<I>Escherichia coli</I> mutants impaired in growth and survival at low external pH in minimal medium were selected and attempts made to identify the disrupted genes. This study suggested that <I>clpX</I>, encoding a heat-shock induced protease and molecular chaperone, was functional in survival of <I>E. coli </I>at pH 3.3. Promoter probe plasmid libraries of <I>Salmonella typhimurium </I>LT2 DNA were created in <I>E. coli </I>and screened for acid-inducible transcriptional elements, and transcriptionally active fragments of degradative amino-acid decarboxylase genes recovered. Chromosomal gene fusions to the reporter gene <I>lacZ </I>in <I>E. coli</I> generated by Mu DII 1734 insertion were screened in a similar way and suggested that the gene encoding adenylate cyclase (<I>cya</I>) could be induced by mild cytoplasmic acidification. The sequence of a gene known to be inducible by cytoplasmic acidification, <I>inaA, </I>became available during the course of this study. The 5' region of this gene was used to generate a set of plasmids carrying fragments of the acid-inducible promoter transcriptionally fused to a luciferase based reporter system. Elements of the sequence required for induction by cytoplasmic acidification were identified. One of these reporter constructs was used to screen an <I>E. coli </I>Tn<I>10</I> chromosomal insertional mutant library for genes involved in the regulation of <I>inaA. </I>One such mutant had a multiple antibiotic resistant (<I>mar</I>) phenotype. The disrupted loci in 2 other mutants were identified by inverse PCR, sequence analysis and database searches. Both were known only as open reading frames (ORFs) discovered during the sequencing of the entire <I>E. coli</I> genome, and were tentatively identified as <I>yddB </I>(closely linked to <I>gadB </I>and <I>gadC</I>; required for glutamate dependent acid resistance) and <I>f300</I> (closely linked to <I>pldA; </I>required for detergent resistance). The promoter of <I>f300</I> was shown to be sensitive to cytoplasmic acidification. The <I>inaA</I> promoter was also demonstrated to be induced at the onset of stationary phase, and to be independent of the stationary phase and weak-acid inducible σ factor RpoS and also of cAMP levels.

Thesis (Ph. D.)--West Virginia University, 2005. / Title from document title page. Document formatted into pages; contains ix, 166 p. : ill. Includes abstract. Includes bibliographical references.

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Thèse (M.A.)--Université Laval, 2007. / Bibliogr.: f. [59]-72. Publié aussi en version électronique dans la Collection Mémoires et thèses électroniques.

Thesis (Ph. D.)--University of Texas at Austin, 1998. / Vita. Includes bibliographical references (leaves 76-81). Available also in a digital version from Dissertation Abstracts.

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Made available in DSpace on 2014-12-17T15:42:13Z (GMT). No. of bitstreams: 1 AlineSM_DISSERT.pdf: 1997460 bytes, checksum: 0ad037938fab17947fef7a249102b4ec (MD5) Previous issue date: 2013-07-31 / Aiming to consumer s safety the presence of pathogenic contaminants in foods must be monitored because they are responsible for foodborne outbreaks that depending on the level of contamination can ultimately cause the death of those who consume them. In industry is necessary that this identification be fast and profitable. This study shows the utility and application of near-infrared (NIR) transflectance spectroscopy as an alternative method for the identification and classification of Escherichia coli and Salmonella Enteritidis in commercial fruit pulp (pineapple). Principal Component Analysis (PCA), Independent Modeling of Class Analogy (SIMCA) and Discriminant Analysis Partial Least Squares (PLS-DA) were used in the analysis. It was not possible to obtain total separation between samples using PCA and SIMCA. The PLS-DA showed good performance in prediction capacity reaching 87.5% for E. coli and 88.3% for S. Enteritides, respectively. The best models were obtained for the PLS-DA with second derivative spectra treated with a sensitivity and specificity of 0.87 and 0.83, respectively. These results suggest that the NIR spectroscopy and PLS-DA can be used to discriminate and detect bacteria in the fruit pulp / Visando ? seguran?a do consumidor, ? de extrema import?ncia identificar a presen?a de contaminantes patog?nicos nos alimentos, pois os mesmos s?o respons?veis por surtos alimentares que dependendo do n?vel de contamina??o pode chegar a causar a morte de quem os consome. Na industria h? uma necessidade de que essa identifica??o de contaminantes seja r?pida e rent?vel. Este estudo mostra a aplica??o e utilidade de medidas espectrais de transflect?ncia no infravermelho pr?ximo (NIR) como um m?todo alternativo para a identifica??o e classifica??o de Escherichia coli e Salmonella Enteritidis em polpa de fruta comercial (abacaxi). An?lise de Componentes Principais (PCA), Modelagem Independente por Analogia Classe (SIMCA) e An?lise Discriminante por M?nimos Quadrados Parciais (PLS-DA) foram utilizados na an?lise. N?o foi poss?vel obter uma separa??o total entre as amostras usando PCA e SIMCA. O PLS-DA apresentou bom desempenho na capacidade de predi??o alcan?ando 87,5% para E. coli e 88,3% para S. Enteritides, respectivamente. Os melhores modelos obtidos para o PLS-DA com os espectros tratados com segunda derivada apresentaram sensibilidade e especificidade de 0,87 e 0,83, repectivamente. Estes resultados sugerem que a espectroscopia NIR e PLS-DA podem ser usados para discriminar e detectar bact?rias na polpa da fruta

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M. Tech (Department of Biotechnology, Faculty of Applied and Computer Sciences) Vaal University of Technology. / Water shortages, water pollution and climate changes are highly interrelated global issues. These have raised immense concerns about serious adverse effects on the quality, treatment and re-use of wastewater. A major role of water is for vitality of life on earth. Water is recognized as source of evolution from origin to degree of civilization, since it is an essential resource its treatment becomes a necessity for day to day for life. Nanoparticles and their application in treatment of wastewater is becoming a major area of research. It is mainly applicable to the removal of major contaminants like microorganisms. This study was carried out with an objective to investigate the antibacterial and antifungal potentials of nanoparticles. Cobalt and cobalt complexes of urea and thiourea were synthesized and characterized using UV-Vs, PL, FTIR, TEM, SEM, XRD and TGA techniques. The Co particles are in a mixture of rod, agglomerates with irregular shape around 50 – 100 nm in diameter. The Co/Thiourea particles appear to be around 10 – 30nm in size. The Co complexed with urea images showed spherical to hexagonal shape with 50 nm size in diameter. The antimicrobial activity was determined using Minimum Inhibitory and bactericidal concentration and the well diffusion method. The antibacterial and antifungal activities of ratios (1:1, 1:2, 1:3, 2:1 μg/mL) of doped cobalt nanoparticles were tested against a panel of five Gramnegative bacteria - (Escherichia coli, Pseudomonas aeruginosa, Shigella enterica, Salmonella typhi and Salmonella sonnei) human pathogenic bacteria; and two fungal strains - Aspergillus niger and Candida albicans. Zones of inhibition as a consequence of nanoparticles were compared with that of different standards like Neomycin for antibacterial activity and Amphotericin B for antifungal activity. The results showed a remarkable inhibition of the bacterial growth against the tested organisms. The most striking feature of this study is that Cobalt, Urea and Thiourea nanoparticles have antifungal activity comparable or more effective (as in case of Thiourea on A. niger) than Amphotericin B and nearly promising antibacterial activity although not comparable to Neomycin.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Enterobactérias produtoras de ESBLs são descritas tanto no ambiente hospitalar quanto na comunidade em todo o mundo. No Brasil, esses microrganismos também têm emergido como uma causa importante de infecções, sendo as enzimas CTX-M as prevalentes. O objetivo deste estudo foi analisar diferentes aspectos genotípicos relacionados à expressão da resistência aos antimicrobianos em cepas Escherichia coli e de Salmonella spp, tais como: a diversidade de ESBLs, os genes de resistência aos antimicrobianos e o conteúdo plasmidial. Os aspectos epidemiológicos das cepas produtoras de ESBLs também foram investigados. Foram estudadas 88 cepas de enterobactérias, sendo 43 E. coli e 45 cepas de Salmonella spp., de origem hospitalar e da comunidade (principalmente alimentos), isoladas na cidade do Rio de Janeiro. A expressão de ESBL foi observada em sete cepas de E. coli (7/43, 16,3%) e em uma cepa de Salmonella Typhimurium (1/45, 2,3%) e as enzimas foram identificadas como variantes de CTX-M e SHV-5, respectivamente. Entre as cepas de E. coli, a enzima CTX-M-2 foi a mais frequente (n = 4), sendo detectada em cepas isoladas de swab retal de pacientes hospitalizados, enquanto as enzimas CTX-M-59 (uma variante de CTX-M) (n = 1) e CTX-M-9 (n = 2) foram identificadas em cepas isoladas a partir de espécimes clínicos. Salmonella Typhimurium produtora de SHV-5 foi isolada do ambiente hospitalar (fórmula infantil). As cepas de E. coli produtoras das enzimas CTX-M pertenceram a grupos filogenéticos (A, B1, D) e STs (ST34, ST69, ST101) diferentes, sendo os genes blaCTX-M identificados em plasmídeos com tipo de replicon IncA/C de cerca de 150 kb (blaCTX-M-2, blaCTX-M-9, blaCTX-M-59) ou 80 kb (blaCTX-M-2). A cepa de S. Typhimurium produtora de SHV-5 pertenceu a um único clone (A-ST19) e o gene blaSHV-5 foi identificado em plasmídeo com o replicon IncL/M com aproximadamente 55Kb. Foi identificado pela primeira vez no Brasil o ST313 em um clone de S. Typhimurium (D-ST313), comumente associado com doenças invasivas severas, particulamente no continente africano. Genes que codificam para a resistência aos antimicrobianos não-beta-lactâmicos e integrons classe 1 foram identificados entre as cepas de E. coli e de Salmonella spp. multirresistentes produtoras ou não de ESBLs. Em conclusão: i) nossos resultados referentes à E. coli confirmaram a disseminação de enzimas CTX-M (principalmente variantes do grupo CTX-M-2) desde, pelo menos, o ano de 2000, em hospitais no Rio de Janeiro; demonstraram a implicação dos plasmídeos IncA/C na disseminação de genes blaCTX-M; indicaram a possível evolução intra-plasmídeo de blaCTX-M-59 a partir de blaCTX-M-2; a observação da diversidade e multiplicidade de plasmídeos poderiam fornecer plataformas genéticas para a dispersão de diferentes genes e/ou elementos de resistência aos antimicrobianos; ii) em relação à Salmonella spp. este estudo descreveu, pela primeira vez, o isolamento, a partir de fórmula infantil, de uma cepa de S. Typhimurium produtora de ESBL; foi demonstrada a associação do gene blaSHV-5 com plasmídeo do tipo IncL/M, que é considerado epidêmico; foi identificado o clone D-ST313 de S. Typhimurium, que está associado a doenças invasivas severas no continente africano, que reuniu cepas isoladas exclusivamente do ambiente hospitalar. / ESBL-producing Enterobacteriaceae have been described in hospitals and in the community worldwide. In Brazil, ESBL-producing Enterobacteriaceae have also emerged as an important cause of infections, being CTX-M enzymes the most prevalent ESBLs. The objective of this study was to analyze different genotypic aspects related to expression of antimicrobial resistance in isolates of Escherichia coli and Salmonella spp., such as: diversity of ESBLs, antibiotic resistance genes and plasmid content. Epidemiological features of ESBL-producing isolates were also investigated. We studied 88 isolates of enterobacteria, 43 E. coli and 45 Salmonella serotypes of hospital and community (mainly food) origin, isolated in the city of Rio de Janeiro. ESBL expression was observed in seven E. coli isolates (7/43; 16,3%) and in one Salmonella Typhimurium (1/45; 2,3%) and the enzymes identified as CTX-M variants and SHV-5, respectively. Among the E. coli isolates, CTX-M-2 was the most frequent (n=4), being detected in isolates recovered from rectal swabs of hospitalized patients, whereas CTX-M-59 (a CTX-M-2-variant) (n=1) and CTX-M-9 (n=2) were identified in E. coli isolated from clinical specimens. SHV-5-producing S. Typhimurium was isolated from the hospital environment (infant formula). CTX-M-producing E. coli belonged to different phylogenetic groups (A, B1, D) and STs (ST34, ST69, ST101), being blaCTX-M genes were identified in IncA/C plasmids of approximately 150 kb (blaCTX-M-2, blaCTX-M-9, blaCTX-M-59) or 80 kb (blaCTX-M-2). SHV-5-producing S. Typhimurium belonged to a single clone (A-ST19) and blaSHV-5 gene was identified in IncL/M plasmids of approximately 55Kb. This study first described in Brazil the isolation of S. Typhimurium belonging to ST313 commonly associated with severe invasive diseases, particularly in Africa. Genes encoding resistance to non-beta-lactams and class 1 integrons were found among ESBL-producers and non-ESBL-producing multidrug-resistant E. coli and Salmonella spp. In conclusion: i) our results related to E. coli confirmed the dissemination of CTX-M-enzymes (especially CTX-M-2-variants) since, at least, the beginning of the last decade in Rio de Janeiro clinical settings; demonstrated the implication of IncA/C plasmids in the spread of blaCTX-M genes; indicated the possible intra-plasmid evolution of blaCTX-M-59 from blaCTX-M-2; observation of the diversity and multiplicity of plasmids would provide genetic platforms for spread of different antibiotic resistance genes and/or elements; ii) in relation to Salmonella spp. this study described for the first time, the isolation, from infant formula, ESBL- producing S. Typhimurium; has been demonstrated the association of blaSHV-5 to plasmids belonging to IncL/M group, that can be considered epidemic plasmids; was identified D-ST313 clone in S. Typhimurium, commonly associated with severe invasive diseases, particularly in Africa, among isolates recovered exclusively from hospital.