Paralab (Portugal)

Biofilm formation is usually associated with antimicrobial treatment failure and the development of chronic infections. Therefore, it is important to develop new strategies to address wounds treatment. Nanocarriers can be engineered with precision, tailored to target specific bacterial strains, or disrupt biofilm architecture. This work aimed to evaluate the potential of quercetin (Q) and/or omega-3 fatty acids (ω 3 )-loaded in hybrid nanoassemblies composed by nanostructured lipid carriers (NLCs) and nanofibers as an innovative strategy to prevent biofilm formation on wounds. NLCs were homogeneous in terms of size and charge, with mean hydrodynamic diameter less than 200 nm and a negative charge. The antibiofilm activity of Q and ω 3 , both free and loaded on NLCs, on S. aureus and E. coli biofilms, was analysed. NLCs have an inhibitory effect, especially those containing ω 3 . The encapsulated compounds also demonstrated increased antibiofilm activity compared to free compounds. Q and/or ω 3 , free or loaded in NLCs, showed no cytotoxicity on cell-cultured fibroblasts. After 7 days, NLCs, especially those containing ω 3 , had a positive effect on cell proliferation, potentially aiding in tissue regeneration. NLC-enriched nanofibers with a porous structure allow for exchange of exudates, gases, and nutrients, while their random orientation mimics the skin's extracellular matrix, aiding cell adhesion and proliferation. The nanofibers present good mechanical strength and an increased surface area that enhances diffusion, enabling rapid and complete Q release, which is beneficial for anti-biofilm activity. Q and ω 3 -loaded NLCs enriched nanofibers may be a promising antibiofilm treatment strategy, allowing for proper delivery of these agents with antimicrobial activity while preventing biofilm formation on wounds. • Q and/or ꞷ 3 -loaded NLCs significantly inhibit S. aureus and E. coli biofilms • Q and/or ꞷ 3 -loaded NLCs promote fibroblast proliferation • NLCs-enriched nanofibers showed adequate mechanical/physical properties • The obtained rapid and complete Q release is critical for anti-biofilm activity.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with a very poor prognosis. Its treatment is hindered by a lack of new therapeutic alternatives and the existence of the blood–brain barrier (BBB), which restricts the access of drugs commonly used in ALS, such as riluzole, to the brain. To overcome these limitations and increase brain targeting, riluzole-loaded nanostructured lipid carriers (NLC) were prepared and functionalized with lactoferrin (Lf), facilitating transport across the BBB by interacting with Lf receptors expressed in the brain endothelium. NLC were characterized with respect to their physicochemical properties (size, zeta potential, polydispersity index) as well as their stability, encapsulation efficiency, morphology, in vitro release profile, and biocompatibility. Moreover, crystallinity and melting behavior were assessed by DSC and PXRD. Nanoparticles exhibited initial mean diameters between 180 and 220 nm and a polydispersity index below 0.3, indicating a narrow size distribution. NLC remained stable over at least 3 months. Riluzole encapsulation efficiency was very high, around 94–98%. FTIR and protein quantification studies confirmed the conjugation of Lf on the surface of the nanocarriers, with TEM images showing that the functionalized NLC presented a smooth surface and uniform spherical shape. An MTT assay revealed that the nanocarriers developed in this study did not cause a substantial reduction in the viability of NSC-34 and hCMEC/D3 cells at a riluzole concentration up to 10 μM, being therefore biocompatible. The results suggest that Lf-functionalized NLC are a suitable and promising delivery system to target riluzole to the brain.

Pickering emulsions are systems composed of two immiscible fluids stabilized by solid organic or inorganic particles. Pickering emulsions are particularly useful in cosmetics, where the surfactants are unwanted, as well as in the pharmaceutical field, where transdermal and/or dermal drug delivery is difficult to achieve and controlled drug release is desired. Here, we studied calcium carbonate particles as stabilizers of Pickering emulsions for topical use. An optimized formulation was obtained using a Quality by Design approach. First, a screening experiment was performed to identify the formulation and process critical variables that affect the quality properties of the Pickering emulsion. The optimization of the production was then studied by establishing the design space. The final formulation was hereinafter investigated regarding the pH, rheological properties, and in vitro cytotoxicity assays. The results showed the formulation had a pH compatible with human skin and a shear thinning behavior. Moreover, this formulation showed a strong network structure, with a suitable spreadability on the skin, allowing an easy application. The in vitro assays were performed to assess the potential cytotoxicity of the calcium carbonate-stabilized emulsion and the particles themselves, and the results revealed that the formulation did not significantly affect the cell viability. In conclusion, the use of calcium carbonate particles as a stabilizer ingredient contributed to achieve an eco-friendly Pickering emulsion.

Portugal is a low incidence country for tuberculosis (TB) disease. Now figuring among TB low incidence countries, it has since the 1990s reported multidrug resistant and extensively drug resistant (XDR) TB cases, driven predominantly by two strain-types: Lisboa3 and Q1. This study describes the largest characterization of the evolutionary trajectory of M/XDR-TB strains in Portugal, spanning a time-period of two decades. By combining whole-genome sequencing and phenotypic susceptibility data for 207 isolates, we report the geospatial patterns of drug resistant TB, particularly the dispersion of Lisboa3 and Q1 clades, which underly 64.2% and 94.0% of all MDR-TB and XDR-TB isolates, respectively. Genomic-based similarity and a phylogenetic analysis revealed multiple clusters (n = 16) reflecting ongoing and uncontrolled recent transmission of M/XDR-TB, predominantly associated with the Lisboa3 and Q1 clades. These clades are now thought to be evolving in a polycentric mode across multiple geographical districts. The inferred evolutionary history is compatible with MDR- and XDR-TB originating in Portugal in the 70’s and 80’s, respectively, but with subsequent multiple emergence events of MDR and XDR-TB particularly involving the Lisboa3 clade. A SNP barcode was defined for Lisboa3 and Q1 and comparison with a phylogeny of global strain-types (n = 28 385) revealed the presence of Lisboa3 and Q1 strains in Europe, South America and Africa. In summary, Portugal displays an unusual and unique epidemiological setting shaped by >40 years of uncontrolled circulation of two main phylogenetic clades, leading to a sympatric evolutionary trajectory towards XDR-TB with the potential for global reach.

The nutritional quality of daphnids diet can influence their growth, reproduction and survival. In aquatic ecosystems, bacteria can contribute significantly to Daphnia diet by supporting, for instances, their high needs for phosphorus. The laboratory feeding of the model organisms Daphnia spp. is algal based, but should be improved to allow their better performance. The aim of this study was to evaluate the potential of two planctomycetes, Gemmata obscuriglobus and Rhodopirellula rubra, from exponential and stationary growth phases as alternative or supplementary food source for Daphnia magna. The actinobacterium Arthrobacter sp. was used for comparison. The feeding with only bacteria showed the inefficacy of both planctomycetes and actinobacteria as the only food source. However, when used in supplement to Raphidocelis subcapitata, a decrease in the age of first reproduction, a significant increase in reproductive output, in somatic growth and in rate of population increase was found for the highest cell densities of bacteria tested. The typical pink coloration of these bacteria present in daphnids body and eggs confirmed bacterial absorption and metabolization of their pigment. Planctomycetes yielded better results than the actinobacteria Arthrobacter but G. obscuriglobus that possesses sterols did not induce a better performance comparatively to R. rubra. No relation could be established between the feeding treatments that allowed improvement of Daphnia performance and the different kind of Daphnia' Fatty Acid Methyl Esters. The use of sonication to separate planctomycetal cells before feeding the daphnids proved to be efficient. We confirmed that R. subcapitata supplemented by bacteria allows a better growth performance of D. magna.

Grapevine-associated microbiota influences wine organoleptic properties. Spoilage due to undesired microorganisms and biogenic amines (BAs) presence are two main constrains that must be seriously considered. In wine, BAs can originate from the grape berries or can be produced during fermentation, ageing or storage. This work aimed to understand if the high BAs levels observed in musts can have its origin in the microbial community present on grapes. The following methodologies were done: bacterial and fungal grapes communities' isolation, BAs quantification in grapes and musts and molecular amplification of the genes related to BAs production. For comparative purposes, microbial communities from grapes and musts from Douro (low BAs levels in musts) and Alentejo (high BAs levels in musts) were used. Higher number and diversity of bacteria were observed in Alentejo grapes comparatively to Douro ones. Filamentous fungi were predominant when compared with yeasts and the diversity was higher in Alentejo. BAs levels mainly due to putrescin were about ten times higher in grapes and musts from Alentejo. As bacteria isolated from Alentejo grapes showed a great BAs-production potential, namely putrescin, our results suggest a bacterial grape origin for the high putrescine levels found in fresh musts of this region.