Detecting mutations and genomic variations is fundamental indiagnosis, isolating disease genes, association studies,functional genomics and pharmacogenomics. The objective hasbeen to use and further develop a variety of tools andtechnologies to analyze these genetic alterations andvariations. The p53 tumor suppressor gene and short arm of chromosome 9have been used as genetic markers to investigate fundamentalquestions concerning early events preceding non-melanoma skincancers, clonal progression and timing of different mutationsand deletions. Conventional gel based DNA sequencing andfragment analysis of microsatellite markers were utilized forthis purpose. In addition, a sequence-specific PCR-mediatedartifact is discussed. Pyrosequencing, a bioluminometric technique based onsequencing-by-synthesis, has been utilized to determinemutation ratios in the p53 gene. In addition, in the case ofmultiple mutations, pyrosequencing was adopted to determineallelic distribution of mutations without the use of cloningprocedures. Exons 5 to 8 of the p53 gene were also sequenced bythis method. The possibility of typing single base variations bypyrosequencing has been evaluated. Two different nucleotidedispensation orders were investigated and data were comparedwith the predicted pattern for each alternative of the variableposition. Analysis of loss of heterozygosity was possible byutilizing single nucleotide polymorphisms. A modified allele-specific extension strategy for genotypingof single nucleotide polymorphisms has been developed. Throughthe use of a real-time bioluminometric assay, it has beendemonstrated that reaction kinetics for a mismatchedprimer-template is slower than the matched configuration,butthe end-point signals are comparable. By introduction ofapyrase, the problems associated with mismatch extensions havebeen circumvented and accurate data has been obtained. Keywords:fragment analysis, microsatellite, loss ofheterozygosity, DNA sequencing, pyrosequencing, cancer,mutation, variation, single nucleotide polymorphism,allele-specific extension, bioluminescence, apyrase. / QC 20100415

Background It is debated whether a low total diversity of the gut microbiota in early childhood is more important than an altered prevalence of particular bacterial species for the increasing incidence of allergic disease. The advent of powerful, cultivation-free molecular methods makes it possible to characterize the total microbiome down to the genus level in large cohorts. Objective We sought to assess microbial diversity and characterize the dominant bacteria in stool during the first year of life in relation to atopic eczema development. Methods Microbial diversity and composition were analyzed with barcoded 16S rDNA 454-pyrosequencing in stool samples at 1 week, 1 month, and 12 months of age in 20 infants with IgE-associated eczema and 20 infants without any allergic manifestation until 2 years of age (ClinicalTrials.gov ID NCT01285830). Results Infants with IgE-associated eczema had a lower diversity of the total microbiota at 1 month (P = .004) and a lower diversity of the bacterial phylum Bacteroidetes and the genus Bacteroides at 1 month (P = .02 and P = .01) and the phylum Proteobacteria at 12 months of age (P = .02). The microbiota was less uniform at 1 month than at 12 months of age, with a high interindividual variability. At 12 months, when the microbiota had stabilized, Proteobacteria, comprising gram-negative organisms, were more abundant in infants without allergic manifestation (Empirical Analysis of Digital Gene Expression in R [edgeR] test: P = .008, q = 0.02). Conclusion Low intestinal microbial diversity during the first month of life was associated with subsequent atopic eczema. / <p>Funding Agencies|BioGaia AB, Stockholm, Sweden||Ekhaga Foundation, the Heart and Lung foundation||Research Council for the South-East Sweden|F2000-106|Olle Engqvist Foundation||Swedish Asthma and Allergy Association||Swedish Research Council||University Hospital of Linkoping||Soderberg Foundation||Vardal Foundation for Health Care Science and Allergy Research, Sweden||BioGaia AB||</p>

Invasive species often threaten biodiversity and environmental processes in their introduced range by extirpating native species due to competition for resources. Nylanderia fulva (formerly N. (=Paratrechina) sp. nr. pubens) is an ecologically dominant and economically important invasive species in the United States. This dissertation addresses aspects of the biology, behavior, management, and collection techniques for N. fulva. Specifically, topics investigated include a modified drip technique for extracting ants from their substrate, the effectiveness of a broadcast ant bait as a stand-alone treatment, the foraging preference and peak activity of workers, the reproductive potential of queens, and the ability of this species to translocate pathogenic microorganisms. The primary goal of these works was to better understand the biological idiosyncrasies of this species that may ultimately lead to the mitigation N. fulva populations. A modified drip technique was developed to quickly and efficiently extract N. fulva from their nesting substrates. Ants and their associated substrates were collected in 18.9 L buckets lined with talcum powder and transported to the laboratory. Substrates were weighted down and a cardboard tower was provided for the immigration of ants as they were forced out of substrates with a slow influx of water. Three applications of Advance Carpenter Ant Bait (ACAB) were applied to a N. fulva population in East Columbia, TX. A series of GIS interpolated maps depict achieved management and subsequent rebound of N. fulva populations. As great as 77% population reduction was achieved by 1 week post treatment, but N. fulva populations rebounded within 3-4 weeks. As a stand-alone treatment, this bait did not provide adequate ant management in treatment plots. Diet preference experiments were performed using artificial diets and food lures. These results of these trials indicated that N. fulva preferred the most carbohydrate rich diet offered through all seasons and that mint apple jelly or hot dog slices were the favored food lures. Diel foraging behavior was observed when temperatures were between 9.95 and 37.26 degrees C. Peak foraging activity occurred at 28.24 +/- 3.12 degrees C. A laboratory investigation of N. fulva suggested that as the number of queens increased, individual queen fecundity increased. This phenomenon is a novel observation among ants and suggests an alternative mechanism for intracolony dominance. Hexagyne colony fecundity of 0.25 +/- 0.12 eggs/queen/hr was the maximum fecundity observed. Results of laboratory experiments showed that N. fulva were capable of transferring E. coli up to 4.5 m in 6 hrs after acquisition from a contaminated source. Pyrosequencing of ectomicrobial assemblages revealed a suite of 518 bacteria and 135 fungi species associated with N. fulva, many of which are known pathogens of plants and animals, including humans. These results suggested that N. fulva should be regarded as both a medically and agriculturally important species.

<p>With the full sequence of the human genome now available, anexciting era in biomedical research has started. The sequenceprovides information about all our genes and greatly increasesthe scope to compare genetic activities in different cells, toanalyze genetic variation between individuals and betweendifferent species and, most importantly, to investigatesystematically the whole genome in a gene-by-gene manner, andthus increase our understanding of gene function.</p><p>This thesis describes studies in which developments weremade in several areas of functional genomics. Messenger RNAlevels were analyzed by the use of an amplification procedure,in which the 3´-ends of the transcripts were selected inorder to amplify the mRNA population in an unbiased fashion. Bysonicating cDNA originating from expressed mRNA, uniformlysized representatives of the transcripts,“signaturetags”, were obtained. The mRNA levels in the original mRNApopulation correlated well with the levels in the amplifiedmaterial, as verified by microarray analysis and realtimequantitative PCR. The expressed transcripts can be identifiedusing pyrosequencing, by comparing the obtained sequenceinformation from the signature tags to information contained invarious sequence databases. In one of the articles, the use ofpyrosequencing is illustrated by efforts to find genes involvedin the disease progression of atherosclerosis.</p><p>More challenging than the study of mRNA levels is to analyzewhen, where and how proteins fulfill their wide-ranging rolesin all the various cellular processes. Proteins are morecomplex biomolecules than mRNA, each having unique properties.Current techniques for studying proteins need much improvement,and are often limited to investigations of a specific portionof the proteome. One approach for studying the whole proteomeis to systematically generate reagents with specific affinityfor the proteins encoded by the genome, one by one. Theaffinity reagents can be used as flags for their targets,providing a flag-specific detection system, so that the targetproteins can be sub-cellularly localized in the majority ofhuman tissues in an array format. One of the articles includedin the thesis presents a pilot project for large-scale affinityreagent production. The aim was to provide a sound basis forwhole proteome studies, but as a pilot study this investigationwas limited to the proteins encoded by human chromosome 21. Allputative genes on the chromosome were subjected to antibodygeneration in a systematic manner. Small, uniform, and easilyproduced representative portions of the full-length proteinswere expressed. These were denoted“Protein EpitopeSignature Tags”and were designed to be unique for theirfull-length counterparts. The antibodies were produced inrabbits and two of the articles in the thesis discuss differentapproaches for affinity purification of the antibodies toachieve the highest possible specificity towards the targets.The resulting“mono-specific”, but still“multi-epitope”, antibodies can be used for a widerange of additional biochemical studies, such as protein arrayand protein pull-out analyses.</p><p><b>Keywords:</b>functional genomics, 3´-end signaturetags, pyrosequencing, amplification, PrEST, chromosome 21,polyclonal antibodies, dual expression, affinitypurification.</p>

Terminal restriction fragment length polymorphism (T-RFLP) analysis and pyrosequencing were used to characterize the microbiota of the tonsil of the soft palate of 126 unfit and 18 healthy pigs. The T-RFLP analysis method was first optimized for the study of the pig tonsil microbiota and the data compared with culture-based identification of common pig pathogens. Putative identifications of the members of the microbiota revealed that the phyla Firmicutes, Proteobacteria and Bacteroidetes were the most prevalent. A comparison of the T-RFLP analysis results grouped into clusters to clinical conditions revealed paleness, abscess, PRRS virus, and Mycoplasma hyopneumoniae to be significantly associated with cluster membership. T-RFLP analysis was also used to select representative tonsil samples for pyrosequencing. These studies confirmed Actinobacteria, Bacteroidetes, Firmicutes, Fusobacteria, and Proteobacteria to be the core phyla of the microbiota of the tonsil of the soft palate of pigs. / OMAFRA Animal Health Strategic Investment

This is a two-part project. Proton pump inhibitors (PPIs) have been associated with increased risk of C. difficile infections and increased toxin production when combined with antimicrobial therapy. The first part of this project involved characterization of a hypervirulent NAP1 C. difficile strain, including genome sequencing and assembly, and the development of methods to study its transcriptomics using RNA-Seq, which will enable future researchers to study different expression patterns when toxigenic C. difficile is challenged with PPIs and/or antimicrobials in vitro. The second part of this project involved characterizing a clinical isolate of a NAP1 C. difficile displaying a markedly elevated MIC to metronidazole (MIC = 16 mg/mL), which initially exhibited MIC of 32 mg/mL. A method of obtaining a metronidazole-susceptible revertant from this isolate was developed and a revertant was obtained. The genomes of both isolates were sequenced, assembled, and aligned, then compared to each other for polymorphisms.

This is a two-part project. Proton pump inhibitors (PPIs) have been associated with increased risk of C. difficile infections and increased toxin production when combined with antimicrobial therapy. The first part of this project involved characterization of a hypervirulent NAP1 C. difficile strain, including genome sequencing and assembly, and the development of methods to study its transcriptomics using RNA-Seq, which will enable future researchers to study different expression patterns when toxigenic C. difficile is challenged with PPIs and/or antimicrobials in vitro. The second part of this project involved characterizing a clinical isolate of a NAP1 C. difficile displaying a markedly elevated MIC to metronidazole (MIC = 16 mg/mL), which initially exhibited MIC of 32 mg/mL. A method of obtaining a metronidazole-susceptible revertant from this isolate was developed and a revertant was obtained. The genomes of both isolates were sequenced, assembled, and aligned, then compared to each other for polymorphisms.

A região anterior do intestino médio de Lepidoptera apresenta uma secreção microapócrina de enzimas digestivas com vesículas migrando pelo interior das microvilosidades. Essas vesículas brotam das microvilosidades intestinais como vesículas de membrana dupla e são descarregadas dentro do lúmen. O objetivo desse trabalho foi identificar as proteínas secretadas e aquelas envolvidas na maquinaria secretória microapócrina em Spodoptera frugiperda. Para isso, vesículas microapócrinas foram preparadas e usadas para a produção de anticorpo policlonal. Esse anticorpo foi utilizado para varrer uma biblioteca de expressão de cDNA do intestino médio de S. frugiperda. Também obtivemos um transcriptoma por pirosequenciamento de uma biblioteca de cDNA proveniente dos transcritos do intestino médio do mesmo inseto. Os clones positivos da varredura foram sequenciados, montados e submetidos a um BLASTN contra as sequências obtidas pelo pirosequenciamento, o que resultou na extensão dessas sequências. Usamos ainda as sequências geradas pelo pirosequenciamento para reanalisar sequências de proteínas presentes nas membranas microvilares, que tinham sido obtidas anteriormente em nosso laboratório (Ferreira et al., 2007). A reanálise das sequências de proteínas microvilares gerou 66 proteínas preditas. Dessas, 18 foram consideradas contaminantes de outros compartimentos celulares e 48 associadas às membranas microvilares. A análise das sequências obtidas das vesículas microapócrinas gerou 50 proteínas preditas que podem ser secretadas por essa rota. As sequências encontradas tanto em membrana microvilar quanto em vesículas microapócrinas podem ser classificadas em 8 grupos, de acordo com sua função: (1) enzimas digestivas; (2) proteínas da membrana peritrófica; (3) envolvidas com proteção; (4) transportadores; (5) receptores; (6) proteínas da maquinaria secretória; (7) proteínas de citoesqueleto; (8) com função desconhecida nesse local. Em ambas as preparações existem uma predominância de sequências de enzimas digestivas. Nas membranas microvilares a maioria das sequências são aminopeptidases, enquanto nas vesículas microapócrinas a maioria são lipases. Os cDNAs correspondentes as proteínas que poderiam estar envolvidas na maquinaria secretória foram clonados e sequenciados. São elas: fimbrina, cofilina, gelsolina-1 e miosina I. RT-PCRs semi-quantitativas dessas proteínas em diferentes tecidos do inseto (intestino, túbulos de Malpighi, corpo gorduroso e carcaça) mostraram que somente gelsolina-1 está presente exclusivamente no intestino. Os domínios G1-G3 da gelsolina característica do intestino (gelsolina-1) foram expressos e usados para produzir anticorpos em coelhos. Esses anticorpos reconhecem a proteína recombinante e uma proteína presente no epitélio intestinal com massa molecular compatível com a massa predita para gelsolina-1. Foi possível diminuir a expressão de gelsolina-1 utilizando RNA interferente. / Lepidoptera anterior midgut presents a microapocrine secretion of digestive enzymes with secretory vesicles migrating inside the microvilli. These vesicles bud from the midgut microvilli as double membrane vesicles and are discharged into the lumen. The aim of this work was to identify the proteins secreted and those involved in the microapocrine secretory machinery in Spodoptera frugiperda larvae. For this, microapocrine vesicles were prepared and used for polyclonal antibody production. This antibody was used to screen a cDNA expression library of S. frugiperda midgut. We also obtained a transcriptome by pyrosequencing a cDNA library derived from transcripts of the midgut. Positive clones from the screening were sequenced, assembled and N-blasted against S. frugiperda sequences obtained by pyrosequencing. This procedure led to the extension of the sequences previously obtained. We also used the sequences generated by pyrosequencing to reanalyze the sequences of microvillar membrane proteins obtained previously by Ferreira et al. (2007). This reanalysis generated 66 predicted proteins that are present in the microvillar membranes. Eighteen were considered to be contaminants from other compartments and 48 associated with the microvillar membranes. Analysing the sequences from microapocrine vesicles we found 50 predicted proteins that should be secreted by microapocrine vesicles. The sequences found in both microvillar membrane and microapocrine vesicles may be classified into 8 groups, according to their function: (1) digestive enzymes; (2) peritrophic membrane proteins; (3) protection; (4) transporters; (5) receptors; (6) secretory machinery; (7) cytoskeleton; (8) with unknown function. In both preparations there is a predominance of sequences of digestive enzymes. In microvillar membranes, there is a remarkable amount of aminopeptidases, while in microapocrine vesicles this is true for lipases. cDNAs coding for proteins that could be involved in the microapocrine secretory machinery were cloned and sequenced. They are: fimbrin, cofilin, gelsolin-1 and myosin I. Using RT-PCR, we showed that mRNAs coding for gelsolin-1 and myosin I are present only in the intestinal tissue. The mRNAs coding for other proteins were found in all tissues. The domains G1-G3 from gelsolin specific from intestinal midgut (gelsolin 1) were expressed and used to raise antibodies in rabbit. These antibodies were able to recognize the recombinant protein and a protein from the midgut epithelium that has a molecular weight similar to the one predicted from gelsolin-1 sequence. We succeed in decreasing the expression of gelsolin-1 by using interfering RNA

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No. of bitstreams: 1 Tese - Marelis Margarita Ruiz.pdf: 48193021 bytes, checksum: 4e7e0fa3715216dbc1c8be5df2d8a197 (MD5) Previous issue date: 2014-02-02 / Não Informada / In the Amazon region, the Urucu Oil Province (Petrobras) has been disposing of hydrocarbon effluents into the surrounding aquatic environments since the beginning of its operations. The goal of this work was to analyze the bacterial biodiversity present in the dike of the effluents and in the stream before and after the disposal of hydrocarbon effluents. A study was conducted in two stages were carried out with the use of culture independent molecular techniques. The full genomic DNA of the sediment and water samples were extracted and used as templates in a PCR reaction with the use of specific oligonucleotides of the 16S rRNA gene bacteria domain. The PCR product was amplified and pyrosequenced, and the generated sequences were analyzed by the free software Mothur. The measurements of the physical-chemical and chromatographical parameters are within the measures established for these types of analyzed water bodies. In the first stage of the study, the taxonomic profiles of the samples have shown that the Proteobacteria phylum proved to be the most abundant together with the Deltaproteobacteria class and the gen known as Candidatus Solibacter. Similarly, in the second stage of the study, the Proteobacteria phylum proved to be the most abundant together with the Alphaproteobacteria class and the Geobacter gen predominant. Both genes are catalogued as bioremediators in contaminated environments, as well as 14% of the total genes. A large proportion of microorganisms were considered “Unclassified” (up to 30%). In first stage of the study the richness and the diversity of species in the Onça stream is greater following the disposal of hydrocarbon effluents, in second stage of the study the abundance and diversity of species in the natural stream (without a name in the region), is greater as compared to the same stream after the mixing of effluents. There is no significant statistical difference between the sample of the community in the natural stream and this effluent-mixed stream; and there is no difference in genetic structure among samples of each community analyzed, suggesting that the discharge of effluents in this body of water has no significant impact on the microbial biodiversity in this aquatic environment. This work, a pioneer in the taxonomic analysis of samples in dikes of effluents and streams with the disposal of effluents in the Urucu oil area, represents a challenge for understanding the relationship between the composition, abundance and diversity of microorganisms in this environment, and a rich source for the discovery of new taxonomic groups, as well as the huge potential for biotechnological exploration of such diversity. / Na região Amazônica, a Província Petrolífera de Urucu (Petrobras) realiza o despejo de efluentes de hidrocarbonetos em ambientes aquáticos ao redor, desde o início de suas operações. O objetivo deste trabalho foi analisar a biodiversidade bacteriana presente no dique de efluentes e no igarapé antes e depois do despejo de efluentes. Foi realizado um estudo em duas etapas, aplicando técnicas moleculares independentes de cultivo. O DNA genômico total das amostras de sedimento e água foi extraído e usado como molde em uma reação de PCR utilizando oligonucleotídeos específicos do gene 16S rRNA para o domínio Bactéria. O produto de PCR foi amplificado e pirosequenciado, e as sequências geradas foram analisadas pelo programa livre Mothur. As medidas dos parâmetros físico-químicos e cromatográficos estão dentro dos valores estabelecidos para os tipos de corpos de água analisados. Na primeira etapa do estudo, os perfis taxonômicos das amostras mostraram que o filo Proteobacteria foi o mais abundante com a classe Deltaproteobacteria e gênero conhecido o Candidatus Solibacter. Assim mesmo, na segunda etapa do estudo, o filo Proteobacteria foi o mais abundante; porém, com a classe Alphaproteobacteria e o gênero Geobacter como predominante. Ambos gêneros são catalogados como biorremediadores de ambientes contaminados, assim como 14% dos gêneros totais. Uma grande parte de microrganismos foram considerados “Não Classificados” (até 30%). Na primeira etapa do estudo a riqueza e diversidade de espécies no Igarapé da Onça, é maior depois do despejo de efluentes de hidrocarboneto, e na segunda etapa do estudo a riqueza e diversidade de espécies no igarapé natural (sem nome na região), é maior com relação a este mesmo igarapé depois da mistura com efluentes. Não existe diferença estatística significativa entre a comunidade do igarapé natural e entre este igarapé misturado com efluentes; e não existe diferença na estrutura genética entre as amostras de cada comunidade analisada, indicando que a descarga de efluentes neste corpo de água não tem impacto significativo na biodiversidade microbiana deste ambiente aquático. Este trabalho, pioneiro em análise taxonômica de amostras de um dique de efluentes e os igarapés com despejo de efluentes na área petrolífera de Urucu, representa uma oportunidade para a compreensão da relação entre a composição, abundância e diversidade dos microrganismos neste ambiente, e uma fonte rica para descoberta de novos grupos taxonômicos, assim como um enorme potencial para exploração biotecnológica desta diversidade.

A imensa diversidade de micro-organismos no solo leva a uma inevitável riqueza de interações entre espécies. Neste intuito, esse projeto é inovador na identificação dos cogumelos da Mata Atlântica, e na descrição da comunidade bacteriana associada às suas micosferas. Usando corpos de frutificação dos fungos (cogumelos) como indicadores para sistemas ricos em nutrientes, as amostras foram coletadas para investigar as interações entre os fungos (maioria do domínio Basidiomycota) e as bactérias presentes no solo em volta das micélios fúngicos (ambiente micosférico). As análises foram feitas com técnicas independentes de cultivo (análise PCR-DGGE, sequenciamento Sanger de fragmentos de ITS/18S e pirosequenciamento de tags da região V4 de 16S DNAr). As famílias fúngicas Marasmiaceae e Lepiotaceae, do domínio Basidiomycota foram as mais abundantes entre os cogumelos amostrados (13 e 5 cogumelos, respectivamente), e estavam presentes entre todas as três parcelas de estudo. As demais amostras foram alocadas dentro das famílias Marasmiaceae, Lepiotaceae, Inocybaceae, Lachnocladiaceae, Bolbitiaceae, Entolomataceae, Hygrophoraceae, Hymenogastraceae, Mycenaceae e Strophariaceae, além de dois do domínio Ascomycota. Baseado na análise de DGGE, é bastante claro que existe uma grande diferença na comunidade bacteriana (de toda a comunidade bacteriana, de ?- proteobacteria e de ?-proteobacteria) entre os solos associados ou não com os corpos de frutificação. Os dados de pirosequenciamento indicaram que dentro dos tratamentos as amostras se agruparam baseado nas famílias fúngicas ou no substrato onde os cocumeglos ocorrem (solo ou serrapilheira), sendo clara em algumas micosferas as alterações na ocorrência de grupos microbianos. O grupo de UTOs mais induzidas na região da micosfera foi composto dos grupos Burkholderia, Acidobacteria Gp1, Comamonadaceae, Sphingobacteriaceae, Burkholderiaceae, Chitinophagaceae), Schlesneria, Acidobacteria Gp3 e Spartobacteria gênero incertae sedis. Isto indica que existe um processo de seleção para bactérias específicas dependendo das diversas variáveis e fatores ambientais presentes no microhabitat micosfera. / The immense microbial diversity in the soil leads to inevitable richness in inter-species interactions. For this reason, this is a novel project that focuses on identifying mushrooms and the associated bacterial community with their mycospheres in the Brazilian Atlantic Rainforest. Using fungal fruiting bodies (mushrooms) as indicators of nutrient-rich systems, samples were taken to investigate the interactions between fungi (mostly of the domain Basidiomycota) and bacteria present in the soil surrounding fungal mycelia (mycosphere environment). The analyses were conducted using culture-independent techniques, where isolating DNA of bacteria and/or mushrooms attempts to provide information on microbial functionality. These culture-independent analyses (PCR-DGGE, Sanger sequencing of ITS/18S fragments, and pyrosequencing of tags from the V4 region of 16S rDNA) generated extensive data on the bacterial diversity selected for in the presence of fungal structures in the soil. The fungal families Maramiaceae and Lepiotaceae, of the domain Basidiomycota were the most abundant among the mushrooms sampled (13 and 5 mushrooms, respectively) and were present in all of the three sampling sites. The rest of the mushrooms were found to be within the families Marasmiaceae, Lepiotaceae, Inocybaceae, Lachnocladiaceae, Bolbitiaceae, Entolomataceae, Hygrophoraceae, Hymenogastraceae, Mycenaceae, and Strophariaceae, in addition to two of the domain Ascomycota. Based on DGGE analysis, it is clear that there is a great difference in the bacterial community (entire bacterial community, of ?- proteobacteria and of ?-proteobacteria) between all fungal fruiting body-associated and non-associated soils. The pyrosequencing data indicated that within each treatment group, the samples did not separate according to fungal families or substrate where the mushrooms were found (in the soil or amoung the leaf litter). However, some mycospheres exhibited clearly altered bacterial communities. The group of OTUs most induced in the mycosphere region consisted of Burkholderia, Acidobacteria Gp1, Comamonadaceae, Sphingobacteriaceae, Burkholderiaceae, Chitinophagaceae), Schlesneria, Acidobacteria Gp3, and Spartobacteria genus incertae sedis. This indicates that there is a selection process for specific bacteria depending on a wide range of variable and environmental factors acting in the mycosphere microhabitat.