Sortase A is a housekeeping enzyme of Gram-positive bacteria that catalyses the anchoring of surface proteins to the bacterial peptidoglycan. The enzyme works to establish an interaction between bacteria and host cells and is essential for pathogenesis. This makes Sortase A a potential suitable target for inhibition, in order to treat bacterial infections. In this degree project Sortase A from Staphylococcus aureus was explored and potential inhibitors were investigated by performing enzyme activity and bacterial binding assays. A robust FRET assay was developed and optimized for a recombinant version of the enzyme and serves as a good starting point for studying inhibition.

Diss. (sammanfattning) Göteborg : Göteborgs universitet, 2010.

Growth of Staphylococcus aureus with the NSAID salicylate increases phenotypic resistance (SAPAR), and the frequency at which heritable resistance occurs to various antibiotics (SAGAR). This study describes the effect of salicylate on heritable and phenotypic resistance to a set of antibiotics for laboratory and multi-drug resistant strains of S. aureus and investigates the link between resistance and SAGAR. Drug gradient plates were used to determine phenotypic resistance to antibiotics targeting DNA replication, transcription, translation and the cell wall in the presence or absence of salicylate. To measure heritable resistance, mutation frequencies were determined for each antibiotic in the presence and absence of salicylate. Salicylate significantly increased mutation frequency of SH1000 to ciprofloxacin 27- fold from 4.9 x 10-8 to 8.5 x 10-7. A significant 8.5- fold increase was observed for LAC from 5.2 x 10-7 to 2.1 x 10-6. Conversely, salicylate significantly decreased mutation frequency for SH1000 to lincomycin 0.035-fold from 3.4 x 10-7 to 1.3 x 10-7. Deletion of the general stress sigma factor sigB encoding σB in SH1000 resulted in decreased heritable and phenotypic resistance, signifying the importance of σB in the full expression of both phenotypes. Metabolite profiling revealed downregulation of glycolysis, TCA, pentose phosphate pathway, and amino acid metabolism. The downregulation of the TCA cycle was confirmed as observed through an increase in NAD+ at growth toxic concentrations of salicylate. Salicylate has been shown to result in ROS accumulation and disruption of proton motive force in mitochondria. SAGAR was only detected for fluoroquinolones, which have been shown to impair TCA cycle and result in ROS accumulation. Examination of ROS under growth-toxic concentrations of salicylate did not reveal a significant increase in ROS levels. Also, the combination of ciprofloxacin and salicylate did not result in an increase in ROS levels. Despite this, addition of the antioxidant glutathione abrogated SAGAR for ciprofloxacin in SH1000 but not for SAPAR. Analysis of SAGAR with NSAIDs benzoate and acetyl salicylic acid revealed a necessity for the ortho hydroxyl group on salicylate to fully express SAGAR. These results suggest that salicylate has pleiotropic effects on S. aureus that include antimicrobial resistance, altered metabolic flux and accumulation of ROS as well as unidentified regulatory genes.

A genetic algorithm was employed to select the optimal combination of preprocessing variables, including data pretreatment, data manipulation and feature extraction procedures, for eventual clustering of a data set consisting of hyperspectral images acquired by a focal plane array Fourier transform infrared (FPA-FTIR) spectrometer. The data set consisted of infrared images of bacterial films, and the classification task investigated was the discrimination between Gram-positive and Gram-negative bacteria. The genetic algorithm evaluated combinations of variables pertaining to bacterial film thickness tolerances, baseline correction, pixel co-addition, outlier removal, smoothing, mean centering, normalization, derivatization, integration and principal component selection. Following numerous iterations of unsupervised processing, the genetic algorithm arrived at a sub-optimal solution yielding a clustering accuracy of 97.8% and a data utilization of 28.6%. The results provided insight into the co-dependencies of the pre-processing variables and their consequential effect on the selected data. The robustness of the classification model was evaluated and reinforced by the successful classification of two distinct validation sets. The overall success of the genetic algorithm suggests that it is an effective time saving resource for the optimization of pre-processing variables that does not require operator intervention.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2008. / Title from first page of PDF file (viewed Feb. 19, 2009). Includes bibliographical references.

Thesis (Ph. D.)--Ohio State University, 2005. / Title from first page of PDF file. Document formatted into pages; contains xv, 139 p.; also includes graphics (some col.) Includes bibliographical references (p. 123-139).

Thesis (Ph. D.)--Florida State University, 2005. / Advisor: Dr. Timothy M. Logan, Florida State University, College of Arts and Sciences, Dept. of Chemistry and Biochemistry. Title and description from dissertation home page (viewed Sept. 14, 2005). Document formatted into pages; contains xix, 166 pages. Includes bibliographical references.

München, Techn. Univ., Diss., 2004.

Made available in DSpace on 2014-06-11T19:24:15Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-06-15Bitstream added on 2014-06-13T19:10:40Z : No. of bitstreams: 1 witzel_cl_me_botfm_parcial.pdf: 120396 bytes, checksum: ec18337e12bca612c7c9844103a8160d (MD5) Bitstreams deleted on 2014-06-27T19:00:45Z: witzel_cl_me_botfm_parcial.pdf,Bitstream added on 2014-06-27T19:02:01Z : No. of bitstreams: 1 witzel_cl_me_botfm.pdf: 1277080 bytes, checksum: c6e53fb083813be017b346181b7be466 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Universidade Estadual Paulista (UNESP) / As Infecções estafilocócicas são grande causa de mortalidade e morbidade. A emergência de isolados de Staphylococcus aureus resistente à meticilina (MRSA) originários da comunidade na década de 1990 trouxe desafios para sua prevenção e abordagem terapêutica. A identificação da prevalência e características de MRSA adquirido em comunidade (CA-MRSA) em populações de alto risco pode fornecer valiosas informações para compreensão de sua epidemiologia. O objetivo deste estudo foi identificar a prevalência, os fatores de risco e as características moleculares de CA-MRSA isolado em nasofaringe de uma população carcerária em Avaré, Brasil. Foram coletados swabs nasais de 302 homens encarcerados no Centro de Ressocialização de Avaré. A caracterização da resistência à meticilina foi feita através de difusão em disco (Oxacilina e Cefoxitina) e identificação do gene mecA (por Reação em Cadeia de Polimerase (PCR). Utilizou-se também PCR para identificação dos genes lukF-PVe lukS-PV que codificam a leucocidina de Panton-Valentine. Fatores de risco para colonização nasal por S. aureus e por MRSA foram avaliados em modelos univariados e multivariados (regressão logística). A prevalência de colonização nasal por S. aureus como um todo foi de 16,5%, e a MRSA (definida como positividade para o gene mecA), de (02) 4,0%. Os testes de difusão com disco de Oxacilina e Cefoxitina identificaram 2 (4%) resistentes simultaneamente a Oxacilina e Cefoxitina. Nenhum dos isolados de S. aureus apresentou genes codificadores da PVL. As amostras positivas para gene mecA apresentaram SCCmec tipo IV, confirmando esta tipagem presente em amostras comunitárias.Uma das amostras positivas para gene mecA apresentou SCCmec IV-d, que pelo método PFGE agrupou em 85,7% com o clone JCSC 4469 de origem japonesa. Os fatores de risco para... / Staphylococci infections are a major cause of morbidity and mortality. The emergence of methicillin-resistant Staphylococcus aureus (MRSA) isolates original from the community in the 1990s brought challenges for prevention and therapeutic approach.Identifying the prevalence and characteristics of community-acquired MRSA (CA-MRSA) in high-risk populations can provide valuable information for understanding its epidemiology. The objective of this study was to identify the prevalence, risk factors and molecular characteristics of CA-MRSA isolates in nasopharynx of a prison population in Avare, Brazil. Nasal swabs were collected from 302 men incarcerated in Central Resocialization of Avaré. The characterization of methicillin resistance was done by disk diffusion (Oxacillin and Cefoxitin) and mecA gene identification (by Polymerase Chain Reaction(PCR). We also used PCR to identify the lukF-PV and lukFS-PV genes that encode the Panton- Valentine Leukocidin. Risk factors for nasal colonization by S. aureus and MRSA were evaluated with univariate and multivariate models (logistic regression). The prevalence of nasal colonization by S. aureus as a whole was 16.5%, and MRSA (defined as positive for the mecA gene), from (02) 4.0%. The diffusion testing with Oxacillin and Cefoxitin identified 2 (4%) isolates resistants to both Oxacillin and Cefoxitin. None of the isolated S.aureus had genes encoding PVL. Positive samples for mecA showed SCCmec type IV, confirming this typing in community samples. One of the positive samples for mecA showed SCCmec IV-d, which by PFGE method grouped in 85.7% with clone 4469 JCSC of Japanese origin. Risk factors for S. aureus acquisition revealed in the survey were: men who... (Complete abstract click electronic access below)

Orientador: Maria de Lourdes Ribeiro de Souza da Cunha / Coorientador: Carlos Magno Castelo Branco Fortaleza / Banca: Paulo Câmara Marques Pereira / Banca: Patrícia Yoshida Faccioli Martins / Resumo: As Infecções estafilocócicas são grande causa de mortalidade e morbidade. A emergência de isolados de Staphylococcus aureus resistente à meticilina (MRSA) originários da comunidade na década de 1990 trouxe desafios para sua prevenção e abordagem terapêutica. A identificação da prevalência e características de MRSA adquirido em comunidade (CA-MRSA) em populações de alto risco pode fornecer valiosas informações para compreensão de sua epidemiologia. O objetivo deste estudo foi identificar a prevalência, os fatores de risco e as características moleculares de CA-MRSA isolado em nasofaringe de uma população carcerária em Avaré, Brasil. Foram coletados swabs nasais de 302 homens encarcerados no Centro de Ressocialização de Avaré. A caracterização da resistência à meticilina foi feita através de difusão em disco (Oxacilina e Cefoxitina) e identificação do gene mecA (por Reação em Cadeia de Polimerase (PCR). Utilizou-se também PCR para identificação dos genes lukF-PVe lukS-PV que codificam a leucocidina de Panton-Valentine. Fatores de risco para colonização nasal por S. aureus e por MRSA foram avaliados em modelos univariados e multivariados (regressão logística). A prevalência de colonização nasal por S. aureus como um todo foi de 16,5%, e a MRSA (definida como positividade para o gene mecA), de (02) 4,0%. Os testes de difusão com disco de Oxacilina e Cefoxitina identificaram 2 (4%) resistentes simultaneamente a Oxacilina e Cefoxitina. Nenhum dos isolados de S. aureus apresentou genes codificadores da PVL. As amostras positivas para gene mecA apresentaram SCCmec tipo IV, confirmando esta tipagem presente em amostras comunitárias.Uma das amostras positivas para gene mecA apresentou SCCmec IV-d, que pelo método PFGE agrupou em 85,7% com o clone JCSC 4469 de origem japonesa. Os fatores de risco para... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Staphylococci infections are a major cause of morbidity and mortality. The emergence of methicillin-resistant Staphylococcus aureus (MRSA) isolates original from the community in the 1990s brought challenges for prevention and therapeutic approach.Identifying the prevalence and characteristics of community-acquired MRSA (CA-MRSA) in high-risk populations can provide valuable information for understanding its epidemiology. The objective of this study was to identify the prevalence, risk factors and molecular characteristics of CA-MRSA isolates in nasopharynx of a prison population in Avare, Brazil. Nasal swabs were collected from 302 men incarcerated in Central Resocialization of Avaré. The characterization of methicillin resistance was done by disk diffusion (Oxacillin and Cefoxitin) and mecA gene identification (by Polymerase Chain Reaction(PCR). We also used PCR to identify the lukF-PV and lukFS-PV genes that encode the Panton- Valentine Leukocidin. Risk factors for nasal colonization by S. aureus and MRSA were evaluated with univariate and multivariate models (logistic regression). The prevalence of nasal colonization by S. aureus as a whole was 16.5%, and MRSA (defined as positive for the mecA gene), from (02) 4.0%. The diffusion testing with Oxacillin and Cefoxitin identified 2 (4%) isolates resistants to both Oxacillin and Cefoxitin. None of the isolated S.aureus had genes encoding PVL. Positive samples for mecA showed SCCmec type IV, confirming this typing in community samples. One of the positive samples for mecA showed SCCmec IV-d, which by PFGE method grouped in 85.7% with clone 4469 JCSC of Japanese origin. Risk factors for S. aureus acquisition revealed in the survey were: men who... (Complete abstract click electronic access below) / Mestre