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Past studies characterizing the feline intestinal microflora have used traditional bacterial culture techniques. However, in recent years it has been recognized that the majority of intestinal bacteria are non cultivable. Therefore, the aim of this study was to describe the microflora along the intestinal tract in healthy cats using comparative 16S ribosomal DNA (16S rDNA) analysis. Intestinal content from the stomach, duodenum, jejunum, ileum, and colon was collected from 4 healthy cats and one specific pathogen free cat (SPF) and the bacterial composition was identified by direct sequencing of bacterial 16S rDNA amplicons. A predominant anaerobic microflora was observed in all evaluated segments of the intestine. Fourteen different bacterial orders were identified with the majority of all sequences classified in the class Clostridiales. Six different Clostridium clusters were identified with the majority of sequences affiliated with Clostridium cluster I. Comparative 16S rDNA analysis was also used to evaluate differences in the fecal microflora between healthy cats (n=6), cats with histopathologically confirmed inflammatory bowel disease (IBD; n=6), and cats with intestinal neoplasia (n=3). Compared to the IBD group, cats in the control group showed a significantly higher number of sequences classified as Firmicutes, Bacteroidetes, and Actinobacteria (p<0.0001). The control group had a significantly higher proportion of clones affiliated with Clostridium cluster XI, and a significantly lower proportion affiliated with cluster I (both p<0.0001). In the neoplasia group, the majority of sequences were classified in the phylum Firmicutes (97.9%) and clones were predominately affiliated with Clostridium clusters I and XI. These data indicate that the feline intestinal microflora is highly diverse and is comprised predominantly of anaerobic bacteria. Further studies are warranted to evaluate the clinical significance of the observed differences in intestinal microflora between healthy cats and cats with gastrointestinal disease.

Abstract This research used molecular biological techniques such as polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) to analyze the dynamic microbial community and biodiversity in the groundwater contaminated with petroleum-hydrocarbons. The 16S rDNA sequences from all water samples were compared with the sequences of relative bacteria in the Ribosomal Database Project Bank to construct a phylogenetic tree. The results allowed us to understand the composition of the microbial communities in the petroleum-hydrocarbon contaminated groundwater. In this study, groundwater samples taken from the Chinese Petroleum Corporation Kaohsiung Refinery (CPCKR), Chinese Petroleum Corporation at Ciaotou fuel Tank Farm (CPCCTF) and China Petrochemical Development Corporation at Kaohsiung Factory (CPDCKF) were analyzed. The contaminated sites at CPDCKR and CPCCTF are remeated by natural attenuation. While the CPDCKF site is remeated by an enhanced air sparging bioremediation. In CPDCKR, we found that the low polluted area contained the richest microbial community, followed by the non-polluted area, and the high polluted area. At the CPCCTF site, the microbial community in the non-polluted area was richer than the high-polluted area. Increased microbial populations and variation in microbial community have beenobserved in non-polluted, less polluted, and highly polluted areas. The microbial community showed a dynamic succession of complexity during the bioremediation process at the CPDCKF site. From the 16S rDNA sequence analysis, it is possible that all samples contained petroleum-hydrocarbon degrading bacteria. These petroleum-hydrocarbon degrading bacteria include Methylobacterium, Xanthobacter, Xanthomonas and Pseudomonas at CPCKR site, Flavobacterium at CPCCTF site, Nocardia, Pseudomonas, Rubrivivax, Methylobacterium, and Candida at CPDCKF site. This study also demonstrates that it is more economic and reliable of using molecular techuiques to analyze the groundwater. Thus, groundwater samples can be used to replace soil samples for future work.

Fifty two cellulolytic thermophilic microorganisms were analyzed for their physiological characterization and phylogenetic systematics. Based on 16S rDNA sequence analysis, 3 strains from DCB and 4 novel isolates from southern Taiwan are close related to the genera of Bacillus and Geobacillus respectively. Among 4 new Geobacillus strains, strain T4, a Gram negative, motile, aerobically growing sporulating rod, can secrete thermostable endoglucanase. When strain T4 was grown in CMC medium, the cellulolytic enzyme activity in culture supernatants was stable up to 70¢XC. Based on 16S rDNA sequence analysis, DNA G+C content, phenotypic and physiological characteristics, as well as DNA-DNA hybridization, strain T4 was classified as Geobacillus thermoleovorans T4 (DSM 14791 = CCRC 17200). Furthermore, a phylogenetic tree of 20 related microorganisms was also constructed based on their thermostable cellulase amino acid sequences. Our sequence analysis shows that cellulases belonging to the large family of glycoside hydrolases (GHs) can be divided into four subfamilies: TC-1 (GH family 12 group), TC-2 (bacterial group £L in which fungal species Thermoascus aurantiacus fits), TC-3 (bacterial group £L£L), and TC-4 (GH family 1 group). Together with the 16S rDNA sequence analysis of strain T4 and 10 related microorganisms, strain T4 has a close phylogenetic relationship with subfamily TC-4 but far from subfamily TC-1.

In this study, we had established a 16S rDNA-DGGE analys is system to detect the microbial community in petroleum polluted soil and assess the feasibility of using this system to monitor the bioremediation process. Three genomic DNA extracted methods, the KIT, the Bead-beating system, and the Freeze-thaw method were used to evaluate the DNA extraction efficiency and purity. These DNA samples were further tested by DGGE to analysis the microbial community in soil samples. The results showed that KIT method performed advantageous not only in the DNA extraction efficiency and purity, but also expressed the richest bacterial community in its PCR products. From the DGGE analysis, our data indicated that composition of bacterial community were different in the soil samples that were taken from the same site but at different time. This indicated that the species and number of microorganisms in a polluted soil were under a dynamic transition. The combination of DGGE and 16S rDNA gene sequence analysis system were also proven useful in identifying the predominant microbes in a soil sample and monitoring its bacterial community.

Submitted by Nathália Neves (nathalia.neves@ufpe.br) on 2015-03-05T18:54:49Z No. of bitstreams: 2 Nelson_Mestrado_Biblioteca.pdf: 2006130 bytes, checksum: b672ac0459557046842a79f276aa4414 (MD5) license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) / Made available in DSpace on 2015-03-05T18:54:49Z (GMT). No. of bitstreams: 2 Nelson_Mestrado_Biblioteca.pdf: 2006130 bytes, checksum: b672ac0459557046842a79f276aa4414 (MD5) license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Previous issue date: 2012-02-15 / CNPq / Ganoderma abrange 80 espécies de grande importância econômica e ecológica, caracterizando-se por possuir basidiosporos de parede dupla e ornamentada de ápice truncado. Sua taxonomia é baseada em caracteres morfológicos, porém a pluralidade de termos e critérios por taxonomistas fazem a identificação desse grupo ser caótica. Nessa perspectiva, as regiões ITS e LSU (rDNA) são as mais utilizadas para diferenciação de táxons morfologicamente semelhantes. No presente estudo, foram utilizados tanto exemplares de Ganoderma frescos quanto amostras herborizadas para extração de DNA genômico, amplificação das regiões ITS e LSU, e posterior sequenciamento. Desse modo, foram sequenciados 27 espécimes de Ganodermataceae para as regiões ITS e LSU, sendo duas pertencentes à Tomophagus colossus e 25 exemplares pertencentes ao gênero Ganoderma (14 do subg. Ganoderma e 11 do subg. Elfvingia), representando seis espécies para este último gênero. Todas as reconstruções filogenéticas realizadas demonstraram que Ganoderma trata-se de um grupo monofilético, distinto de Tomophagus, porém pertencentes à mesma família. Para o subgênero Ganoderma baseada em exemplares brasileiros, observou-se a delimitação filogenética das cinco espécies estudadas (G. chalceum, G. multiplicatum, G. orbiforme, G. parvulum e G. resinaceum), todas distintas em relação a G. lucidum. No subgênero Elfvingia, os representantes brasileiros momentaneamente identificados como G. tornatum, alocaram-se em subclados distintos de acordo com a distribuição geográfica (Mata Atlântica e Floresta Amazônica). De modo geral, para Ganoderma observa-se uma forte correlação entre os agrupamentos formados e a distribuição geográfica. Esses resultados demonstram a utilidade das regiões do rDNA, associadas aos métodos tradicionais, nos estudos taxonômicos e sua importância para uma identificação mais confiável das espécies.

Submitted by Haroudo Xavier Filho (haroudo.xavierfo@ufpe.br) on 2016-02-26T16:26:29Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) dissertacaofinal19ABR12.pdf: 6509323 bytes, checksum: 3ea20960f438629f3ce90be45103b653 (MD5) / Made available in DSpace on 2016-02-26T16:26:29Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) dissertacaofinal19ABR12.pdf: 6509323 bytes, checksum: 3ea20960f438629f3ce90be45103b653 (MD5) Previous issue date: 2012-03-15 / CAPES / O gênero Cyathus Haller: Pers., junto com mais quatro gêneros, formou, por muito tempo, um grupo conhecido como “Bird’s nest fungi” devido a morfologia cônica com estrutura lenticulares em seu interior, lembrando ovos em seus ninhos. Os estudos morfológicos, durante muito tempo, forneceram a base para as diferentes construções de classificação infragenérica e, como muitas espécies se diferenciavam de forma tênue, isso gerava muita polêmica. Sequências de DNA vêm sendo usadas para esclarecer melhor a história evolutiva do grupo. No Brasil, o grupo é pouquíssimo estudado, de forma que mais trabalhos envolvendo a união da taxonomia molecular com a tradicional são necessários para uma melhor compreensão do gênero. Assim, o presente trabalho teve como objetivos inferir a filogenia de espécimes de Cyathus oriundas do nordeste brasileiro e, com a associação dos caracteres morfológicos, propor uma classificação melhor para o táxon. Para isso, foram examinadas 46 exsicatas, das quais seis eram do herbário JPB, cinco do Herbário HUEFS e 37 do herbário de Fungos UFRN, sendo que este último apresenta o maior número de exsicatas de Cyathus no Nordeste, somando um total de 19 espécies. Dos 46 espécimes, 33 foram utilizados para extração de DNA, resultando no sequenciamento da região ITS de seis espécies e da região LSU de oito espécies. As árvores filogenéticas mostram que essas espécies se encontram em clados separados das demais espécies oriundas de regiões não tropicais, no entanto, não altera a topologia geral dos principais clados sugeridos em trabalhos anteriores. De acordo com os estudos taxonômicos tradicionais, temos duas espécies que constituem segundo registro para o mundo: C. pedicelatus e C. setosus; uma primeira referencia para o Brasil de C. olivaceobrunneus, e a primeira referência para o Nordeste de C. bekerleyanus. A análise dos dados moleculares aponta para a delimitação de cinco novas espécies para a ciência. Todas as sequências de DNA geradas foram depositadas no GenBank e são novas para o banco de dados. / The genus Cyathus Haller: Pers., together with other four genera, formed a group known as bird's nest fungi due to their conic morphology with lenticular structures inside resembling bird’s eggs in their nests. Morphological studies, for a long time, provided the basis to the different constructions of infrageneric classification and, as many species differed weakly from one another, that generated much controversy. DNA sequences have been used to better clarify the evolutionary history of the group. In Brazil, the group is as yet poorly studied, hence, more work involving the union of the traditional and molecular taxonomy is needed for better understanding of the genus. Therefore, this work had the objectives of inferring the phylogeny of specimens of Cyathus from the Brazilian Northeast and, in association with morphological characters, to propose a better classification for the taxon. Forty six exsiccates were examined, being six from JPB herbarium, five from HUEFS herbarium and 37 of UFRN-Fungos herbarium. The latter has the largest number of exsiccates of this genus in the Northeast, totaling 19 species. Thirty-three specimens were chosen for DNA extraction, resulting in the sequencing of the ITS region of six species and the LSU region of eight species. The phylogenetic trees show that these species sampled in this work are positioned in clades separated from those reported from non-tropical regions, however, this does not change the general topology of the main clades suggested in the works published previously. According to the traditional taxonomy studies we have two species that constitute the second record to the world, C. pedicelatus and C. setosus; one first record to Brazil, C. olivaceobrunneus; and one first record to the Northeast region of C. bekerleyanus. Analysis of the molecular data suggests five new species to Science. All DNA sequences of the tropical species sampled were deposited in the GenBank and are new records for the database.

Abstract The genetic diversity and identification of slow- and fast- growing soybean root nodule bacterial isolates from different agro-climatic regions in Mpumalanga, Limpopo and Gauteng Provinces of South Africa were evaluated. The 16S-rDNA-RFLP analysis of 100 rhizobial isolates and eight reference type strains placed the isolates into six major clusters, and revealed their site-dependent genomic diversity. Sequence analysis of single and concatenated housekeeping genes (atpD, glnII and gyrB), as well as the symbiotic gene nifH captured a considerably higher level of genetic diversity and indicated the dominance of Bradyrhizobium diazoefficiens and Bradyrhizobium japonicum in Mpumalanga, Limpopo and Gauteng Provinces. Gene sequence similarities of isolates with type strains of Bradyrhizobium ranged from 97.3 to 100% for the 16S rDNA, and 83.4 to 100% for the housekeeping genes. The glnII gene phylogeny showed discordance with the other genes, suggesting lateral gene transfer or recombination events. Concatenated gene sequence analysis showed that most of the isolates did not align with known type strains and might represent new species from South Africa. This underscores the high genetic variability associated with soybean Bradyrhizobium in South African soils, and the presence of an important reservoir of novel soybean-nodulating bradyrhizobia in the country. In this study, the grouping of isolates was influenced by site origin, with Group I isolates originating from Limpopo Province and Group II and III from Mpumlanga Province in the 16S rDNA-RFLP analysis.

Submitted by Daniella Sodre (daniella.sodre@ufpe.br) on 2015-03-17T11:47:51Z No. of bitstreams: 2 Dissertação Lidia Silva.pdf: 4103104 bytes, checksum: eb5e0fe6bb40a7f4698d819d1e3c3755 (MD5) license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) / Made available in DSpace on 2015-03-17T11:47:51Z (GMT). No. of bitstreams: 2 Dissertação Lidia Silva.pdf: 4103104 bytes, checksum: eb5e0fe6bb40a7f4698d819d1e3c3755 (MD5) license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Previous issue date: 2013-02-22 / CNPq / Os Agaricomycetes de solo são representados pelos fungos clavarioides, estereoides, agaricoides e algumas espécies poroides. Esses fungos apresentam um papel fundamental na manutenção desses ecossistemas através da ciclagem de nutrientes e suas diferentes relações ecológicas (saprofitismo, parasitismo e micorrização). Com o objetivo de avaliar a diversidade e as interações ecológicas de Agaricomycetes terrícolas (clavarioides, estereoides e poroides), esses fungos foram investigados em quatro áreas de Mata Atlântica. Das 18 visitas realizadas nas áreas (oito na Reserva Biológica de Saltinho, quatro no Parque Estadual Dois Irmãos, quatro no Jardim Botânico de Recife e duas na RPPN Frei Caneca) entre 2011 e 2012, foram coletados 54 espécimes de Agaricomycetes terrícolas. Além das coletas, foram revisadas 105 exsicatas depositadas do Herbário URM, resultando em um total de 159 espécimes. Foram identificadas 54 espécies distribuídas em 19 gêneros e 10 famílias. Foram encontrados 12 novos registros para a Região Nordeste e um para o bioma Mata Atlântica. Dos 54 espécimes coletados, foram obtidos sete isolados em meio de cultura, distribuídos em quatro gêneros e cinco espécies. Esses isolados foram testados para celulase, amilase, protease, lacase e peroxidase. Todas as cinco espécies foram negativas para a produção de celulase, amilase e protease e positivas para lacase e peroxidase. Em relação aos aspectos ecológicos, verificou-se que o número de espécimes e espécies coletados foi significativamente maior no período chuvoso. Dos 54 espécimes coletados e dos sete isolados, foram obtidas 17 sequencias da região ITS: cinco sequencias de Amauroderma schomburgkii, duas de A. praetervisum, uma de A. sprucei, uma de Clavulina amazonensis, uma de Clavulinopsis aff. flavella, uma de Clavariaceae, três de Hymenochaete damicornis, uma de Lachnocladium schweinfurthianum, uma de Podoscypha sp1, uma de Ramaria aff. tubulosa e duas de Podoscypha cf. tomentipes (substrato de madeira).

Brassica napus (AACC, 2n = 38) is an allotetraploid species derived from the parentel diploid species Brassica rapa (AA, 2n = 20) and Brassica oleracea (CC, 2n = 18). The aim of thesis was to carry out the genetic and epigenetic analysis of high-copy rRNA genes (or rDNA) in several varieties of hybrid species B. napus. The experiments involved determining the ratio of parental genes in hybrids, sequencing and methylation analysis of the promoter region of rDNA. Using Southern hybridization, it was revealed significant variability in the number of parental rDNA units between each variety. Data from sequence analysis were in good agreement with the results of Southern blot. Genetic recombination between parental rDNA units was revealed in one variety by DNA sequencing of promotor region. To study methylation, bisulfite sequencing was performed. It was found out that rDNA units of B. rapa origin have a higher value of methylation than units originated from B. oleracea.