Thesis (M.S.)--University of Michigan, 1976.

Thesis (M.S.)--University of Tennessee Health Science Center, 2009. / Title from title page screen (viewed on July 29, 2009 ). Research advisor: Stephen White. Document formatted into pages (vii, 47 p. : ill.). Vita. Abstract. Includes bibliographical references (p. 41-46).

Thesis (Ph. D.)--University of Washington, 1999. / Vita. Includes bibliographical references (leaves 143-181).

Thesis (Ph. D.)--University of Washington, 1996. / Vita. Includes bibliographical references (leaves [141]-172).

Thesis (doctoral)--Rijksuniversiteit te Utrecht, 1963. / "Appendices" (1 v.) inserted in pocket at end.

Thesis (doctoral)--Rijksuniversiteit te Utrecht, 1963. / "Appendices" (1 v.) inserted in pocket at end. eContent provider-neutral record in process. Description based on print version record.

It was demonstrated that induction of a major outer protein, HI, was associated with increased resistance to chelators of divalent cations such as EDTA and to the cationic antibiotics polymyxins and aminoglycosides. Outer membrane protein HI was the major cellular protein in cells grown in Mg²⁺-deficient medium (0.02 mM Mg²⁺) and in mutants selected for resistance to polymyxin. Increase in protein HI was associated with decrease in cell envelope Mg²⁺. Induction of protein HI was prevented by supplementation of Mg²⁺-deficient medium with 0.5 mM Mg²⁺, Ca²⁺, Mn²⁺ or Sr²⁺, but not by Zn²⁺, Ba²⁺, or Sn²⁺. Cells grown in Ca²⁺, Mn²⁺ or Zn²⁺ showed enhanced levels of these cations as main major cell envelope associated cation. Only cells grown in the presence of those cations which failed to prevent HI induction were resistant to chelators, polymyxin B and gentamicin. Protein HI overproducing cells also demonstrated altered streptomycin uptake. It was further demonstrated that aminoglycosides could interact with the outer membrane so as to make it more permeable to other substances. Mg²⁺ inhibited aminoglycoside-mediated permeabilization. Both aminoglycosides and polymyxin B could be shown to displace a small amount of Mg²⁺ from the cell envelope. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

No description available.

No description available.

Acinetobacter spp é um dos mais freqüentes agentes de infecções nosocomiais nos hospitais brasileiros. Vários surtos hospitalares causados por Acinetobacter resistente aos carbapenêmicos já foram descritos no Brasil. O presente estudo verificou a presença de alteração de proteínas da membrana externa e produção de carbapenemases em cepas de Acinetobacter spp. resistentes aos carbapenêmicos isoladas de corrente sanguínea de pacientes internados no HC-FMUSP. Os isolados foram identificados pelo método miniaturizado API20NE. As concentrações inibitórias mínimas dos carbapenêmicos foram realizadas através da técnica de microdiluição em caldo. As carbapenemases foram estudas através de testes fenotípicos, detecção dos genes foi realizada pela reação de amplificação em cadeia da polimerase (PCR), sequenciamento e hidrólise de imipenem. As proteínas de membrana externa foram caracterizadas através da técnica de SDS-Page e PCR. A tipagem molecular foi realizada usando a técnica de eletroforese em campo pulsado. Noventa e oito por cento dos isolados apresentaram genes codificadores da enzima Oxa-51-like, sendo que a única amostra que não apresentou essa enzima, após seqüenciamento do gene codificador da porção 16S do RNA ribossomal, foi considerada da espécie A. calcoaceticus. A presença do gene blaoxa-51-like não foi encontrada adjacente a seqüência ISAba1. Em dezoito por cento do total das amostras foi detectada pela reação de PCR o gene blaoxa-51-like/oxa-23- like, sendo que sete desses isolados pertenciam ao mesmo clone. Somente em um isolado não foi detectada a presença do grupamento ISAba-1 adjacente ao gene codificador da enzima Oxa-23. Dessas amostras apenas uma hidrolisou o antimicrobiano imipenem. Cinco isolados apresentaram o gene blaimp (IMP-1 pelo seqüenciamento). Apenas dois desses isolados se mostraram capazes de hidrolisar imipenem e através da inibição com o EDTA confirmou-se a presença da produção de enzimas Metalo-- lactamase. Vários testes fenotípicos foram utilizados para a detecção de carbapenemase como DDST, CD e Etest. Quando comparados com a PCR (metodologia padrão-ouro), os melhores resultados foram observados com a aproximação de disco de imipenem (10g) com um disco contendo 3L do ácido -mercaptoetanol não diluído (sensibilidade de 100% e especificidade de 71%). Observou-se que na maioria dos isolados, as proteínas de membrana externa analisadas estavam diminuídas ou ausentes. Três isolados apresentaram ausência dessas proteínas, suas CIM variaram de 32 a 128g/mL e de 64 a 256g/mL para o imipenem e meropenem respectivamente. O presente estudo verificou que a resistência aos carbapenêmicos está relacionada à presença de carbapenemases e alterações de membrana externa. Além disso, a importância das carbapenemases como principal mecanismo de resistência em isolados de Acinetobacter deve ser melhor investigada, já que esses genes podem não estar sendo expressos / Acinetobacter spp is an important etiologic agent causing nasocomial infection in Brazilian hospitals. Carbapenem resistance among this agent has been increasing in the last decade, and several outbreaks due to resistant Acinetobacter had been identified in Brazil. This study verified the presence of altered outer membrane proteins and production of carbapenemase in carbapenem-resistant Acinetobacter spp. strains isolated from bloodstream infections of patients hospitalized in the HC-FMUSP. This study verified the presence of altered outer membrane proteins and production of carbapenemase in Acinetobacter spp. carbapenem-resistant strains isolated from bloodstream of patients hospitalized in the HC-FMUSP. The isolates were identified by miniatured method API20NE and the miminal inhibitory concentration of carbepenem was performed by broth microdilution. Carbapenemases were studied by phenotypic screnning tests, detection of its genes coder by amplification polymerase chain reaction (PCR), sequence and imipenem hydrolise. The proteins of external membrane were studied by SDS-PAGE and PCR. Molecular typing was performed using the technique of pulsed field gel electrophoresis. About 98% of these isolates were positives for genes encoding the enzyme Oxa-51-like, only one strain did not exhibit this enzyme. After sequencing of the gene encoding portion 16S ribosomal RNA this strain was considered the species A. calcoaceticus. The presence of the sequence adjacent ISAba1 was not found in none of these isolates with the gene blaOXA-51-like. Eighteen percent of the total of strains demonstrated by PCR the gene blaoxa-51-like/oxa-23-like. Seven of these strains belonged to the same clone. Only one isolate did not show the presence of ISAba1 adjacent to the gene encoding Oxa-23. Only one of this strain hydrolyzed imipenem. Five isolates had the gene blaimp, (IMP-1 by sequencing). Two of these isolates had proved capable of hydrolyzing the antibiotic imipenem and the inhibition with EDTA confirmed the presence of MBL-producing enzymes. Several phenotypic tests were used to screnning of carbapenemase as DDST, CD and Etest. When compared with PCR, the gold standard, the best results were seen with the next disk of imipenem (10 mg) with another disk containing 3L acid -mercaptoethanol pure (100% sensitivity and specificity 71%). In most isolates the outer membrane proteins were decreased or absent. Three isolate showed total absence of these proteins, their MIC ranged from 32 to 64 to 128g/mL and 256g/mL to imipenem and meropenem respectively. This study has shown that carbapenem resistance was linked to the presence of carbapemenases and alteration of the outer membrane. Moreover, the significance of carbapenemase as the main mechanism of resistance in isolates of Acinetobacter should be better investigated, since these genes might not be cast