Orientador: Angelo Berchieri Junior / Resumo: Salmonella Enteritidis e Salmonella Typhimurium causam infecções em seres humanos e animais que são frequentemente associadas à extensa colonização intestinal e excreção fecal. A presença de estrutura flagelar no patógeno está relacionada à indução de inflamação intestinal e atenuação de infecção sistêmica no hospedeiro. Por outro lado, a infecção por estirpes aflageladas resulta em pouca inflamação e consequente infecção sistêmica grave. No presente estudo, foi avaliada a hipótese de que a síntese de maior quantidade de flagelina em estirpes de Salmonella geneticamente modificadas poderia levar a infecção sistêmica menos intensa em aves. Para investigar as conseqüências da superprodução de flagelina, foram construídas estirpes de Salmonella Enteritidis e Typhimurium contendo deleções nos genes clpP e fliD (que levam à superexpressão de flagelina) e patogenicidade e imunogenicidade foram comparadas com as respectivas estirpes selvagens em aves infectadas. Os resultados indicaram que o aumento da síntese de flagelina por SE ΔclpPΔfliD e STM ΔclpPΔfliD culmina em déficit da taxa de multiplicação bacteriana. Porém, tais alterações não interferiram na capacidade de colonização cecal e excreção fecal das estirpes mutantes. O mesmo foi observado em fígado e baço, mas após 14 dpi as estirpes mutantes tendem a serem eliminadas destes órgãos. Mesmo com síntese mais elevada de flagelina, as estirpes mutantes recrutaram quantidades semelhantes de linfócitos e macrófagos em tonsila cecal... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Salmonella Enteritidis and Salmonella Typhimurium cause infections in humans and animals that are often associated with extensive intestinal colonization and faecal shedding. The presence of flagellar structure in the pathogen is related to the induction of intestinal inflammation and attenuation of systemic infection in the host. On the other hand, the absence of flagellin results in severe systemic infection as a result of mild inflammatory intestinal responses provoked by aflagellated strains. The hypothesis that higher flagellin production by Salmonella strains could induce immunogenic response during infection in chickens was evaluated in the present study. To investigate the consequences of flagellin overproduction, strains of Salmonella Enteritidis and Typhimurium containing clpP and fliD deletions (which lead to flagellin overexpression) were constructed, and pathogenicity and immunogenicity were compared with their respective wild-type strains in infected chickens. The results suggested that the increase in flagellin synthesis by SE ΔclpPΔfliD and STM ΔclpPΔfliD culminates in a deficit in the bacterial multiplication rate. However, that changes did not interfere with the capacity for caecal colonization and faecal excretion of mutant strains. The same was observed in the liver and spleen, but after 14 dpi, mutant strains tend to be eliminated from these organs. Even with higher flagellin synthesis, the mutant strains recruited similar amounts of lymphocytes and macro... (Complete abstract click electronic access below) / Doutor

Orientador: Ticiana Sidorenko de Oliveira Capote / Resumo: A doença periodontal (DP) é uma desordem imunoinflamatória, disbiótica, multifatorial e com suscetibilidade genética. Estudos demonstraram a associação entre periodontite e diabetes mellitus (DM). Considerando que tanto o DM como a DP estão associadas a um estado de inflamação crônica e a uma produção aumentada de espécies reativas de oxigênio, elas podem estar relacionadas à produção de danos no DNA. Portanto, o objetivo deste projeto foi investigar a relação da periodontite crônica com perfil glicêmico e danos no DNA. Foram realizados exames bioquímicos (Hemoglobina glicada - HbA1c; Glicemia jejum; Insulina; Colesterol total; Triglicérides) e periodontais, avaliação de indicadores antropométricos de obesidade (IMC e relação quadril/cintura), e avaliação do dano ao DNA pelo ensaio de micronúcleo pelo bloqueio da citocinese (CBMN) em linfócitos, observando a frequência de células binucleadas com micronúcleo (FBMN) e a frequência de micronúcleo (FMN). Os pacientes foram divididos conforme seus níveis glicêmicos e seu perfil periodontal em três grupos: grupo A, n=37, sem periodontite e sem DM tipo 2 - DM2; grupo B, n=47, com periodontite, sem DM2; grupo C, n=19, com periodontite e com DM2. Regressão multivariada de Poisson foi realizada utilizando a FBMN ou a FMN como variáveis dependentes. Os valores foram estimados em razão de taxa com seus respectivos intervalos de confiança definidos em 95% (IC 95%). Os parâmetros periodontais dos pacientes diabéticos e com periodontite est... (Resumo completo, clicar acesso eletrônico abaixo) / Mestre

Aspergillus niger DSM 26641 was exposed to 60Co g-radiation to enhance the b-1,4-endoxylanase activity, restrict colony growth and improve robustness of pellets. The first promising mutant obtained after g-radiation of the fungal spores at 50-2000 Gy showed a restricted colony growth and an 82% enhancement in b-1,4-endoxylanase activity. The mutant was subjected to a second round of g-radiation at 1400 Gy generating a mutant with double the b-1,4-endoxylanase activity compared to the native strain. The selected final mutant, deposited as Aspergillus niger DSM 28712, showed a maximal saccharification activity of 26 U·ml-1 on xylan based broth, 48 U·ml-1 on lignocellulose hydrolysate and 375 U·ml-1 on lignocellulose hydrolysate supplemented with yeast extract and mineral salts.

English Abstract Shortly after the identification of HIV as a causative agent of AIDS, an aspartic protease was identified in the viral genetic information. The very same time protease has become one of the dominant therapeutical targets in AIDS therapy. The introduction of protease inhibitors into the antiretroviral therapy has led to a significant improvement in the quality and length of life of HIV patients. However, the virus is still able to effectively prevent the impact of an inhibitor via generating inhibitor-resistant mutated protease variants. Thus, there is a constant need for novel types of inhibitors that would be capable of effectively blocking these resistant variants and simultaneously not supporting the development of novel resistant viral strains. One way to identify such inhibitors could be searching for compounds interacting with the enzyme at different sites than the active cavity, via the mechanisms of noncompetitive or uncompetitive inhibition. The group of compounds called metallacarboranes - inorganic compounds consisting of carbon, boron, hydrogen and metall ion - were shown to exhibit such an activity against HIV-1 protease. However, for further optimization of these inhibitors, detailed biophysical investigation of the enzyme-inhibitor complex is needed. This work focuses on the...

Staphylococcus aureus has developed resistance to several antibiotics including vancomycin, which is often used as a “last resort” treatment. There is an ever-increasing need to develop novel antimicrobial treatments to combat S. aureus and other drug resistant bacteria. Microorganisms are most often found in polymicrobial communities where they either exhibit synergistic or antagonistic relationships. Competition between microorganisms can lead to the discovery of new antimicrobial targets as the specific mechanisms of resistance are elucidated. In addition, synergistic treatments are being evaluated for their combined effect and potential to decrease the concentration of drugs needed, and thus the side effects also. Alcaligenes faecalis is a microorganism that our lab has previously shown to inhibit S. aureus and other various bacterial species. In this study, we found that A. faecalis reduces the planktonic growth of S. aureus by 94.5% and biofilm growth by 76.6%. A. faecalis also has a synergistic effect when paired with bacitracin to reduce the planktonic growth by 99.9% and biofilm growth by 99.7%. Transposon mutagenesis was successfully performed on A. faecalis, and loss of function mutations were attained. Two mutants were no longer able to inhibit the growth of Staphylococcus aureus, Candida albicans, or Bacillus megaterium. Further analysis and genomic sequencing of these mutants is needed to determine the gene(s) that were interrupted and the mechanism of A. faecalis’ antimicrobial activity. The findings of this study may aid in the identification of new therapeutic targets for novel S. aureus treatments.

Chapter II Abstract Human positive cofactor 4 (PC4) is a transcriptional coactivator with a highly conserved single-strand DNA (ssDNA) binding domain of unknown function. We identified PC4 as a suppressor of the oxidative mutator phenotype of the Escherichia coli fpg mutY mutant and demonstrate that this suppression requires its ssDNA binding activity. Saccharomyces cerevisiae mutants lacking their PC4 ortholog Sub1 are sensitive to hydrogen peroxide and exhibit spontaneous and peroxide-induced hypermutability. PC4 expression suppresses the peroxide sensitivity of the yeast sub1Δ mutant, suggesting that the human protein has a similar function. A role for yeast and human proteins in DNA repair is suggested by the demonstration that Sub1 acts in a peroxide resistance pathway involving Rad2 and by the physical interaction of PC4 with the human Rad2 homolog XPG. We show that XPG recruits PC4 to a bubble-containing DNA substrate with a resulting displacement of XPG and formation of a PC4-DNA complex. We discuss the possible requirement for PC4 in either global or transcription-coupled repair of oxidative DNA damage to mediate the release of XPG bound to its substrate. Chapter III Abstract Previously I established that (1) PC4 significantly suppresses oxidative mutagenesis via its single-strand DNA binding activity, (2) a partial suppression of H2O2-induced lethality was observed in a sub1Δ rad2Δ yeast double mutant compared to the sub1Δ mutant, and (3) PC4 interacts with XPG physically and functionally. These results led me to believe that suppression of oxidative mutagenesis and lethality by PC4 is partially due to its function in an XPG/Rad2-dependent pathway and through additional unidentified mechanism(s). In this chapter, I present studies aimed at investigating different DNA repair pathways in which PC4/Sub1 might participate. I address the possible roles of PC4/Sub1 in transcription-coupled repair (TCR) in terms of its binding specificity to oxidative DNA lesions and its ability to allow efficient resumption of transcription after oxidative DNA damaging treatment. To ask if PC4/Sub1 interacts with other DNA repair proteins to protect cells from oxidative DNA damage, I analyzed spontaneous mutation rates among a series of isogenic, haploid yeast mutant strains deficient of SUB1, base excision repair (BER) and/or nucleotide excision repair (NER) functions. I further analyzed genetic interactions between SUB1 and genes critical to various DNA damage avoidance/tolerance mechanisms, such as mismatch repair (MMR), homologous recombination (HR) and translesion synthesis (TLS).

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第18316号 / 農博第2041号 / 新制||農||1020(附属図書館) / 学位論文||H26||N4823(農学部図書室) / 31174 / 京都大学大学院農学研究科食品生物科学専攻 / (主査)教授 保川 清, 教授 安達 修二, 教授 伏木 亨 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第19016号 / 農博第2094号 / 新制||農||1029(附属図書館) / 学位論文||H27||N4898(農学部図書室) / 31967 / 京都大学大学院農学研究科食品生物科学専攻 / (主査)教授 保川 清, 教授 河田 照雄, 教授 谷 史人 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(人間・環境学) / 甲第19069号 / 人博第722号 / 新制||人||173(附属図書館) / 26||人博||722(吉田南総合図書館) / 32020 / 京都大学大学院人間・環境学研究科相関環境学専攻 / (主査)准教授 土屋 徹, 教授 宮下 英明, 教授 川本 卓男 / 学位規則第4条第1項該当 / Doctor of Human and Environmental Studies / Kyoto University / DFAM

No description available.