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The present study examined the biosynthesis and characterization of selenium nanoparticles (SeNPs) using two contrasting endophytic selenobacteria, one Gram-positive (Bacillus sp. E5 identified as Bacillus paranthracis) and one Gram-negative (Enterobacter sp. EC5.2 identified as Enterobacter ludwigi), for further use as biofortifying agents and/or for other biotechnological purposes. We demonstrated that, upon regulating culture conditions and selenite exposure time, both strains were suitable “cell factories” for producing SeNPs (B-SeNPs from B. paranthracis and E-SeNPs from E. ludwigii) with different properties. Briefly, dynamic light scattering (DLS), transmission electron microscopy (TEM), and atomic force microscopy (AFM) studies revealed that intracellular E-SeNPs (56.23 ± 4.85 nm) were smaller in diameter than B-SeNPs (83.44 ± 2.90 nm) and that both formulations were located in the surrounding medium or bound to the cell wall. AFM images indicated the absence of relevant variations in bacterial volume and shape and revealed the existence of layers of peptidoglycan surrounding the bacterial cell wall under the conditions of biosynthesis, particularly in the case of B. paranthracis. Raman spectroscopy, Fourier transform infrared spectroscopy (FTIR), energy-dispersive X-ray (EDS), X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS) showed that SeNPs were surrounded by the proteins, lipids, and polysaccharides of bacterial cells and that the numbers of the functional groups present in B-SeNPs were higher than in E-SeNPs. Thus, considering that these findings support the suitability of these two endophytic stains as potential biocatalysts to produce high-quality Se-based nanoparticles, our future efforts must be focused on the evaluation of their bioactivity, as well as on the determination of how the different features of each SeNP modulate their biological action and their stability.

Background To evaluate the clinical and histopathological effects of natural extracts in the treatment of oral ulcers induced in animal experimental models. Material and methods We carried out a search in the Medline, Scopus, WoS and Embase databases from the start of the databases to December 2020, and also made a manual search of the references. The search and selection were carried out by two researchers independently. The inclusion criteria were: experimental studies in animal models, in english, which complied with the study object. Results A total of 705 articles were identified. After selection by title, abstract and full text, 19 articles were finally included. Natural extracts of Jasminum grandiflorum, Ficus deltoidea, curcumin and Bixina orellana provoked a significantly greater reduction in the size of the ulcer. Extracts of Salvatora persica, Musa acuminate, Ganoderma lucidum mycelia and Bixina Orellana, as well as preparations of Kouyanqing Granule and curcumin, were able to reduce levels of pro-inflammatory cytokines and increase the expression and serum levels of growth factors and anti-inflammatory cytokines. Extracts of Piper sarmentosus, Cannabis sativa and Bletilla striata provoked a reduction in the severity of the histological inflammation. No significant differences were observed compared to controls in the treatments with extracts of Cannabis sativa, Aloe barbadensus Miller and Malva sylvestris in reducing the area of the oral ulcers. Conclusions Most of the natural extracts described in this review presented a positive clinical and histological effect on the cicatrisation of oral ulcers induced in animal models. Key words:Recurrent aphthous stomatitis, oral ulcer, plants, herbs, extracts, medicine, treatment.

Despite decades of therapeutic application of aminoglycosides, it is still a matter of debate if porins contribute to the translocation of the antibiotics across the bacterial outer membrane. Here, we quantified the uptake of kanamycin across the major porin channels OmpF and OmpC present in the outer membrane of

Despite decades of therapeutic application of aminoglycosides, it is still a matter of debate if porins contribute to the translocation of the antibiotics across the bacterial outer membrane. Here, we quantified the uptake of kanamycin across the major porin channels OmpF and OmpC present in the outer membrane of E. coli. Our analysis revealed that, despite its relatively large size, about 10 - 20 kanamycin molecules per second permeate through OmpF and OmpC under a 10 mM concentration gradient, whereas OmpN does not allow the passage. Molecular simulations elucidate the uptake mechanism of kanamycin through these porins. Whole-cell studies with a decisive set of E. coli porin mutants provide evidence that translocation of kanamycin via porins is relevant for antibiotic potency.

Despite decades of therapeutic application of aminoglycosides, it is still a matter of debate if porins contribute to the translocation of the antibiotics across the bacterial outer membrane. Here, we quantified the uptake of kanamycin across the major porin channels OmpF and OmpC present in the outer membrane of Escherichia coli. Our analysis revealed that, despite its relatively large size, about 10-20 kanamycin molecules per second permeate through OmpF and OmpC under a 10 ?M concentration gradient, whereas OmpN does not allow the passage. Molecular simulations elucidate the uptake mechanism of kanamycin through these porins. Whole-cell studies with a defined set of E. coli porin mutants provide evidence that translocation of kanamycin via porins is relevant for antibiotic potency. The values are discussed with respect to other antibiotics.

In the last decades, the encapsulation of antibiotics into nanoparticulate carriers has gained increasing attention for the treatment of infectious diseases. Sodium colistimethate-loaded solid lipid nanoparticles (Colist-SLNs) and nanostructured lipid carriers (Colist-NLCs) were designed aiming to treat the pulmonary infection associated to cystic fibrosis patients. The nanoparticles were freeze-dried using trehalose as cryoprotectant. The stability of both nanoparticles was analysed over one year according to the International Conference of Harmonisation (ICH) guidelines by determining the minimum inhibitory concentration (MIC) against clinically isolated Pseudomonas aeruginosa strains and by studying their physico-chemical characteristics. The results showed that Colist-SLNs lost their antimicrobial activity at the third month; on the contrary, the antibacterial activity of Colist-NLCs was maintained throughout the study within an adequate range (MIC ≤16 μg/mL). In addition, Colist-NLCs exhibited suitable physico-chemical properties at 5 °C and 25 °C/60% relative humidity over one year. Altogether, Colist-NLCs proved to have better stability than Colist-SLNs.

Among the pathogens that affect cystic fibrosis (CF) patients, Pseudomonas aeruginosa is the most prevalent. As a way to fight against this infection, nanotechnology has emerged over the last decades as a promising alternative to overcome resistance to antibiotics in infectious diseases. The goal of this work was to elaborate and characterize lipid nanoparticles for pulmonary delivery of tobramycin. Tobramycin-loaded nanostructured lipid carriers (Tb-NLCs) were prepared by hot melt homogenization technique. In addition, nanoparticles labeled with infrared dye (IR-NLCs) were used to investigate their in vivo performance after pulmonary administration. Tb-NLCs displayed a mean diameter size around 250 nm, high drug encapsulation (93%) and sustained release profile. Tb-NLCs showed to be active against clinically isolated P. aeruginosa. Moreover, Tb-NLCs did not decrease cell viability and were able to overcome an artificial mucus barrier in the presence of mucolytics agents. During the in vivo assay, IR-NLCs were administered to several mice by the intratracheal route using a Penn Century device. Next, the biodistribution of the nanoparticles was analyzed at different time points showing a wide nanosystem distribution in the lungs. Altogether, tobramycin-loaded NLCs seem to us an encouraging alternative to the currently available CF therapies.

Fosfomycin is a frequently prescribed drug in the treatment of acute urinary tract infections. It enters the bacterial cytoplasm and inhibits the biosynthesis of peptidoglycans by targeting the MurA enzyme. Despite extensive pharmacological studies and clinical use, the permeability of fosfomycin across the bacterial outer membrane is largely unexplored. Here, we investigate the fosfomycin permeability across the outer membrane of Gram-negative bacteria by electrophysiology experiments as well as by all-atom molecular dynamics simulations including free-energy and applied-field techniques. Notably, in an electrophysiological zero-current assay as well as in the molecular simulations, we found that fosfomycin can rapidly permeate the abundant Escherichia coli porin OmpF. Furthermore, two triple mutants in the constriction region of the porin have been investigated. The permeation rates through these mutants are slightly lower than that of the wild type but fosfomycin can still permeate. Altogether, this work unravels molecular details of fosfomycin permeation through the outer membrane porin OmpF of E. coli and moreover provides hints for understanding the translocation of phosphonic acid antibiotics through other outer membrane pores.

© 2014 Elsevier B.V. All rights reserved. Lung impairment is the most life-threatening factor for cystic fibrosis patients. Indeed, Pseudomonas aeruginosa is the main pathogen in the pulmonary infection of these patients. In this work, we developed sodium colistimethate loaded lipid nanoparticles, namely, solid lipid nanoparticles (SLN) and nanostructured lipid carriers (NLC), as a strategy to enhance the antimicrobial therapy against P. aeruginosa in cystic fibrosis patients. The nanoparticles obtained displayed a 200-400 nm size, high drug entrapment (79-94%) and a sustained drug release profile. Moreover, both SLN and NLC presented antimicrobial activity against clinically isolated P. aeruginosa. The integrity of the nanoparticles was not affected by nebulization through a mesh vibrating nebulizer. Moreover, lipid nanoparticles appeared to be less toxic than free sodium colistimethate in cell culture. Finally, an in vivo distribution experiment showed that nanoparticles spread homogenously through the lung and there was no migration of lipid nanoparticles to other organs, such as liver, spleen or kidneys., This work was carried out under the TERFIQEC Project, Cromprehensive Research On Effective Therapies For The Treatment Of Cystic Fibrosis And Associated Diseases; IPT-2011-1402-900000 was funded by the Ministry of Economy and Competitiveness. Technical and human support provided by SGIker (UPV/EHU, MICINN, GV/EJ, ERDF and ESF) is gratefully acknowledged. M. Moreno-Sastre thanks the University of the Basque Country for the ZabaldUz fellowship grant. The authors gratefully acknowledge the support of University of the Basque Country UPV/EHU (UFI11/32), of University of Barcelona (UB) as to the CSIC and FISIB

Cystic fibrosis (CF) is a genetic disorder in which frequent pulmonary infections develop secondarily. One of the major pulmonary pathogens colonizing the respiratory tract of CF patients and causing chronic airway infections is Pseudomonas aeruginosa. Although tobramycin was initially effective against P. aeruginosa, tobramycin-resistant strains have emerged. Among the strategies for overcoming resistance to tobramycin and other antibiotics is encapsulation of the drugs in nanoparticles. In this study, we explored the antimicrobial activity of nanoencapsulated tobramycin, both in solid lipid nanoparticles (SLN) and in nanostructured lipid carriers (NLC), against clinical isolates of P. aeruginosa obtained from CF patients. We also investigated the efficacy of these formulations in biofilm eradication. In both experiments, the activities of SLN and NLC were compared with that of free tobramycin. The susceptibility of planktonic bacteria was determined using the broth microdilution method and by plotting bacterial growth. The minimal biofilm eradication concentration (MBEC) was determined to assess the efficacy of the different tobramycin formulations against biofilms. The activity of tobramycin-loaded SLN was less than that of either tobramycin-loaded NLC or free tobramycin. The minimum inhibitory concentration (MIC) and MBEC of nanoencapsulated tobramycin were slightly lower (1-2 logs) than the corresponding values of the free drug when determined in tobramycin-susceptible isolates. However, in tobramycin-resistant strains, the MIC and MBEC did not differ between either encapsulated form and free tobramycin. Our results demonstrate the efficacy of nanoencapsulated formulations in killing susceptible P. aeruginosa from CF and from other patients. http://www.mdpi.com/2076-2607/5/3/35

Pseudomonas aeruginosa frequently infects the respiratory tract of cystic fibrosis (CF) patients. Multidrug-resistant phenotypes and high capacity to form stable biofilms are common. Recent studies have described the emergence of colistin-resistant isolates in CF patients treated with long-term inhaled colistin. The use of nanoparticles containing antimicrobials can contribute to overcome drug resistance mechanisms. The aim of this study was to explore antimicrobial activity of nanoencapsulated colistin (SLN-NLC) versus free colistin against P. aeruginosa clinical isolates from CF patients and to investigate their efficacy in biofilm eradication. Susceptibility of planktonic bacteria to antimicrobials was examined by using the broth microdilution method and growth curve assay. Minimal biofilm eradication concentration (MBEC) and biofilm prevention concentration (BPC) were determined to assess antimicrobial susceptibility of sessile bacteria. We used atomic force microscopy (AFM) to visualize treated and untreated biofilms and to determine surface roughness and other relevant parameters. Colistin nanoparticles had the same antimicrobial activity as free drug against planktonic bacteria. However, nanoencapsulated colistin was much more efficient in the eradication of biofilms than free colistin. Thus, these formulations have to be considered as a good alternative therapeutic option to treat P. aeruginosa infections.

P13 is one of the major outer membrane proteins of Borrelia burgdorferi. Previous studies described P13 as a porin. In the present study some structure and function aspects of P13 were studied. P13 showed according to lipid bilayer studies a channel-forming activity of 0.6 nanosiemens in 1 m KCl. Single channel and selectivity measurements demonstrated that P13 had no preference for either cations or anions and showed no voltage-gating up to ±100 mV. Blue native polyacrylamide gel electrophoresis was used to isolate and characterize the P13 protein complex in its native state. The complex had a high molecular mass of about 300 kDa and was only composed of P13 monomers. The channel size was investigated using non-electrolytes revealing an apparent diameter of about 1.4 nm with a 400-Da molecular mass cut-off. Multichannel titrations with different substrates reinforced the idea that P13 forms a general diffusion channel. The identity of P13 within the complex was confirmed by second dimension SDS-PAGE, Western blotting, mass spectrometry, and the use of a p13 deletion mutant strain. The results suggested that P13 is the protein responsible for the 0.6-nanosiemens pore-forming activity in the outer membrane of B. burgdorferi.