The goal of this research was to develop a novel biosensor for detecting and eliminating pathogenic E. coli. Traditionally, identifying pathogenic E. coli and distinguishing it from harmless environmental strains includes serotyping and DNA sequencing, which can take days or weeks. Our biosensor platform makes use of a material that is part of the immune system from single- multi- cellular organisms that target viruses, fungi, and bacteria called antimicrobial peptides (AMPs). Using the quartz crystal microbalance with dissipation monitoring (QCM-D), we characterized non-specific binding between CP1 to silicon nitride and gold, and covalent binding of cysteine-terminated CP1 (CP1-cys) to gold. QCM-D monitors frequency and dissipative changes resulting from adsorbed mass, and peptide film thickness and density can be calculated using Voigt Viscoelastic modeling. Viability of the E. coli was monitored using a live/dead kit consisting of nucleic acid stains Syto 9 and Propidium Iodide. Successfully immobilizing peptide to a substrate is particularly important if CP1 would be applied on a food processing surface. By immobilizing CP1 to silicon nitride, we measured the binding forces between bacteria and peptides with the atomic force microscope (AFM), and explored important bacterial features such as LPS composition and length that influence binding affinity with CP1. The structure of the LPS is comprised of 3 sections: lipid A, core group, and O-antigen. We are mostly interested in the initial binding between AMP and LPS since our goal is to develop a novel biosensor that can detect pathogenic bacteria within seconds of exposure. Considering the short exposure period, the AMP would only be exposed to the O-antigen and outer core groups, which are repeating sugar chains that are essential for bacterial pathogenicity and adhesion to substrates. Although geared for use as a novel biosensor, results of this study can also be applied to the use of AMPs for replacing or enhancing the activity of antibiotics. Our work suggests that CP1 may not be serotype-specific, but targets the O-antigen before interfering with phospholipid groups of the bacterial membrane. Other factors that assist in pathogenicity, such as LPS length, may also be important for the consideration of CP1 potency.

Orientador: Marcelo Coelho Goiato / Coorientador: Daniela Micheline dos Santos / Banca: Aldiéris Alves Pesqueira / Banca: Marcelle Danelon / Resumo: Durante a reabilitação com próteses fixas, o uso de provisórios é importante para que se alcance o sucesso clínico e a longevidade da prótese definitiva, mantendo a saúde e protegendo os tecidos dentais e periodontais. O objetivo deste estudo foi avaliar a estabilidade de cor, microdureza, rugosidade e energia livre de superfície de resinas de restaurações provisórias, com e sem a aplicação de glaze fotopolimerizável, e avaliar a eficácia antibiofilme do digluconato de clorexidina a 0,12% sobre as mesmas. Para isso, foram confeccionadas amostras de resina de Bis-Acril (Protemp 4, 3M ESPE, São Paulo, Brasil) e de resina acrílica ativada quimicamente (RAAQ) (Dencor, Artigos Odontológicos Clássico LTDA, São Paulo, Brasil), de forma que, apenas metade dessas amostras foi submetida ao polimento químico com glaze fotopolimerizável. Essas amostras foram posicionadas em dispositivos palatinos contendo 4 nichos (1 RAAQ sem glaze, 1 RAAQ com glaze, 1 Bis-Acril sem glaze, 1 Bis-Acril com glaze), utilizados por voluntários (n=20). As fases experimentais foram: Os voluntários usaram o dispositivo contendo as amostras do grupo controle, durante sete dias, sendo realizada aplicação de sacarose 30% sobre as amostras. Posteriormente, usaram o dispositivo contendo novas amostras do grupo teste, durante mais sete dias, sendo realizado tratamento com digluconato de clorexidina a 0,12% e aplicação de sacarose 30%. A seguir, determinou-se a alteração de cor, microdureza, rugosidade e energia livre... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: During rehabilitation with ceramic fragments, it is important to use provisionalrestorations, which are essential to achieve clinical success and longevity of the definitive prosthesis, maintaining the health and protecting dental and periodontal tissues. The aimof this study was to evaluate colorstability, microhardness, roughness, and surface free energy of provisional prosthesissamples, with and without the application of light-curing glaze on the surface, and to evaluate the antibiofilm efficacy of 0.12% chlorhexidine digluconate in these samples.For this, resin samples of Bis-Acryl(Protemp, 3M ESPE, Sao Paulo, Brazil) and chemically-activated acrylic resin (CAAR)(Dencor, Artigos Odontológicos Clássico LTDA, Sao Paulo, Brazil)were made. Half of these samples were submitted to chemical polishing with light-curing glaze. These samples were placed in intraoral devices, whichcontained 4 niches(1 CAARwithout glaze, 1 CAARwith glaze, 1 Bis-Acrylwithout glaze, 1 Bis-Acrylwith glaze), used by volunteers (n=20). The experimental pases were: Thevolunteersinitially used the device containing the samples from the control group for seven days, in which 30% sucrose were applied. Then, theyused the device containing the samples from the test group for anotherseven days, in which 0.12% chlorhexidine digluconate and 30% sucrose wereapplied. Next, the tests of color change, microhardness, roughness and surface energy, initially and after theexperimentais phases, of all samples, and the bio... (Complete abstract click electronic access below) / Mestre

In this study photocatalytic antibacterial and self-cleaning activities of TiO2-SiO2 thin films as a function of TiO2/SiO2 ratios were investigated. TiO2-SiO2 mixed oxides were synthesized by sol-gel method and coated over soda-lime glass plates by dip coating technique. Escherichia coli was used as a model microorganism for the photocatalytic antibacterial tests. Degradation rate of methylene blue (MB) molecules was used to characterize photocatalytic self-cleaning activities of thin film surfaces. The maximum antibacterial activity was achieved over 92 wt% SiO2 containing thin films. However, when the SiO2 content exceeds 92 wt%, photocatalytic antibacterial activity decreased considerably, which was explained by the dilution of TiO2 phase and inaccessibility of TiO2. Increase in photocatalytic antibacterial activity was attributed to increases in the relative surface area, roughness, hydroxyl (OH-) groups and bacterial adhesion. The favored bacterial adhesion enhanced direct contact of bacteria with TiO2 particles and surface reactive oxygen species. The highest initial decomposition rate of MB was obtained for 60 wt% SiO2 and the activity decreases as SiO2 concentration increases. The increase in photocatalytic activity by the SiO2 addition can be explained by the increase of the amount of MB per unit area of TiO2-SiO2 thin films. Different adsorption capability of thin films against MB molecule and E. coli cell was explained as the first reason why the antibacterial and self-cleaning activities reached their maximum values at different SiO2 ratios. The second reason could be related with the different control mechanisms of self-cleaning and antibacterial activities by different textural and surface properties.

Ph.D.

Nosocomial, or hospital acquired, infections, are ubiquitous within the modern clinical setting leading to over $5 billion annually of related healthcare costs in North America. All indwelling devices are highly susceptible to bacterial colonization where physico-chemical interactions between bacteria and biomaterial surfaces have been implicated as determinant factors in the fate of the initial adhesion processes. It has been proposed that by exploiting interference strategies within this critical step of infection the ability to create 'non-infective' biomaterials may be developed. / This thesis demonstrates the effectivity of North American cranberry (Vaccinium macrocarpon) derived proanthocyanidins in preventing the adhesion of pathogenic bacteria to biomaterial surfaces. Specifically, using a model of catheter associated urinary tract infection, significant reductions in initial adhesion of uropathogenic Escherichia coli and Enterococcus faecalis to PVC and PTFE were observed. With the application of colloidal theory, a mechanism of steric interference was determined as responsible for these effects. / The evidence presented implicates PAC as a molecule of interest for the development of novel biomaterials with increased resistance to bacteria colonization.

Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2008. / Title from first page of PDF file (viewed June 6, 2008). Includes bibliographical references.

Thesis (M.S.)--University of Michigan, 1995. / Includes bibliographical references.

Na natureza, os microorganismos podem apresentar forma de vida planctônica ou podem estar aderidos a superfícies formando comunidades conhecidas como biofilmes. A formação de biofilmes na indústria alimentícia é uma constante preocupação visto que os microorganismos aderidos podem causar contaminações persistentes, levando a deterioração do alimento e a transmissão de doenças. Uma alternativa para evitar a adesão bacteriana e a formação de biofilmes é o pré-condicionamento de superfícies com biosurfatantes, que são compostos tensoativos de origem microbiana capazes de alterar as propriedades físico-químicas e conseqüentemente modificar as interações entre a bactéria e a superfície. Os biosurfatantes, surfactina obtida de Bacillus subtilis e ramnolipídeo de Pseudomonas aeruginosa, foram testados quanto a capacidade de evitar a adesão e remover biofilmes de bactérias patogênicas de interesse alimentar. Foram avaliadas culturas individuais e mistas de Staphylococcus aureus, Listeria monocytogenes, e Salmonella Enteritidis utilizando-se como modelo superfícies de poliestireno. O pré-condicionamento da superfície com surfactina na concentração de 0,25% reduziu a adesão de Salmonella Enteritidis e Listeria monocytogenes em 42%, enquanto que o tratamento com ramnolipídeo a 1% reduziu a adesão de Listeria monocytogenes e Staphylococcus aureus ao poliestireno em 57,8% e 67,8% respectivamente. O condicionamento com os biosurfatantes não se mostrou eficiente na redução da adesão das culturas mistas das bactérias se comparado aos resultados obtidos para as culturas individuais. O poliestireno condicionado com os biosurfatantes apresentou redução na hidrofobicidade devido ao caráter aniônico destas moléculas. A repulsão eletrostática e a redução das interações hidrofóbicas promovidas pelo condicionamento do poliestireno com ramnolipídeo foram fatores determinantes na atividade antiadesiva observada para L. monocytogenes e S. aureus, entretanto os resultados obtidos para a superfície tratada com surfactina sugerem que outros parâmetros influenciaram nos resultados observados. Após 2 h de contato a surfactina na concentração de 0,1% promoveu a remoção de 63,7% do biofilme de S. aureus, 95,9% do biofilme de L. monocytogenes, 35,5% do biofilme de S. Enteritidis e 58,5% do biofilme da cultura mista das três bactérias. Já o ramnolipídeo na concentração de 0,25% removeu 58,5% do biofilme de S. aureus, 26,5% do biofilme de L. monocytogenes, 23,0 % do biofilme de S. Enteritidis e 24% do biofilme da cultura mista após 2 h contato. De modo geral, o aumento do tempo de contato e da concentração dos biosurfatantes reduziu a remoção dos biofilmes. A surfactina e o ramnolipídeo demonstraram potencial para uso como agentes anti-adesivos assim como para a remoção de biofilmes de bactérias patogênicas de importância alimentar. / In nature, microrganisms can live as planktonic cells or can be found living in communities attached in surfaces forming biofilms. Biofilm represents a great concern for food industry, since it can be a source of persistent contamination that can lead to food spoilage and the transmission of diseases. To avoid the adhesion of bacteria and the formation of biofilms, an alternative is the pre-conditioning of surfaces using biosurfactants that are microbial compounds that can modify the physico-chemical properties of the surfaces changing bacterial interactions and consequently adhesion. The biosurfactants, surfactin obtained from Bacillus subtilis and rhamnolipids from Pseudomonas aeruginosa, were evaluated as agents to avoid the adhesion and to disrupt biofilms of food-borne pathogenic bacteria. Individual cultures and mixed cultures of Staphylococcus aureus, Listeria monocytogenes e Salmonella Enteritidis were studied using polystyrene as the model surface. The pre-conditioning with surfactin 0,25% reduces in 42,0% the adhesion of L. monocytogenes and S. Enteritidis, whereas the treatment using rhamnolipids 1,0% reduced in 57,8% the adhesion of L. monocytogenes and in 67,8% the adhesion of S. aureus to polystyrene. The conditioning of surface with biosurfactants was less effective to avoid adhesion of mixed cultures of the bacteria when compared with the results obtained for individual cultures. The polystyrene surface conditioned with the biosurfactants showed a reduction in the hydrophobicity due to the anionic character of the molecules. The electrostatic repulsion and the reduction on hydrophobic interactions promoted by the conditioning of surface with rhamnolipids were determinant factors to explain the anti-adhesive activity observed for L. monocytogenes and S. aureus, however, the data obtained with surfactin suggest that other parameters have influenced the results observed. After 2 h contact with surfactin at 0,1% concentration, the pre-formed biofilms of S. aureus were reduced by 63,7%, L. monocytogenes biofilms were reduce by 95,9% , S. Enteritidis biofilms by 35,5% and the mixed culture biofilm by 58,5%. The rhamnolipids at 0,25% concentration removed 58,5% of biofilm of S. aureus, 26,5% of the biofilm of L. monocytogenes, 23,0% the biofilm of S. Enteritidis and 24,0% the biofilm of the mixed culture after 2 h of contact. In general, the increase in concentration of biosurfactants and in the time of contact decreases the biofilm remove percentage. These results demonstrate that surfactin and rhamnolipids present potential to be used as agents to control the attachment and to disrupt biofilms of food-borne pathogens.