Enterococcus and biocides: mechanisms of tolerance and selection for vancomycin resistance

Dissertation presented to obtain a Doctoral degree in Biology, Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa.

Cancer theranostics: multifunctional gold nanoparticles for diagnostics and therapy

Doctorate in Biology, Specialty in Biotechnology

Gold nanoparticles for plasmid delivery

Gene therapy presents an ideal strategy for the treatment of genetic as well as acquired diseases, such as cancer and typically involves the insertion of a functioning gene into cells to correct a cellular dysfunction or to provide a new cellular function. Gene delivery vectors are based in two models: viral and non-viral. Viral vectors have high transfection efficiency but their major barrier is immunogenicity. Since the non-viral vectors have no immunogenicity, these have been widely studied. Gold nanoparticles have been proposed as optimal delivery systems of genetic material, due their small size, high surface-to-volume ratio and the ability to be functionalized with multiple molecules. In the present work, an AuNP-based formulation was developed to deliver a plasmid in a colorectal cancer cell line, containing as reporter gene the gene encoding to EGFP. The delivery system resulted from the functionalization of 14 nm AuNP with a PEG layer (4300114 PEG chains/AuNP), which increases stability and biocompatibility of AuNPs; quaternary ammonium groups which provide positive charges that allow electrostatic binding of plasmid, which is considered the therapeutic agent to be transported into cells. The system developed was characterized by UV-vis spectroscopy, DLS, TEM and by electrophoretic mobility, yielding a formulation with 113.5 nm.Transfection efficiency of the formulation developed was evaluated through PCR and through EGFP expression by fluorescence microscopy and fluorescence spectroscopy. The internalization was observed 3h post transfection; however a low level of EGFP expression was achieved. After 24h of incubation, EGFP expression increases just 3 times compared to non-transfected cells. The commercial system (Lipofectamine) expressed EGFP 5 times more than the system developed AuNP@PEG@R4N+@pEGFP. This difference could be related to lower translocation to the nucleus.

Quaternary elephants in Portugal: new data

In this paper, a synopsis on the already known data, and the study of further specimens either new (*) or hither to undescribed are presented. The specimens dealt with have been collected at the Condeixa travertines; in the Tagus terraces near Carregado(*) and at Foz do Enxarrique(*); in the mousterian, about 31000 BP infilling from the Figueira Brava cave(*); and at Algar de Joao Ramos(*). Some specimens are ascribed to Elephas antiquus, the best represented form. Material from Figueira Brava cave and Algar de João Ramos is ascribed to Mammuthus primigenius, a species recognized here for the first time in Portugal.

Ostracodes; witnesses of the Plio-Quaternary boundary in the North Eastern Atlantic preliminary results

Proceedings of the 1" R.C.A.N.S. Congress, Lisboa, October 1992

Pliocene-quaternary upwelling in the SoutheasternAtlantic may reflect changes in water mass production

Proceedings of tile 1" R.C.A.N.S. Congress, Lisboa, October 1992

Paleoambiental reconstruction based on mineralogical data of quaternary littoral deposits occuring at Cortegaça and Maceda beaches (Ovar, Portugal)

The main goal of the present work is the use of mineralogical data corresponding to sediment fine fractions (silt and clay) of Quaternary littoral deposits for the definition of a more detailed vertical zonography and to discriminate the most significant morphoclimatic changes concerned with sediment source areas and sediment deposition areas. The analysis of the available mineralogical data reveals a vertical evolution of the mineral composition. The following aspects deserve particular reference: 1) fine fractions (<38 nm) are composed of quartz and phyllosilicates associated to feldspars, prevailing over other minerals; however in certain sections iron hydroxides and evaporitic minerals occur in significant amounts; 2) clay fractions (<2 nm) show a general prevalence of illite associated with kaolinite and oscillations, in relative terms, of kaolinite and illite contents. Qualitative and quantitative lateral and vertical variations of clay and non clay minerals allow the discrimination of sedimentary sequences and the establishment of the ritmicity and periodicity of the morphoclimatic Quaternary episodes that occurred in the Cortegaça and Maceda beaches. To each one of the sedimentary sequences corresponds, in a first stage, a littoral environment that increasingly became more continental. Climate would be mild to cold, sometimes with humidity - aridity oscillations. Warmer and moister episodes alternated with cooler and dryer ones.

Late Quaternary warm marine mollusks from Santa Maria (Azores); paleoecologic and paleobiogeographic considerations

The sub-fossil fauna from the Late Quaternary marine deposits of Santa Maria is made of more than 50 species of gastropods and bivalves, 19 of them collected recently and for the first time in the northern coast of the island (Lagoinhas Bay). The sub-fossil shells are found in deposits of beach sands, situated 2-3 meters above the present low tide. The carbonated sands from the basal part of the succession yield an autochthonous association of borers dominated by the bivalve Myoforceps aristata (Dillwin, 1817). Upwards, the marine sands contain concentrations of beach drift shells, including well-preserved supratidal and intertidal gastropods, among them a large number of Rissoidae. The bivalve fauna is dominated by disarticulated valves of Ervilia castanea (Montagu, 1803), a small infaunal coloniser of mobile sandy substrates. The composition of the fauna is made essentially of West European species, many of them common to the West Coast of Portugal. However, a few "warm guests" with West African or Caribbean affinities were also found, suggesting a close relation with some of the "Tyrrhenian" warm associations found in the Western Mediterranean.

Rediscovery of Leptaxis vetusta (Morelet & Drouet, 1857), a subfossil land snail from the Quaternary of Santa Maria (Azores)

During a recent field work on the southern coast of the island of Santa Maria (Azores) a bulk sample of 37 shells and 25 fragments of Leptaxis vetusta was assembled from Late Pleistocene and Holocene slope deposits outcropping in the area. These specimens are the first of this rare subfossil species to be mentioned since the original descriptions of Arthur Morelet and Henri Drouet (1857). The purposes of our paper are a systematic and biometric description of L. vestuta. For the first time, the original type: locality was localized with accuracy over the southern downslopes of Pico do Facho, between Figueiral and Prainha. The subfossil specimens were collected in slope deposits and detritic fans, overlying a fossiliferous marine deposit situated over the 2-3 m abrasion platform of Praia and Prainha bay. The age and factors associated to the extinction of this species are discussed, including the destruction of the original laurel cover and the colonization by Otala lactea (Muller, 1774), a continental helicid introduced and widespread in Santa Maria.

The removal of xenobiotic compounds from wastewater through the use of biological processes and advanced oxidation technologies

Dissertação apresentada para obtenção do Grau de Doutor em Engenharia Química e Bioquímica pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

The inhibitory activity of synthetic compounds and ions against transporters of multi-drug resistant bacteria

RESUMO: Sessenta e três derivados de hidantoína foram utilizados para avaliar possíveis efeitos de modulação na actividade das bombas de efluxo (BE) na Salmonella NCTC 13349 utilizando um método fluorimétrico semi-automático. Nenhum dos compostos apresentaram actividade anti-bacteriana até concentrações de 240 mg/L. Entre todos os compostos, SZ-7 demonstrou possuir propriedades de modulação de effluxo na presença de glucose. Para testar esta actividade, estirpes de Salmonella resistentes à ciprofloxacina, induzidas a elevados níveis de resistência com sobre-expressão de BE, foram expostas ao SZ-7. Este derivado afectou a susceptibilidade das estirpes à ciprofloxacina. Uma vez que os 63 compostos estudados apresentaram pouco efeito inibitório /cumulativo, apesar de serem conhecidos pelos seus efeitos moduladores de BE-dependentes de iões em eucariotas, foi questionado o papel dos iões na regulação de BE bacterianas, que poderão influenciar a eficácia de novos compostos. Para este estudo, utilizamos a Escherichia coli AG100 como modelo, devido ao extenso conhecimento no que respeita a estrutura e actividade das BE. Devido à importância de iões de cálcio (Ca2+) nos canais de transporte membranar e na actividade de ATPases, a sua actividade na modulação do efluxo foi investigada. De resultados anteriormente obtidos concluiu-se que a pH 5 o efluxo é independente de energia metabólica; contudo, a pH 8 é absolutamente dependente, sendo que o Ca2+ é indispensável para manter a actividade das ATPases bacterianas. A acumulação/effluxo de EtBr pela E. coli AG100 foi determinada na presença/ausência de Ca2+, clorpromazina (inibidor de ligação de Ca2+ a proteínas), e ácido etilenodiamino tetra-acético (quelante de Ca2+). Acumulação/effluxo aumentou a pH 8, contudo o Ca2+ reverte estes efeitos evidenciando a sua importância no funcionamento das BE bacterianas. Em resumo este trabalho colocou em evidência que muitos aspectos bioquímicos e bioenergéticos devem ser tomados em consideração no estudo da resistência bacteriana mediada por BE.------- ABSTRACT: Sixty-three hydantoin derivatives were evaluated for their modulating effects on efflux pump (EP) activity of Salmonella NCTC 13349 utilizing a semi-automatic fluorometric method. None of the compounds presented antibacterial activities at concentrations as high as 240 mg/L. Among all compounds, SZ-7 showed possible efflux modulating activity in the presence of glucose, indicative of a potential EP inhibitor. To verify its potential effects, ciprofloxacin-resistant Salmonella strains, induced to high level resistance with over-expressing EPs, were exposed to SZ-7. This derivative affected the susceptibility of the ciprofloxacin-resistant strains. Since the 63 compounds studied had very low inhibitory/accumulative effects, even though their known for being efficient in modulating ion-driven eukaryotic EPs, we questioned whether ions had a leading role in regulating bacterial EPs, influencing the effectiveness of new compounds. For this study we used Escherichia coli AG100 as a model, due to the extensive knowledge on its EPs structure and activity. Owing the importance of calcium ions (Ca2+) for membrane transport channels and activity of ATPases, the role of Ca2+ was investigated. From previous results we concluded that at pH 5 efflux is independent of metabolic energy; however, at pH 8 it is entirely dependent of metabolic energy and the Ca2+ ions are essential to maintain the activity of bacterial ATPases. Accumulation and efflux of ethidium bromide (EtBr) by E. coli AG100 was determined in the presence and absence of Ca2+, chlorpromazine (inhibitor of Ca2+-binding to proteins), and ethylenediaminetetraacetic acid (Ca2+ chelator). Accumulation of EtBr increased at pH 8; however Ca2+ reversed these effects providing information as to the importance of this ion in the regulation of bacterial EP systems. Overall this work puts in evidence that many biochemical and bioenergetic aspects related to the strains physiology need to be taken into consideration in bacterial drug resistance mediated by EPs.

Sarcoplasmic reticulum calcium ATPase is inhibited by organic vanadium coordination compounds: pyridine-2,6-dicarboxylatodioxovanadium(V), BMOV, and an amavadine analogue

Inorg Chem. 2008 Jul 7;47(13):5677-84. doi: 10.1021/ic702405d

Study on solubility of pharmaceutical compounds in ionic liquids

Dissertation to obtain the degree of master in Chemical and Biochemical Engineering

Study of membrane emulsification process as a pre-step for the microencapsulation of lipid compounds by spray drying

Dissertation presented to Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa for obtaining the master degree in Membrane Engineering

Quantification and characterization of the phenolic compounds of kenaf leaves

Dissertation for the Master Degree in Technology and Food Security