Molecular monitoring of viral infections in immunocompromised patients in a large university hospital in Italy: reflections after thirteen years of real-life activity.

PURPOSE: This study aimed to investigate the prevalence and viral reactivations of clinical interest in the immunocompromised patient with particular focus on hematologic and solid organ transplant recipients. METHODS: Molecular screening data of CMV, EBV, JCV and BKV from 2011 to 2023 were analyzed. This extensive time span allowed the access to more than 100,000 samples from over 20,000 patients treated at Policlinico Umberto I. It was possible to temporally investigate patient attendance patterns, average age distribution, seasonality of infections, and positivity rates of the analyzed viruses. RESULTS: Between 2019 and 2022 a significant reduction in organ transplants performed and in the positive molecular detection of EBV, JCV and BKV was observed. Additionally, there has been a noteworthy decrease in CMV reactivations, with a reduction of up to 50% starting in 2019. A remarkable reduction of 39% in the rate of CMV viral reactivation has been also achieved in SOT between 2016 and 2023. CONCLUSION: The years following 2019 were profoundly impacted by the COVID-19 pandemic era. This period resulted in a substantial reduction in healthcare services and hospital visits. Furthermore, the introduction of the drug Letermovir in Italy in 2019 demonstrated remarkable efficacy, evidenced by a reduction in CMV reactivations. Additionally, the adoption of a novel clinical approach centered on personalized therapy facilitated improved management of immunocompromised patients.

Insights on Stability Constants and Structures of Complexes between Coumarin Derivatives and Pb(II) in Aqueous Media.

In the frame of a systematic study on the sequestering ability of natural antioxidants towards metal cations, here the complexation of coumarin-3-carboxilic acid (HCCA) with Pb(II) and the overall stability constants of the resulting complexes, at 37 °C and in 0.16 M NaClO4, are discussed. Reaction of Pb(ClO4)2 with HCCA in an aqueous medium at a pH range from 2 to 6 and various ratios (1:1-1:10) yielded the Pb-CCA complexes, which were characterized spectrometrically by laser desorption ionization mass spectrometry (LD-MS). LD-MS has provided the composition and structure of Pb-CCA species according to the speciation model proposed on the basis of the potentiometric data. The graphic representation of the complex's concentration curves is given by the distribution diagram, which provides a whole depiction of the species present in the solution at the selected pH ranges.

Torque teno virus (TTV): A gentle spy virus of immune status, predictive marker of seroconversion to COVID-19 vaccine in kidney and lung transplant recipients.

To date, no comprehensive marker to monitor the immune status of patients is available. Given that Torque teno virus (TTV), a known human virome component, has previously been identified as a marker of immunocompetence, it was retrospectively investigated whether TTV viral load may also represent a marker of ability to develop antibody in response to COVID-19-BNT162B2 vaccine in solid organ transplant recipients (SOT). Specifically, 273 samples from 146 kidney and 26 lung transplant recipients after successive doses of vaccine were analyzed. An inverse correlation was observed within the TTV copy number and anti-Spike IgG antibody titer with a progressive decrease in viremia the further away from the transplant date. Analyzing the data obtained after the second dose, a significant difference in TTV copy number between responsive and nonresponsive patients was observed, considering a 5 log10 TTV copies/mL threshold to discriminate between the two groups. Moreover, for 86 patients followed in their response to the second and third vaccination doses a 6 log10 TTV copies/mL threshold was used to predict responsivity to the booster dose. Although further investigation is necessary, possibly extending the analysis to other patient categories, this study suggests that TTV can be used as a good marker of vaccine response in transplant patients.

A Picrocrocin-Enriched Fraction from a Saffron Extract Affects Lipid Homeostasis in HepG2 Cells through a Non-Statin-like Mode.

Dyslipidemia is a lipid metabolism disorder associated with the loss of the physiological homeostasis that ensures safe levels of lipids in the organism. This metabolic disorder can trigger pathological conditions such as atherosclerosis and cardiovascular diseases. In this regard, statins currently represent the main pharmacological therapy, but their contraindications and side effects limit their use. This is stimulating the search for new therapeutic strategies. In this work, we investigated in HepG2 cells the hypolipidemic potential of a picrocrocin-enriched fraction, analyzed by high-resolution 1H NMR and obtained from a saffron extract, the stigmas of Crocus sativus L., a precious spice that has already displayed interesting biological properties. Spectrophotometric assays, as well as expression level of the main enzymes involved in lipid metabolism, have highlighted the interesting hypolipidemic effects of this natural compound; they seem to be exerted through a non-statin-like mechanism. Overall, this work provides new insights into the metabolic effects of picrocrocin, thus confirming the biological potential of saffron and paving the way for in vivo studies that could validate this spice or its phytocomplexes as useful adjuvants in balancing blood lipid homeostasis.

Edible Insects an Alternative Nutritional Source of Bioactive Compounds: A Review.

Edible insects have the potential to become one of the major future foods. In fact, they can be considered cheap, highly nutritious, and healthy food sources. International agencies, such as the Food and Agriculture Organization (FAO), have focused their attention on the consumption of edible insects, in particular, regarding their nutritional value and possible biological, toxicological, and allergenic risks, wishing the development of analytical methods to verify the authenticity, quality, and safety of insect-based products. Edible insects are rich in proteins, fats, fiber, vitamins, and minerals but also seem to contain large amounts of polyphenols able to have a key role in specific bioactivities. Therefore, this review is an overview of the potential of edible insects as a source of bioactive compounds, such as polyphenols, that can be a function of diet but also related to insect chemical defense. Currently, insect phenolic compounds have mostly been assayed for their antioxidant bioactivity; however, they also exert other activities, such as anti-inflammatory and anticancer activity, antityrosinase, antigenotoxic, and pancreatic lipase inhibitory activities.

Quality of T-Cell Response to SARS-CoV-2 mRNA Vaccine in ART-Treated PLWH.

We investigated specific humoral and T-cell responses in people living with HIV (PLWH) before (T0), after two (T1) and after six months (T2) from the third dose of the BNT162b2 vaccine. Healthy donors (HD) were enrolled. The specific humoral response was present in most PLWH already after the second dose, but the third dose increased both the rate of response and its magnitude. Collectively, no significant differences were found in the percentage of responding T-cells between PLWH and HD. At T0, stratifying PLWH according to CD4 cell count, a lower percentage of responding T-cells in <200 cells/µL subgroup compared to >200 cells/µL one was observed. At T1, this parameter was comparable between the two subgroups, and the same result was found at T2. However, the pattern of co-expression of IFNγ, IL2 and TNFα in PLWH was characterized by a higher expression of TNFα, independently of CD4 cell count, indicating a persistent immunological signature despite successful ART. mRNA vaccination elicited a specific response in most PLWH, although the cellular one seems qualitatively inferior compared to HD. Therefore, an understanding of the T-cell quality dynamic is needed to determine the best vaccination strategy and, in general, the capability of immune response in ART-treated PLWH.

Mass Spectrometry-Based Peptide Profiling of Haemolymph from Pterostichus melas Exposed to Pendimethalin Herbicide.

Pendimethalin-based herbicides are used worldwide for pre-emergence selective control of annual grasses and weeds in croplands. The endurance of herbicides residues in the environment has an impact on the soil biodiversity and fertility, also affecting non-target species, including terrestrial invertebrates. Carabid beetles are known as natural pest control agents in the soil food web of agroecosystems, and feed on invertebrates and weed seeds. Here, a mass spectrometry untargeted profiling of haemolymph is used to investigate Pterostichus melas metabolic response after to pendimethalin-based herbicide exposure. Mass spectrometric data are examined with statistical approaches, such as principal component analysis, for possible correlation with biological effects. Those signals with high correlation are submitted to tandem mass spectrometry to identify the associated biomarker. The time course exposure showed many interesting findings, including a significant downregulation of related to immune and defense peptides (M-lycotoxin-Ls4a, Peptide hormone 1, Paralytic peptide 2, and Serine protease inhibitor 2). Overall, the observed peptide deregulations concur with the general mechanism of uptake and elimination of toxicants reported for Arthropods.

Analytical Strategy for MS-Based Thanatochemistry to Estimate Postmortem Interval.

An analytical strategy for a matrix-assisted laser desorption mass spectrometry-based untargeted metabolomic study on vitreous humor (VH) was developed, looking for statistically significant parameters correlated to death time estimation. Five incubation stages of VH, 0, 24, 48, 72, and 96 h, at physiological pH and controlled temperature, were adopted to monitor time-dependent changes and correlate them with the postmortem interval (PMI). Using two multivariate statistical approaches, principal component regression (PCR) and partial least squares regression (PLSR), the PMI was assessed, considering the m/z values from mass spectra and the incubation time (ISt) as predictors. An independent validation set was used to evaluate the predictive capability of the models through the coefficient of determination (R2) and the root-mean-square error (RMSE). Different pre-treatments were applied to the raw mass spectra, and their performance in assessing PMI was evaluated. Based on the best outcomes in terms of both R2 and RMSE, multiplicative scatter correction combined with a logarithmic transformation was chosen. The results of PCR and PLSR based on the selected pre-treatment are encouraging because validation R2 is about 0.95 for both models. Moreover, the prediction error is 6 h for both models, when PMI is lower than 1 day. Although these results are obtained by the uncritical application of the models, they are comparable to or even better than those reported in the literature. Notwithstanding, we consider that many in situ influences, such as passive diffusion, functional loss of tissues, and advanced autolytic processes, could not get captured in vitro. However, the developed approach was optimized using VH samples and overcomes the limitations of the vast majority of methods that require validation for serum and/or urine samples.

Synchronizing Two Superconducting Qubits through a Dissipating Resonator.

A system consisting of two qubits and a resonator is considered in the presence of different sources of noise, bringing to light the possibility of making the two qubits evolve in a synchronized way. A direct qubit-qubit interaction turns out to be a crucial ingredient, as well as the dissipation processes involving the resonator. The detrimental role of the local dephasing of the qubits is also taken into account.

Protein Extraction, Enrichment and MALDI MS and MS/MS Analysis from Bitter Orange Leaves (Citrus aurantium).

Citrus aurantium is a widespread tree in the Mediterranean area, and it is mainly used as rootstock for other citrus. In the present study, a vacuum infiltration centrifugation procedure, followed by solid phase extraction matrix-assisted laser desorption ionization tandem mass spectrometry (SPE MALDI MS/MS) analysis, was adopted to isolate proteins from leaves. The results of mass spectrometry (MS) profiling, combined with the top-down proteomics approach, allowed the identification of 78 proteins. The bioinformatic databases TargetP, SignalP, ChloroP, WallProtDB, and mGOASVM-Loc were used to predict the subcellular localization of the identified proteins. Among 78 identified proteins, 20 were targeted as secretory pathway proteins and 36 were predicted to be in cellular compartments including cytoplasm, nucleus, and cell membrane. The largest subcellular fraction was the secretory pathway, accounting for 25% of total proteins. Gene Ontology (GO) of Citrus sinensis was used to simplify the functional annotation of the proteins that were identified in the leaves. The Kyoto Encyclopedia of Genes and Genomes (KEGG) showed the enrichment of metabolic pathways including glutathione metabolism and biosynthesis of secondary metabolites, suggesting that the response to a range of environmental factors is the key processes in citrus leaves. Finally, the Lipase GDSL domain-containing protein GDSL esterase/lipase, which is involved in plant development and defense response, was for the first time identified and characterized in Citrus aurantium.

Evanescent Wave Approximation for Non-Hermitian Hamiltonians.

The counterpart of the rotating wave approximation for non-Hermitian Hamiltonians is considered, which allows for the derivation of a suitable effective Hamiltonian for systems with some states undergoing decay. In the limit of very high decay rates, on the basis of this effective description we can predict the occurrence of a quantum Zeno dynamics, which is interpreted as the removal of some coupling terms and the vanishing of an operatorial pseudo-Lamb shift.

Bounds on Mixed State Entanglement.

In the general framework of d 1 × d 2 mixed states, we derive an explicit bound for bipartite negative partial transpose (NPT) entanglement based on the mixedness characterization of the physical system. The derived result is very general, being based only on the assumption of finite dimensionality. In addition, it turns out to be of experimental interest since some purity-measuring protocols are known. Exploiting the bound in the particular case of thermal entanglement, a way to connect thermodynamic features to the monogamy of quantum correlations is suggested, and some recent results on the subject are given a physically clear explanation.

Complexation of As(III) by phosphonate ligands in aqueous fluids: Thermodynamic behavior, chemical binding forms and sequestering abilities.

In recent years, the contamination of water by arsenic reached alarming levels in many countries of the world, attracting the interest of many researchers engaged in testing methodologies able to remove this harmful pollutant. An important aspect that must be taken into consideration is the possibility to find arsenic in different chemical forms which could require different approaches for its removal. At this aim, a speciation analysis appears to be crucial for better understanding the behavior of arsenic species in aqueous solutions, especially in presence of compounds with marked chelating properties. Phosphonates can be identified as good sequestering agents and, at this purpose, this manuscript intends to investigate the interaction of As(III) with three phosphonic acids derived from nitrilotriacetic acid (NTA) by replacements of one (N-(Phosphonomethyl) iminodiacetic acid, NTAP), two (N,N-Bis-(phosphonomethyl) glycine, NTA2P) and three (Nitrilotri(methylphosphonic acid), NTA3P) carboxylic groups with the same number of phosphonate groups. An in-depth potentiometric and calorimetric investigation allowed to determine speciation models featured by simple ML, MLHi and ML(OH) species. A complete thermodynamic characterization of the systems is reported together with the definition of coordination mode by mass spectrometry measurements. On the light of the speciation models, the possibility of using these ligands in arsenic removal techniques was assessed by determining the pL0.5 (the concentration of ligand able to remove the 50% of metal ion present in trace). All ligands show a good sequestering ability, in particular under the conditions of fresh water, following the trend NTA3P > NTA2P > NTAP.

High-throughput determination of vitamin E in extra virgin olive oil by paper spray tandem mass spectrometry.

Extra virgin olive oil provides an important intake of α-tocopherol, which is part of vitamin E complex. A fast analytical method for its quantification, based on paper spray mass spectrometry, has been developed. The methodology possesses the ability to record mass spectra without sample preparation or preseparation steps. The experiments were performed in Multiple Reaction Monitoring scan mode; in particular, the transitions m/z 429 → m/z 163 for α-tocopherol and m/z 435 → m/z 169 for the labeled internal standard were monitored, in order to obtain the greatest specificity and the best sensitivity. The accuracy of the method was tested analyzing spiked samples prepared at concentrations within the dynamic range of the calibration curve, which returned values near 100%. Furthermore, good values of LOQ and LOD were obtained, demonstrating that this approach can be applied for a rapid screening of tocopherols in different vegetable oils. The results were compared with analyses performed by traditional chromatographic methods. Graphical abstract.

A comprehensive evaluation of tyrosol and hydroxytyrosol derivatives in extra virgin olive oil by microwave-assisted hydrolysis and HPLC-MS/MS.

A rapid and reliable method to assay the total amount of tyrosol and hydroxytyrosol derivatives in extra virgin olive oil has been developed. The methodology intends to establish the nutritional quality of this edible oil addressing recent international health claim legislations (the European Commission Regulation No. 432/2012) and changing the classification of extra virgin olive oil to the status of nutraceutical. The method is based on the use of high-performance liquid chromatography coupled with tandem mass spectrometry and labeled internal standards preceded by a fast hydrolysis reaction step performed through the aid of microwaves under acid conditions. The overall process is particularly time saving, much shorter than any methodology previously reported. The developed approach represents a mix of rapidity and accuracy whose values have been found near 100% on different fortified vegetable oils, while the RSD% values, calculated from repeatability and reproducibility experiments, are in all cases under 7%. Graphical abstract Schematic of the methodology applied to the determination of tyrosol and hydroxytyrosol ester conjugates.

Electrospray ion mobility mass spectrometry of positively and negatively charged (1R,2S)-dodecyl(2-hydroxy-1-methyl-2-phenylethyl)dimethylammonium bromide aggregates.

RATIONALE: Self-assembling processes of surfactants in the gas phase constitute a developing research field of interest since they allow information to be gained on the peculiar structural organization of these aggregates, on their ability to incorporate from small molecules up to proteins and on their possible use as carriers of drugs in the gas phase or as cleaning agents and exotic reaction media. METHODS: The mass spectra of charged aggregates of the chiral surfactant (1R,2S)-dodecyl(2-hydroxy-1-methyl-2-phenylethyl)dimethylammonium bromide (DMEB) in the gas phase have been recorded using a Synapt G2-Si mass spectrometer in the positive and negative ion mode. For comparison purposes, the mass spectra of sodium bis(2-ethylhexyl)sulfosuccinate and sodium octane sulfonate aggregates have also been recorded under the same experimental conditions. The collisional cross sections of positively and negatively charged DMEB aggregates were obtained through an appropriate calibration of the measured drift times. RESULTS: For all the surfactants investigated, it has been found that there is a lowest and a highest limit of the aggregation number at each charge state: no aggregates are found outside this range. Moreover, the occurrence at each aggregation number and extra charge of a unique value of drift time points toward aggregates whose conformations do not show discernible shape change in the experiment time scale. The analysis of the collisional cross sections emphasizes that the DMEB aggregates are nearly spherical clusters somewhat affected by the charge state and constituted by interlaced polar and apolar domains. CONCLUSIONS: The analysis of all the experimental findings indicates that in the gas phase DMEB forms supramolecular aggregates characterized by an internal organization whose stability is triggered by the charge state. The comparison of the behavior of DMEB aggregates with that of sodium bis(2-ethylhexyl)sulfosuccinate and sodium octane sulfonate aggregates allows us to highlight the effects on the aggregate organization in gas phase due to nature of the head group and alkyl chain steric hindrance.

Fast analysis of caffeine in beverages and drugs by paper spray tandem mass spectrometry.

A simple and fast method based on paper spray mass spectrometry for the determination of caffeine in commercial beverages and drugs has been developed; the analyses were carried out in MRM mode, monitoring the transitions m/z 195 → m/z 138 for caffeine and m/z 198 → m/z 140 for the labeled internal standard. To verify the reliability of the proposed approach, a spiked sample (soda drink and paracetamol tablet) with a known amount of caffeine has been prepared and analyzed by PS-MS, providing accuracy values about 100 %; the LOQ and LOD values were calculated at 1.2 and 1.6 µg/mL, respectively. Both beverages and drugs were also analyzed with the classic analytical method based on LC-UV measurements, showing consistent results between the two approaches, thus confirming the reliability of the developed ambient MS determination. Graphical Abstract The assay of caffeine by paper mass spectrometry.

Clusterin transcript variants expression in thyroid tumor: a potential marker of malignancy?

BACKGROUND: Clusterin (CLU) is a ubiquitous multifunctional factor involved in neoplastic transformation. The CLU transcript variants and protein forms play a crucial role in balancing cells proliferation and death. METHODS: We investigated the regulation of CLU transcript variants expression in an in vivo model system consisting of both neoplastic tissues and fine needle aspiration biopsy (FNAB) samples isolated from patients undergoing thyroidectomy. RESULTS: The immunohistochemical analyses showed an overall CLU up-regulation in papillary carcinoma. A specific CLU2 transcript variant increase was registered using qPCR in papillary carcinomas while CLU1 decreased. In addition, the analysis of CLU transcripts expression level showed an increase of the CLU2 transcript in the TIR 3 patients with histologically confirmed thyroid cancer. CONCLUSIONS: Our results suggest the existence of a specific alteration of CLU2:CLU1 ratio towards CLU2, thus providing the first circumstantial evidence for the potential use of CLU transcript variants as effective biomarkers for a more accurate assessment of the so called "indeterminate" thyroid nodules.

Determination of ketosteroid hormones in meat by liquid chromatography tandem mass spectrometry and derivatization chemistry.

A method for the determination and quantification of ketosteroid hormones in meat by mass spectrometry, based on the derivatization of the carbonyl moiety of steroids by O-methylhydroxylamine, is presented. The quantitative assay is performed by means of multiple-reaction-monitoring (MRM) scan mode and using the corresponding labelled species, obtained by reaction with d 3-methoxylamine, as internal standard. The accuracy of the method was established by evaluating artificially spiked samples, obtaining values in the range 90-110%. Recovery tests were performed on blank matrix samples spiked with non-natural steroids including trenbolone and melengestrol acetate. The latter experiment revealed that the yield of the extraction processes was approximately 60%. Good values of LOQ and LOD were achieved, making this method competitive with current hormone assay methods.

Mass spectrometry-based proteomic approach in Oenococcus oeni enological starter.

A simple procedure is proposed for selective protein solubilization and trypsin digestion, followed by off-line liquid chromatography-matrix assisted laser desorption ionization mass spectrometry (LC-MALDI MS) analysis of Oenococcus oeni (O. oeni) bacterium. Peptides were identified from tryptic digests using sequencing by tandem mass spectrometry and database searches. Cytoplasmic and membrane related proteins (MRP) were identified in the O. oeni bacterium. MS/MS data analysis points out 13 peptides having one point mutation from 9 proteins. The major microheterogeneity was found for Zn-dependent alcohol dehydrogenase (Zn-ADH, Q04GE6) and 60 kDa chaperonin (GroEL, Q04E64) that are involved in methionine catabolism and post-translational protein folding, respectively. MS/MS data processing also leads to the identification of 34 unique phosphorylation sites from 19 phosphoproteins.