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The authors regret the incorrect publication of legend for Table 1 and Table 3 in black/white in the original article. The complete legend for Table 1 and Table 3 with colours, indicating degree of significance, are presented below.

In this work, we describe the preparation and characterization of supramolecular ionic liquid gels based on binary mixtures of biopolymers, comprising chitosan, chitin, cellulose and lignin. The gels were obtained in ionic liquids differing in the cation or the anion, with no need for a cross-linking agent or acid treatment. The materials obtained were characterized for the minimum gelation concentration, porosity, swelling and rheological properties, finding a prominent influence of the nature of the ionic liquid anion. Then, we investigated the ability of the gels to scavenge free radicals, finding that the gels exhibit a higher antioxidant ability than their individual components. Moreover, our gels showed a bactericidal activity toward Gram-positive and Gram-negative bacterial strains like Streptomyces coelicolor and Escherichia coli, respectively. The kinetics of bacterial cell viability showed that the gel based on a chitosan : chitin mixture in the IL [bmim][Cl] was the fastest acting against E. coli, with a practically total killing of bacterial cells within 30 min. Once again, a major influence of the ionic liquid anion was detected. Finally, we found an inverse relationship between the antioxidant and antimicrobial activities of our gels.

The reaction of octakis(3-chloropropyl)octasilsesquioxane with four equivalents of 1-hexylimidazole or 1-decylimidazole gave two products labelled as HQ-POSS (hexyl-imidazolium quaternized POSS) and DQ-POSS (decyl-imidazolium quaternized POSS) as regioisomer mixtures. An investigation of the biological activity of these two compounds revealed the higher antimicrobial performances of HQ-POSS against Gram-positive and Gram-negative microorganisms, proving its broad-spectrum activity. Due to its very viscous nature, HQ-POSS was adsorbed in variable amounts on the surface of biologically active oxides to gain advantages regarding the expendability of such formulations from an applicative perspective. Titania and 5 wt% Cu on titania were used as supports. The materials 10HQ-POSS/Ti and 15HQ-POSS/5CuTi strongly inhibited the ability of Pseudomonas PS27 cells—a bacterial strain described for its ability to handle very toxic organic solvents and perfluorinated compounds—to grow as planktonic cells. Moreover, the best formulations (i.e., 10HQ-POSS/Ti and 15HQ-POSS/5CuTi) could prevent Pseudomonas PS27 biofilm formation at a certain concentration (250 μg mL−1) which greatly impaired bacterial planktonic growth. Specifically, 15HQ-POSS/5CuTi completely impaired cell adhesion, thus successfully prejudicing biofilm formation and proving its suitability as a potential antifouling agent. Considering that most studies deal with quaternary ammonium salts (QASs) with long alkyl chains (>10 carbon atoms), the results reported here on hexylimidazolium-based POSS further deepen the knowledge of QAS formulations which can be used as antifouling compounds.

12 páginas.- 5 figuras.- 46 referencias.- Supplementary Materials http://dx.doi.org/10.1089/ast.2022.0062, Antarctic deserts are among the driest and coldest ecosystems of the planet; there, some microbes survive under these extreme conditions inside porous rocks, forming the so-called endolithic communities. Yet the contribution of distinct rock traits to support complex microbial assemblies remains poorly determined. Here, we combined an extensive Antarctic rock survey with rock microbiome sequencing and ecological networks and found that contrasting combinations of microclimatic and rock traits such as thermal inertia, porosity, iron concentration, and quartz cement can help explain the multiple complex microbial assemblies found in Antarctic rocks. Our work highlights the pivotal role of rocky substrate heterogeneity in sustaining contrasting groups of microorganisms, which is essential to understand life at the edge on Earth and for the search for life on other rocky planets such as Mars., CC and LS wish to thank the Italian National Program for Antarctic Research (PNRA) for funding sampling campaigns and research activities in Italy in the frame of PNRA projects. The Italian Antarctic National Museum (MNA) is kindly acknowledged for financial support to the Mycological Section of the MNA and for providing rock samples used in this study stored in the Culture Collection of Antarctic fungi (MNA-CCFEE), University of Tuscia, Italy. LS acknowledges funding from the PNRA. MD-B is supported by a project from the Spanish Ministry of Science and Innovation (PID2020- 115813RA-I00), and a project of the Fondo Europeo de Desarrollo Regional (FEDER) and the Consejería de Transformación Económica, Industria, Conocimiento y Universidades of the Junta de Andalucía (FEDER Andalucía 2014-2020 Objetivo temático ‘01 – Refuerzo de la investigación, el desarrollo tecnológico y la innovación’) associated with the research project P20_00879 (ANDABIOMA). This paper was previously submitted to BioXRiv as a preprint. The preprint can be found at https://doi.org/ 10.1101/2021.08.04.455018

Lipopolysaccharide (LPS) is an essential component of the outer membrane (OM) of many Gram-negative bacteria, providing a barrier against the entry of toxic molecules. In Escherichia coli, LPS is exported to the cell surface by seven essential proteins (LptA-G) that form a transenvelope complex. At the inner membrane, the ATP-binding cassette (ABC) transporter LptB

Glaciers are ecosystems and they host active biological communities. Despite their remoteness, glaciers act as cold condensers where high precipitation rates and cold temperatures favor the deposition of pollutants. These contaminants include a broad range of substances, including legacy pollutants, but also compounds still largely used. Some of these compounds are monitored in the environment and their effects on the ecosystems are known, in contrast others can be defined as emerging pollutants since their presence and their impact on the environment are still poorly understood (e.g. microplastics, radionuclides). This review aim to provide an overview of the studies that have investigated the effects of pollutants on the supraglacial ecosystem so far. Despite the distribution of the pollutants in glacier environments has been discussed in several studies, no review paper has summarized the current knowledge on the effects of these substances on the ecological communities living in glacier ecosystems.

IntroductionAs we approach the post-antibiotic era, the development of innovative antimicrobial strategies that carry out their activities through non-specific mechanisms could limit the onset and spread of drug resistance. In this context, the use of nanogranular coatings of multielement nanoparticles (NPs) conjugated to the surface of implantable biomaterials might represent a strategy to reduce the systemic drawbacks by locally confining the NPs effects against either prokaryotic or eukaryotic cells.MethodsIn the present study, two new multielement nanogranular coatings combining Ag and Cu with either Ti or Mg were synthesized by a gas phase physical method and tested against pathogens isolated from periprosthetic joint infections to address their potential antimicrobial value and toxicity in an in vitro experimental setting.ResultsOverall, Staphylococcus aureus, Staphylococcus epidermidis and Escherichia coli displayed a significantly decreased adhesion when cultured on Ti-Ag-Cu and Mg-Ag-Cu coatings compared to uncoated controls, regardless of their antibiotic resistance traits. A dissimilar behavior was observed when Pseudomonas aeruginosa was cultured for 30 and 120 minutes upon the surface of Ti-Ag-Cu and Mg-Ag-Cu-coated discs. Biofilm formation was mainly reduced by the active effect of Mg-Ag-Cu compared to Ti-Ag-Cu and, again, coatings had a milder effect on P. aeruginosa, probably due to its exceptional capability of attachment and matrix production. These data were further confirmed by the evaluation of bacterial colonization on nanoparticle-coated discs through confocal microscopy. Finally, to exclude any cytotoxic effects on eukaryotic cells, the biocompatibility of NPs-coated discs was studied. Results demonstrated a viability of 95.8% and 89.4% of cells cultured in the presence of Ti-Ag-Cu and Mg-Ag-Cu discs, respectively, when compared to negative controls.ConclusionIn conclusion, the present study demonstrated the promising anti-adhesive features of both Ti-Ag-Cu and Mg-Ag-Cu coatings, as well as their action in hampering the biofilm formation, highlighting the safe use of the tested multi-element families of nanoparticles as new strategies against bacterial attachment to the surface of biomedical implants.

IntroductionImplant-related infections and infected fractures are significant burdens in orthopedics. Staphylococcus epidermidis is one of the main causes of bone infections related to biofilm formation upon implants. Current antibiotic prophylaxis/therapy is often inadequate to prevent biofilm formation and results in antibiotic resistance. The development of bioactive materials combining antimicrobial and osteoconductive properties offers great potential for the eradication of microorganisms and for the enhancement of bone deposition in the presence of infections. The purpose of this study is to prevent the development of methicillin-resistant S. epidermidis (MRSE)-infected nonunion in a rat model.MethodsTo this end, a recently developed in our laboratories bioactive material consisting of antibiotic-loaded nanoparticles based on carboxylic acid functionalized hyperbranched aliphatic polyester (CHAP) that are integrated into peptide-enriched silk fibroin sponges with osteoconductive properties (AFN-PSF) was employed, whose biocompatibility and microbiological tests provided proof of its potential for the treatment of both orthopedic and dental infections. In particular, non-critical femoral fractures fixed with plates and screws were performed in Wistar rats, which were then randomly divided into three groups: 1) the sham control (no infection, no treatment); 2) the control group, infected with MRSE and treated with peptide-enriched silk fibroin sponges incorporating non-drug-loaded functionalized nanoparticles (PSF); 3) the treated group, infected with MRSE and treated with peptide-enriched silk fibroin sponges incorporating vancomycin-loaded functionalized nanoparticles (AFN-PSF). After 8 weeks, bone healing and osteomyelitis were clinically assessed and evaluated by micro-CT, microbiological and histological analyses.ResultsThe sham group showed no signs of infection and complete bone healing. The PSF group failed to repair the infected fracture, displaying 75% of altered bone healing and severe signs of osteomyelitis. The AFN-PSF treated group reached 70% of fracture healing in the absence of signs of osteomyelitis, such as abscesses in the cortical and intraosseous compartments and bone necrosis with sequestra.DiscussionAFN-PSF sponges have proven effective in preventing the development of infected nonunion in vivo. The proposed nanotechnology for local administration of antibiotics can have a significant impact on patient health in the case of orthopedic infections.

I solventi clorurati sono composti organici di sintesi appartenenti alla classe degli idrocarburi alifatici clorurati, largamente impiegati in diversi settori industriali e, come conseguenza, ampiamente diffusi nell’ambiente come inquinanti e particolarmente pericolosi anche per la salute umana. Tra i metodi di bonifica di siti contaminati da solventi clorurati, quelli basati sul biorisanamento sfruttano uno dei peculiari metabolismi batterici coinvolti nella biodegradazione di tali composti: la declorurazione riduttiva anaerobia in cui il composto clorurato è utilizzato come accettore terminale di elettroni in un vero e proprio processo di respirazione anaerobia, detta dealorespirazione; il cometabolismo aerobio in cui il composto clorurato viene fortuitamente ossidato ad opera di ossigenasi batteriche normalmente utilizzate per l’uptake del substrato di crescita preferenziale; l’ossidazione aerobia diretta in cui il composto clorurato funge da donatore di elettroni e substrato di crescita primario per il microrganismo. Il presente lavoro di tesi di dottorato si è occupato della caratterizzazione microbiologica di un’acqua contaminata da solventi clorurati, prevalentemente da 1,2-dicloroetano (1,2-DCA), e della valutazione del suo potenziale di biodegradazione, con l’obiettivo di sfruttare consorzi microbici autoctoni alla matrice contaminata per la definizione di un eventuale intervento di biorisanamento in situ. Le comunità microbiche autoctone dell’acqua contaminata sono state dapprima caratterizzate in termini di struttura, diversità e composizione, mediante ARISA (Automated Ribosomal Intergenic Spacer Analysis) e 16S rRNA metabarcoding, evidenziando una grande variabilità: campioni di acqua raccolti da diversi punti di prelievo e in diversi periodi sono risultati infatti piuttosto eterogenei, sia da un punto di vista chimico-fisico che da un punto di vista microbiologico. Abbastanza limitata è risultata la presenza di specie batteriche note in grado di degradare solventi clorurati. La biodegradazione di 1,2-DCA è stata successivamente valutata mediante analisi gascromatografiche in spazio di testa effettuate in microcosmo. Allo scopo di indagare tutti i metabolismi biodegradativi noti nei confronti di 1,2-DCA, sono stati allestiti e opportunamente biostimolati microcosmi in aerobiosi e anaerobiosi. I microcosmi sono stati realizzati all’interno di vials con acqua contaminata, un opportuno terreno di coltura e una quota aggiunta di 1,2-DCA. Il monitoraggio della concentrazione di 1,2-DCA nel tempo ha mostrato elevati abbattimenti nei microcosmi anaerobi biostimolati in presenza/assenza di lattato e nei microcosmi biostimolati in presenza di O2. In entrambi, la caratterizzazione della comunità microbica ha evidenziato la presenza di batteri in grado di svolgere declorurazione riduttiva di solventi clorurati e dealogenazione idrolitica di 1,2-DCA. Nel DNA metagenomico dei suddetti microcosmi è stata anche rilevata la presenza di sequenze dei geni per una dealogenasi riduttiva specifica per 1,2-DCA (dcaA) e per la aloalcano dealogenasi (dhlA). Un’efficiente biodegradazione di 1,2-DCA è stata evidenziata anche in una simulazione di sistema di Barriera Reattiva Permeabile in colonna biostimolato con poli-β-idrossibutirrato (PHB), quale substrato fermentabile e donatore di elettroni a lento rilascio per i processi di declorurazione riduttiva. Il sistema in colonna è stato alimentato con l’acqua contaminata, inoculata con un consorzio batterico precedentemente arricchito su PHB. Infine, è stata avviata una sperimentazione che vede l’utilizzo di carrier biopolimerici biodegradabili come innovativa tecnologia di immobilizzazione di batteri biodegradatori direttamente reclutati dall’acqua contaminata, allo scopo di ottenere una maggiore efficienza di biodegradazione di solventi clorurati, a basso costo e in maniera ecocompatibile. Supporti in acido polilattico hanno dato preliminari evidenze sia di assorbimento del contaminante 1,2-DCA, sia di adesione microbica quando immersi nell’acqua contaminata in un sistema a scala di microcosmo.

The urban plan of Palermo (Sicily, Italy) has evolved throughout Punic, Roman, Byzantine, Arab, and Norman ages until it stabilized within the borders that correspond to the current historic center. During the 2012 to 2013 excavation campaign, new remains of the Arab settlement, directly implanted above the structures of the Roman age, were found. The materials investigated in this study derived from the so-called Survey No 3, which consists of a rock cavity of subcylindrical shape covered with calcarenite blocks: it was probably used to dispose of garbage during the Arabic age and its content, derived from daily activities, included grape seeds, scales and bones of fish, small animal bones, and charcoals. Radiocarbon dating confirmed the medieval origin of this site. The composition of the bacterial community was characterized through a culture-dependent and a culture-independent approach. Culturable bacteria were isolated under aerobic and anaerobic conditions and the total bacterial community was characterized through metagenomic sequencing. Bacterial isolates were tested for the production of compounds with antibiotic activity: a Streptomyces strain, whose genome was sequenced, was of particular interest because of its inhibitory activity, which was due to the Type I polyketide aureothin. Moreover, all strains were tested for the production of secreted proteases, with those belonging to the genus Nocardioides having the most active enzymes. Finally, protocols commonly used for ancient DNA studies were applied to evaluate the antiquity of isolated bacterial strains. Altogether these results show how paleomicrobiology might represent an innovative and unexplored source of novel biodiversity and new biotechnological tools. IMPORTANCE One of the goals of paleomicrobiology is the characterization of the microbial community present in archaeological sites. These analyses can usually provide valuable information about past events, such as occurrence of human and animal infectious diseases, ancient human activities, and environmental changes. However, in this work, investigations about the composition of the bacterial community of an ancient soil sample (harvested in Palermo, Italy) were carried out aiming to screen ancient culturable strains with biotechnological potential, such as the ability to produce bioactive molecules and secreted hydrolytic enzymes. Besides showing the biotechnological relevance of paleomicrobiology, this work reports a case of germination of putatively ancient bacterial spores recovered from soil rather than extreme environments. Moreover, in the case of spore-forming species, these results raise questions about the accuracy of techniques usually applied to estimate antiquity of DNA, as they could lead to its underestimation.

CFTR modulators have been developed to correct and/or enhance CFTR activity in patients with specific cystic fibrosis (CF) genotypes. However, it is of great importance to identify potential off-targets of these novel therapies to understand how they affect lung physiology in CF.

Chlorinated solvents, belonging to the class of chlorinated aliphatic hydrocarbons (CAHs), are synthetic organohalide chemicals frequently found as contaminants of groundwater and soil, due to their widespread use in several industrial processes and improper disposal methods. These compounds pose serious health threats because of their toxic and sometimes carcinogenic effects. Among these compounds, 1,2-dichloroethane (1,2-DCA) is one of the most common aquifer contaminants, considered toxic and classified as a possible human carcinogen. Remediation approaches toward these contaminants include conventional cleanup technologies based on physical/chemical methods, and bioremediation, considered an eco-friendly and low-cost alternative. Bioremediation relies on biodegradation of CAHs, that is carried out by specialized bacteria under anaerobic and aerobic conditions. Biodegradation processes include direct pathways, associated with the growth of specialized microorganisms, and indirect (or cometabolic) pathways, fortuitously mediated by non-specific enzymes produced by microorganisms to grow on other substrates. In contaminated aquifers, chlorinated solvents, including 1,2-DCA, are generally biodegraded by anaerobic specialized bacteria called OHRBs (Organohalide Respiring Bacteria) in a reductive process called dehalorespiration. Biostimulation with appropriate amendments can allow the exploitation of these processes in bioremediation strategies. The aim of this work is to investigate the intrinsic biodegradation potential of a CAHs (mainly 1,2-DCA) contaminated aquifer by evaluating the response of the autochthonous microbial communities to anaerobic and aerobic biostimulation treatments in microcosm, to provide knowledge in the perspective of eventual site-specific bioremediation strategies. The preliminary microbiological characterization of the autochthonous bacterial communities of the aquifer, performed by 16S rRNA Illumina sequencing, revealed a notable difference between the bacterial communities from different sampling sites of the aquifer. Low relative abundances of known anaerobic and aerobic dechlorinating taxa were observed, suggesting a poor intrinsic bioremediation potential of the site. In order to enrich the dechlorinating components of the autochthonous bacterial communities, different anaerobic and aerobic biostimulation treatments in microcosm were tested, using groundwater samples and appropriate culture media amended with 1,2-DCA and additional substrates, if required, to stimulate different biodegradative processes. Chemical monitoring of 1,2-DCA concentration by Gas Chromatography-Mass Spectrometry, revealed high removal efficiency in different anaerobic and aerobic microcosm conditions. The analysis of microcosms bacterial communities, performed by 16S rRNA Illumina sequencing, revealed the enrichment in anaerobic or aerobic dechlorinating taxa. Reductive dehalogenase (dcaA) or haloalkane dehalogenase (dhlA) genes were detected in anaerobic and aerobic microcosms, respectively, by PCR probing the metagenomic DNA extracted from microcosms. Isolation attempts from the aerobic biostimulation treatments led to the achievement of stable aerobic 1,2-DCA dechlorinating oligospecfic consortia, that were analyzed by a Next Generation Sequencing approach. The enriched dechlorinating bacterial communities obtained from biostimulation trials were used in a pilot study to test the feasibility of a polyhydroxybutyrate-based Permeable Reactive Barrier (PRB) and of an aerobic bioremediation approach in two column simulations fed with real contaminated groundwater. Both approaches led to an effective 1,2-DCA removal. Finally, innovative biopolymeric tridimensional scaffolds were tested to evaluate their suitability as carriers either for the immobilization of groundwater autochthonous bacteria or for bioaugmentation with specialized dehalogenating biofilms to be eventually exploited in enhanced bioremediation. Laboratory scale studies and a preliminary field experiment confirmed scaffold could uptake 1,2-DCA and be colonized by a bacterial biofilm from groundwater bacterial communities, also made up of dechlorinating taxa.

The human microbiome seeding starts at birth, when pioneer microbes are acquired mainly from the mother. Mode of delivery, antibiotic prophylaxis, and feeding method have been studied as modulators of mother-to-infant microbiome transmission, but other key influencing factors like modern westernized lifestyles with high hygienization, high-calorie diets, and urban settings, compared with non-westernized lifestyles have not been investigated yet. In this study, we explored the mother-infant sharing of characterized and uncharacterized microbiome members via strain-resolved metagenomics in a cohort of Ethiopian mothers and infants, and we compared them with four other cohorts with different lifestyles. The westernized and non-westernized newborns' microbiomes composition overlapped during the first months of life more than later in life, likely reflecting similar initial breast-milk-based diets. Ethiopian and other non-westernized infants shared a smaller fraction of the microbiome with their mothers than did most westernized populations, despite showing a higher microbiome diversity, and uncharacterized species represented a substantial fraction of those shared in the Ethiopian cohort. Moreover, we identified uncharacterized species belonging to the Selenomonadaceae and Prevotellaceae families specifically present and shared only in the Ethiopian cohort, and we showed that a locally produced fermented food, injera, can contribute to the higher diversity observed in the Ethiopian infants' gut with bacteria that are not part of the human microbiome but are acquired through fermented food consumption. Taken together, these findings highlight the fact that lifestyle can impact the gut microbiome composition not only through differences in diet, drug consumption, and environmental factors but also through its effect on mother-infant strain-sharing patterns., This work was supported by the European Research Council (ERC-STG project MetaPG-716575) to N.S., by MIUR “Futuro in Ricerca” (grant no. RBFR13EWWI_001) to N.S., by the European H2020 program (ONCOBIOME-825410 project and MASTER-818368 project) to N.S., by the National Cancer Institute of the National Institutes of Health (1U01CA230551) to N.S., by the Premio Internazionale Lombardia e Ricerca 2019 to N.S., by the Italian Ministry of Health with Ricerca Corrente and 5x1000 funds, and by the EMBO ALTF 593-2020 to M.V.-C. The authors would like to acknowledge support from the European Research Council (ERC-STG project MAMI, ref. 639226) and PROMETEO GVA (ref. 012/2020) to M.C.C. and M.S.-R., respectively, and from Plan GenT project (CDEIGENT 2020) to R.C.-R. Finally, M.C.C., R.C.-R., and M.S.-R. acknowledge the award of the Spanish government MCIN/AEI to the IATA-CSIC as Center of Excellence Accreditation Severo Ochoa (CEX2021-001189-S/MCIN/AEI/10.13039/501100011033).This article is part of a project that has received funding from the European Union’s Horizon 2020 Research and Innovation Programme (GA no. 949742 ERC-HealthXCross) to R.R.

Helianthemum lippii is a perennial shrubby plant growing in the sandy environments of Italy, Mediterranean countries of North Africa and Middle East. H. lippii is used in traditional medicine but there are very few reports referring to the phytochemical characterization, the ethnopharmacology, and the biological activity of H. lippii. The goal of this study was todetermine the phytochemical composition of different H. lippii extracts, cold (CME) and hot (HME) methanol, cold (CPEE) and hot (HPEE) petroleum ether, as well as to evaluate their anticancer and antimicrobial activities and biofilm formation reduction. Fifty-fours phytocompounds have been determined by HPLC-UV-ESI-QTOF-MS analysis. All the four extracts reduced the viability of human MDA-MB231 and HCT116 cells, being cold and hot methanol extracts the most effective. The antimicrobial activity against S. aureus, P. aeruginosa, E. faecalis, E. coli, C. albicans was also evaluated. Data showed the greater susceptibility of S. aureus to hot methanolic extract. Concerning antifungal activity, C. albicans resulted more susceptible to petroleum ether. Moreover, some of the samples exhibited a good antibiofilm activity both on immature and on mature biofilms. The four extracts showed interesting antimicrobial and cytotoxic activities and can be considered good candidates for new therapeutic applications

Artemisia arborescens (Vaill.) L. is a perennial shrubby plant growing along the coastal strips of the Mediterranean region. It is used in traditional medicine. Its essential oil and solvent extracts exhibit a very interesting chemotherapeutic potential, which makes this plant useful in maintaining human health. The goal of this study was to determine the phytochemical composition of the petroleum ether and methanol extracts, as well as to evaluate anticancer activities and antimicrobial and biofilm formation reduction. Thirty-nine phytochemical compounds in negative ion mode, and 25 in positive ion mode were identified by HPLC-ESI-QTOF-MS. All four extracts reduced the viability of human MDA-MB231 and HCT116 cancer cells suggesting a similar cytotoxic efficacy of the different extracts in MDA-MB231 and more pronounced antiproliferative effects on HCT116 cells treated with HPEE and CPEE. Antimicrobial activity was exhibited by the hot petroleum ether extract against all tested bacteria (MIC 0.15-2.5 mg/mL), except for S. agalactiae. Both methanol extracts showed activity against C. albicans with a MIC value of 2.5 mg/mL, and a lack of antibacterial activity. To whom concern the anti-biofilm activity, hot methanol extract shows the highest activity in inhibiting biofilm formation among all extract.

Mucosal vaccination is regarded as a promising alternative to classical, intramuscular vaccine delivery. However, only a limited number of vaccines have been licensed for mucosal administration in humans. Here we propose Leishmania tarentolae, a protozoan parasite, as a potential antigen vehicle for mucosal vaccination, for administration via the rectal or oral routes. To test this hypothesis, we exploited L. tarentolae for the production and delivery of SARS-CoV-2 antigens. Two antigens were assayed in BALB/c mice: Lt-spike, a L. tarentolae clone engineered for the surface expression of the SARS-CoV-2 spike protein; RBD-SD1, a purified portion of the spike protein, produced by another engineered clone of the protozoon. Immune response parameters were then determined at different time points. Both antigens, administered either separately or in combination (Lt-spike + RBD-SD1, hereafter LeCoVax-2), determined significant IgG seroconversion and production of neutralizing antibodies after subcutaneous administration, but only in the presence of adjuvants. After rectal administration, the purified RBD-SD1 antigen did not induce any detectable immune response, in comparison with the intense response observed after administration of LeCoVax-2 or Lt-spike alone. In rectal administration, LeCoVax-2 was also effective when administered without adjuvant. Our results show that L. tarentolae is an efficient and safe scaffold for production and delivery of viral antigens, to be used as vaccines. In addition, rectal vaccination experiments prove that L. tarentolae is suitable as a vaccine vehicle and adjuvant for enteral vaccination. Finally, the combined preparation LeCoVax-2 can be considered as a promising candidate vaccine against SARS-CoV-2, worthy of further investigation.

Seed-associated microbiota are believed to play a crucial role in seed germination, seedling establishment, and plant growth and fitness stimulation, due to the vertical transmission of a core microbiota from seeds to the next generations. It might be hypothesized that medicinal and aromatic plants could use the seeds as vectors to vertically transfer beneficial endophytes, providing plants with metabolic pathways that could influence phytochemicals production. Here, we investigated the localization, the structure and the composition of the bacterial endophytic population that resides in Origanum heracleoticum L. seeds. Endocellular bacteria, surrounded by a wall, were localized close to the aleurone layer when using light and transmission electron microscopy. From surface-sterilized seeds, cultivable endophytes were isolated and characterized through RAPD analysis and 16S RNA gene sequencing, which revealed the existence of a high degree of biodiversity at the strain level and the predominance of the genus Pseudomonas. Most of the isolates grew in the presence of six selected antibiotics and were able to inhibit the growth of clinical and environmental strains that belong to the Burkholderia cepacia complex. The endophytes production of antimicrobial compounds could suggest their involvement in plant secondary metabolites production and might pave the way to endophytes exploitation in the pharmaceutical field.

The intense use of tellurium (Te) in industrial applications, along with the improper disposal of Te-derivatives, is causing their accumulation in the environment, where oxyanion tellurite (TeO32−) is the most soluble, bioavailable, and toxic Te-species. On the other hand, tellurium is a rare metalloid element whose natural supply will end shortly with possible economic and technological effects. Thus, Te-containing waste represents the source from which Te should be recycled and recovered. Among the explored strategies, the microbial TeO32− biotransformation into less toxic Te-species is the most appropriate concerning the circular economy. Actinomycetes are ideal candidates in environmental biotechnology. However, their exploration in TeO32− biotransformation is scarce due to limited knowledge regarding oxyanion microbial processing. Here, this gap was filled by investigating the cell tolerance, adaptation, and response to TeO32− of a Micromonospora strain isolated from a metal(loid)-rich environment. To this aim, an integrated biological, physical-chemical, and statistical approach combining physiological and biochemical assays with confocal or scanning electron (SEM) microscopy and Fourier-transform infrared spectroscopy in attenuated total reflectance mode (ATR-FTIR) was designed. Micromonospora cells exposed to TeO32− under different physiological states revealed a series of striking cell responses, such as cell morphology changes, extracellular polymeric substance production, cell membrane damages and modifications, oxidative stress burst, protein aggregation and phosphorylation, and superoxide dismutase induction. These results highlight this Micromonospora strain as an asset for biotechnological purposes.

Sauerkraut is a traditionally fermented cabbage, and recent evidence suggests that it has beneficial properties for human health. In this work, a multi-disciplinary approach was employed to characterize the fermentation process and gut health-promoting properties of locally produced, organic sauerkraut from two distinct producers, SK1 and SK2. 16S rRNA metataxonomics showed that bacterial diversity gradually decreased as fermentation progressed. Differences in sauerkraut microbiota composition were observed between the two producers, especially at the start of fermentation. Lactic acid bacteria (LAB) dominated the microbiota after 35 days, with Lactiplantibacillus being the dominant genus in both sauerkraut products, together with Leuconostoc and Paucilactobacillus in SK1, and with Pediococcus, Levilactibacillus, and Leuconostoc in SK2. LAB reached between 7 and 8 Log CFU/mL brine at the end of fermentation (35 days), while pH lowering happened within the first week of fermentation. A total of 220 LAB strains, corresponding to 133 RAPD-PCR biotypes, were successfully isolated. Lactiplantibacillus plantarum and Lactiplantibacillus pentosus accounted for 67% of all SK1 isolates, and Lactiplantibacillus plantarum/paraplantarum and Leuconostoc mesenteroides represented 72% of all the isolates from SK2. 1H-NMR analysis revealed significant changes in microbial metabolite profiles during the fermentation process, with lactic and acetic acids, as well as amino acids, amines, and uracil, being the dominant metabolites quantified. Sauerkraut brine did not affect trans-epithelial electrical resistance through a Caco-2 cell monolayer as a measure of gut barrier function. However, significant modulation of inflammatory response after LPS stimulation was observed in PBMCs-Caco-2 co-culture. Sauerkraut brine supported a robust inflammatory response to endotoxin, by increasing TNF-α and IL-6 production while also stimulating the anti-inflammatory IL-10, therefore suggesting positive resolution of inflammation after 24 h and supporting the potential of sauerkraut brine to regulate intestinal immune function.

Membrane vesicles (MVs) are spherical particles with nanoscale dimensions and characterized by the presence of diverse cargos, such as nucleic acids, proteins, lipids, and cellular metabolites. Many examples of (micro)organisms producing MVs are reported in literature. Among them, bacterial MVs are of particular interest because they are now considered as the fourth mechanism of horizontal gene transfer. Streptomyces bacteria are well-known for their ecological roles and ability to synthesize bioactive compounds, with Streptomyces coelicolor being the model organism. It was previously demonstrated that it can produce distinct populations of MVs characterized by different protein and metabolite cargos. In this work we demonstrated for the first time that MVs of S. coelicolor carry both DNA and RNA and that their DNA content represents the entire chromosome of the bacterium. These findings suggest that MV DNA could have a role in the evolution of Streptomyces genomes and that MVs could be exploited in new strain engineering strategies.

Multi drug resistant (MDR) bacteria are insensitive to the most common antibiotics currently in use. The spread of antibiotic-resistant bacteria, if not contained, will represent the main cause of death for humanity in 2050. The situation is even more worrying when considering patients with chronic bacterial infections, such as those with Cystic Fibrosis (CF). The development of alternative approaches is essential and novel therapies that combine exogenous and host-mediated antimicrobial action are promising. In this work, we demonstrate that asymmetric phosphatidylserine/phosphatidic acid (PS/PA) liposomes administrated both in prophylactic and therapeutic treatments, induced a reduction in the bacterial burden both in wild-type and cftr-loss-of-function (cftr-LOF) zebrafish embryos infected with Pseudomonas aeruginosa (Pa) PAO1 strain (PAO1). These effects are elicited through the enhancement of phagocytic activity of macrophages. Moreover, the combined use of liposomes and a phage-cocktail (CKΦ), already validated as a PAO1 “eater”, improves the antimicrobial effects of single treatments, and it is effective also against CKΦ-resistant bacteria. We also address the translational potential of the research, by evaluating the safety of CKΦ and PS/PA liposomes administrations in in vitro model of human bronchial epithelial cells, carrying the homozygous F508del-CFTR mutation, and in THP-1 cells differentiated into a macrophage-like phenotype with pharmacologically inhibited CFTR. Our results open the way to the development of novel pharmacological formulations composed of both phages and liposomes to counteract more efficiently the infections caused by Pa or other bacteria, especially in patients with chronic infections such those with CF.

The proliferation and transmission of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or the (COVID-19) disease, has become a threat to worldwide biosecurity. Therefore, early diagnosis of COVID-19 is crucial to combat the ongoing infection spread. In this study we propose a flexible aptamer-based electrochemical sensor for the rapid, label-free detection of SARS-CoV-2 spike protein (SP). A platform made of a porous and flexible carbon cloth, coated with gold nanoparticles, to increase the conductivity and electrochemical performance of the material, was assembled with a thiol functionalized DNA aptamer via S-Au bonds, for the selective recognition of the SARS-CoV-2 SP. The various steps for the sensor preparation were followed by using scanning electron microscopy, cyclic voltammetry and differential pulse voltammetry (DPV). The proposed platform displayed good mechanical stability, revealing negligible changes on voltammetric responses to bending at various angles. Quantification of SARS-CoV-2 SP was performed by DPV and chronopotentiometry (CP), exploiting the changes of the electrical signals due the [Fe(CN)6]3-/4- redox probe, when SARS-CoV-2 SP binds to the aptamer immobilized on the electrode surface. Current density, in DPV, and square root of the transition time, in CP, varied linearly with the log[ SARS-CoV-2 SP], providing lower limits of detection (LOD) of 0.11 ng/mL and 37.8 ng/mL, respectively. The sensor displayed good selectivity, repeatability, and was tested in diluted human saliva, spiked with different SARS-CoV-2 SP concentrations, providing LODs of 0.167 ng/mL and 46.2 ng/mL for DPV and CP, respectively.

Pseudomonas phages are increasingly important biomedicines for phage therapy, but little is known about how these viruses package DNA. This paper explores the terminase subunits from the Myoviridae E217, a Pseudomonas-phage used in an experimental cocktail to eradicate P. aeruginosa in vitro and in animal models. We identified the large (TerL) and small (TerS) terminase subunits in two genes ~58 kbs away from each other in the E217 genome. TerL presents a classical two-domain architecture, consisting of an N-terminal ATPase and C-terminal nuclease domain arranged into a bean-shaped tertiary structure. A 2.05 Å crystal structure of the C-terminal domain revealed an RNase H-like fold with two magnesium ions in the nuclease active site. Mutations in TerL residues involved in magnesium coordination had a dominant-negative effect on phage growth. However, the two ions identified in the active site were too far from each other to promote two-metal-ion catalysis, suggesting a conformational change is required for nuclease activity. We also determined a 3.38 Å cryo-EM reconstruction of E217 TerS that revealed a ring-like decamer, departing from the most common nonameric quaternary structure observed thus far. E217 TerS contains both N-terminal helix-turn-helix motifs enriched in basic residues and a central channel lined with basic residues large enough to accommodate double-stranded DNA. Overexpression of TerS caused a more than a four-fold reduction of E217 burst size, suggesting a catalytic amount of the protein is required for packaging. Together, these data expand the molecular repertoire of viral terminase subunits to Pseudomonas-phages used for phage therapy.

New data from sites in Spain, Sicily and North America establish that the spread of the European legume Cytisus scoparius (Scotch broom) across North America involved the concurrent invasion of a European mobile genetic element (symbiosis island [SI]) into North American Bradyrhizobium nodule bacteria. At four SI loci, bacteria from nine C. scoparius populations across the continent were all identical to haplotypes present in European Bradyrhizobium strains. At seven non-symbiotic (housekeeping) loci, these American C. scoparius bacteria grouped into eight diverse lineages related to, or in some cases identical to, symbionts from several native North American legumes. Inoculation experiments showed that American Bradyrhizobium strains with the European SI variant conferred superior growth to C. scoparius plants compared to strains with SI variants found indigenously in North America. Cytisus scoparius invasion in North America thus enabled proliferation of a European SI variant that spread to multiple indigenous American Bradyrhizobium lineages via horizontal gene transfer. Spread of this SI variant, in turn, likely facilitated C. scoparius invasion of North American habitats.

Pectin recovered via hydrodynamic cavitation (IntegroPectin) from lemon and grapefruit agri-food waste intrinsically containing antimicrobial bioactive substances (flavonoids, phenolic acids, terpenes, and terpenoids) was used to generate innovative and eco-compatible films that efficiently inhibit the growth of Gram-negative pathogens. Extensive characterization of films confirmed the presence of these substances, which differently interact with the polysaccharide polymer (pectin), plasticizer (glycerol), surfactant (Tween 60), and cross-linker (Ca2+), conferring to these films a unique structure. Besides, IntegroPectin-based films constitute versatile systems for the sustained, controlled, and slow-release (up to 72 h) of bioactive substances in an aqueous environment. This feature is crucial for the good in vitro antimicrobial activity exerted by IntegroPectin films against three Gram-negative bacteria (two indicator pathogen strains Pseudomonas aeruginosa ATCC 10145, P. aeruginosa PAO1, and the clinical isolate Klebsiella pneumoniae) that are involved in the global emergence of the antimicrobial resistance. Graphical abstract

Diatoms are microalgae well known for their high variability and high primary productivity, being responsible for about 20% of the annual global carbon fixation. Moreover, they are interesting as potential feedstocks for the production of biofuels and high-value lipids and carotenoids. Diatoms exhibit trophic flexibility and, under certain conditions, they can grow mixotrophically by combing photosynthesis and respiration. So far, only a few species of diatoms have been tested for their mixotrophic metabolism; in some cases, they produced more biomass and with higher lipid content when grown under this condition. Phaeodactylum tricornutum is the most studied diatom species for its mixotrophic metabolism due to available genome sequence and molecular tools. However, studies in additional species are needed to better understand the conservation of this process in diatoms and its potential in industrial applications. Here, we describe the photosynthetic and respiratory pathways involved in mixotrophy and provide an overview of the trophic variability in diatoms. This review also highlights promising areas of industrial applications for diatoms when cultivated under mixotrophy.

Chronic hepatitis B (CHB) is a major worldwide public health problem and novel anti-HBV therapies preventing liver disease progression to cirrhosis and hepatocellular carcinoma are urgently needed. Over the last several years, capsid assembly modulators (CAM) have emerged as clinically effective anti-HBV agents which can inhibit HBV replication in CHB patients. As part of a drug discovery program aimed at obtaining novel CAM endowed with high in vitro and in vivo antiviral activity, we identified a novel series of sulfamoylbenzamide (SBA) derivatives. Compound 10, one of the most in vitro potent SBA-derived CAM discovered to date, showed excellent pharmacokinetics in mice suitable for oral dosing. When studied in a transgenic mouse model of hepatic HBV replication, it was considerably more potent than NVR 3-778, the first sulfamoylbenzamide (SBA) CAM that entered clinical trials for CHB, at reducing viral replication in a dose-dependent fashion. We present herein the discovery process, the SAR analysis and the pre-clinical profile of this novel SBA CAM.

The aim of this systematic review is to collect clinical trials conducted using essential oils (EOs) in obstetric symptoms by evaluating if and in which context the aromatherapy practice is effective in obstetrics. The research was conducted by using the databases of EMBASE, Medline, Biosis and Toxcenter, PubMed, and Google Scholar search engine, selecting articles from January 2004 to July 2020. This study was performed according to the MOOSE and PRISMA guidelines. Only the randomized clinical trials were considered, and in cases of multiple publications, it was considered the most up to date information. Biases were highlighted. In the presence of homogeneous data, pooling statistics and meta-analysis were applied. The research led to 71 articles, 17 of which were eligible. Among the trials selected, eight investigated the effectiveness of EOs on anxiety, depression, and stress. Two concerned the treatment of nausea and vomiting, six evaluated the application of EOs on labor for pain treatment, and two showed the effectiveness in the treatment of episiotomy. The heterogeneity of works carried out so far has made it possible to develop a meta-analysis only in the field of pain treatment during childbirth, identifying the effectiveness of the EOs Lavandula spp. and Rosa damascena.

The antibacterial activity of methanolic extracts of eight Mediterranean brown algae belonging to the genera Cystoseira sensu lato, Dictyopteris and Padina was investigated. Extracts from Sargassaceae showed antibacterial activity. Specifically, Carpodesmia crinita and Carpodesmia brachycarpa extracts inhibited the growth of the Gram-positive strain Kocuria rhizophila whereas Cystoseira compressa and Carpodesmia amentacea extracts showed antibacterial activity against the Gram-positive strains, Kocuria rhizophila and Staphylococcus aureus. None of them inhibited the growth of the Gram-negative bacterium Escherichia coli. The extracts of Cystoseira sp. pl. and Treptacantha ballesterosii did not show any antibacterial activity. HPLC-MS analysis of the extracts allowed identification of polar compounds. Taxa of the genus Cystoseira sensu lato showed a higher number of compounds than Dictyopteris poly- podioides and Padina pavonica. Some differences found among and within taxa can be attributed to the different depths where these algae live, to thallus morphology and herbivore pressure. Particularly interesting were the phenol compounds, which showed differences among the genera but also within the genus Cystoseira sensu lato. Noteworthy, the almost exclusive presence of the fukiik acid isomer and of bromo-phloroglucinol in Carpodesmia amentacea, and the presence of exifone, able to bind iron metal ions, in Carpodesmia amentacea, Cystoseira compressa and Treptacantha ballesterosii.

As a reference laboratory for measles and rubella surveillance in Lombardy, we evaluated the association between SARS-CoV-2 infection and measles-like syndromes, providing preliminary evidence for undetected early circulation of SARS-CoV-2. Overall, 435 samples from 156 cases were investigated. RNA from oropharyngeal swabs (N = 148) and urine (N = 141) was screened with four hemi-nested PCRs and molecular evidence for SARS-CoV-2 infection was found in 13 subjects. Two of the positive patients were from the pandemic period (2/12, 16.7%, March 2020-March 2021) and 11 were from the pre-pandemic period (11/44, 25%, August 2019-February 2020). Sera (N = 146) were tested for anti-SARS-CoV-2 IgG, IgM, and IgA antibodies. Five of the RNA-positive individuals also had detectable anti-SARS-CoV-2 antibodies. No strong evidence of infection was found in samples collected between August 2018 and July 2019 from 100 patients. The earliest sample with evidence of SARS-CoV-2 RNA was from September 12, 2019, and the positive patient was also positive for anti-SARS-CoV-2 antibodies (IgG and IgM). Mutations typical of B.1 strains previously reported to have emerged in January 2020 (C3037T, C14408T, and A23403G), were identified in samples collected as early as October 2019 in Lombardy. One of these mutations (C14408T) was also identified among sequences downloaded from public databases that were obtained by others from samples collected in Brazil in November 2019. We conclude that a SARS-CoV-2 progenitor capable of producing a measles-like syndrome may have emerged in late June-late July 2019 and that viruses with mutations characterizing B.1 strain may have been spreading globally before the first Wuhan outbreak. Our findings should be complemented by high-throughput sequencing to obtain additional sequence information. We highlight the importance of retrospective surveillance studies in understanding the early dynamics of COVID-19 spread and we encourage other groups to perform retrospective investigations to seek confirmatory proofs of early SARS-CoV-2 circulation.

Purpose Tyro is a deep hypersaline anoxic basin (DHAB) located at the seafloor of the Eastern Mediterranean sea. Tyro hosts a stratified eukaryotic microbiome moving from seawater to the brine, but no reports are available on its prokaryotic community. We provide the first snapshot of the bacterial community structure in Tyro brine, seawater-brine interface, and the overlaying deep seawater. Methods In this study, we combined the use of molecular analyses, i.e., DNA fingerprinting and 16S rRNA pyrosequencing for the description of the bacterial community structure and taxonomy. PiCRUST2 was used to infer information on the prokaryotes functional diversity. A culture-dependent approach was applied to enrich bacteria of interest for marine biotechnology. Results Bacterial communities sharply clustered moving from the seawater to the Tyro brine, in agreement with the abrupt increase of salinity values. Moreover, specific taxonomic groups inhabited the seawater-brine interface compared to the overlaying seawater and their identification revealed converging taxonomy with other DHABs in the Eastern Mediterranean sea. Functional traits inferred from the prokaryote taxonomy in the upper interface and the overlaying seawater indicated metabolic pathways for the synthesis of osmoprotectants, likely involved in bacterial adaptation to the steep increasing salinity. Metabolic traits related to methane and methylated compounds and to hydrocarbon degradation were also revealed in the upper interface of Tyro. The overall capability of the Tyro microbiome for hydrocarbon metabolism was confirmed by the isolation of hydrocarbonoclastic bacteria in the sediments. Conclusions Our results suggest that Tyro seawater-brine interface hosts a specific microbiome adapted to the polyextreme condition typical of DHABs with potential metabolic features that could be further explored for the characterization of the metabolic network connecting the brine with the deep seawater through the chemocline. Moreover, Tyro could be a reservoir of culturable microbes endowed with functionalities of interest for biotechnological applications like hydrocarbon bioremediation.

Despite the increasing interest in using microbial-based technologies to support human space exploration, many unknowns remain not only on bioprocesses but also on microbial survivability and genetic stability under non-Earth conditions. Here the desert cyanobacterium Chroococcidiopsis sp. CCMEE 029 was investigated for robustness of the repair capability of DNA lesions accumulated under Mars-like conditions (UV radiation and atmosphere) simulated in low Earth orbit using the EXPOSE-R2 facility installed outside the International Space Station. Genomic alterations were determined in a space-derivate of Chroococcidiopsis sp. CCMEE 029 obtained upon reactivation on Earth of the space-exposed cells. Comparative analysis of whole-genome sequences showed no increased variant numbers in the space-derivate compared to triplicates of the reference strain maintained on the ground. This result advanced cyanobacteria-based technologies to support human space exploration.

The composition of the gut microbiome has been associated with clinical responses to immune checkpoint inhibitor (ICI) treatment, but there is limited consensus on the specific microbiome characteristics linked to the clinical benefits of ICIs. We performed shotgun metagenomic sequencing of stool samples collected before ICI initiation from five observational cohorts recruiting ICI-naive patients with advanced cutaneous melanoma (n = 165). Integrating the dataset with 147 metagenomic samples from previously published studies, we found that the gut microbiome has a relevant, but cohort-dependent, association with the response to ICIs. A machine learning analysis confirmed the link between the microbiome and overall response rates (ORRs) and progression-free survival (PFS) with ICIs but also revealed limited reproducibility of microbiome-based signatures across cohorts. Accordingly, a panel of species, including Bifidobacterium pseudocatenulatum, Roseburia spp. and Akkermansia muciniphila, associated with responders was identified, but no single species could be regarded as a fully consistent biomarker across studies. Overall, the role of the human gut microbiome in ICI response appears more complex than previously thought, extending beyond differing microbial species simply present or absent in responders and nonresponders. Future studies should adopt larger sample sizes and take into account the complex interplay of clinical factors with the gut microbiome over the treatment course.