Design and Validation of a Custom NGS Panel Targeting a Set of Lysosomal Storage Diseases Candidate for NBS Applications

Lysosomal storage diseases (LSDs) are a heterogeneous group of approximately 70 monogenic metabolic disorders whose diagnosis represents an arduous challenge for clinicians due to their variability in phenotype penetrance, clinical manifestations, and high allelic heterogeneity. In recent years, the approval of disease-specific therapies and the rapid emergence of novel rapid diagnostic methods has opened, for a set of selected LSDs, the possibility for inclusion in extensive national newborn screening (NBS) programs. Herein, we evaluated the clinical utility and diagnostic validity of a targeted next-generation sequencing (tNGS) panel (called NBS_LSDs), designed ad hoc to scan the coding regions of six genes (GBA, GAA, SMPD1, IDUA1, GLA, GALC) relevant for a group of LSDs candidate for inclusion in national NBS programs (MPSI, Pompe, Fabry, Krabbe, Niemann Pick A-B and Gaucher diseases). A standard group of 15 samples with previously known genetic mutations was used to test and validate the entire flowchart. Analytical accuracy, sensitivity, and specificity, as well as turnaround time and costs, were assessed. Results showed that the Ion AmpliSeq and Ion Chef System-based high-throughput NBS_LSDs tNGS panel is a fast, accurate, and cost-effective process. The introduction of this technology into routine NBS procedures as a second-tier test along with primary biochemical assays will allow facilitating the identification and management of selected LSDs and reducing diagnostic delay.     

Synaptic Alterations in a Transgenic Model of Tuberous Sclerosis Complex: Relevance to Autism Spectrum Disorders

Tuberous sclerosis complex (TSC) is a rare, multi-system genetic disease with serious neurological and mental symptoms, including autism. Mutations in the TSC1/TSC2 genes lead to the overactivation of mTOR signalling, which is also linked to nonsyndromic autism. Our aim was to analyse synaptic pathology in a transgenic model of TSC: two-month-old male B6;129S4-Tsc2^tm1Djk/J mice with Tsc2 haploinsufficiency. Significant brain-region-dependent alterations in the expression of several synaptic proteins were identified. The most prominent changes were observed in the immunoreactivity of presynaptic VAMP1/2 (ca. 50% increase) and phospho-synapsin-1 (Ser62/67) (ca. 80% increase). Transmission electron microscopy demonstrated serious ultrastructural abnormalities in synapses such as a blurred structure of synaptic density and a significantly increased number of synaptic vesicles. The impairment of synaptic mitochondrial ultrastructure was represented by excessive elongation, swelling, and blurred crista contours. Polyribosomes in the cytoplasm and swollen Golgi apparatus suggest possible impairment of protein metabolism. Moreover, the delamination of myelin and the presence of vacuolar structures in the cell nucleus were observed. We also report that Tsc2^+/− mice displayed increased brain weights and sizes. The behavioural analysis demonstrated the impairment of memory function, as established in the novel object recognition test. To summarise, our data indicate serious synaptic impairment in the brains of male Tsc2^+/− mice.     

Volatile Compound and Sensory Analysis for the Characterization of an Italian White Wine from “Inzolia” Grapes

The research evidences the importance of linking chemical and sensory data for the wine characterization; in particular, the Italian white wine made from Inzolia grapes has been considered. Inzolia is one of the most widespread native white grapes in Sicily (Italy), and wine samples from two different areas (Monreale and Sambuca di Sicilia) have been analyzed. A headspace solid-phase microextraction/gas chromatography–mass spectroscopy method has been developed, and 56 volatile components, esters, fatty acids, alcohols, and terpenes have been identified; the method allows also the quantification of the main components, namely, ethyl octanoate (banana, fruit, fat) (257.2–541.6 mg/l) and ethyl decanoate (fruity, oily, floral; 171.8–272.0 mg/l). A good repeatability in terms of retention times and peak areas resulted. Sensory analysis was performed by ten trained judges that evaluated eight attributes: two referring to appearance (yellow color and yellow reflex), four referring to aroma (fruity, banana, ripened apple, and floral), and two referring to oral perception (acid and pungent). The volatile constituents and the aroma sensory attributes were in agreement.     

Relationship between interferon-gamma, interleukin-4 and IgE production of lymphocytes from hen's egg-sensitive patients

We measured in vitro interferon-gamma, interleukin-4 and IgE production of peripheral blood mononuclear cells stimulated with ovalbumin. Interferon-gamma in culture supernatants of ovalbumin-stimulated peripheral blood mononuclear cells from hen's egg- sensitive patients with atopic dermatitis was significantly higher than that of healthy children or hen's egg-sensitive patients with immediate symptoms. Furthermore, in patients with atopic dermatitis who were sensitive to hen's egg and patients with immediate symptoms, there was an inverse relationship between interferon-gamma and IgE production (r = -0.535, p <0.05), and significant correlation was found between interleukin-4 and IgE production (r = 0.802, p <0.05). For the above reasons, IgE synthesis of ovalbumin-stimulated peripheral blood mononuclear cells may be suppressed by interferon-gamma in patients with atopic dermatitis. In contrast, in patients with immediate symptoms, interleukin-4 may play a major role in the pathogenesis of increased IgE production.     

Thermal performance analysis on a two composite material honeycomb heat regenerators used for HiTAC burners

Honeycomb heat regenerators do not only reduce the fuel consumption in a high temperature air combustion (HiTAC) burning system but also provide the necessary high temperature of combustion air. A two-dimensional simulation model was developed to numerically determine the dynamic temperature and velocity profiles of gases and solid heat-storing materials in a composite material honeycomb regenerator. Consequently, the energy storage and the pressure drop are calculated and the thermal performance of honeycomb heat regenerator is evaluated at different switching times and loading. The model takes into account the thermal conductivity parallel and perpendicular to flow direction of solid and flowing gases. It considers the variation of all thermal properties of solid material and gases with temperature. Moreover, the radiation from combustion flue gases to the storage materials was considered in the analysis. The results are presented in a non-dimensional form in order to be a design tool as well. These analyses were applied on a regenerator made of two layers of ceramic materials, one is pure alumina and other is cordierite. This regenerator is contained in a 100 kW twin-type regenerative-burning system used for HiTAC. The effectiveness and the energy recovery rate were 88% and 72% respectively at nominal operating range of the regenerator and the pressure drop across the twin regenerator system was 1.16 kPa. The periodic steady state condition is reached after about 11 min and it takes only 2 min of operation until the temperature of combustion air remains above the self-ignition temperature that is required for HiTAC. Furthermore, these mathematical analyses show good agreement with experiments made on the same regenerator. In the experiments, the dynamic behavior of the heat regenerator operation was considered in order to compensate measurement readings for this effect.     

Preparation of Pectin-Based Biosorbents for Cadmium and Lead Ions Removal

The optimal formation conditions of pectin-based, as well as new hybrid pectin-guar gum, biosorbents were found. The lead affinity for obtained biosorbents was very strong even in solution of pH = 1, in opposite to cadmium, which was adsorbed from solutions of pH ≥ 2. The sorption capacities in the studied conditions were about 0.6 and 0.8 mmol/g for cadmium and lead, respectively. The cadmium removal could be fast and it is almost quantitatively desorbed using 0.25 M HNO₃, while desorption of lead required 1 M HNO₃ to achieve a removal efficiency of ca. 75% from pectin-based beads. The possibility of biosorbents’ reuse was also proved. Addition of guar gum to pectin biosorbent did not change its sorption properties significantly.     

Tracking residual organochlorine pesticides (OCPs) in green,herbal, and black tea leaves and infusions of commercially available tea products marketed in Poland

The content of residual organochlorine pesticides (OCPs) was examined in green, herbal, and black tea leaves as well as in their infusions prepared from tea products marketed in the main supermarkets in Poland. It was found that the detected mean levels of organochlorine residues in tea leaves ranged from ?1 dry weight. Among hexachlorocyclohexane isomers, γ-HCH in green tea occurred in the highest concentrations. Among dichlorodiphenyltrichloroethane (DDT) metabolites the highest level of p,p′DDT (1.96 ng g^?1 dw) was in green tea samples. The transfer of OCPs from tea leaves to brew was investigated. The present study revealed that during the infusion process, a significant percentage of the residues, particularly pesticides with high water solubility, were transferred to the infusions. The obtained results show that the percentage transfer of each pesticides from tea to the tea infusions ranged from 6.74% (heptachlor) to 86.6% (endrin). The detected residues were below current MRLs for these pesticides.     

Slaughter value and flesh characteristics of European catfish (Silurus glanis) fed natural and formulated feed under different rearing conditions

The experimental material was comprised of European catfish (Silurus glanis) individuals aged 2.5 years that were reared on natural feed in earthen ponds and individuals aged 1.5 years cultivated intensively on formulated feed in recirculating systems. Weight gain and morphometric parameters were measured along with physical and color characteristics and proximate composition of fillets. Despite the different age and diets of the fish, no differences were observed in the linear biometric indices. The total weight of the fish of both groups and their condition coefficients were also similar. In addition, no differences were reported in their body weight or head and gutted (H&G) carcass and fillet yields. Proximate composition analysis demonstrated a lack of differences in the contents of dry matter. Of the dry matter, differences were reported in the fat content, which was higher in the catfish fed formulated feed. The color of the fillets was characterized by high values of lightness, negative values of redness, and positive values of yellowness, and the fish fed formulated feed displayed higher saturation of green. The flesh of both groups of catfish demonstrated similar pH values and water retention capacity, whereas the values of thermal drip appeared to be higher in the fish fed formulated feed.     

Kinetics of extracted bioactive components from mushrooms in artificial digestive juices

This article presents a study on fruiting bodies of Boletus badius, Boletus edulis, Cantharellus cibarius, Lactarius deliciosus, Leccinum scabrum, Suillus bovinus, Suillus luteus, Pleurotus ostreatus, Tricholoma equestre, Armillaria mellea, Agaricus bisporus, and Auricularia polytricha. The amounts of their health-promoting bioelements and physiologically significant indole and phenolic compounds were determined. The significance of edible mushrooms in the human diet and in disease prevention were also established. The high-performance liquid chromatography was used to assess the contents of indole compounds and phenolic acids. The atomic absorption spectroscopy was applied to determine bioelements. In all samples, the content of 5-hydroxy-L-tryptophan was studied. The highest extraction amount of serotonin was found for P. ostreatus species. Phenolic compounds were determined in the conditions described in the present study for the first time. Additionally, protocatechuic, p-hydroxybenzoic, syringic, and gallic acids were determined.