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Andrea Fulgione,1,* Flora Ianniello,1,* Marina Papaianni,1 Felice Contaldi,1 Tiziana Sgamma,2 Cinzia Giannini,3 Stella Pastore,3 Raffaele Velotta,4 Bartolomeo Della Ventura,5 Norberto Roveri,6 Marco Lelli,6 Federico Capuano,7 Rosanna Capparelli1 1Department of Agriculture, University of Naples “Federico II”, Portici, Naples, Italy; 2Biomolecular Technology Group, School of Allied Health Sciences, De Montfort University, Leicester, UK; 3Istituto di Cristallografia, Consiglio Nazionale delle Ricerche, Bari, Italy; 4Department of Physics “Ettore Pancini”, University of Naples “Federico II”, Portici, Naples, Italy; 5Department of Physics, Politecnico di Milano, Milano, Italy; 6Chemical Center Srl, Granarolo dell’Emilia, Bologna, Italy; 7Department of Food Microbiology, Istituto Zooprofilattico Sperimentale del Mezzogiorno, Portici, Naples, Italy *These authors contributed equally to this work Purpose: The use of bacteriophages represents a valid alternative to conventional antimicrobial treatments, overcoming the widespread bacterial antibiotic resistance phenomenon. In this work, we evaluated whether biomimetic hydroxyapatite (HA) nanocrystals are able to enhance some properties of bacteriophages. The final goal of this study was to demonstrate that biomimetic HA nanocrystals can be used for bacteriophage delivery in the context of bacterial infections, and contribute – at the same time – to enhance some of the biological properties of the same bacteriophages such as stability, preservation, antimicrobial activity, and so on. Materials and methods: Phage isolation and characterization were carried out by using Mitomycin C and following double-layer agar technique. The biomimetic HA water suspension was synthesized in order to obtain nanocrystals with plate-like morphology and nanometric dimensions. The interaction of phages with the HA was investigated by dynamic light scattering and Zeta potential analyses. The cytotoxicity and intracellular killing activities of the phage–HA complex were evaluated in human hepatocellular carcinoma HepG2 cells. The bacterial inhibition capacity of the complex was assessed on chicken minced meat samples infected with Salmonella Rissen. Results: Our data highlighted that the biomimetic HA nanocrystal–bacteriophage complex was more stable and more effective than phages alone in all tested experimental conditions. Conclusion: Our results evidenced the important contribution of biomimetic HA nanocrystals: they act as an excellent carrier for bacteriophage delivery and enhance its biological characteristics. This study confirmed the significant role of the mineral HA when it is complexed with biological entities like bacteriophages, as it has been shown for molecules such as lactoferrin. Keywords: biomimetic hydroxyapatite nanocrystals, Salmonella Rissen, bacteriophages, antibiotic resistance

Nunzia Nocerino,1 Andrea Fulgione,1 Marco Iannaccone,1 Laura Tomasetta,1 Flora Ianniello,1 Francesca Martora,1 Marco Lelli,2 Norberto Roveri,2 Federico Capuano,3 Rosanna Capparelli1 1Department of Agriculture Special Biotechnology Center Federico II, CeBIOTEC Biotechnology, University of Naples Federico II, Naples, 2Department of Chemistry, G Ciamician, Alma Mater Studiorum, University of Bologna, Bologna, 3Department of Food Inspection IZS ME, Naples, Italy Abstract: The emergence of bacterial strains resistant to antibiotics is a general public health problem. Progress in developing new molecules with antimicrobial properties has been made. In this study, we evaluated the biological activity of a hybrid nanocomposite composed of synthetic biomimetic hydroxyapatite surface-functionalized by lactoferrin (LF-HA). We evaluated the antimicrobial, anti-inflammatory, and antioxidant properties of LF-HA and found that the composite was active against both Gram-positive and Gram-negative bacteria, and that it modulated proinflammatory and anti-inflammatory responses and enhanced antioxidant properties as compared with LF alone. These results indicate the possibility of using LF-HA as an antimicrobial system and biomimetic hydroxyapatite as a candidate for innovative biomedical applications. Keywords: lactoferrin, hydroxyapatite nanocrystals, biomimetism, biological activity, drug delivery

Marco Lelli,1 Olivia Marchisio,2 Ismaela Foltran,1 Annamaria Genovesi,2 Giulia Montebugnoli,1 Massimo Marcaccio,1 Ugo Covani,2 Norberto Roveri11Alma Mater Studiorum, University of Bologna, Department of Chemistry, Bologna, Italy; 2University of Pisa, Istituto Stomatologico Tirreno, Lido di Camaiore, Lucca, ItalyAbstract: Titanium plates treated in vitro with a mouthwash containing amine fluoride (100 ppm F-) and another containing zinc-substituted carbonate–hydroxyapatite have been analyzed by scanning electron microscopy and atomic force microscopy to evaluate the modification of the surface roughness induced by treatment with these two different mouthwashes. The treatment with F--based mouthwash produces a roughness characterized by higher peaks and deeper valleys in the streaks on the titanium bracket surface compared with those observed in the reference polished titanium plates. This effect causes a mechanical weakness in the metallic dental implant causing bacterial growth and therefore promotes infection and prosthesis contamination. However, the in vitro treatment with a mouthwash containing zinc-substituted carbonate–hydroxyapatite reduced the surface roughness by filling the streaks with an apatitic phase. This treatment counteracts the surface oxidative process that can affect the mechanical behavior of the titanium dental implant, which inhibits the bacterial growth contaminating prostheses.Keywords: mouthwash, titanium brackets, corrosion, hydroxyapatite, aminic fluoride

The quadrupolar interaction experienced by the spin-1 14N nucleus is known to be extremely sensitive to local structure and dynamics. Furthermore, the 14N isotope is 99.6% naturally abundant, making it an attractive target for characterisation of nitrogen-rich biological molecules by solid-state NMR. In this study, dynamic nuclear polarization (DNP) is used in conjunction with indirect 14N detected solid-state NMR experiments to simultaneously characterise the quadrupolar interaction at multiple 14N sites in the backbone of the microcrystalline protein, GB3. Considerable variation in the quadrupolar interaction (＞700 kHz) is observed throughout the protein backbone. The distribution in quadrupolar interactions observed reports on the variation in local backbone conformation and subtle differences in hydrogen-bonding; demonstrating a new route to the structural and dynamic analysis of biomolecules. [ABSTRACT FROM AUTHOR]

A new unit for continuous monitoring of CO 2 flux diffused from soil was developed and preliminary tested in the field at Lipari Island (Sicily, Italy). The developed device (CO 2 -S-POT), based on the accumulation chamber method, is an assembled unit easy to install and to manage. Both dimensions (104 × 61 × 75 cm) and weight (32 Kg) of CO 2 -S-POT are lower than the equipment which are commonly used for continuous monitoring of diffused CO 2 flux. Besides CO 2 flux measurements, it is able to measure several environmental parameters [air temperature, soil temperature, atmospheric pressure, Volumetric Water Content (soil VWC%) and soil conductivity]. Laboratory tests showed a very good performance of the assembled system in terms of detection limit (0.1 mol m −2 ·day −1 ), reproducibility (RSD better than 3.5%) and linearity (up to 4 order of magnitude). Field tests (3 months) were performed at Lipari Island (Sicily, Italy), where no continuous monitoring of diffuse CO 2 flux using the accumulation chamber method is ongoing. During field tests, the CO 2 -S-POT performed flux measurements every hour, together with monitoring of the environmental parameters. During field tests, no ordinary or extraordinary maintenance was necessary and no missing data were detected. The soil CO 2 flux showed strong influence operated by air pressure, air temperature, soil temperature, soil conductivity, Volumetric Water Content and rain. Statistical methods based on Time Series Fourier analysis (TSFA), Principal Component Analysis (PCA) and also Multiple Linear Regression Analysis (MLRA) are used in order to filter the influence of the environmental parameters on the flux data, and, at the same time, to highlight the contribution which could be due from deep system. Raw flux data (in ppm·sec −1 ) are used in time series analysis, since these represent the real physical variable measured in the field, without transformations. In fact, the conversion from ppm·sec −1 to another dimension (i.e. mol·m −2 ·day −1 ) affects the correlation between the measured parameters (i.e. CO 2 flux and environmental parameters). The results of the statistical procedure showed that 1) the oscillation at low period (hours and days) are due to variations of environmental parameters and 2) the soil CO 2 flux measured at Lipari is characterized also by long period variations (compared to the previous ones, i.e. weeks), difficult to explain just in terms of environmental parameters. The applied methodology, based on multivariate statistical techniques, has allowed to identify residual anomalies, following a suitable procedure to handle the multicollinearity. In the period from August 20 to August 23, 2014, when there were not both significant air pressure variations and rain events, anomalous positive residuals were identified. These residual anomalies are those interesting from volcanic surveillance point of view. However, to assess whether the residual anomalies are effectively controlled by variations induced by the deep system (i.e. seismic and/or volcanic activity) others parameters, such as wind speed and wind direction, and more time of observation should be taken into consideration. [ABSTRACT FROM AUTHOR]

The order parameter, Szz, where z is the para axis of the difluoroterphenyl groups in DTC5C9, has been obtained from chemical shift anisotropies measured by 13C – {1H} NMR experiments at temperatures throughout the nematic, NU, and twist-bend nematic, NTB, phases shown by this compound. The order parameter temperature profiles are unusual in having a maximum value in the NU phase and then decreasing until the NTB phase is reached. There is a small discontinuity (∼2%) in Szz at TNNTB and then a gradual decrease until a new phase appears. This behaviour is interpreted as revealing a temperature-dependent tilting of local directors in both phases away from the applied magnetic field direction. In the enantiomorphic twist-bend phase this tilt is consistent with the structure of the phase as a helical arrangement of local directors, whilst in the high-temperature non-chiral nematic the tilt must involve a non-chiral arrangement. It is proposed that in both phases the tilting of directors has a common origin in the bent shape of the molecules. [ABSTRACT FROM AUTHOR]

The orientational order of the molecules in the bent mesogen CB6OCB has been studied throughout the range of temperature stability of both the NU and NTB liquid-crystal phases by 13C NMR spectroscopy. These spectra provide local order parameters for the para axes of both of the nonequivalent cyanobiphenyl groups and show how they change on entering the twist-bend nematic phase. A key feature of the order parameters is a weak, but clear maximum in the temperature variation of the order parameter prior to the NTB phase. This suggests that the directors in both the NU and NTB phases are tilted with respect to the magnetic field of the spectrometer. Significantly the conformational states of the spacer are comparable in both phases, although the low temperature nematic is chiral but not that at high temperature. It is proposed that the higher temperature, tilted phase could be the splay-bend nematic phase. [ABSTRACT FROM AUTHOR]

Abstract The Colli Albani volcanic complex (Lazio, Italy) hosts areas characterized by anomalously high emissions of CO 2 -rich gases (e.g. Tivoli, Cava dei Selci, Tor Caldara, Solforata). The source of these gases is a regional aquifer within the Mesozoic carbonate rock sequences. These degassing zones release significant concentrations of H 2 S and other toxic gases (e.g. GEM: Gaseous Elemental Mercury, and Rn) and represent a serious hazard for local inhabitants, especially for those living at Cava dei Selci (near Rome, Italy), where the emitting areas are nested inside residential neighborhoods. In April 2016, a comprehensive geochemical survey was carried out in an abandoned stone quarry nearby the urban settlement aimed to: (i) investigate the gas composition from both punctual discharges and anomalously high diffuse soil degassing sites, and (ii) evaluate their environmental impact on the local air quality. The spatial distribution of the soil CO 2 fluxes was mainly dependent on the local geostructural setting, whereas shallow secondary processes (e.g. oxidation and gas-water interaction) likely represent the main controlling factor on reactive and/or water-soluble gas species, such as CH 4 and H 2 S. The total output of CO 2 from the abandoned stone quarry accounted for 0.53% of total CO 2 discharged from the whole Colli Albani volcanic district. The naturally emitted toxic gases (e.g. CO 2 , H 2 S, CH 4 , GEM) largely affect the air quality and pose a serious threat for the health of the local residents. A mobile multi-instrumental station able to continuously and simultaneously acquire CO 2 , H 2 S, SO 2 , CH 4 , GEM and CO was deployed to verify the concentrations of both the main deep-originated gas compounds and potential secondary gaseous contaminants (i.e. SO 2) around and inside the urban settlement most exposed to the lethal gases. Hydrogen sulfide was found to be the most impacting gas, occasionally exceeding the 24-h air quality guideline for ambient air and causing odor annoyance at a distance up to more than 250 m downwind from the emitting area. In poorly ventilated basements, toxic gas accumulations up to hazardous levels were measured, producing anomalous outdoor air concentrations at the street level in front of the descending vehicular access to private garages and relatively far from the main emitting area. The geochemical survey, carried out via mobile station and soil gas measurements, resulted to be particularly efficient for evaluating the potential effects caused by gas emissions in inhabited areas. The multi-measurement approach adopted in the present study is of paramount importance for managing future urban development plans. Highlights • Natural gas emissions and air quality at Cava dei Selci were investigated. • Processes in the soil affected diffuse emissions of endogenous gases. • A mobile multi-instrumental station (MMS) was deployed for air quality measurement. • H 2 S was the most impacting gas in the residential area. • The MMS is a pivotal tool for managing future monitoring and intervention plans. [ABSTRACT FROM AUTHOR]

Dynamic Nuclear Polarization (DNP) can significantly enhance the sensitivity of solid-state NMR. In DNP, microwave irradiation induces polarization transfer from unpaired electron spins to 1 H nuclear spins via hyperfine couplings and spin-diffusion. The structure of the polarizing agents that host the electron spins is key for DNP efficiency. Currently, only a handful of structures perform well at very high magnetic fields (≥18.8 T), and enhancements are significantly lower than those obtained at lower fields. Here, we introduce a new series of water-soluble nitroxide biradicals with a scaffold augmented by dihydroxypropyl antenna chains that perform significantly better than previous dinitroxides at 18.8 T. The new radical M-TinyPol(OH) 4 yields enhancement factors of ∼220 at 18.8 T and 60 kHz MAS, which is a nearly factor 2 larger than for the previous best performing dinitroxides. The performance is understood through 2 H ESEEM measurements to probe solvent accessibility, supported by Molecular Dynamics simulations, and by experiments on deuterated samples. We find that the deuterated glycerol molecules in the matrix are located mainly in the second solvation shell of the NO bond, limiting access for protonated water molecules, and restricting spin diffusion pathways. This provides a rational understanding of why the dihydroxypropyl chains present in the best-performing structures are essential to deliver the polarization to the bulk solution., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Antimicrobial resistance is a leading cause of mortality, calling for the development of new antibiotics. The fungal antibiotic plectasin is a eukaryotic host defence peptide that blocks bacterial cell wall synthesis. Here, using a combination of solid-state nuclear magnetic resonance, atomic force microscopy and activity assays, we show that plectasin uses a calcium-sensitive supramolecular killing mechanism. Efficient and selective binding of the target lipid II, a cell wall precursor with an irreplaceable pyrophosphate, is achieved by the oligomerization of plectasin into dense supra-structures that only form on bacterial membranes that comprise lipid II. Oligomerization and target binding of plectasin are interdependent and are enhanced by the coordination of calcium ions to plectasin's prominent anionic patch, causing allosteric changes that markedly improve the activity of the antibiotic. Structural knowledge of how host defence peptides impair cell wall synthesis will likely enable the development of superior drug candidates., (© 2024. The Author(s).)

Proteorhodopsins are widely distributed photoreceptors from marine bacteria. Their discovery revealed a high degree of evolutionary adaptation to ambient light, resulting in blue- and green-absorbing variants that correlate with a conserved glutamine/leucine at position 105. On the basis of an integrated approach combining sensitivity-enhanced solid-state nuclear magnetic resonance (ssNMR) spectroscopy and linear-scaling quantum mechanics/molecular mechanics (QM/MM) methods, this single residue is shown to be responsible for a variety of synergistically coupled structural and electrostatic changes along the retinal polyene chain, ionone ring, and within the binding pocket. They collectively explain the observed color shift. Furthermore, analysis of the differences in chemical shift between nuclei within the same residues in green and blue proteorhodopsins also reveals a correlation with the respective degree of conservation. Our data show that the highly conserved color change mainly affects other highly conserved residues, illustrating a high degree of robustness of the color phenotype to sequence variation.

Cabotegravir is an integrase strand transfer inhibitor (INSTI) for HIV treatment and prevention. Cabotegravir-based long-acting pre-exposure prophylaxis (PrEP) presents an emerging paradigm for infectious disease control. In this scheme, a combination of a high efficacy and low solubility of anti-infection drugs permits the establishment of a pharmaceutical firewall in HIV-vulnerable groups over a long period. Although the structure-activity-relationship (SAR) of cabotegravir as an INSTI is known, the structural determinants of its low solubility have not been identified. In this work, we have integrated multiple experimental and computational methods, namely X-ray diffraction, solid-state NMR (SSNMR) spectroscopy, solution NMR spectroscopy, automated fragmentation (AF)-QM/MM and density functional theory (DFT) calculations, to address this question. The molecular organization of cabotegravir in crystal lattice has been determined. The combination of very-fast magic-angle-sample-spinning (VF MAS) SSNMR and solution NMR, as supported by AF-QM/MM and DFT calculations, permits the identification of structural factors that contribute to the low aqueous solubility of cabotegravir. Our study reveals the multitasking nature of pharmacophores in cabotegravir, which controls the drug solubility and, meanwhile, the biological activity. By unraveling these function-defining molecular features, our work could inspire further development of long-acting HIV PrEP drugs.

Antimicrobial resistance is a leading mortality factor worldwide. Here, we report the discovery of clovibactin, an antibiotic isolated from uncultured soil bacteria. Clovibactin efficiently kills drug-resistant Gram-positive bacterial pathogens without detectable resistance. Using biochemical assays, solid-state nuclear magnetic resonance, and atomic force microscopy, we dissect its mode of action. Clovibactin blocks cell wall synthesis by targeting pyrophosphate of multiple essential peptidoglycan precursors (C 55 PP, lipid II, and lipid III WTA ). Clovibactin uses an unusual hydrophobic interface to tightly wrap around pyrophosphate but bypasses the variable structural elements of precursors, accounting for the lack of resistance. Selective and efficient target binding is achieved by the sequestration of precursors into supramolecular fibrils that only form on bacterial membranes that contain lipid-anchored pyrophosphate groups. This potent antibiotic holds the promise of enabling the design of improved therapeutics that kill bacterial pathogens without resistance development., Competing Interests: Declaration of interests The following authors, A.J.P., C.A., A.N., A.L.S., L.L.L., D.H., and K.L., declare competing financial interests because they are employees and consultants of NovoBiotic Pharmaceuticals. A patent US 11,203,616 B2 was issued on 12/21/2021 and describes the use of clovibactin (Novo29) and as an antibiotic, as well as the pharmaceutical composition and antibiotic use of derivatives., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Proton-decoupled deuterium NMR spectra were obtained for an asymmetric liquid crystal dimer 1-(4-cyanobiphenyl-4'-yloxy)-6-(4-cyanobiphenyl-4'-yl)hexane (CB6OCB) containing a single -CD_{2}- group. The sample has two nematic liquid crystal phases: a twist-bend nematic, N_{TB}, at the lowest temperature followed by a uniaxial nematic, N_{U}, on increasing the temperature. Proton decoupling reduces the linewidths of the peaks in the deuterium spectrum from kHz to ∼100Hz, enabling quadrupolar splittings, Δν, to be obtained with enhanced precision as well as the dipolar coupling between deuterium nuclei within the CD_{2} group, hence enhancing the information content.

Low sensitivity is the primary limitation to extending nuclear magnetic resonance (NMR) techniques to more advanced chemical and structural studies. Photochemically induced dynamic nuclear polarization (photo-CIDNP) is an NMR hyperpolarization technique where light is used to excite a suitable donor-acceptor system, creating a spin-correlated radical pair whose evolution drives nuclear hyperpolarization. Systems that exhibit photo-CIDNP in solids are not common, and this effect has, up to now, only been observed for 13 C and 15 N nuclei. However, the low gyromagnetic ratio and natural abundance of these nuclei trap the local hyperpolarization in the vicinity of the chromophore and limit the utility for bulk hyperpolarization. Here, we report the first example of optically enhanced solid-state 1 H NMR spectroscopy in the high-field regime. This is achieved via photo-CIDNP of a donor-chromophore-acceptor molecule in a frozen solution at 0.3 T and 85 K, where spontaneous spin diffusion among the abundant strongly coupled 1 H nuclei relays polarization through the whole sample, yielding a 16-fold bulk 1 H signal enhancement under continuous laser irradiation at 450 nm. These findings enable a new strategy for hyperpolarized NMR beyond the current limits of conventional microwave-driven DNP.

The aggregation of amyloid-β 42 (Aβ42) is directly related to the pathogenesis of Alzheimer's disease. Here, we have investigated the early stages of the aggregation process, during which most of the cytotoxic species are formed. Aβ42 aggregation kinetics, characterized by the quantification of Aβ42 monomer consumption, were tracked by real-time solution NMR spectroscopy (RT-NMR) allowing the impact that low-molecular-weight (LMW) inhibitors and modulators exert on the aggregation process to be analysed. Distinct differences in the Aβ42 kinetic profiles were apparent and were further investigated kinetically and structurally by using thioflavin T (ThT) and transmission electron microscopy (TEM), respectively. LMW inhibitors were shown to have a differential impact on early-state aggregation. Insight provided here could direct future therapeutic design based on kinetic profiling of the process of fibril formation., (© 2023 The Authors. ChemBioChem published by Wiley-VCH GmbH.)

Background: Aggregation of α-synuclein (α-syn) is a prominent feature of Parkinson's disease (PD) and other synucleinopathies. Currently, α-syn seed amplification assays (SAAs) using cerebrospinal fluid (CSF) represent the most promising diagnostic tools for synucleinopathies. However, CSF itself contains several compounds that can modulate the aggregation of α-syn in a patient-dependent manner, potentially undermining unoptimized α-syn SAAs and preventing seed quantification., Methods: In this study, we characterized the inhibitory effect of CSF milieu on detection of α-syn aggregates by means of CSF fractionation, mass spectrometry, immunoassays, transmission electron microscopy, solution nuclear magnetic resonance spectroscopy, a highly accurate and standardized diagnostic SAA, and different in vitro aggregation conditions to evaluate spontaneous aggregation of α-syn., Results: We found the high-molecular weight fraction of CSF (> 100,000 Da) to be highly inhibitory on α-syn aggregation and identified lipoproteins to be the main drivers of this effect. Direct interaction between lipoproteins and monomeric α-syn was not detected by solution nuclear magnetic resonance spectroscopy, on the other hand we observed lipoprotein-α-syn complexes by transmission electron microscopy. These observations are compatible with hypothesizing an interaction between lipoproteins and oligomeric/proto-fibrillary α-syn intermediates. We observed significantly slower amplification of α-syn seeds in PD CSF when lipoproteins were added to the reaction mix of diagnostic SAA. Additionally, we observed a decreased inhibition capacity of CSF on α-syn aggregation after immunodepleting ApoA1 and ApoE. Finally, we observed that CSF ApoA1 and ApoE levels significantly correlated with SAA kinetic parameters in n = 31 SAA-negative control CSF samples spiked with preformed α-syn aggregates., Conclusions: Our results describe a novel interaction between lipoproteins and α-syn aggregates that inhibits the formation of α-syn fibrils and could have relevant implications. Indeed, the donor-specific inhibition of CSF on α-syn aggregation explains the lack of quantitative results from analysis of SAA-derived kinetic parameters to date. Furthermore, our data show that lipoproteins are the main inhibitory components of CSF, suggesting that lipoprotein concentration measurements could be incorporated into data analysis models to eliminate the confounding effects of CSF milieu on α-syn quantification efforts., (© 2023. The Author(s).)

Introduction: There are no recent data on primary ciliary dyskinesia (PCD) distribution, diagnosis and treatment in Italy., Methods: A descriptive study based on a survey questionnaire. It consisted of three sections (patients, diagnosis, and treatment), and sent to all the Italian PCD Centers., Results: Questionnaires obtained from 20/22 centers in 12/20 regions showed that the total number of PCD patients treated at the participating centers was of 416. Out of all centers, 55% follow <20 patients, two centers have >40 patients, and 75% follow both pediatric and adults. Age at diagnosis was between 4 and 8 years in 45% of the centers, <3 years in three centers. Nasal nitric oxide, transmission electron microscopy and ciliary high-speed video microscopy are performed in 75%, 90%, and 40% of centers, respectively. Immunofluorescence is available in five centers. Genetic analysis is offered in 55% of the centers, and in seven centers >50% of the patients have a known genetic profile. Patients treated at all centers receive inhaled saline solutions, corticosteroids and chest physiotherapy. Prophylactic antibiotics and mucolytics are prescribed in 95% and 50% of the centers, respectively. Pseudomonas infection is treated with oral or inhaled antibiotics., Conclusions: Many Italian centers care for a small number of pediatric and adult patients, and diagnosis is often delayed. We found a great variability in the available diagnostic procedures, as well in the prescribed therapies. Our study will help to uniform diagnostic algorithm and share treatments protocols for PCD in Italy and allowed to set specific national goals., (© 2023 Wiley Periodicals LLC.)

Stain-removing domiciliary protocols are focused on the elimination of dental extrinsic pigmentations by the application of abrasive toothpastes, extensively available in commerce. The goal of the present study is to evaluate the efficacy of two different stain removal molecule-formulated toothpastes by the reduction of clinical parameters: the micro-cleaning crystals and activated charcoal. A total of 40 participants with extrinsic dental pigmentations were enrolled and divided into two groups: a Control group, assigned to a toothpaste with micro-cleaning crystals (Colgate Sensation White); and a Trial group, with microparticle-activated charcoal toothpaste (Coswell Blanx Black). At T0 (baseline), T1 (10 days), T2 (1 month), and T3 (3 months), clinical parameters, including Lobene stain index calculated for intensity and extension, plaque control record, and bleeding on probing, were measured. Statistically significant differences were found in both groups ( p < 0.05): a reduction of extrinsic pigmentation, both in intensity and extension, was obtained in the Control group, but their total elimination could be achieved only in the Trial group with the activated charcoal molecule, though without significant difference between the groups ( p > 0.05). No intergroup differences were found for each timeframe for PCR, BoP, LSI-I, and LSI-E. Both tested toothpastes can be recommended for domiciliary oral hygiene of patients with extrinsic pigmentations.

Biomimetic zinc-carbonate hydroxyapatite technology was developed to realize materials that mimic the natural hydroxyapatite of enamel and dentin and possess good activity in terms of affinity to adhere to these biological tissues. The chemical and physical characteristics of this active ingredient allows the hydroxyapatite itself to be particularly similar to dental hydroxyapatite, enhancing the bond between biomimetic hydroxyapatite and dental hydroxyapatite. The aim of this review is to assess the efficacy of this technology in terms of benefits for enamel and dentin and reduction of dental hypersensitivity., Materials and Methods: A literature search (Pubmed/MEDLINE and Scopus) of articles from 2003 to 2023 was conducted to analyze studies focused on the use of zinc-hydroxyapatite products. Duplicates were eliminated from the 5065 articles found, leaving 2076 articles. Of these, 30 articles were analyzed based on the use of products with zinc-carbonate hydroxyapatite in these studies., Results: 30 articles were included. Most of the studies showed benefits in terms of remineralization and prevention of enamel demineralization in terms of occlusion of the dentinal tubules and reduction of dentinal hypersensitivity., Conclusion: Oral care products such as toothpaste and mouthwash with biomimetic zinc-carbonate hydroxyapatite were shown to provide benefits according to the aims of this review.

Antibiotics that use novel mechanisms are needed to combat antimicrobial resistance 1-3 . Teixobactin 4 represents a new class of antibiotics with a unique chemical scaffold and lack of detectable resistance. Teixobactin targets lipid II, a precursor of peptidoglycan 5 . Here we unravel the mechanism of teixobactin at the atomic level using a combination of solid-state NMR, microscopy, in vivo assays and molecular dynamics simulations. The unique enduracididine C-terminal headgroup of teixobactin specifically binds to the pyrophosphate-sugar moiety of lipid II, whereas the N terminus coordinates the pyrophosphate of another lipid II molecule. This configuration favours the formation of a β-sheet of teixobactins bound to the target, creating a supramolecular fibrillar structure. Specific binding to the conserved pyrophosphate-sugar moiety accounts for the lack of resistance to teixobactin 4 . The supramolecular structure compromises membrane integrity. Atomic force microscopy and molecular dynamics simulations show that the supramolecular structure displaces phospholipids, thinning the membrane. The long hydrophobic tails of lipid II concentrated within the supramolecular structure apparently contribute to membrane disruption. Teixobactin hijacks lipid II to help destroy the membrane. Known membrane-acting antibiotics also damage human cells, producing undesirable side effects. Teixobactin damages only membranes that contain lipid II, which is absent in eukaryotes, elegantly resolving the toxicity problem. The two-pronged action against cell wall synthesis and cytoplasmic membrane produces a highly effective compound targeting the bacterial cell envelope. Structural knowledge of the mechanism of teixobactin will enable the rational design of improved drug candidates., (© 2022. The Author(s).)

The engineering of the surface of nanomaterials with bioactive molecules allows controlling their biological identity thus accessing functional materials with tuned physicochemical and biological profiles suited for specific applications. Then, the manufacturing process, by which the nanomaterial surface is grafted, has a significant impact on their development and innovation. In this regard, we report herein the grafting of sugar headgroups on a graphene oxide (GO) surface by exploiting a green manufacturing process that relies on the use of vibrational ball mills, a grinding apparatus in which the energy is transferred to the reacting species through collision with agate spheres inside a closed and vibrating vessel. The chemical composition and the morphology of the resulting glyco-graphene oxide conjugates (glyco-GO) are assessed by the combination of a series of complementary advanced techniques ( i.e. UV-vis and Raman spectroscopy, transmission electron microscopy, and Magic Angle Spinning (MAS) solid-state NMR (ssNMR) providing in-depth insights into the chemical reactivity of GO in a mechanochemical route. The conjugation of monosaccharide residues on the GO surface significantly improves the antimicrobial activity of pristine GO against P. aeruginosa . Indeed, glyco-GO conjugates, according to the monosaccharide derivatives installed into the GO surface, affect the ability of sessile cells to adhere to a polystyrene surface in a colony forming assay. Scanning electron microscopy images clearly show that glyco-GO conjugates significantly disrupt an already established P. aeruginosa biofilm.

The structural characterization of supported molecular catalysts is challenging due to the low density of active sites and the presence of several organic/organometallic surface groups resulting from the often complex surface chemistry associated with support functionalization. Here, we provide a complete atomic-scale description of all surface sites in an N-heterocyclic carbene based on iridium and supported on silica, at all stages of its synthesis. By combining a suitable isotope labeling strategy with the implementation of multinuclear dipolar recoupling DNP-enhanced NMR experiments, the 3D structure of the Ir-NHC sites, as well as that of the synthesis intermediates were determined. As a significant fraction of parent surface fragments does not react during the multistep synthesis, site-selective experiments were implemented to specifically probe proximities between the organometallic groups and the solid support. The NMR-derived structure of the iridium sites points to a well-defined conformation. By interpreting EXAFS spectroscopy and chemical analysis data augmented by computational studies, the presence of two coordination geometries is demonstrated: Ir-NHC fragments coordinated by a 1,5-cyclooctadiene and one Cl ligand, as well as, more surprisingly, a fragment coordinated by two NHC and two Cl ligands. This study demonstrates a unique methodology to disclose individual surface structures in complex, multisite environments, a long-standing challenge in the field of heterogeneous/supported catalysts, while revealing new, unexpected structural features of metallo-NHC-supported substrates. It also highlights the potentially large diversity of surface sites present in functional materials prepared by surface chemistry, an essential knowledge to design materials with improved performances.

Dynamic nuclear polarization (DNP) at cryogenic temperatures has proved to be a valuable technique to enhance the sensitivity of solid-state NMR spectroscopy. Over the years, sample formulations have been optimized for experiments at cryogenic temperatures. At 9.4 T, the best performing polarizing agents are dinitroxides such as AMUPol and TEKPol that lead to enhancement factors of around 250 at 100 K. However, the performance of these radicals plummets at higher temperatures. Here we introduce trehalose-based DNP polarizing matrices, suitable to embed biomolecular assemblies. Several formulation protocols are investigated, in combination with various polarizing agents, including a new biradical structure chemically tethered to a trehalose molecule. The DNP efficiency of these new polarizing media is screened as a function of the radical concentration, the hydration level of the matrix and the protein content. Sizeable enhancement factors are reported at 100 K and 9.4 T. More importantly, we show that the DNP performance of these new polarizing media outperform the conventionally used water/glycerol mixture at temperatures above 180 K. This study establishes trehalose matrices as a promising DNP medium for experiments at temperatures >150 K where conventional water-based formulations soften and are no longer viable, thus opening new avenues for DNP enhanced solid-state NMR spectroscopy at temperatures close to ambient temperature.

Recently, it has been shown that methods based on the dynamics of 1 H nuclear hyperpolarization in magic angle spinning (MAS) NMR experiments can be used to determine mesoscale structures in complex materials. However, these methods suffer from low sensitivity, especially since they have so far only been feasible with indirect detection of 1 H polarization through dilute heteronuclei such as 13 C or 29 Si. Here we combine relayed-DNP (R-DNP) with fast MAS using 0.7 mm diameter rotors at 21.2 T. Fast MAS enables direct 1 H detection to follow hyperpolarization dynamics, leading to an acceleration in experiment times by a factor 16. Furthermore, we show that by varying the MAS rate, and consequently modulating the 1 H spin diffusion rate, we can record a series of independent R-DNP curves that can be analyzed jointly to provide an accurate determination of domain sizes. This is confirmed here with measurements on microcrystalline l-histidine·HCl·H 2 O at MAS frequencies up to 60 kHz, where we determine a Weibull distribution of particle sizes centered on a radius of 440 ± 20 nm with an order parameter of k = 2.2., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Several urea-inserted organo-polyoxometalates (POMs) derived from polyoxotungstovanadate [P 2 V 3 W 15 O 61 ] 9- were prepared. The insertion of the carbonyl into the polyoxometallic framework activates the urea toward Hydrogen-bond catalysis. This was shown on the Friedel-Crafts arylation of trans-β-nitrostyrene. Modelling shows that the most stable form of the organo-POMs features a cis-trans arrangement of the two N-H bonds, but that the likely catalytically active trans-trans form is accessible at room temperature. Finally, it is possible that the oxo substituents next to the vanadium atoms may help the approach of the nucleophile via H-bonding., (© 2021 Wiley-VCH GmbH.)

Pairing the spectral resolution provided by high magnetic fields at ambient temperature with the enhanced sensitivity offered by dynamic nuclear polarization (DNP) is a major goal of modern solid-state NMR spectroscopy, which will allow one to unlock ever-challenging applications. This study demonstrates that, by combining HyTEK2, a hybrid BDPA-nitroxide biradical polarizing agent, with ortho -terphenyl (OTP), a rigid DNP matrix, enhancement factors as high as 65 can be obtained at 230 K, 40 kHz magic angle spinning (MAS), and 18.8 T. The temperature dependence of the DNP enhancement and its behavior around the glass transition temperature ( T g ) of the matrix is investigated by variable-temperature EPR measurements of the electron relaxation properties and numerical simulations. A correlation is suggested between the decrease in enhancement at the passage of the T g and the concomitant drop of both transverse electron relaxation times in the biradical.

Toothpastes containing biomimetic hydroxyapatite have been investigated in recent years; the behavior of this material in the oral environment has been evaluated directly on dental enamel showing a marked remineralizing activity. To propose microRepair ® -based toothpastes (Zn-carbonate hydroxyapatite) for the domiciliary oral hygiene in patients with dental composite restorations, the aim of this study was to evaluate the deposition of Zn-carbonate hydroxyapatite on a polymeric composite resin with Scanning Electron Microscopy/Energy-Dispersive X-ray Spectrometry (SEM/EDS) analysis. Twenty healthy volunteers underwent the bonding of 3 orthodontic buttons on the vestibular surfaces of upper right premolars and first molar. On the surface of the buttons, a ball-shaped mass of composite resin was applied and light-cured. Then, the volunteers were randomly divided into two groups according to the toothpaste used for domiciliary oral hygiene: the Control toothpaste containing stannous fluoride and the Trial toothpaste containing microRepair ® . The buttons were debonded after 7 days (T1-first premolar), after 15 days (T2-second premolar), and after 30 days (T3-first molar) to undergo the SEM/EDS analysis. The deposition of calcium, phosphorus, and silicon was assessed through EDS analysis and data were submitted to statistical analysis ( p < 0.05). SEM morphologic evaluation showed a marked deposition of the two toothpastes on the surfaces of the buttons. EDS quantitative analysis showed an increase of calcium, phosphorus, and silicon in both the groups, with a statistically significant difference of calcium deposition at T3 for the Trial group. Therefore, the use of toothpaste containing Zn-carbonate hydroxyapatite could be proposed as a device for domiciliary oral hygiene because the deposition of hydroxyapatite on polymeric composite resin could prevent secondary caries on the margins of restorations.

Both ^{1}H and ^{13}C NMR spectra have been obtained in a static magnetic field of 23.5 T on a bent-shaped dimer molecule, 1^{''},7^{''}-bis(4-cyanobiphenyl-4'-yl) nonane (CB9CB), which shows the sequence of liquid crystal phases twist-bend nematic, N&#95;{TB}, and uniaxial nematic, N&#95;{U}, before entering the isotropic phase. The ^{1}H spectra are used to locate the temperature at which the sample melts to form a twist-bend nematic, T&#95;{CrN&#95;{TB}}, and then T&#95;{N&#95;{U}I} when the isotropic phase is entered, both in a magnetic field of 23.5 T, and to compare these with those measured at the Earth's field. The differences between these transition temperatures are found to be zero within the error in their measurement, in stark contrast to previous measurements by Salili et al. [Phys. Rev. Lett. 116, 217801 (2016)10.1103/PhysRevLett.116.217801]. In the isotropic phase in the presence of the field the sample exists in a paranematic phase in which the molecules of CB9CB are partially ordered. The ^{1}H and ^{13}C NMR spectra in the paranematic phase are used to measure the critical temperature T* below which this phase is unstable. The spectra are also used to study the structure, molecular orientational order, and distribution of molecular conformations in the paranematic phase.

Solid-state nuclear magnetic resonance under magic angle spinning (MAS) enhanced with dynamic nuclear polarization (DNP) is a powerful approach to characterize many important classes of materials, allowing access to previously inaccessible structural and dynamic parameters. Here, we present the first DNP MAS experiments using a 0.7 mm MAS probe, which allows us to reach spinning frequencies of 65 kHz, with microwave irradiation, at 100 K. At the highest magnetic field available for DNP today (21.1 T), we find that the polarizing agent HyTEK2 provides DNP enhancements as high as 200 at a spinning rate of 65 kHz at 100 K, and BDPA yields an enhancement of 106 under the same conditions. Fast spinning rates enable excellent DNP performance, but they also yield unprecedented 1 H resolution under DNP conditions. We report well-resolved 1 H-detected 1 H- 13 C and 1 H- 15 N correlation spectra of microcrystalline histidine·HCl·H 2 O.

The development of magic-angle spinning dynamic nuclear polarization (MAS DNP) has allowed atomic-level characterization of materials for which conventional solid-state NMR is impractical due to the lack of sensitivity. The rapid progress of MAS DNP has been largely enabled through the understanding of rational design concepts for more efficient polarizing agents (PAs). Here, we identify a new design principle which has so far been overlooked. We find that the local geometry around the unpaired electron can change the DNP enhancement by an order of magnitude for two otherwise identical conformers. We present a set of 13 new stable mono- and dinitroxide PAs for MAS DNP NMR where this principle is demonstrated. The radicals are divided into two groups of isomers, named open (O-) and closed (C-), based on the ring conformations in the vicinity of the N-O bond. In all cases, the open conformers exhibit dramatically improved DNP performance as compared to the closed counterparts. In particular, a new urea-based biradical named HydrOPol and a mononitroxide O-MbPyTol yield enhancements of 330 ± 60 and 119 ± 25, respectively, at 9.4 T and 100 K, which are the highest enhancements reported so far in the aqueous solvents used here. We find that while the conformational changes do not significantly affect electron spin-spin distances, they do affect the distribution of the exchange couplings in these biradicals. Electron spin echo envelope modulation (ESEEM) experiments suggest that the improved performance of the open conformers is correlated with higher solvent accessibility.

Identifying and characterizing systems that generate well-defined states with large electron spin polarization is of high interest for applications in molecular spintronics, high-energy physics, and magnetic resonance spectroscopy. The generation of electron spin polarization on free-radical substituents tethered to pentacene derivatives has recently gained a great deal of interest for its applications in molecular electronics. After photoexcitation of the chromophore, pentacene-radical derivatives can rapidly form spin-polarized triplet excited states through enhanced intersystem crossing. Under the right conditions, the triplet spin polarization, arising from m S -selective intersystem crossing rates, can be transferred to the tethered stable radical. The efficiency of this spin polarization transfer depends on many factors: local magnetic and electric fields, excited-state energetics, molecular geometry, and spin-spin coupling. Here, we present transient electron paramagnetic resonance (EPR) measurements on three pentacene derivatives tethered to Finland trityl, BDPA, or TEMPO radicals to explore the influence of the nature of the radical on the spin polarization transfer. We observe efficient polarization transfer between the pentacene excited triplet and the trityl radical but do not observe the same for the BDPA and TEMPO derivatives. The polarization transfer behavior in the pentacene-trityl system is also investigated in different glassy matrices and is found to depend markedly on the solvent used. The EPR results are rationalized with the help of femtosecond and nanosecond transient absorption measurements, yielding complementary information on the excited-state dynamics of the three pentacene derivatives. Notably, we observe a 2 orders of magnitude difference in the time scale of triplet formation between the pentacene-trityl system and the pentacene systems tethered with the BDPA and TEMPO radicals.

The natural antibiotic teixobactin kills pathogenic bacteria without detectable resistance. The difficult synthesis and unfavourable solubility of teixobactin require modifications, yet insufficient knowledge on its binding mode impedes the hunt for superior analogues. Thus far, teixobactins are assumed to kill bacteria by binding to cognate cell wall precursors (Lipid II and III). Here we present the binding mode of teixobactins in cellular membranes using solid-state NMR, microscopy, and affinity assays. We solve the structure of the complex formed by an improved teixobactin-analogue and Lipid II and reveal how teixobactins recognize a broad spectrum of targets. Unexpectedly, we find that teixobactins only weakly bind to Lipid II in cellular membranes, implying the direct interaction with cell wall precursors is not the sole killing mechanism. Our data suggest an additional mechanism affords the excellent activity of teixobactins, which can block the cell wall biosynthesis by capturing precursors in massive clusters on membranes.

In bacteria, nucleoid associated proteins (NAPs) take part in active chromosome organization by supercoil management, three-dimensional DNA looping and direct transcriptional control. Mycobacterial integration host factor (mIHF, rv1388) is a NAP restricted to Actinobacteria and essential for survival of the human pathogen Mycobacterium tuberculosis. We show in vitro that DNA binding by mIHF strongly stabilizes the protein and increases its melting temperature. The structure obtained by Nuclear Magnetic Resonance (NMR) spectroscopy characterizes mIHF as a globular protein with a protruding alpha helix and a disordered N-terminus, similar to Streptomyces coelicolor IHF (sIHF). NMR revealed no residues of high flexibility, suggesting that mIHF is a rigid protein overall that does not undergo structural rearrangements. We show that mIHF only binds to double stranded DNA in solution, through two DNA binding sites (DBSs) similar to those identified in the X-ray structure of sIHF. According to Atomic Force Microscopy, mIHF is able to introduce left-handed loops of ca. 100 nm size (~300 bp) in supercoiled cosmids, thereby unwinding and relaxing the DNA., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019. Published by Elsevier Inc.)