluzije.Koristeći TEM i tehniku bojenja ćelija sa RR i ultratankih preseka utvrdili smo prisustvo omotača od fimbrija kod tri soja (A.314, A.azollae i N.302) i tipične fimbrije kod dva soja (N.311 i N.9229). Metodom negativnog bojenja NS PTA uočili smo takođe tipične fimbrije, igličastog-dlakastog izgleda, jasnih granica niti kod tri soja (N.302, N.7901 i N.9229), međutim uočili smo i atipične sluzne fimbrije, koje nemaju jasno izražene granice, ali su veoma moćno raširene oko vegetativnih ćelija, kod tri soja (A.314, A.azollae, N.311).Kod simbiotskih-infektivnih sojeva N.7901 i N.9229 javljaju se samo tipične fimbrije iz prve klase, a kod diazotofnih sojeva i simbiotskog - neinfektivnog soja A.azollae javljaju se atipične-sluzne fimbrije iz druge klase.Za ispitivanje sličnosti cijanobakterija metodom PCR-a pomoću STRR konzervativnih sekvenci DNA genoma korišćeno je 39 sojeva azotofiksirajućih cijanobakterija i kod 38 je utvrđeno njihovo prisustvo. Svi sojevi se mogu podeliti u tri grupe, klastera. Prvi klaster je najveći i obuhvata 24 soja i deli se na dva podklastera: Ia koji obuhvata 12 sojeva gde dominiraju sojevi Nostoc-a (8), i podklaster Ib koji obuhvata takođe 12 sojeva, od čega 6 sojeva pripada rodu Anabaena. Podklaster Ia i podklaster Ib pokazuju različitost od 90%. Sva tri simbiozna, infektivna, soja Nostoc-a se nalaze u klasteru I: N.7901, N.9229 i N. 8001. Svaki simbiozni soj Nostoc-a ima genetske sličnosti sa po jednim diazotrofnim sojem Nostoc-a izolovanim iz zemljišta Vojvodine.Klaster II obuhvata sedam (7) sojeva među kojima dominiraju sojevi Cylindrospermum-a, dok klaster III obuhvata 7 sojeva od čega 6 pripadaju rodu Nostoc, a jedan rodu Rivularia.Detaljno poznavanje svojstava izolovanih azotofiksirajućih cijanobakterija doprineće njihovoj budućoj primeni kako u proizvodnji ratarskih i povrtarskih kultura, tako i u biotehnološkoj proizvodnji
In this study examined is the frequency of nitrogen-fixing cyanobacteria, total number of algae and total number of bacteria in different soil types on eleven localities in the Vojvodina Province. Seven out of those eleven localities are found in protected nature reserves. Actually, studied was the number of the cyanobacteria and algae depending on the depth of pedological characterization as well as on season. First, isolated were the types of nitrogen-fixing cyanobacteria, determined was their taxonomic origin and basic cytological characteristics. Also examined was the ultrastructure of vegetative cells, heterocysts and other outer structures on fimbriae and pili cells using TEM, transmission electron microscope. Finally, performed was the genetic characterization of isolated types of nitrogen-fixing cyanobacteria using the PCR method and analyzing STRR fragments of DNA.The presence of nitrogen-fixing cyanobacteria and total number of algae was significantly higher with hydromorphic and halomorphic soils than with authomorphic ones. Highest annual average number of nitrogen-fixing cyanobacteria in the topsoil (0-5 cm) was reported with fluvisol soil in Special Nature Reserve „Koviljsko petrovaradinski rit” (Swamp) and there were 150864 units of bacteria per gram of absolutely dry soil. The soil with the lowest presence of nitrogen-fixing cyanobacteria recorded was cambisol in National Park “Fruska gora” with 1582 units per gram of soil in the topsoil.With all the researched types of soils the number of nitrogen-fixing cyanobacteria was in the topsoil, 0-5 cm of depth and decreased in line with the depthof soil and lowest was at the deepest layer, 30-60cm. The highest frequency of nitrogen-fixing cyanobacteria was found during the winter season with most of the examined soils. Nitrogen-fixing cyanobacteria are the dominant type of bacteria in our soils and are presented with 56, 27% compared to other types of algae.30 strains of nitrogen-fixing cyanobacteria were isolated: 19 types of Nostoc sp., 4 of Anabaena sp. and one in each genus of Calothrix, Tolypothrix and Phormidium.Using ultrastructural examination and TEM microscope when studying vegetative cells of nitrogen-fixing cyanobacteria observed were the inclusions of regular frequency: carboxysomes (Cs), cyanophycin granules (CG) , polyphosphate granules (PG), ribosomes (R), lipid granules (SS -granule ) and thylakoids ( T ) as well as the inclusion of irregular occurrence: a membrane-bound crystal inclusions.Using TEM technique and staining the cells with the RR and ultra thin cross section, we determined the presence of depletion of the fimbriae with three strains (A.314, A.azollae and N.302) and typical fimbriae with the two strains (N.311 and N.9229). Applying the method of negative staining NS PTA also noticed were a typical fimbriae, needle-hairy like looks with clear boundaries with the three strains (N.302, N.7901, N.9229). However, also observed were atypical mucous fimbriae, which do not have clearly expressed borders, but they are very strongly spread around the vegetative cells, with the three strains (A.314, A.azollae, N.311).With symbiotic - infective strains N.7901 and N.9229 only typical fimbriae of first class occurred, and in diazotroph strains and symbiotic – non infectious strain A. azollae found were atypical mucous fimbriae of second class.To test the similarity of cyanobacteria by PCR method and using a STRR - conservative DNA sequence of the genome used were 39 strains fixing cyanobacteria and with 38 established was their presence. All strains can be divided into three groups of clusters. The first cluster is the largest and covers 24 strains, and is divided into two subclusters: Ia which includes 12 strains,where predominant are Nostoc strains ( 8 ) , and subcluster Ib , which also implies 12 strains , out of which 6 strains belong to the genus Anabaena. Subcluster Ia and Ib show a difference of 90 %. All three symbiotic , infectious Nostoc strains are classified in a cluster I: N.7901, N.9229 and N. 8001. Each symbiotic Nostoc strain has a genetic similarity with one di-nitrogen Nostoc strain isolated from a lot of different soils in Vojvodina.Cluster II includes seven (7) strains , including strains among which the predominant are Cylindrospermum ones , while cluster III includes 7 strains of which 6 belong to the genus Nostoc and one to genus Rivularia.Detailed knowledge of the properties of isolated fixing cyanobacteria could contribute to their future application both in the production of field crops and vegetables, as well as in biotechnological production.

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Sow, Mamadou Dia. "Rôle fonctionnel de l'épigénétique (Méthylation de l’ADN) dans la réponse du peuplier à des variations de disponibilité en eau du sol". Thesis, Orléans, 2019. http://www.theses.fr/2019ORLE3046.

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Un dépérissement du foret est observé depuis plusieurs années à l’échelle mondiale en lien avec les changements climatiques en cours. Ainsi, les arbres qui sont des organismes fixes et pérennes doivent développer des stratégies leur permettant de s’adapter constamment aux variations environnementales. Récemment, les mécanismes épigénétiques et notamment les modifications de la chromatine telles que la méthylation de l’ADN ou les modifications des histones ont été proposés comme une source possible de flexibilité lors de l’adaptation des organismes vivants aux changements climatiques.Cette thèse avait pour objectif d’évaluer le rôle de l’épigénétique (méthylation de l’ADN) dans la réponse du peuplier à la sécheresse, à trois échelles de temps : à court terme (plasticité développementale), à moyen terme (mémoire et priming) et à long terme (adaptation). Ces études se sont focalisées sur les méristèmes (apical et cambium) qui sont les centres de la morphogenèse. Pour cela, trois différentes approches ont été utilisées : corrélative, épigénétique inverse et épigénomique des populations. Le peuplier qui est un arbre modèle très sensible aux variations de disponibilité en eau du sol a été étudié aussi bien avec des populations naturelles que des hybrides et des lignées RNAi sur des dispositifs expérimentaux en serre ou pépinière.Les résultats majeurs obtenus sont : i) La méthylation de l’ADN est affectée lors du déficit hydrique dans les méristèmes, ii) les variations de la méthylation de l’ADN induites sont stables au cours du temps, iii) la méthylation de l’ADN pourrait être utilisée comme un marqueur génétique de la différentiation des populations en condition hydrique limitante, iv) la méthylation de l’ADN en contexte CG et dans une moindre mesure en CHG pourrait être utilisée comme un marqueur de la différentiation des individus et/ou des populations à l’image des marqueurs génétiques (i.e. SNPs), v) les peupliers hypométhylés (par mutation DDM1) sont plus tolérantes à la sécheresse et présentent un phénotype de réponse aux pathogènes suggérant une résistance accrue à l’infection, vi) La méthylation de l’ADN peut moduler la réponse hormonale et favoriser un effet de priming. vii) la réduction par une stratégie RNAi du niveau de méthylation de l’ADN favorise la réactivation des éléments transposables qui peuvent s’insérer dans de nouvelles régions génomiques altérant la stabilité du génome,Ces résultats proposent un rôle de la méthylation de l’ADN à l’interface entre la réponse physiologique notamment hormonale et la variation génétique en réponse à des contraintes environnementales. Ces travaux ouvrent des perspectives en amélioration et conservation des ressources génétiques forestières notamment dans le cadre du projet ANR EPITREE
Forest decline has been observed around the world due to climate change. As sessile and long-lived organisms subject to repeated environmental constraints, trees need to develop strategies enabling them to cope with fluctuating environmental conditions. Epigenetic mechanisms have recently emerged as a valuable source of flexibility for adaptation to climate changes.This thesis aimed to evaluate the role of epigenetics (DNA methylation) in poplar, in response to drought, at three time scales: at short term (developmental plasticity), middle term (memory, priming) and long term (adaptation). These studies focused on meristems which are the center of morphogenesis.The main results are : i) DNA methylation is affected during drought stress in meristems, ii) Induced-DNA methylation variations are stable in time, iii) DNA methylation could be used as a genetic marker for populations differentiation under drought, iv) DNA methylation in CG and in a lesser extent in CHG contexts could be used as a marker of individuals and/or populations differentiation like the genetic markers, v) Hypomethylated poplar trees (RNAi DDM1) are more tolerant to drought and show a pathogens responsive phenotype suggestive of enhanced resistance, vi) DNA methylation could alter hormonal balance and promotes priming, vii) Decrease in DNA methylation level (RNAi DDM1) favors the reactivation of transposable elements and their integration in new genomic regions during the drought stress leading to genome instability.These results propose a role of DNA methylation at the interface between the physiological response (hormonal response) and the genetic variation under environmental constraints. These works open up new perspectives in Breeding and forest conservation management, particularly in the frame of the ANR EPITREE project

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