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Raman spectroscopy was applied to study the structural differences between herpes simplex virus Type I (HSV-1) and Epstein–Barr virus (EBV). Raman spectra were first collected with statistical validity on clusters of the respective virions and analyzed according to principal component analysis (PCA). Then, average spectra were computed and a machine-learning approach applied to deconvolute them into sub-band components in order to perform comparative analyses. The Raman results revealed marked structural differences between the two viral strains, which could mainly be traced back to the massive presence of carbohydrates in the glycoproteins of EBV virions. Clear differences could also be recorded for selected tyrosine and tryptophan Raman bands sensitive to pH at the virion/environment interface. According to the observed spectral differences, Raman signatures of known biomolecules were interpreted to link structural differences with the viral functions of the two strains. The present study confirms the unique ability of Raman spectroscopy for answering structural questions at the molecular level in virology and, despite the structural complexity of viral structures, its capacity to readily and reliably differentiate between different virus types and strains. [ABSTRACT FROM AUTHOR]

The aim of this study was to elucidate the chemistry of cellular degeneration in human neuroblastoma cells upon exposure to outer-membrane vesicles (OMVs) produced by Porphyromonas gingivalis (Pg) oral bacteria by monitoring their metabolomic evolution using in situ Raman spectroscopy. Pg-OMVs are a key factor in Alzheimer's disease (AD) pathogenesis, as they act as efficient vectors for the delivery of toxins promoting neuronal damage. However, the chemical mechanisms underlying the direct impact of Pg-OMVs on cell metabolites at the molecular scale still remain conspicuously unclear. A widely used in vitro model employing neuroblastoma SH-SY5Y cells (a sub-line of the SK-N-SH cell line) was spectroscopically analyzed in situ before and 6 h after Pg-OMV contamination. Concurrently, Raman characterizations were also performed on isolated Pg-OMVs, which included phosphorylated dihydroceramide (PDHC) lipids and lipopolysaccharide (LPS), the latter in turn being contaminated with a highly pathogenic class of cysteine proteases, a key factor in neuronal cell degradation. Raman characterizations located lipopolysaccharide fingerprints in the vesicle structure and unveiled so far unproved aspects of the chemistry behind protein degradation induced by Pg-OMV contamination of SH-SY5Y cells. The observed alterations of cells' Raman profiles were then discussed in view of key factors including the formation of amyloid β (Aβ) plaques and hyperphosphorylated Tau neurofibrillary tangles, and the formation of cholesterol agglomerates that exacerbate AD pathologies. [ABSTRACT FROM AUTHOR]

Hydrolytic reactions taking place at the surface of a silicon nitride (Si3N4) bioceramic were found to induce instantaneous inactivation of Human herpesvirus 1 (HHV-1, also known as Herpes simplex virus 1 or HSV-1). Si3N4 is a non-oxide ceramic compound with strong antibacterial and antiviral properties that has been proven safe for human cells. HSV-1 is a double-stranded DNA virus that infects a variety of host tissues through a lytic and latent cycle. Real-time reverse transcription (RT)-polymerase chain reaction (PCR) tests of HSV-1 DNA after instantaneous contact with Si3N4 showed that ammonia and its nitrogen radical byproducts, produced upon Si3N4 hydrolysis, directly reacted with viral proteins and fragmented the virus DNA, irreversibly damaging its structure. A comparison carried out upon testing HSV-1 against ZrO2 particles under identical experimental conditions showed a significantly weaker (but not null) antiviral effect, which was attributed to oxygen radical influence. The results of this study extend the effectiveness of Si3N4's antiviral properties beyond their previously proven efficacy against a large variety of single-stranded enveloped and non-enveloped RNA viruses. Possible applications include the development of antiviral creams or gels and oral rinses to exploit an extremely efficient, localized, and instantaneous viral reduction by means of a safe and more effective alternative to conventional antiviral creams. Upon incorporating a minor fraction of micrometric Si3N4 particles into polymeric matrices, antiherpetic devices could be fabricated, which would effectively impede viral reactivation and enable high local effectiveness for extended periods of time. [ABSTRACT FROM AUTHOR]

This study probed in vitro the mechanisms of competition/coexistence between Streptococcus sanguinis (known for being correlated with health in the oral cavity) and Streptococcus mutans (responsible for aciduric oral environment and formation of caries) by means of quantitative Raman spectroscopy and imaging. In situ Raman assessments of live bacterial culture/coculture focusing on biofilm exopolysaccharides supported the hypothesis that both species engaged in antagonistic interactions. Experiments of simultaneous colonization always resulted in coexistence, but they also revealed fundamental alterations of the biofilm with respect to their water-insoluble glucan structure. Raman spectra (collected at fixed time but different bacterial ratios) showed clear changes in chemical bonds in glucans, which pointed to an action by Streptococcus sanguinis to discontinue the impermeability of the biofilm constructed by Streptococcus mutans. The concurrent effects of glycosidic bond cleavage in water-insoluble α − 1,3–glucan and oxidation at various sites in glucans' molecular chains supported the hypothesis that secretion of oxygen radicals was the main "chemical weapon" used by Streptococcus sanguinis in coculture. [ABSTRACT FROM AUTHOR]

This study targets on-site/real-time taxonomic identification and metabolic profiling of seven different Candida auris clades/subclades by means of Raman spectroscopy and imaging. Representative Raman spectra from different Candida auris samples were systematically deconvoluted by means of a customized machine-learning algorithm linked to a Raman database in order to decode structural differences at the molecular scale. Raman analyses of metabolites revealed clear differences in cell walls and membrane structure among clades/subclades. Such differences are key in maintaining the integrity and physical strength of the cell walls in the dynamic response to external stress and drugs. It was found that Candida cells use the glucan structure of the extracellular matrix, the degree of α-chitin crystallinity, and the concentration of hydrogen bonds between its antiparallel chains to tailor cell walls' flexibility. Besides being an effective ploy in survivorship by providing stiff shields in the α–1,3–glucan polymorph, the α–1,3–glycosidic linkages are also water-insoluble, thus forming a rigid and hydrophobic scaffold surrounded by a matrix of pliable and hydrated β–glucans. Raman analysis revealed a variety of strategies by different clades to balance stiffness, hydrophobicity, and impermeability in their cell walls. The selected strategies lead to differences in resistance toward specific environmental stresses of cationic/osmotic, oxidative, and nitrosative origins. A statistical validation based on principal component analysis was found only partially capable of distinguishing among Raman spectra of clades and subclades. Raman barcoding based on an algorithm converting spectrally deconvoluted Raman sub-bands into barcodes allowed for circumventing any speciation deficiency. Empowered by barcoding bioinformatics, Raman analyses, which are fast and require no sample preparation, allow on-site speciation and real-time selection of appropriate treatments. [ABSTRACT FROM AUTHOR]

This study presents a set of vibrational characterizations on a nanogel-cross-linked porous freeze-dried gel (NanoCliP-FD gel) scaffold for tissue engineering and regenerative therapy. This scaffold is designed for the in vitro culture of high-quality cartilage tissue to be then transplanted in vivo to enable recovery from congenital malformations in the maxillofacial area or crippling jaw disease. The three-dimensional scaffold for in-plate culture is designed with interface chemistry capable of stimulating cartilage formation and maintaining its structure through counteracting the dedifferentiation of mesenchymal stem cells (MSCs) during the formation of cartilage tissue. The developed interface chemistry enabled high efficiency in both growth rate and tissue quality, thus satisfying the requirements of large volumes, high matrix quality, and superior mechanical properties needed in cartilage transplants. We characterized the cartilage tissue in vitro grown on a NanoCliP-FD gel scaffold by human periodontal ligament-derived stem cells (a type of MSC) with cartilage grown by the same cells and under the same conditions on a conventional (porous) atelocollagen scaffold. The cartilage tissues produced by the MSCs on different scaffolds were comparatively evaluated by immunohistochemical and spectroscopic analyses. Cartilage differentiation occurred at a higher rate when MSCs were cultured on the NanoCliP-FD gel scaffold compared to the atelocollagen scaffold, and produced a tissue richer in cartilage matrix. In situ spectroscopic analyses revealed the cell/scaffold interactive mechanisms by which the NanoCliP-FD gel scaffold stimulated such increased efficiency in cartilage matrix formation. In addition to demonstrating the high potential of human periodontal ligament-derived stem cell cultures on NanoCliP-FD gel scaffolds in regenerative cartilage therapy, the present study also highlights the novelty of Raman spectroscopy as a non-destructive method for the concurrent evaluation of matrix quality and cell metabolic response. In situ Raman analyses on living cells unveiled for the first time the underlying physiological mechanisms behind such improved chondrocyte performance. [ABSTRACT FROM AUTHOR]

Oral candidiasis, a common opportunistic infection of the oral cavity, is mainly caused by the following four Candida species (in decreasing incidence rate): Candida albicans, Candida glabrata, Candida tropicalis, and Candida krusei. This study offers in-depth Raman spectroscopy analyses of these species and proposes procedures for an accurate and rapid identification of oral yeast species. We first obtained average spectra for different Candida species and systematically analyzed them in order to decode structural differences among species at the molecular scale. Then, we searched for a statistical validation through a chemometric method based on principal component analysis (PCA). This method was found only partially capable to mechanistically distinguish among Candida species. We thus proposed a new Raman barcoding approach based on an algorithm that converts spectrally deconvoluted Raman sub-bands into barcodes. Barcode-assisted Raman analyses could enable on-site identification in nearly real-time, thus implementing preventive oral control, enabling prompt selection of the most effective drug, and increasing the probability to interrupt disease transmission. [ABSTRACT FROM AUTHOR]