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Nanašanje zdravilnih učinkovin na biokompatibilne podlage s tehnologijo 2D tiskanja predstavlja inovativen in učinkovit način prilagajanja odmerka zdravilnih učinkovin vsakemu posamezniku. S takim načinom zdravljenja bi lahko uspeli rešiti izzive, ki se pojavljajo zlasti pri zdravljenju kroničnih bolezni, pri zdravljenju otrok in starostnikov ter pri visokopotentnih učinkovinah in/ali učinkovinah z ozkim terapevtskim oknom. Pomembna prednost tehnologije 2D tiskanja se kaže tudi v možnosti priprave farmacevtske oblike neposredno pred izdajo bolniku z dispergiranjem suhih delcev v osnovnem mediju črnila in njihovim tiskanjem na biokompatibilne podlage. Zato je bil cilj našega raziskovalnega dela pripraviti suhe delce s simvastatinom z metodo sušenja z razprševanjem, ki bi bili primerni za dispergiranje v mediju črnil in bi ostali fizikalno-kemijsko dovolj stabilni tekom celotnega obdobja uporabe. Izdelali smo tri različne emulzije z laktozo, kalcijevim silikatom in kombinacijo laktoze in nanoceluloze kot antiadhezivi. Delce smo pripravili z metodo sušenja z razprševanjem omenjenih emulzij, pri čemer je prišlo do pretvorbe simvastatina v amorfno obliko, kar smo potrdili z diferenčno dinamično kalorimetrijo in z infrardečo spektroskopijo s Fourierjevo transformacijo. Pripravili smo delce brez in z dodatkom askorbinske kisline, da bi preverili vpliv antioksidanta na kemijsko stabilnost simvastatina v delcih. Delce smo dispergirali v treh medijih, ki so vsebovali različne deleže propilen glikola in vode. Opravili smo enomesečno stabilnostno študijo pri čemer smo suhe in dispergirane delce shranili pri 25 °C in 40 % RV ter pri 40 °C in 75 % RV. Vzorce smo analizirali s tekočinsko kromatografijo ultra visoke ločljivosti ter določili vsebnost simvastatina v delcih. Ugotovili smo, da dodatek askorbinske kisline pomembno vpliva na kemijsko stabilnost delcev, ki smo jih pripravili z uporabo laktoze ter s kombinacijo laktoze in nanoceluloze. Omenjeni delci po enem mesecu shranjevanja izkazujejo zadovoljivo kemijsko stabilnost. Svežim in dispergiranim delcem smo z lasersko difrakcijo izmerili velikost ter z vrstičnim elektronskim mikroskopom ovrednotili njihove morfološke lastnosti. Zanimal nas je vpliv lokacije vzorčenja, uporabljenega antiadheziva, medija ter temperature shranjevanja na velikost dispergiranih delcev, saj je le-ta ključna, da ne prihaja do neželenega mašenja šob glave tiskalnika. Potrdimo lahko, da ustrezno velikost in fizikalno-kemijsko stabilnost izkazujejo predvsem delci s kombinacijo laktoze in nanoceluloze, ki vsebujejo antioksidant in so dispergirani v najbolj viskoznem mediju. Applying active pharmaceutical ingredients to biocompatible substrates using 2D printing technology is an innovative and effective way to customise the dosage of active pharmaceutical ingredients to each individual. This treatment modality could address the challenges that arise in particular in the treatment of chronic diseases, in the treatment of children and the elderly, and in the treatment of high-potency active substances and/or active substances with a narrow therapeutic window. Another important advantage of 2D printing technology is the possibility to prepare the dosage form immediately prior to dispensing to the patient by dispersing the dry particles in the base ink medium and printing them on biocompatible substrates. Therefore, the aim of our research work was to prepare simvastatin dry particles by spray-drying which are suitable for dispersion in the ink media and remain sufficiently physicochemically stable over the entire shelf life. Three different emulsions were produced with lactose, calcium silicate and a combination of lactose and nanocellulose as anti-adhesives. The particles were prepared by spray-drying of the emulsions and the conversion of simvastatin to an amorphous form which was verified by differential scanning calorimetry and Fourier transform infrared spectroscopy. Particles were prepared without and with the addition of ascorbic acid in order to verify the effect of the antioxidant on the chemical stability of simvastatin in the particles. The particles were dispersed in three media containing different proportions of propylene glycol and water. A stability study was carried out for one month, where both dry and dispersed particles were stored at 25 °C and 40 % RH and at 40 °C and 75 % RH. The samples were analysed by ultra-high performance liquid chromatography and the particle content of simvastatin was determined. We found that the addition of ascorbic acid had a significant effect on the chemical stability of the particles prepared using lactose and a combination of lactose and nanocellulose. These particles show satisfactory chemical stability after 1 month of storage. The fresh and dispersed particles were measured for size by laser diffraction and their morphological properties were evaluated by scanning electron microscopy. We were interested in the influence of the sampling location, the used anti-adhesive, the medium and the storage temperature on the size of the dispersed particles, since it is crucial to avoid unwanted lubrication of the printer head nozzles. It can be confirmed that the particles with the combination of lactose and nanocellulose, which contain an antioxidant and are dispersed in the most viscous medium, show the appropriate size and physico-chemical stability.

Staphylococcus aureus adhesion to the host's skin and mucosae enables asymptomatic colonization and the establishment of infection. This process is facilitated by cell wall-anchored adhesins that bind to host ligands. Therapeutics targeting this process could provide significant clinical benefits; however, the development of anti-adhesives requires an in-depth knowledge of adhesion-associated factors and an assay amenable to high-throughput applications. Here, we describe the development of a sensitive and robust whole cell assay to enable the large-scale profiling of S. aureus adhesion to host ligands. To validate the assay, and to gain insight into cellular factors contributing to adhesion, we profiled a sequence-defined S. aureus transposon mutant library, identifying mutants with attenuated adhesion to human-derived fibronectin, keratin, and fibrinogen. Our screening approach was validated by the identification of known adhesion-related proteins, such as the housekeeping sortase responsible for covalently linking adhesins to the cell wall. In addition, we also identified genetic loci that could represent undescribed anti-adhesive targets. To compare and contrast the genetic requirements of adhesion to each host ligand, we generated a S. aureus Genetic Adhesion Network, which identified a core gene set involved in adhesion to all three host ligands, and unique genetic signatures. In summary, this assay will enable high-throughput chemical screens to identify anti-adhesives and our findings provide insight into the target space of such an approach.

peer-reviewed Streptococcus pneumoniae is a Gram-positive pathogen, which is regularly found in the upper respiratory tract of healthy individuals. Increased numbers of S. pneumoniae have been observed colonising the upper respiratory tract of children affected by respiratory tract infections. Galβ1-4GlcNAcβ1-3Gal has been previously identified as one of the receptors involved in the adherence and translocation of S. pneumoniae. As this structure is similar to the milk oligosaccharide lacto-N-neoTetraose, many studies have investigated if free milk oligosaccharides can inhibit the adhesion of S. pneumoniae to epithelial cells of the respiratory tract. Here, we demonstrate that bovine oligosaccharides, which were extracted from demineralised whey, using a combination of membrane filtration and chromatography, were capable of reducing S. pneumoniae adhesion to pharynx and lung cells in vitro when tested at physiological concentrations. This study strengthens the potential use of bovine derived milk oligosaccharides as functional ingredients to reduce the incidence of infectious diseases.

A comparison of the different conformations adopted by C-glycosylated mannosides in FimH crystal structures is reported together with the investigation of new interactions., Selective inhibitors of the type 1 fimbrial adhesin FimH are recognized as attractive alternatives for antibiotic therapies and prophylaxes against Escherichia coli infections such as urinary-tract infections. To construct these inhibitors, the α-d-mannopyranoside of high-mannose N-glycans, recognized with exclusive specificity on glycoprotein receptors by FimH, forms the basal structure. A hydrophobic aglycon is then linked to the mannose by the O1 oxygen inherently present in the α-anomeric configuration. Substitution of this O atom by a carbon introduces a C-glycosidic bond, which may enhance the therapeutic potential of such compounds owing to the inability of enzymes to degrade C-glycosidic bonds. Here, the first crystal structures of the E. coli FimH adhesin in complex with C-glycosidically linked mannopyranosides are presented. These findings explain the role of the spacer in positioning biphenyl ligands for interactions by means of aromatic stacking in the tyrosine gate of FimH and how the normally hydrated C-glycosidic link is tolerated. As these new compounds can bind FimH, it can be assumed that they have the potential to serve as potent new antagonists of FimH, paving the way for the design of a new family of anti-adhesive compounds against urinary-tract infections.

Summary Biosurfactants comprise a wide group of surface active compounds that are produced by microorganisms. Based on their unique features, these compounds find applications in many industrial fields including the food tech industry. These features and their usefulness for particular food applications will be discussed. Biosurfactants can be used as emulsifiers, texturizers, wetting and cleaning agents, antimicrobials, and anti-adhesives. Moreover, these compounds can be used to prepare microemulsion systems or to aid the production of nanoparticles. Such systems may be designed to encapsulate food odors, colorants, functional ingredients, and micronutrients, which could lead to the development of innovative food products or new functional characteristics of existing ones. Although biosurfactants are very promising, still few examples of their industrial application can be highlighted, mainly due to the scalability of the production processes and to the regulation. These issues, as well as the food tech industry opportunities regarding potential applications of the biosurfactants, will be presented in this chapter., The authors acknowledge the Portuguese Foundation for Science and Technology (FCT) for the financial support under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684), and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 – Programa Operacional Regional do Norte. The authors also thank the FCT for the financial support under the scope of the Project MultiBiorefinery – multi-purpose strategies for broadband agro-forest and fisheries byproducts valorisation: a step forward for a truly integrated biorefinery (POCI-01-0145-FEDER-016403). E. J. Gudiña was supported by the Post-Doctoral grant CEB-BPD/01/2015/07 from the project UID/BIO/04469/2013, funded by FCT., info:eu-repo/semantics/publishedVersion

Background: Escherichia coli strains adhere to the normally sterile human uroepithelium using type 1 pili, that are long, hairy surface organelles exposing a mannose-binding FimH adhesin at the tip. A small percentage of adhered bacteria can successfully invade bladder cells, presumably via pathways mediated by the high-mannosylated uroplakin-Ia and a3b1 integrins found throughout the uroepithelium. Invaded bacteria replicate and mature into dense, biofilm-like inclusions in preparation of fluxing and of infection of neighbouring cells, being the major cause of the troublesome recurrent urinary tract infections. Methodology/Principal Findings: We demonstrate that a-D-mannose based inhibitors of FimH not only block bacterial adhesion on uroepithelial cells but also antagonize invasion and biofilm formation. Heptyl a-D-mannose prevents binding of type 1-piliated E. coli to the human bladder cell line 5637 and reduces both adhesion and invasion of the UTI89 cystitis isolate instilled in mouse bladder via catheterization. Heptyl a-D-mannose also specifically inhibited biofilm formation at micromolar concentrations. The structural basis of the great inhibitory potential of alkyl and aryl a-D-mannosides was elucidated in the crystal structure of the FimH receptor-binding domain in complex with oligomannose-3. FimH interacts with Mana1,3Manb1,4GlcNAcb1,4GlcNAc in an extended binding site. The interactions along the a1,3 glycosidic bond and the first b1,4 linkage to the chitobiose unit are conserved with those of FimH with butyl a-D-mannose. The strong stacking of the central mannose with the aromatic ring of Tyr48 is congruent with the high affinity found for synthetic inhibitors in which this mannose is substituted for by an aromatic group. Conclusions/Significance: The potential of ligand-based design of antagonists of urinary tract infections is ruled by the structural mimicry of natural epitopes and extends into blocking of bacterial invasion, intracellular growth and capacity to fluxing and of recurrence of the infection.