The relevance of bacterial interactions on oral health

Dissertação de mestrado integrado em Engenharia Biomédica (área de especialização em Engenharia Clínica)

Acetylated bacterial cellulose coated with urinary bladder matrix as a substrate for retinal pigment epithelium

This work evaluated the effect of acetylated bacterial cellulose (ABC) substrates coated with urinary bladder matrix (UBM) on the behavior of Retinal Pigment Epithelium (RPE), as assessed by cell adhesion, proliferation and development of cell polarity exhibiting transepithelial resistance and polygonal shaped-cells with microvilli. Acetylation of bacterial cellulose (BC) generated a moderate hydrophobic surface (around 65°) while the adsorption of UBM onto these acetylated substrates did not affect significantly the surface hydrophobicity. The ABS substrates coated with UBM enabled the development of a cell phenotype closer to that of native RPE cells. These cells were able to express proteins essential for their cytoskeletal organization and metabolic function (ZO-1 and RPE65), while showing a polygonal shaped morphology with microvilli and a monolayer configuration. The coated ABC substrates were also characterized, exhibiting low swelling effect (between 1.52.0 swelling/mm3), high mechanical strength (2048 MPa) and non-pyrogenicity (2.12 EU/L). Therefore, the ABC substrates coated with UBM exhibit interesting features as potential cell carriers in RPE transplantation that ought to be further explored.

High-level of viral genomic diversity in cervical cancers: a Brazilian study on human papillomavirus type 16

Invasive cervical cancer (ICC) is the third most frequent cancer among women worldwide and is associated with persistent infection by carcinogenic human papillomaviruses (HPVs). The combination of large populations of viral progeny and decades of sustained infection may allow for the generation of intra-patient diversity, in spite of the assumedly low mutation rates of PVs. While the natural history of chronic HPVs infections has been comprehensively described, within-host viral diversity remains largely unexplored. In this study we have applied next generation sequencing to the analysis of intra-host genetic diversity in ten ICC and one condyloma cases associated to single HPV16 infection. We retrieved from all cases near full-length genomic sequences. All samples analyzed contained polymorphic sites, ranging from 3 to 125 polymorphic positions per genome, and the median probability of a viral genome picked at random to be identical to the consensus sequence in the lesion was only 40%. We have also identified two independent putative duplication events in two samples, spanning the L2 and the L1 gene, respectively. Finally, we have identified with good support a chimera of human and viral DNA. We propose that viral diversity generated during HPVs chronic infection may be fueled by innate and adaptive immune pressures. Further research will be needed to understand the dynamics of viral DNA variability, differentially in benign and malignant lesions, as well as in tissues with differential intensity of immune surveillance. Finally, the impact of intralesion viral diversity on the long-term oncogenic potential may deserve closer attention.

Hydrocarbonoclastic bacteria: from bioremediation to bioenergy feedstock

Tese de Doutoramento em Engenharia Química e Biológica.

Diversity of non-clinical Acinetobacter species in a sub-saharan Africa region: evidence of carbapenem-hydrolysing class D-β-lactamase producers

[Excerpt] Although Acinetobacter baumannii has been the main agent for healthcare infections, recent reports suggest that some Acinetobacter environmental species should be considered as a potential cause of disease. In Angola, there are no previous data on its environmental reservoirs and resistance features. We aimed to unveil the occurrence and diversity of Acinetobacter species and the presence of resistance mechanisms in different non-clinical settings in Angola.

A spin-offs journey into achieving marketable products from bacterial cellulose

[Excerpt] Academic spin-offs, technological ventures born inside Universities, have increasingly strengthen the connections between the scholarship and the economy, by fostering the role of technology transfer and knowledge commercialization. This presentation will outline the major steps in taking an idea or a technology to market, growing the venture and aiming at securing a successful exit. Also, it will present BCTechnologies (Bacterial Cellulose Technologies), a spin-off from the University of Minho (Portugal). (...)

Structural and enzymatic characterization of ABgp46, a novel phage endolysin with broad anti-Gram-negative bacterial activity

The present study demonstrates the antibacterial potential of a phage endolysin against Gram-negative pathogens, particularly against multidrug resistant strains of Acinetobacter baumannii. We have cloned, heterologously expressed and characterized a novel endolysin (ABgp46) from Acinetobacter phage vb_AbaP_CEB1 and tested its antibacterial activity against several multidrug-resistant A. baumannii strains. LC-MS revealed that ABgp46 is an N-acetylmuramidase, that is also active over a broad pH range (4.0-10.0) and temperatures up to 50°C. Interestingly, ABgp46 has intrinsic and specific anti-A. baumannii activity, reducing multidrug resistant strains by up to 2 logs within 2 hours. By combining ABgp46 with several organic acids that act as outer membrane permeabilizing agents, it is possible to increase and broaden antibacterial activity to include other Gram-negative bacterial pathogens. In the presence of citric and malic acid, ABgp46 reduces A. baumannii below the detection limit (> 5 log) and more than 4 logs P. aeruginosa and Salmonella Typhimurium strains. Overall, this globular endolysin exhibits a broad and high activity against Gram-negative pathogens, that can be enhanced in presence of citric and malic acid, and be used in human and veterinary medicine.

Microbial respiration with chlorine oxyanions: diversity and physiological and biochemical properties of chlorate- and perchlorate-reducing microorganisms

Chlorine oxyanions are valuable electron acceptors for microorganisms. Recent findings have shed light on the natural formation of chlorine oxyanions in the environment. These suggest a permanent introduction of respective compounds on Earth, long before their anthropogenic manufacture. Microorganisms that are able to grow by the reduction of chlorate and perchlorate are affiliated with phylogenetically diverse lineages, spanning from the Proteobacteria to the Firmicutes and archaeal microorganisms. Microbial reduction of chlorine oxyanions can be found in diverse environments and different environmental conditions (temperature, salinities, pH). It commonly involves the enzymes perchlorate reductase (Pcr) or chlorate reductase (Clr) and chlorite dismutase (Cld). Horizontal gene transfer seems to play an important role for the acquisition of functional genes. Novel and efficient Clds were isolated from microorganisms incapable of growing on chlorine oxyanions. Archaea seem to use a periplasmic Nar-type reductase (pNar) for perchlorate reduction and lack a functional Cld. Chlorite is possibly eliminated by alternative (abiotic) reactions. This was already demonstrated for Archaeoglobus fulgidus, which uses reduced sulfur compounds to detoxify chlorite. A broad biochemical diversity of the trait, its environmental dispersal, and the occurrence of relevant enzymes in diverse lineages may indicate early adaptations of life toward chlorine oxyanions on Earth.

Chlamydia pneumoniae and atherosclerosis. Identification of bacterial DNA in the arterial wall

OBJECTIVE: The intracellular Gram-negative bacterium Chlamydia pneumoniae has been associated with atherosclerosis. The presence of Chlamydia pneumoniae has been investigated in fragments of the arterial wall with a technique for DNA identification. METHODS: Arterial fragments obtained from vascular surgical procedures in 58 patients were analyzed. From these patients, 39 were males and the mean age was 65±6 years. The polymerase chain reaction was used to identify the bacterial DNA with a pair of primers that codify the major outer membrane protein (MOMP) of Chlamydia pneumoniae. The amplified product was visualized by electrophoresis in the 2% agarose gel stained with ethidium bromide, and it was considered positive when migrating in the band of molecular weight of the positive controls. RESULTS: Seven (12%) out of the 58 patients showed positive results for Chlamydia pneumoniae. CONCLUSION: DNA from Chlamydia pneumoniae was identified in the arterial wall of a substantial number of patients with atherosclerosis. This association, which has already been described in other countries, corroborates the evidence favoring a role played by Chlamydia pneumoniae in atherogenesis.

Antibiofilm strategies in the food industry

Biofilms in food processing plants represent not only a problem to human health but also cause economic losses by technical failure in several systems. In fact, many foodborne outbreaks have been found to be associated with biofilms. Biofilms may be prevented by regular cleaning and disinfection, but this does not completely prevent biofilm formation. Besides, due to their diversity and to the development of specialized phenotypes, it is well known that biofilms are more resistant to cleaning and disinfection than planktonic microorganisms. In recent years, a considerable effort has been made in the prevention of microbial adhesion and biofilm formation on food processing surfaces and novel technologies have been introduced. In this context, this chapter discusses the main conventional and emergent strategies that have been employed to prevent bacterial adhesion to food processing surfaces and thus to efficiently maintain good hygiene throughout the food industries.

Bacterial nanocellulose as a structured platform for conductive biopolymers

[Exert] Since the discovery that polyacetylene could be doped to the metallic state more than 3 decades ago, an ever-growing body of a multidisciplinary approach to material design, synthesis, and system integration has been evidenced. The present chapter will primarily review the emerging field of intrinsically conducting polymer and conductive polymer blends, with polyaniline and polypyrrole as the major representatives of conducting polymers. This survey will also address some of the potential areas for applications of such conductive polymer blends. Also, current results concerning the chemical polymerization of conducting polymers on bacterial nanocellulose (BNC) will be presented, including brief remarks on the rationale for the use of conductive BNC blends. This will be followed by a discussion on their properties and potential applications (...).

Bacterial cellulose surface modification

The unique properties of bacterial nanocellulose (BNC) provide the basis for a wide range of applications in human and veterinary medicine, odontology, pharmaceuticals, acoustic and filter membranes, biotechnological devices, and in the food and paper industry. In this chapter, an overview of surface modifications of bacterial cellulose is presented. Depending on the envisaged applications, chemical modifications, incorporation of bioactive molecules, modification of the porosity, crystallinity, and biodegradability may be obtained, further enlarging the potential of BNC.

All different or all the same? Exploring the diversity of professional practices in Portuguese school psychology

"Published online: 29 March 2016"

Laccase immobilization on bacterial nanocellulose membranes: antimicrobial, kinetic and stability properties

This work studied the physical immobilization of a commercial laccase on bacterial nanocellulose (BNC) aiming to identify the laccase antibacterial properties suitable for wound dressings. Physico-chemical analysis demonstrates that the BNC structure is manly formed by pure crystalline I cellulose. The pH optimum and activation energy of free laccase depends on the substrate employed corresponding to pH 6, 7, 3 and 57, 22, 48 kJ mol1 for 2,6-dimethylphenol (DMP), catechol and 2,2 -azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), respectively. The Michaelis-Menten constant (Km) value for the immobilized laccase (0.77 mM) was found to be almost double of that of the free enzyme (0.42 mM). However, the specific activities of immobilized and free laccase are similar suggesting that the cage-like structure of BNC allows entrapped laccase to maintain some flexibility and favour substrate accessibility. The results clearly show the antimicrobial effect of laccase in Gram-positive (92%) and Gram-negative (26%) bacteria and cytotoxicity acceptable for wound dressing applications.

Bacterial cellulose-lactoferrin as an antimicrobial edible packaging

Bacterial cellulose (BC) films from two distinct sources (obtained by static culture with Gluconacetobacter xylinus ATCC 53582 (BC1) and from a commercial source (BC2)) were modified by bovine lactoferrin (bLF) adsorption. The functionalized films (BC+bLF) were assessed as edible antimicrobial packaging, for use in direct contact with highly perishable foods, specifically fresh sausage as a model of meat products. BC+bLF films and sausage casings were characterized regarding their water vapour permeability (WVP), mechanical properties, and bactericidal efficiency against two food pathogens, Escherichia coli and Staphylococcus aureus. Considering their edibility, an in vitro gastrointestinal tract model was used to study the changes occurring in the BC films during passage through the gastrointestinal tract. Moreover, the cytotoxicity of the BC films against 3T3 mouse embryo fibroblasts was evaluated. BC1 and BC2 showed equivalent density, WVP and maximum tensile strength. The percentage of bactericidal efficiency of BC1 and BC2 with adsorbed bLF (BC1+bLF and BC2+bLF, respectively) in the standalone films and in inoculated fresh sausages, was similar against E. coli (mean reduction 69 % in the films per se versus 94 % in the sausages) and S. aureus (mean reduction 97 % in the films per se versus 36 % in the case sausages). Moreover, the BC1+bLF and BC2+bLF films significantly hindered the specific growth rate of both bacteria. Finally, no relevant cytotoxicity against 3T3 fibroblasts was found for the films before and after the simulated digestion. BC films with adsorbed bLF may constitute an approach in the development of bio-based edible antimicrobial packaging systems.