http://hdl.handle.net/10362/3394 Wed, 22 May 2013 03:45:58 GMT 2013-05-22T03:45:58Z http://run.unl.pt:80/retrieve/57623/bio-chem.jpg http://hdl.handle.net/10362/3394 http://hdl.handle.net/10362/8593 Title: Epidemiological studies of Streptococcus pneumoniae carriage in the post-vaccination era among two risk groups: children and the elderly Authors: Nunes, Sónia Abstract: Streptococcus pneumoniae is a global cause of disease including pneumonia, otitis media, conjunctivitis, sepsis, and bacterial meningitis. These infections are not essential to the transmission or long-term survival of the bacterium; indeed, S. pneumoniae depends on asymptomatic colonization of the human nasopharynx for its dissemination to additional hosts. Considering this, colonization studies are a good way to monitor changes in the pneumococcal epidemiology that may result from the use of antibiotics and vaccines. The molecular characterization of pneumococci is crucial to assess these changes which highlight the need for the development and validation of easier and faster methods of molecular typing. Since 1996 our group has been monitoring the pneumococcal population colonizing children attending day care centers. However, for several years these studies have been confined to the Lisbon area. In this PhD we have addressed this situation by including other regions of Portugal in our study. In addition, we have started to study pneumococcal colonization in the elderly, the other age group where the incidence of pneumococcal infections is high. This thesis summarizes five studies conducted during this PhD. The first four studies were focused on the pneumococcal epidemiology among the two age groups where the rates of pneumococcal disease are highest: children up to six years old and adults older than 60 years. The fifth and last study describes the evaluation and validation of a new genotyping strategy for pneumococci.(...) Description: Dissertation presented to obtain the Ph.D. degree in Biology/ Molecular Biology Thu, 01 Nov 2012 00:00:00 GMT http://hdl.handle.net/10362/8593 2012-11-01T00:00:00Z http://hdl.handle.net/10362/8592 Title: Insights into cell wall synthesis and cell division in Staphylococcus aureus Authors: Jorge, Ana Maria Abstract: Staphylococcus aureus is a gram-positive bacterial pathogen that besides persistently colonizing healthy individuals, is responsible for a large number of hospital-associated bacterial infections. The extraordinary capacity of S. aureus to acquire resistance to antibiotics led to the emergence of highly resistant strains, mainly methicillin-resistant S. aureus (MRSA) strains, that are a major cause of soft skin and tissue infections and bacteremia. In one third of European countries, including Portugal, more than 25% of S. aureus infections are caused by MRSA strains. The capacity of MRSA strains to resist β-lactam antibiotics (such as penicillin) is mainly due to the acquisition of an extra-species penicillin-binding protein (PBP), PBP2A. PBPs are bacterial enzymes involved in the synthesis of the cell wall polymer peptidoglycan. Besides PBP2A, which is present only in MRSA strains, S. aureus has 4 native PBPs (PBP1-4), which catalyze the polymerization (transglycosylation) and the cross-linking (transpeptidation) of glycan chains, forming a strong yet flexible structure that protects the cell from the high internal osmotic pressure. Peptidoglycan is unique to the bacterial kingdom and its biosynthesis is the target of a vast number of clinically important antibiotics such as β-lactams and glycopeptides. β-lactam antibiotics target the transpeptidase domain of PBPs, halting peptidoglycan synthesis and eventually leading to cell lysis. However, in MRSA strains the existence of PBP2A, which has a low affinity for β-lactams, enables cell wall synthesis to continue even in the presence of these antibiotics. Under these conditions, the transpeptidase domain of PBP2A functionally cooperates with the transglycosylase domain of the unique bifunctional PBP, PBP2, to ensure continued cell wall synthesis and cell survival.(...) Description: Dissertation presented to obtain the Ph.D degree in Biology Sun, 01 Apr 2012 00:00:00 GMT http://hdl.handle.net/10362/8592 2012-04-01T00:00:00Z http://hdl.handle.net/10362/8574 Title: The role of Rac1-modulated gene transcription in tumorigenesis Authors: Barros, Patrícia Abstract: Gene expression regulation is a dynamic and multi-step process, in which transcription plays a major role. Transcription initiation depends on binding of transcription regulators to DNA elements located in promoter or enhancer regions, a process often controlled by signalling pathways. One such pathway is regulated by Rac1, a member of the Rho family of small GTPases involved in cell proliferation, adhesion and migration. In this work, novel links between Rac1 signalling and transcriptional regulation in colorectal tumour cells are described. First, it is shown that Rac1 activation leads to PAK1-mediated phosphorylation of the transcriptional repressor BCL-6 in colorectal cancer cells, inactivating its repressor function. In the presence of active Rac1, BCL-6 redistribution within the nucleus, a reduction in its affinity to chromatin and increased expression of the endogenous target genes NFKB1 and CD44, and of a BCL-6-controlled luciferase reporter construct were observed. Next, it was found that Rac1 signalling promotes gene transcription by inducing a transcriptional switch from the repressor BCL-6 to the activator STAT5A at the promoter of certain target genes. Using chromatin immunoprecipitation, it is demonstrated in different colorectal cell lines that active Rac1 promotes release of BCL-6 with concomitant nuclear translocation and binding of STAT5A at the same promoter site. Three endogenous cell-cycle-related genes (CCND2, CDKN2B, SUMO1) were identified to be inversely regulated by BCL-6 and STAT5A and shown to respond to Rac1 signalling with promoter occupancy switches that correlate directly with changes in their expression levels.(...) Description: Dissertation presented to obtain the Ph.D degree in Biology Mon, 01 Oct 2012 00:00:00 GMT http://hdl.handle.net/10362/8574 2012-10-01T00:00:00Z http://hdl.handle.net/10362/8573 Title: Understanding the relationship between central metabolism and virulence in the human pathogen Streptococcus pneumoniae Authors: Carvalho, Sandra M. Abstract: Streptococcus pneumoniae is a normal inhabitant of the human nasopharynx, but it is better known for its role in a plethora of human diseases. Growing emergence of antibiotic-resistant streptococci and non-type vaccine strains increases the urgency of finding new targets for the development of novel therapeutic and preventive drugs. As a major concern for global public health, S. pneumoniae has always attracted great attention from the scientific community, which has translated into knowledge on pathogenesis and virulence and the development of a considerable “toolbox” for genetic manipulation and genomic analysis, as well as a large number of deciphered genome sequences. Interestingly, genome-wide studies have consistently pinpointed genes involved in carbohydrate uptake and metabolism as essential for the virulence of S. pneumoniae. These global studies offered the opportunity to investigate in greater depth the potential connections between basic physiology, and in particular central metabolism, and pneumococcal virulence and pathogenesis. The general goal of this thesis is to achieve a deeper understanding of the molecular mechanisms underlying sugar metabolism and their relation to virulence factors in S. pneumoniae, with a special focus on capsule production. In the present work, glucose (Glc) and galactose (Gal) were used as carbon sources for the study of pneumococcal sugar metabolism. This choice was made for two reasons: Firstly, Glc is a common preferred sugar and is also found as a major carbon source in niches potentially occupied by S. pneumoniae during host inflammation or hyperglycaemia. Secondly, Gal, generally a slowly metabolized non-preferred sugar, is a major carbohydrate in the human nasopharynx, the non-pathological colonization niche of S. pneumoniae. Description: Dissertation presented to obtain the Ph.D. degree in Biochemistry Sat, 01 Sep 2012 00:00:00 GMT http://hdl.handle.net/10362/8573 2012-09-01T00:00:00Z