Immunomodulatory Activity on Human Macrophages by Cell-Free Supernatants to Explore the Probiotic and Postbiotic Potential of Lactiplantibacillus plantarum Strains of Plant Origin.

Upon dietary administration, probiotic microorganisms can reach as live cells the human gut, where they interact with the microbiota and host cells, thereby exerting a beneficial impact on host functions, mainly through immune-modulatory activities. Recently, attention has been drawn by postbiotics, i.e. non-viable probiotic microbes, including their metabolic products, which possess biological activities that benefit the host. Lactiplantibacillus plantarum is a bacterial species that comprises recognised probiotic strains. In this study, we investigated in vitro the probiotic (and postbiotic) potential of seven L. plantarum strains, including five newly isolated from plant-related niches. The strains were shown to possess some basic probiotic attributes, including tolerance to the gastrointestinal environment, adhesion to the intestinal epithelium and safety. Besides, their cell-free culture supernatants modulated cytokine patterns in human macrophages in vitro, promoting TNF-α gene transcription and secretion, while attenuating the transcriptional activation and secretion of both TNF-α and IL-8 in response to a pro-inflammatory signal, and enhancing the production of IL-10. Some strains induced a high IL-10/IL-12 ratio that may correlate to an anti-inflammatory capacity in vivo. Overall, the investigated strains are good probiotic candidates, whose postbiotic fraction exhibits immunomodulatory properties that need further in vivo studies. The main novelty of this work consists in the polyphasic characterisation of candidate beneficial L. plantarum strains obtained from relatively atypical plant-associated niches, by an approach that explores both probiotic and postbiotic potentials, in particular studying the effect of microbial culture-conditioned media on cytokine pattern, analysed at both transcriptional and secretion level in human macrophages.

Antimicrobial Properties, Functional Characterisation and Application of Fructobacillus fructosus and Lactiplantibacillus plantarum Isolated from Artisanal Honey.

Honey is a valuable reservoir of lactic acid bacteria (LAB) and, particularly, of fructophilic LAB (FLAB), a relatively novel subgroup of LAB whose functional potential for human and food application has yet to be explored. In this study, FLAB and LAB strains have been isolated from honeys of different floral origins and selected for their broad antimicrobial activity against typical foodborne pathogenic bacteria and spoilage filamentous fungi. The best candidates, two strains belonging to the species Lactiplantibacillus plantarum and Fructobacillus fructosus, were submitted to partial characterisation of their cell free supernatants (CFS) in order to identify the secreted metabolites with antimicrobial activity. Besides, these strains were examined to assess some major functional features, including in vitro tolerance to the oro-gastrointestinal conditions, potential cytotoxicity against HT-29 cells, adhesion to human enterocyte-like cells and capability to stimulate macrophages. Moreover, when the tested strains were applied on table grapes artificially contaminated with pathogenic bacteria or filamentous fungi, they showed a good ability to antagonise the growth of undesired microbes, as well as to survive on the fruit surface at a concentration that is recommended to develop a probiotic effect. In conclusion, both LAB and FLAB honey-isolated strains characterised in this work exhibit functional properties that validate their potential use as biocontrol agents and for the design of novel functional foods. We reported antimicrobial activity, cytotoxic evaluation, probiotic properties and direct food application of a F. fructosus strain, improving the knowledge of this species, in particular, and on FLAB, more generally.

Monofunctionalized Fluorinated Bambusurils and Their Conjugates for Anion Transport and Extraction.

Bambusurils are macrocyclic molecules that are known for their high binding affinity and selectivity toward anions. Here, we present the preparation of two bambusurils bearing fluorinated substituents and one carboxylic function. These monofunctionalized bambusurils were conjugated with crown ether and cholesterol units. The resulting conjugates were successfully tested in liquid-liquid extraction of inorganic salts and chloride/bicarbonate transport across lipid bilayers.

A single change in the aptamer of the Lactiplantibacillus plantarum rib operon riboswitch severely impairs its regulatory activity and leads to a vitamin B2 - overproducing phenotype.

Manufacturing of probiotics and functional foods using lactic acid bacteria (LAB) that overproduce vitamin B2 has gained growing interest due to ariboflavinosis problems affecting populations of both developing and affluent countries. Two isogenic Lactiplantibacillus plantarum strains, namely a riboflavin-producing parental strain (UFG9) and a roseoflavin-resistant strain (B2) that carries a mutation in the FMN-aptamer of the potential rib operon riboswitch, were analysed for production and intra- and extracellular accumulation of flavins, as well as for regulation of the rib operon expression. Strain B2 accumulated in the medium one of the highest levels of riboflavin+FMN ever reported for LAB, exceeding by ~ 25 times those accumulated by UFG9. Inside the cells, concentration of FAD was similar in both strains, while that of riboflavin+FMN was ~ 8-fold higher in B2. Mutation B2 could decrease the stability of the aptamer's regulatory P1 helix even in the presence of the effector, thus promoting the antiterminator structure of the riboswitch ON state. Although the B2-mutant riboswitch showed an impaired regulatory activity, it retained partial functionality being still sensitive to the effector. The extraordinary capacity of strain B2 to produce riboflavin, together with its metabolic versatility and probiotic properties, can be exploited for manufacturing multifunctional foods.

Autochthonous Biological Resources for the Production of Regional Craft Beers: Exploring Possible Contributions of Cereals, Hops, Microbes, and Other Ingredients.

Selected biological resources used as raw materials in beer production are important drivers of innovation and segmentation in the dynamic market of craft beers. Among these resources, local/regional ingredients have several benefits, such as strengthening the connection with territories, enhancing the added value of the final products, and reducing supply costs and environmental impacts. It is assumed that specific ingredients provide differences in flavours, aromas, and, more generally, sensory attributes of the final products. In particular, of interest are ingredients with features attributable and/or linked to a specific geographical origin. This review encompasses the potential contribution and exploitation of biodiversity in the main classes of beer inputs, such as cereals, hops, microbes, and adjuncts, with a specific emphasis on autochthonous biological resources, detailing the innovative paths already explored and documented in the scientific literature. This dissertation proposes an overview of the impact on beer quality for each raw material category, highlighting the benefits and limitations that influence its concrete applications and scale-up, from the field to the stain. The topics explored promote, in the sector of craft beers, trends already capitalised in the production of other alcoholic beverages, such as the preservation and revalorisation of minor and autochthonous varieties, the exploitation of yeast and bacteria strains isolated from specific sites/plant varieties, and the valorisation of the effects of peculiar terroirs on the quality of agricultural products. Finally, the examined tendencies contribute toward reducing the environmental impacts of craft beer manufacturing, and are in line with sustainable development of food systems, increasing the economic driver of biodiversity preservation.

Transmembrane Transport of Bicarbonate Unravelled.

Invited for the cover of this issue are Dr. Stephen Butler, Dr. Hennie Valkenier and co-workers at Université Libre de Bruxelles, Loughborough University, Masaryk University, and the University of Bristol. The image depicts the transport of bicarbonate anions versus the spontaneous diffusion of CO2 across the lipid bilayer of a liposome. Read the full text of the article at 10.1002/chem.202100491.

Transmembrane Transport of Bicarbonate Unravelled*.

Anion receptors can be used to transport ions across lipid bilayers, which has potential for therapeutic applications. Synthetic bicarbonate transporters are of particular interest, as defects in transmembrane transport of bicarbonate are associated with various diseases. However, no convenient method exists to directly observe bicarbonate transport and study the mechanisms involved. Here, an assay is presented that allows the kinetics of bicarbonate transport into liposomes to be monitored directly and with great sensitivity. The assay utilises an encapsulated europium(III) complex, which exhibits a large increase in emission intensity upon binding bicarbonate. Mechanisms involving CO2 diffusion and the dissipation of a pH gradient are shown to be able to lead to an increase in bicarbonate concentration within liposomes, without transport of the anion occurring at all. By distinguishing these alternative mechanisms from actual bicarbonate transport, this assay will inform the future development of bicarbonate transporters.

Screening of Lactic Acid Bacteria for the Bio-Control of Botrytis cinerea and the Potential of Lactiplantibacillus plantarum for Eco-Friendly Preservation of Fresh-Cut Kiwifruit.

Botrytis cinerea, responsible for grey mold, represents the first biological cause of fruit and vegetable spoilage phenomena in post-harvest. Kiwifruit is a climacteric fruit particularly prone to this mold infestation during storage. Lactic acid bacteria (LAB) are food-grade bacteria that can synthesize several metabolites with antimicrobial activity and are, therefore, suggested as promising and eco-friendly resources for the bio-control of molds on fruits and vegetables. In this work, we propose the screening of a collection of 300 LAB previously isolated from traditional sourdoughs for their ability to counteract in vitro the growth of Botrytis cinerea CECT 20973. Only 2% of tested LAB strains belonging to Lactiplantibacillus plantarum species, exerted a strong antagonism against B. cinerea. The cell-free supernatants were partially characterized and results clearly indicated that high levels of lactic acid contributed to the antagonistic activity. PAN01 and UFG 121 cell-free supernatants were investigated as potential bio-control agents in a preliminary in vivo assay using freshly cut kiwifruits as a food model. The application of cell-free supernatants allowed to delay the growth of B. cinerea on artificially contaminated kiwifruits until two weeks. The antagonistic activity was greatly affected by the storage temperature (25 °C and 4 °C) selected for the processed fruits, suggesting the importance to include microbial-based solution in a broader framework of hurdle technologies.

Selection of Riboflavin Overproducing Strains of Lactic Acid Bacteria and Riboflavin Direct Quantification by Fluorescence.

Riboflavin (vitamin B2) is a vitamin of the B group involved in essential biological pathways, including redox reactions and the electron transport chain. Some lactic acid bacteria (LAB) can synthesize riboflavin and this capability is strain-dependent. In the last years, a growing interest has focused on the selection of riboflavin-overproducing food-grade LAB for the vitamin biofortification of fermented foods, as well as for the formulation of innovative functional products.In this chapter we report fast and inexpensive techniques in order to (1) screen LAB isolates able to produce riboflavin from different matrices, (2) select spontaneous roseoflavin-resistant riboflavin overproducing strains, and (3) quantify vitamin B2 in culture media by fluorescence detection.These protocols could be useful to select new overproducing strains and/or species from different ecological niches, as well as to optimize the conditions for vitamin bioproduction.

Microbial-based Biocontrol Solutions for Fruits and Vegetables: Recent Insight, Patents, and Innovative Trends.

BACKGROUND: Fruits and vegetables are susceptible to colonisation by undesired microflora, which, in pre- and post-harvest conditions, negatively impact the quality of these products, leading to a reduction of yield, shelf-life, and marketability. In the few last years, the use of microbial Biological Control Agents (BCAs) has assumed international relevance in order to control harmful microorganisms, as a promising alternative to chemical interventions. OBJECTIVE: The purpose of this review is to discuss the microbial-based solutions applicable for the biocontrol of the main microbial spoilers, phytopathogens, and human food-borne pathogens affecting fruits and vegetables during their production and storage. RESULTS: A comprehensive overview of the scientific literature investigating the effectiveness of BCA-based products available on the market is provided, as well as of the most recent patents protecting biotechnological applications in this field. Innovative trends are discussed, with a particular focus on the integration of BCAs to minimise spoilage phenomena and microbiological risks adopting combined approaches. CONCLUSION: This study underlines the growing interest about biocontrol strategies to counteract the growth of spoilage and/or pathogenic microorganisms indicating that in the next years a considerable increase of commercial products and patents will be developed worldwide to exploit innovative biotechnological solutions in the sector.

Botrytis cinerea and Table Grapes: A Review of the Main Physical, Chemical, and Bio-Based Control Treatments in Post-Harvest.

Consumers highly appreciate table grapes for their pleasant sensory attributes and as good sources of nutritional and functional compounds. This explains the rising market and global interest in this product. Along with other fruits and vegetables, table grapes are considerably perishable post-harvest due to the growth of undesired microorganisms. Among the microbial spoilers, Botrytis cinerea represents a model organism because of its degrading potential and the huge economic losses caused by its infection. The present review provides an overview of the recent primary physical, chemical, and biological control treatments adopted against the development of B. cinerea in table grapes to extend shelf life. These treatments preserve product quality and safety. This article also focuses on the compliance of different approaches with organic and sustainable production processes. Tailored approaches include those that rely on controlled atmosphere and the application of edible coating and packaging, as well as microbial-based activities. These strategies, applied alone or in combination, are among the most promising solutions in order to prolong table grape quality during cold storage. In general, the innovative design of applications dealing with hurdle technologies holds great promise for future improvements.

Effect of different anaesthetic techniques on gene expression profiles in patients who underwent hip arthroplasty.

OBJECTIVES: To investigate the modulation of genes whose expression level is indicative of stress and toxicity following exposure to three anaesthesia techniques, general anaesthesia (GA), regional anaesthesia (RA), or integrated anaesthesia (IA). METHODS: Patients scheduled for hip arthroplasty receiving GA, RA and IA were enrolled at Rizzoli Orthopaedic Institute of Bologna, Italy and the expression of genes involved in toxicology were evaluated in peripheral blood mononuclear cells (PBMCs) collected before (T0), immediately after surgery (T1), and on the third day (T2) after surgery in association with biochemical parameters. RESULTS: All three anaesthesia methods proved safe and reliable in terms of pain relief and patient recovery. Gene ontology analysis revealed that GA and mainly IA were associated with deregulation of DNA repair system and stress-responsive genes, which was observed even after 3-days from anaesthesia. Conversely, RA was not associated with substantial changes in gene expression. CONCLUSIONS: Based on the gene expression analysis, RA technique showed the smallest toxicological effect in hip arthroplasty. TRIAL REGISTRATION: ClinicalTrials.gov number NCT03585647.

Solvent-Free Enantioselective Michael Reactions Catalyzed by a Calixarene-Based Primary Amine Thiourea.

An upper-rim functionalized calix[4]arene-based thiourea installed onto the ( R, R)-1,2-cyclohexanediamine scaffold was synthesized with a view to investigate its catalytic ability in enantioselective Michael additions. The reactions were found to conveniently proceed under solvent-free conditions, observing good to high enantioselectivities. From this preliminary study, the calix[4]arene unit is likely to play a role in affecting the conversion and to a lesser extent to the stereochemical outcome of the reactions through van der Waals contacts and C-H···π interactions with the substrates.

In-Plane Ultrasound-Guided Lumbar Plexus Block Using Catheter-Over-Needle Technique in a 14-Month-Old Baby.

This case report details the feasibility of a continuous lumbar plexus block obtained with a paramedian transverse scan of the lumbar paravertebral region, using a catheter-over-needle system, as a main postoperative orthopedic analgesia, in a 14-month-old patient undergoing surgical treatment of a localized Ewing family tumor of the right proximal femur. After surgery, the patient remained in the intensive care unit for 3 days; pain was assessed regularly using the Children's and Infants' Postoperative Pain Scale, and comfort was monitored. Analgesia was provided with continuous levobupivacaine 0.2% infusion 0.2 mL kg h, rectal paracetamol 15 mg kg, and oral ibuprofen 60 mg twice per day, with a dose of morphine 0.05 mg kg intravenously as rescue. Pain relief was excellent, and no additional morphine was needed during the postoperative course. Nausea, vomiting, and disturbed sleep were not reported. Oral diet was resumed on the first postoperative day. In conclusion, this case represents successful postoperative management of orthopedic pain in a child using continuous posterior lumbar plexus infusion.

Calix[6]arene threading with weakly interacting tertiary ammonium axles: generation of chiral pseudorotaxane architectures.

The TFPB "superweak anion" effectively induces the threading of a scarcely preorganized calix[6]arene (1) with weakly interacting tertiary ammonium axles (2(+)-3(+)). The pseudorotaxanes (4(+)-5(+)) thus formed show an unexpected chirality due to an unprecedented formation of a threading-induced stereogenic ammonium center.

[Non-hemorrhagic stroke, of possible aortoembolic origin, in the course of acute myocardial infarct treated with thrombolysis]. / Stroke non emorragico, di possibile origine aortoembolica, in corso di infarto miocardico acuto trattato con trombolisi.

We describe the case of a middle-aged man with nonhemorrhagic stroke occurred during thrombolysis for acute myocardial infarction. The ischemic etiology of stroke was demonstrated by cerebral computed tomography scan. Transthoracic echocardiogram excluded left ventricular thrombosis. Transesophageal echocardiogram displayed complicated atheromas of the aortic arch. The reported case emphasizes that aortic atheromas may be the source of emboli during thrombolytic therapy.

Detection of HTLV-I tax-rex and pol gene sequences of thymus gland in a large group of patients with myasthenia gravis.

We report in this study the use of polymerase chain reaction (PCR) for the amplification of the genomic DNA, isolated from thymic tissue, using the primers flanking HTLV-I/II tax-rex genes and the sequence method to analyze the HTLV-I pol sequence of 27 Italian patients with myasthenia gravis. These molecular methods showed that 92.5% of patients tested positive for tax gene and 55% for pol genes; 55.5% samples were positive for both the tax gene of HTLV-I/II, and the pol gene of HTLV-I. Histologic investigation of the thymus showed that 15 samples had thymic hyperplasia, 93% tested positive for the tax gene, and 40% tested positive for both the tax and pol genes of HTLV-I. In contrast, 91.6% of thymoma-positive samples were positive for tax gene I/II and 75% positive for both genes, tax and pol type I. The sequence analysis of PCR product for tax and pol genes confirmed that these amplified products were HTLV-I, with minimal variations. Our date suggested that either HTLV-I or part of the virus genome is involved in the etiopathogenesis of myasthenia gravis.