Antimicrobial and antioxidant activity of encapsulated tea polyphenols in chitosan/alginate-coated zein nanoparticles: a possible supplement against fish pathogens in aquaculture.

Regulation of antibiotic use in aquaculture calls for the emergence of more sustainable alternative treatments. Tea polyphenols (GTE), particularly epigallocatechin gallate (EGCG), have various biological activities. However, tea polyphenols are susceptible to degradation. In this work, EGCG and GTE were encapsulated in zein nanoparticles (ZNP) stabilized with alginate (ALG) and chitosan (CS) to reduce the degradation effect. ALG-coated ZNP and ALG/CS-coated ZNP encapsulating EGCG or GTE were obtained with a hydrodynamic size of less than 300 nm, an absolute ζ-potential value >30 mV, and an encapsulation efficiency greater than 75%. The antioxidant capacity of the encapsulated substances, although lower than that of the free ones, maintained high levels. On the other hand, the evaluation of antimicrobial activity showed greater efficiency in terms of growth inhibition for ALG/CS-ZNP formulations, with average overall values of around 60%, reaching an inhibition of more than 90% for Photobacterium damselae. These results support encapsulation as a good strategy for tea polyphenols, as it allows maintaining significant levels of antioxidant activity and increasing the potential for antimicrobial activity, in addition to increasing protection against sources of degradation.

[First Hungarian report of Geotrichum capitatum/Saprochaete capitata infection in an immunocompromised child]. / Geotrichum capitatum/Saprochaete capitata fertozés elso hazai esete egy immunszupprimált gyermekben.

Patients with leukemia may occasionally suffer from rare opportunistic fungal infections with poor prognosis. Fungal infection caused by Geotrichum captitatum has not yet been described in Hungary. With this case report, we would like to draw attention to the fungal infection caused by G. capitatum. The 1.5-year-old girl with acute myeloid leukemia was treated for relapse diagnosed +120 days after a sibling donor bone marrow transplantation. High-grade, fluctuating fever began 11 days after the start of chemotherapy which did not decrease despite combined treatment with broad-spectrum antibiotics and antifungals (posaconasole). Due to worsening respiratory symptoms, a chest CT-scan was performed, raising suspicion of an invasive fungal infection. Blood culture confirmed G. capitatum infection. Initial empiric treatment with liposomal amphotericin B was combined with voriconazole based on international experience. However, we did not observe any improvement, and a few days later the patient passed away due to progression of the underlying disease. G. capitatum (presently known as Saprochaete capitata) is an ubiquitous yeast that can cause an infection with a poor prognosis, mainly in patients with leukemia. Its symptoms primarily appear in the skin and respiratory tract. The accurate identification of this pathogen is essential because the standard diagnostic tests do not give a specific reaction. Based on the limited international experience, the combination of amphotericin B and voriconazole can play a fundamental role in the treatment, however, even with adequate therapy 50% of the cases are fatal. By describing the first Hungarian case caused by G. capitatum, we draw attention to the importance of this rare, opportunistic fungal species with a poor prognosis that develops in immunosuppressed patients. Orv Hetil. 2023; 164(26): 1034-1038.

Phosphonate Derivatives of Paracetamol and Valproic Acid.

Paracetamol and valproic acid are standalone drugs with leading position in the world drug market. The phosphonate analogues of these drugs were synthesized and were tested in vivo. N-(4-hydroxyphenylcarbamoyl)phosphonic acid was four times more potent than paracetamol in preventing acetic acid-induced writhing. Phosphonate derivative of valproic acid, (2-propylpentanoyl)phosphonic acid, had similar in vivo activity to valproic acid in the pentylenetetrazole-induced kindling mouse model.

Evaluation of In Vitro Protein Hydrolysis in Seven Insects Approved by the EU for Use as a Protein Alternative in Aquaculture.

Rapid population growth is leading to an increase in the demand for high-quality protein such as fish, which has led to a large increase in aquaculture. However, fish feed is dependent on fishmeal. It is necessary to explore more sustainable protein alternatives that can meet the needs of fish. Insects, due to their high protein content and good amino acid profiles, could be a successful alternative to fishmeal and soybean meal traditionally used in sectors such as aquaculture. In this work, seven species of insects (Hermetia illucens, Tenebrio molitor, Acheta domestica, Alphitobius diaperinus, Gryllodes sigillatus, Gryllus assimilis, and Musca domestica) approved by the European Union (UE) for use as feed for farmed animals (aquaculture, poultry, and pigs) were studied. Their proximate composition, hydrolysis of organic matter (OMd), hydrolysis of crude protein (CPd), degree of hydrolysis (DH/NH2 and DH/100 g DM), and total hydrolysis (TH) were analyzed. The results showed that Tenebrio molitor had digestibility similar to that of fishmeal, while Acheta domestica and Hermetia illucens provided similar digestibility to that of soybean meal. The acid detergent fiber (ADF) data were negatively correlated with all protein digestibility variables. The differences in the degree of hydrolysis (DH) results and the similarity in total hydrolysis (TH) results could indicate the slowing effects of ADF on protein digestibility. Further in vivo studies are needed.

Pre-Treatment of Fish By-Products to Optimize Feeding of Tenebrio molitor L. Larvae.

Fish discards are organic waste with high and good-quality protein levels, as well as a fatty acid profile rich in n-3 LCPUFAs, mainly eicosapentaenoic acid and docosahexaenoic acid. These discards can be used as food for Tenebrio molitor (Linnaeus, 1758) larvae, thus increasing the nutritional value of this insect. This study focused on increasing larval acceptance of fish through different pre-treatments of the diets provided, as well as increasing the accumulation of EPA and DHA in fish-fed larvae. Four different diets were prepared: control (broiler feed), DGF50: 50% dried ground fish (Pagellus bogaraveo, Brünnich, 1768) + 50% broiler feed, for different periods, FGF100: 100% fresh ground P. bogaraveo and DUF100: 100% dried whole unground P. bogaraveo. Growth, mortality, proximate composition, fatty acid profile and lipid nutritional indices were determined. Larvae fed with FGF100 displayed better results among treatments, doubling the initial weight, as well as increasing their protein level and decreasing fat levels. Regarding fatty acids, eicosapentaenoic acid and docosahexaenoic acid were only detected in larvae fed with a fish-based diet for a period longer than 5 days. These results show that pre-treatment of fish-based diets causes changes in the growth and compositional parameters of T. molitor larvae.

Personalized First-Line Treatment of Metastatic Pancreatic Neuroendocrine Carcinoma Facilitated by Liquid Biopsy and Computational Decision Support.

BACKGROUND: We present the case of a 50-year-old female whose metastatic pancreatic neuroendocrine tumor (pNET) diagnosis was delayed by the COVID-19 pandemic. The patient was in critical condition at the time of diagnosis due to the extensive tumor burden and failing liver functions. The clinical dilemma was to choose between two registered first-line molecularly-targeted agents (MTAs), sunitinib or everolimus, or to use chemotherapy to quickly reduce tumor burden. METHODS: Cell-free DNA (cfDNA) from liquid biopsy was analyzed by next generation sequencing (NGS) using a comprehensive 591-gene panel. Next, a computational method, digital drug-assignment (DDA) was deployed for rapid clinical decision support. RESULTS: NGS analysis identified 38 genetic alterations. DDA identified 6 potential drivers, 24 targets, and 79 MTAs. Everolimus was chosen for first-line therapy based on supporting molecular evidence and the highest DDA ranking among therapies registered in this tumor type. The patient's general condition and liver functions rapidly improved, and CT control revealed partial response in the lymph nodes and stable disease elsewhere. CONCLUSION: Deployment of precision oncology using liquid biopsy, comprehensive molecular profiling, and DDA make personalized first-line therapy of advanced pNET feasible in clinical settings.

Effect of bacterial inoculation on co-composting of lavender (Lavandula angustifolia Mill.) waste and cattle manure.

The primary purpose of this study was to investigate the influence of Cellulomonas flavigena and Streptomyces viridosporus, as a bacterial inoculant, on the compostability of post-extraction lavender waste. The major physicochemical, microbiological, and biological properties of the composting materials were monitored for 161 days. The technology developed was shown to improve the compostability of recalcitrant herbal residues. The use of lavender waste beneficially affected the composting process by extending the thermophilic phase, accelerating the degradation of organic matter, and elevating the viable counts of useful microorganisms; however, adverse effects were also observed, including an increased carbon-to-nitrogen ratio (19.05) and a decreased germination index (93.4%). Bacterial inoculation was found to preserve the nitrogen content (2.50%) and improve the efficiency of biodegradation. The Salmonella- and Escherichia coli-free final composting products were mature, stable, and ready for soil application. To the authors' knowledge, no previous research has investigated the compostability of lavender waste. Likewise, this is the first study that has used strains of C. flavigena and S. viridosporus in combination to facilitate a composting process.

The Warburg Micro Syndrome-associated Rab3GAP-Rab18 module promotes autolysosome maturation through the Vps34 Complex I.

Warburg micro syndrome (WMS) is a hereditary autosomal neuromuscular disorder in humans caused by mutations in Rab18, Rab3GAP1, or Rab3GAP2 genes. Rab3GAP1/2 forms a heterodimeric complex, which acts as a guanosine nucleotide exchange factor and activates Rab18. Although the genetic causes of WMS are known, it is still unclear whether loss of the Rab3GAP-Rab18 module affects neuronal or muscle cell physiology or both, and how. In this work, we characterize a Rab3GAP2 mutant Drosophila line to establish a novel animal model for WMS. Similarly to symptoms of WMS, loss of Rab3GAP2 leads to highly decreased motility in Drosophila that becomes more serious with age. We demonstrate that these mutant flies are defective for autophagic degradation in multiple tissues including fat cells and muscles. Loss of Rab3GAP-Rab18 module members leads to perturbed autolysosome morphology due to destabilization of Rab7-positive autophagosomal and late endosomal compartments and perturbation of lysosomal biosynthetic transport. Importantly, overexpression of UVRAG or loss of Atg14, two alternative subunits of the Vps34/PI3K (vacuole protein sorting 34/phosphatidylinositol 3-kinase) complexes in fat cells, mimics the autophagic phenotype of Rab3GAP-Rab18 module loss. We find that GTP-bound Rab18 binds to Atg6/Beclin1, a permanent subunit of Vps34 complexes. Finally, we show that Rab3GAP2 and Rab18 are present on autophagosomal and autolysosomal membranes and colocalize with Vps34 Complex I subunits. Our data suggest that the Rab3GAP-Rab18 module regulates autolysosomal maturation through its interaction with the Vps34 Complex I, and perturbed autophagy due to loss of the Rab3GAP-Rab18 module may contribute to the development of WMS.

Degradation of arouser by endosomal microautophagy is essential for adaptation to starvation in Drosophila.

Hunger drives food-seeking behaviour and controls adaptation of organisms to nutrient availability and energy stores. Lipids constitute an essential source of energy in the cell that can be mobilised during fasting by autophagy. Selective degradation of proteins by autophagy is made possible essentially by the presence of LIR and KFERQ-like motifs. Using in silico screening of Drosophila proteins that contain KFERQ-like motifs, we identified and characterized the adaptor protein Arouser, which functions to regulate fat storage and mobilisation and is essential during periods of food deprivation. We show that hypomorphic arouser mutants are not satiated, are more sensitive to food deprivation, and are more aggressive, suggesting an essential role for Arouser in the coordination of metabolism and food-related behaviour. Our analysis shows that Arouser functions in the fat body through nutrient-related signalling pathways and is degraded by endosomal microautophagy. Arouser degradation occurs during feeding conditions, whereas its stabilisation during non-feeding periods is essential for resistance to starvation and survival. In summary, our data describe a novel role for endosomal microautophagy in energy homeostasis, by the degradation of the signalling regulatory protein Arouser.

Measuring sociability of mice using a novel three-chamber apparatus and algorithm of the LABORAS™ system.

BACKGROUND: The social approach and social novelty tests utilizing the three-chamber apparatus are widely accepted to measure social behavior of rodents. The LABORAS™ system offers a possibility to assess sociability of mice in a reliable and objective manner. NEW METHOD: We assessed the capability of the LABORAS™ sociability cage and algorithm (2.6.6) to detect social behaviors in mice. Furthermore, we investigated whether the system is able to detect various levels of sociability due to genetic background or after pharmacological treatments. RESULTS: By comparing manual scoring with various detection zone settings of the automated registration, the most fitting algorithm with a detection zone radius of 90 mm was identified. When different strains were investigated, C57Bl/6 J and NMRI mice proved to be social, while CD1 mice were found asocial. The system was able to detect the sociability increasing effect of R-baclofen (0.5 mg/kg i.p.) and oxytocin (12 ng i.c.v.) in asocial CD1 mice. The negative control PCP impaired social behavior of C57Bl/6 J mice (1 mg/kg i.p.) and increased social avoidance in CD1 mice (0.3 mg/kg i.p.). COMPARISON WITH EXISTING METHOD(S): This setup, in contrast to video frame analysis softwares, determines signal changes caused by movements of rodents allowing accurate detection and analysis of trajectories. Parallel automated measurements also allow replacing time and labor intensive, highly subjective human observational work. CONCLUSIONS: The set-up provides a fast and reliable method to examine social behavior of mice in the three-chamber apparatus. The system is capable of detecting pro or antisocial activity of pharmacological agents.

[Treatment of Theory of Mind deficits in schizophrenia by using Virtual Reality, the VR-ToMIS training (Virtual Reality based Theory of Mind Intervention in Schizophrenia)].

Rapid development in information technology has been observed recently and has led to valuable developments also in healthcare. 3D-bio-printing or the virtual simulations that help the acquisition of anatomical and pathological knowledge and testing the acquired knowledge are just some of the examples. This progress can be recognized also in psychiatry. One of the most spectacular ways of using these technologies in psychiatry might be the therapeutic techniques associated with Virtual Reality (VR) simulations, which are currently available for anxiety disorders, eating disorders and addictions. A research team of the Psychiatric and Psychotherapeutic Department of the Semmelweis University has developed a Virtual Reality-based intervention that fits in with this perspective. The intervention mainly aims at Theory of Mind deficit and pragmatic language impairment in schizophrenia. In this article the current status of our research team's work will be presented. The article reviews the literature that provides the basis for the development, leads the reader through the main stages of the development process, and finally the program itself will be introduced. Process and mechanism of change associated with the intervention and the potential risks of the use of VR will also be discussed.

Vps8 overexpression inhibits HOPS-dependent trafficking routes by outcompeting Vps41/Lt.

Two related multisubunit tethering complexes promote endolysosomal trafficking in all eukaryotes: Rab5-binding CORVET that was suggested to transform into Rab7-binding HOPS. We have previously identified miniCORVET, containing Drosophila Vps8 and three shared core proteins, which are required for endosome maturation upstream of HOPS in highly endocytic cells (Lorincz et al., 2016a). Here, we show that Vps8 overexpression inhibits HOPS-dependent trafficking routes including late endosome maturation, autophagosome-lysosome fusion, crinophagy and lysosome-related organelle formation. Mechanistically, Vps8 overexpression abolishes the late endosomal localization of HOPS-specific Vps41/Lt and prevents HOPS assembly. Proper ratio of Vps8 to Vps41 is thus critical because Vps8 negatively regulates HOPS by outcompeting Vps41. Endosomal recruitment of miniCORVET- or HOPS-specific subunits requires proper complex assembly, and Vps8/miniCORVET is dispensable for autophagy, crinophagy and lysosomal biogenesis. These data together indicate the recruitment of these complexes to target membranes independent of each other in Drosophila, rather than their transformation during vesicle maturation.

Comorbidity of localized scleroderma and primary biliary cholangitis.

BACKGROUND AND OBJECTIVES: Morphea is frequently associated with other autoimmune disorders. Little is known about the association of morphea and primary biliary cholangitis (PBC). The objective of this case series was to study the possible association of morphea with PBC and to identify risk factors. PATIENTS AND METHODS: Patients with morphea were screened for anti-mitochondrial antibodies (AMA) by indirect immunofluorescence and/or immunoblot. Human leucocyte antigen (HLA) genotyping and deep sequencing for the HLA DRB1 subgroup were confirmed in AMA-positive patients. RESULTS: 6 of 91 patients (6.6 %) showed positivity for AMA, and 4 (4.39 %) had PBC. The mean age of AMA-positive patients was 69.0 years. Of 6 AMA-positive patients, common predisposing alleles (HLA DRB1*15: 01 and HLA DRB1*08) were detectable in two patients. One patient had predisposing alleles for both diseases (HLA DRB1*03: 01 and HLA DRB1*14). One patient had a PBC-conferring allele. Female gender, menopause and tendency for remission of morphea were common in all patients. CONCLUSION: The coexistence of morphea, AMA positivity and PBC is a rare but possible association. Common predisposing HLA alleles might interact in such a simultaneous manifestation. We suggest AMA screening for female patients with generalized morphea before the initiation of methotrexate.

Analysis of Complications After Button Battery Ingestion in Children.

AIM: Button battery ingestion (BBI) in children may cause severe complications. This analysis is a literature review of complications after pediatric BBI. METHODS: Literature was searched on PubMed (1995-2015) using the terms "button battery," "ingestion," and "children." End points were age, type and diameter of battery, complications, affected organ, and fatality. RESULTS: A total of 31 publications were analyzed. Patients from 4 months to 19 years old were included (n = 136,191, with n = 102,143 or 75% aged <6 y). In 6262, the diameter of the battery was documented. Batteries of 20 mm or greater in size were more prone to complications (n = 226). With regard to the anatomy, BBI caused complications mainly in the esophagus (n = 88, 38.94%). Sixty-one fatal outcomes were reported. CONCLUSIONS: Children younger than 6 years are the most prone to BBI, with lithium batteries of 20 mm or greater in size associated with complications. Complications have been estimated at 0.165%, with lethality of 0.04%. The esophagus is the most affected organ, but vascular involvement is often fatal.

MiniCORVET is a Vps8-containing early endosomal tether in Drosophila.

Yeast studies identified two heterohexameric tethering complexes, which consist of 4 shared (Vps11, Vps16, Vps18 and Vps33) and 2 specific subunits: Vps3 and Vps8 (CORVET) versus Vps39 and Vps41 (HOPS). CORVET is an early and HOPS is a late endosomal tether. The function of HOPS is well known in animal cells, while CORVET is poorly characterized. Here we show that Drosophila Vps8 is highly expressed in hemocytes and nephrocytes, and localizes to early endosomes despite the lack of a clear Vps3 homolog. We find that Vps8 forms a complex and acts together with Vps16A, Dor/Vps18 and Car/Vps33A, and loss of any of these proteins leads to fragmentation of endosomes. Surprisingly, Vps11 deletion causes enlargement of endosomes, similar to loss of the HOPS-specific subunits Vps39 and Lt/Vps41. We thus identify a 4 subunit-containing miniCORVET complex as an unconventional early endosomal tether in Drosophila.

A novel cell-based duplex high-throughput screening assay combining fluorescent Ca(2+) measurement with homogeneous time-resolved fluorescence technology.

Cell-based assays for G-protein-coupled receptor (GPCR) activation applied in high-throughput screening (HTS) monitor various readouts for second messengers or intracellular effectors. Recently, our understanding of diverging signaling pathways downstream of receptor activation and the capability of small molecules to selectively modulate signaling routes has increased substantially, underlining the importance of selecting appropriate readouts in cellular functional screens. To minimize the rate of false negatives in large-scale screening campaigns, it is crucial to maximize the chance of a ligand being detected, and generally applicable methods for detecting multiple analytes from a single well might serve this purpose. The few assays developed so far based on multiplexed GPCR readouts are limited to only certain applications and usually rely on genetic manipulations hindering screening in native or native-like cellular systems. Here we describe a more generally applicable and HTS-compatible homogeneous assay based on the combination of fluorometric detection of [Ca(2+)] with subsequent homogeneous time-resolved fluorescence (HTRF) cAMP readout in the same well. Besides describing development and validation of the assay, using a cell line recombinantly expressing the human PTH1 receptor screening of a small library is also presented, demonstrating the robustness and HTS compatibility of the novel paradigm.

Utility of different outcome measures for the nitroglycerin model of migraine in mice.

INTRODUCTION: Majority of the work for establishing nitroglycerin (NTG)-induced migraine models in animals was done in rats, though recently some studies in mice were also reported. Different special formulations of NTG were investigated in various studies; however, NTG treated groups were often compared to simple saline treated control groups. The aim of the present studies was to critically assess the utility of a panel of potential outcome measures in mice by revisiting previous findings and investigating endpoints that have not been tested in mice yet. METHODS: We investigated two NTG formulations, Nitrolingual and Nitro Pohl, at an intraperitoneal dose of 10mg/kg, in comparison with relevant vehicle controls, and evaluated the following outcome measures: light aversive behaviour, cranial blood perfusion by laser Doppler imaging, number of c-Fos- and neuronal nitrogen monoxide synthase (nNOS)-immunoreactive neurons in the trigeminal nucleus caudalis (TNC) and trigeminal ganglia, thermal hyperalgesia and tactile allodynia of the hind paw and orofacial pain hypersensitivity. RESULTS: We could not confirm previous reports of significant NTG-induced changes in light aversion and cranial blood perfusion of mice but we observed considerable effects elicited by the vehicle of Nitrolingual. In contrast, the vehicle of Nitro Pohl was apparently inert. Increased c-Fos expression in the TNC, thermal hyperalgesia, tactile allodynia and orofacial hypersensitivity were apparently good endpoints in mice that were increased by NTG-administration. The NTG-induced increase in c-Fos expression was prevented by topiramate but not by sumatriptan treatment. However, the NTG-induced orofacial hypersensitivity was dose dependently attenuated by sumatriptan. DISCUSSION: Our results pointed to utilisable NTG formulations and outcome measures for NTG-induced migraine models in mice. Pending further cross-validation with positive and negative control drugs in these mouse models and in the human NTG models of migraine, these tests might be valuable translational research tools for development of new anti-migraine drugs.

Loss of Drosophila Vps16A enhances autophagosome formation through reduced Tor activity.

The HOPS tethering complex facilitates autophagosome-lysosome fusion by binding to Syx17 (Syntaxin 17), the autophagosomal SNARE. Here we show that loss of the core HOPS complex subunit Vps16A enhances autophagosome formation and slows down Drosophila development. Mechanistically, Tor kinase is less active in Vps16A mutants likely due to impaired endocytic and biosynthetic transport to the lysosome, a site of its activation. Tor reactivation by overexpression of Rheb suppresses autophagosome formation and restores growth and developmental timing in these animals. Thus, Vps16A reduces autophagosome numbers both by indirectly restricting their formation rate and by directly promoting their clearance. In contrast, the loss of Syx17 blocks autophagic flux without affecting the induction step in Drosophila.

How and why to study autophagy in Drosophila: it's more than just a garbage chute.

During the catabolic process of autophagy, cytoplasmic material is transported to the lysosome for degradation and recycling. This way, autophagy contributes to the homeodynamic turnover of proteins, lipids, nucleic acids, glycogen, and even whole organelles. Autophagic activity is increased by adverse conditions such as nutrient limitation, growth factor withdrawal and oxidative stress, and it generally protects cells and organisms to promote their survival. Misregulation of autophagy is likely involved in numerous human pathologies including aging, cancer, infections and neurodegeneration, so its biomedical relevance explains the still growing interest in this field. Here we discuss the different microscopy-based, biochemical and genetic methods currently available to study autophagy in various tissues of the popular model Drosophila. We show examples for results obtained in different assays, explain how to interpret these with regard to autophagic activity, and how to find out which step of autophagy a given gene product is involved in.