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Objective: to analyze survival and factors associated with increased risk of death for older adults diagnosed with COVID-19, living in the Northeast region of Brazil. Method: retrospective observational study developed with secondary data provided by the Brazilian Ministry of Health, between June 14 and December 26, 2020. The Kaplan-Meyer method, the time-dependent cox regression model was used, including covariates (age, sex, skin color, comorbidities, admission to the ICU, ventilatory support). Results: out of 9,306 individuals analyzed, 55.9% died and 44.1% survived. The highest risk of death was observed for those aged 80-89 (HR=1.95), brown-skinned (HR=1.99), with immunodeficiency (HR=1.259) or kidney disease (HR=1.147), admitted to the ICU (HR=1,795) and in use of ventilatory support (HR=1606). Conclusion: among older adults residing in the Northeast region of Brazil, there was a higher risk of death from COVID-19 for octogenarians, brown-skinned, with comorbidities, hospitalization in the ICU, followed by the use of ventilatory support. The creation of health prevention strategies that identify older adults with these profiles is suggested to prevent deaths in future pandemic situations.

Does oligometastatic breast cancer (OMBC) deserve a dedicated treatment? Although some authors recommend multidisciplinary management of OMBC with a curative intent, there is no evidence proving this strategy beneficial in the absence of a randomized trial. The existing literature sheds little light on OMBC. Incidence is unknown; data available are either obsolete or biased; there is no consensus on the definition of OMBC and metastatic sites, nor on necessary imaging techniques. However, certain proposals merit consideration. Knowledge of eventual specific OMBC biological characteristics is limited to circulating tumor cell (CTC) counts. Given the data available for other cancers, studies on microRNAs (miRNAs), circulating tumor DNA (ctDNA) and genomic alterations should be developed Finally, safe and effective therapies do exist, but results of randomized trials will not be available for many years. Prospective observational cohort studies need to be implemented., Highlights • The incidence of oligometastatic breast cancer is unknown. • Only one publication provides information regarding the biology of these cancers. • Oligometastatic breast cancer and metastatic site definitions should be harmonized. • Prospective observational cohort studies are needed.

CD40 ligand (CD40L) mRNA stability is dependent on an activation-induced pathway that is mediated by the binding complexes containing the multifunctional RNA-binding protein, polypyrimidine tract-binding protein 1 (PTBP1) to a 3′ untranslated region of the transcript. To understand the relationship between regulated CD40L and the requirement for variegated expression during a T-dependent response, we engineered a mouse lacking the CD40L stability element (CD40LΔ5) and asked how this mutation altered multiple aspects of the humoral immunity. We found that CD40LΔ5 mice expressed CD40L at 60% wildtype levels, and lowered expression corresponded to significantly decreased levels of T-dependent Abs, loss of germinal center (GC) B cells and a disorganized GC structure. Gene expression analysis of B cells from CD40LΔ5 mice revealed that genes associated with cell cycle and DNA replication were significantly downregulated and genes linked to apoptosis upregulated. Importantly, somatic hypermutation was relatively unaffected although the number of cells expressing high-affinity Abs was greatly reduced. Additionally, a significant loss of plasmablasts and early memory B cell precursors as a percentage of total GL7+ B cells was observed, indicating that differentiation cues leading to the development of post-GC subsets was highly dependent on a threshold level of CD40L. Thus, regulated mRNA stability plays an integral role in the optimization of humoral immunity by allowing for a dynamic level of CD40L expression on CD4 T cells that results in the proliferation and differentiation of pre-GC and GC B cells into functional subsets.

Heat shock proteins (HSP) are critical elements for the preservation of cellular homeostasis by participating in an array of biological processes. In addition, HSP play an important role in cellular protection from various environmental stresses. HSP are part of a large family of different molecular mass polypeptides, displaying various expression patterns, subcellular localizations, and diversity functions. An unexpected observation was the detection of HSP on the cell surface. Subsequent studies have demonstrated that HSP have the ability to interact and penetrate lipid bilayers by a process initiated by the recognition of phospholipid heads, followed by conformational changes, membrane insertion, and oligomerization. In the present study, we described the interaction of HSPA8 (HSC70), the constitutive cytosolic member of the HSP70 family, with lipid membranes. HSPA8 showed high selectivity for negatively charged phospholipids, such as phosphatidylserine and cardiolipin, and low affinity for phosphatidylcholine. Membrane insertion was mediated by a spontaneous process driven by increases in entropy and diminished by the presence of ADP or ATP. Finally, HSPA8 was capable of driving into the lipid bilayer HSP90 that does not display any lipid biding capacity by itself. This observation suggests that HSPA8 may act as a membrane chaperone.

PE_PGRS proteins ofMycobacterium tuberculosis(Mtb) constitute a large family of complex modular proteins whose role is still unclear. Among those, we have previously shown, using the heterologous expression inMycobacterium smegmatis, that PE_PGRS3 containing a unique arginine-rich C-terminal domain, promotes adhesion to host cells. In this study, we investigate the role of PE_PGRS3 and its C-terminal domain directly inMtbusing functional deletion mutants. The results obtained here show that PE_PGRS3 is localized on the mycobacterial cell wall and its arginine-rich C-terminal region protrudes from the mycobacterial membrane and mediatesMtbentry into epithelial cells. Most importantly, this positively charged helical domain specifically binds phosphorylated phosphatidylinositols and cardiolipin, whereas it is unable to bind other phospholipids. Interestingly, administration of cardiolipin and phosphatidylinositol but no other phospholipids was able to turn-off expression ofpe_pgrs3 activated by phosphate starvation conditions. These findings suggest that PE_PGRS3 has the key role to serve as a bridge between mycobacteria and host cells by interacting with specific host phospholipids and extracting them from host cells, for their direct integration or as a source of phosphate, during phases of TB pathogenesis whenMtbis short of phosphate supply.

Introduction: gut microbiota (GM) seems to be involved in the pathophysiology and progression of both metabolic syndrome (MS) and obesity. The aim was to investigate GM's composition in patients with severe obesity, candidates for bariatric/metabolic surgery BMS. Materials and Methods: Multicentre, prospective, cohort study, enrolling 84 patients with BMI 40–55 kg/m2, divided by metabolic status (MS) in healthy (group A), pre-MS (B), or MS (C). Results: No differences were found regarding anthropometric, nutritional parameters, except for vitamin D. As a whole the alpha and beta diversity examinations showed no statistical differences in GM profile. A total of 5/7 phyla with relative frequencies were identified above 0.1% (Actinobacteria, Bacteroidetes, Firmicutes, Proteobacteria, Verrucomicrobia). Fusobacteria and Patescibacteria represented the less abundant. There were no significant differences in the top ten genera. Data on Bacteroidetes (inversely related to triglycerides and LDL and directly related to HDL levels) and on Firmicutes (opposite trend) relative abundances suggest no differences among the three conditions. No correlation between the relative abundance of the main phyla and plasmatic glucose levels was observed. Conclusions: In a selected cohort of patients with obesity, MS did not affect the preoperative GM's profile. Severe obesity, per se, seems to be an independent condition affecting GM.

Heat shock proteins (HSPs) are ubiquitous polypeptides expressed in all living organisms that participate in several basic cellular processes, including protein folding, from which their denomination as molecular chaperones originated. There are several HSPs, including HSPA5, also known as 78-kDa glucose-regulated protein (GRP78) or binding immunoglobulin protein (BIP) that is an ER resident involved in the folding of polypeptides during their translocation into this compartment prior to the transition to the Golgi network. HSPA5 is detected on the surface of cells or secreted into the extracellular environment. Surface HSPA5 has been proposed to have various roles, such as receptor-mediated signal transduction, a co-receptor for soluble ligands, as well as a participant in tumor survival, proliferation, and resistance. Recently, surface HSPA5 has been reported to be a potential receptor of some viruses, including the novel SARS-CoV-2. In spite of these observations, the association of HSPA5 within the plasma membrane is still unclear. To gain information about this process, we studied the interaction of HSPA5 with liposomes made of different phospholipids. We found that HSPA5 has a high affinity for negatively charged phospholipids, such as palmitoyl-oleoyl phosphoserine (POPS) and cardiolipin (CL). The N-terminal and C-terminal domains of HSPA5 were independently capable of interacting with negatively charged phospholipids, but to a lesser extent than the full-length protein, suggesting that both domains are required for the maximum insertion into membranes. Interestingly, we found that the interaction of HSPA5 with negatively charged liposomes promotes an oligomerization process via intermolecular disulfide bonds in which the N-terminus end of the protein plays a critical role. Electronic supplementary material The online version of this article (10.1007/s12192-020-01134-9) contains supplementary material, which is available to authorized users.

This Policy Brief succinctly presents the Ecological Observing System of the Adriatic Sea (ECOAdS), aimed at integrating the ecological and oceanographic dimensions within the conservation strategy of the Natura 2000 network, and to propose a way to go for its future development and maintenance. After a definition of marine ecological observatories, we describe the current structure of ECOAdS, its key components and potential relevance in relation to the main European strategies for biodiversity and marine observation for the next decade. Finally, we suggest some actions that could be undertaken for the future development of ECOAdS, targeting possible perspectives in different regional, macro-regional, national and European strategic contexts. This Policy Brief is one of the outcomes of the Interreg Italy-Croatia Project ECOSS (ECological Observing System in the Adriatic Sea: oceanographic observations for biodiversity; https://www.italy-croatia.eu/web/ecoss), which had the main purpose to design and carry out the first steps for the establishment of ECOAdS.

Funder: European Molecular Biology Laboratory; funder-id: http://dx.doi.org/10.13039/100013060, Funder: University of Cambridge, Many complex genomic rearrangements arise through template switch errors, which occur in DNA replication when there is a transient polymerase switch to an alternate template nearby in three-dimensional space. While typically investigated at kilobase-to-megabase scales, the genomic and evolutionary consequences of this mutational process are not well characterised at smaller scales, where they are often interpreted as clusters of independent substitutions, insertions and deletions. Here we present an improved statistical approach using pair hidden Markov models, and use it to detect and describe short-range template switches underlying clusters of mutations in the multi-way alignment of hominid genomes. Using robust statistics derived from evolutionary genomic simulations, we show that template switch events have been widespread in the evolution of the great apes’ genomes and provide a parsimonious explanation for the presence of many complex mutation clusters in their phylogenetic context. Larger-scale mechanisms of genome rearrangement are typically associated with structural features around breakpoints, and accordingly we show that atypical patterns of secondary structure formation and DNA bending are present at the initial template switch loci. Our methods improve on previous non-probabilistic approaches for computational detection of template switch mutations, allowing the statistical significance of events to be assessed. By specifying realistic evolutionary parameters based on the genomes and taxa involved, our methods can be readily adapted to other intra- or inter-species comparisons.

This letter takes a cue from a recently published study, which looked at diagnostic tests currently available for SARS-CoV-2 infection, to present and discuss the results of a re-evaluation of 122 positive serum samples from COVID-19 patients using an in-house serological assay for SARSCoV-2 antibodies detection. While serology targeting the SARS-CoV-2 S protein should be preferable, IgA testing is proposed in all situations for which serology is the solely practicable diagnostic strategy for SARSCoV-2 infection.

The present research was conducted on fish mesocosms using a non-lethal cadmium concentration as a trace metal pollutant to verify, respectively, whether post-remediation by Ulva lactuca reduces the need for activation of cellular antioxidative defenses and DNA repair mechanisms. Spectrophotometric evaluation of the antioxidant Glutathione S-Transferase and total soluble and fat-soluble antioxidant capacity, molecular detection of genotoxicity by mobility shift tests, and radio detection of DNA repair by PolyADP-ribosilation (PARP) were performed on the Lates calcarifer liver. Cadmium represents an environmental risk but its toxic effect seems mitigated by the bioaccumulation properties of Ulva lactuca. We detected high total soluble antioxidant capacity, low Glutathione S-Transferase levels as well as low levels of PARP and undamaged DNA in the fish liver when seaweeds were added as compared to standard feeding. These results suggest a safe and sustainable implementation of Lates calcarifer sea farming using Ulva lactuca.

The evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus leads to new variants that warrant timely epidemiological characterization. Here we use the dense genomic surveillance data generated by the COVID-19 Genomics UK Consortium to reconstruct the dynamics of 71 different lineages in each of 315 English local authorities between September 2020 and June 2021. This analysis reveals a series of subepidemics that peaked in early autumn 2020, followed by a jump in transmissibility of the B.1.1.7/Alpha lineage. The Alpha variant grew when other lineages declined during the second national lockdown and regionally tiered restrictions between November and December 2020. A third more stringent national lockdown suppressed the Alpha variant and eliminated nearly all other lineages in early 2021. Yet a series of variants (most of which contained the spike E484K mutation) defied these trends and persisted at moderately increasing proportions. However, by accounting for sustained introductions, we found that the transmissibility of these variants is unlikely to have exceeded the transmissibility of the Alpha variant. Finally, B.1.617.2/Delta was repeatedly introduced in England and grew rapidly in early summer 2021, constituting approximately 98% of sampled SARS-CoV-2 genomes on 26 June 2021., A study of the evolution of the SARS-CoV-2 virus in England between September 2020 and June 2021 finds that interventions capable of containing previous variants were insufficient to stop the more transmissible Alpha and Delta variants.

BackgroundHealthcare-associated wastewater reservoirs and asymptomatic gastrointestinal patient colonisation by carbapenemase-producing Enterobacterales (CPE) can be important in nosocomial CPE dissemination and infection. We characterised these niches and within-niche diversity in a blaKPC-associated CPE (KPC-E) endemic healthcare setting, to better understand transmission potential.MethodsWe systematically sampled wastewater sites and patients across three units (six wards) over 6-12 months in 2016 in a KPC-E endemic hospital. We used Illumina sequencing to characterise up to five isolates per sample. Recombination-adjusted phylogenies were used to define genetically related strains; assembly and mapping-based typing approaches were used to characterise antimicrobial resistance gene and insertion sequence profiles, and Tn4401 types/target site sequences. The wider accessory genome was evaluated in a subset of the largest clusters, and those crossing niches.FindingsWastewater site KPC-E-positivity was substantial (101/349 sites [28.9%] positive, 319/4,488 [7.1%] sampling events positive); 183/4,425 (4.1%) of patients were CPE culture-positive over the same timeframe. 13 species and 109 KPC-E strains were observed across niches, and 24% of wastewater and 26% of patient KPC-E-positive samples harboured ≥1 strain. Most diversity was explained by the individual niche, suggesting highly localised factors are important in selection and spread. Tn4401+target site sequence (TSS) diversity was greater in wastewater sites (p4401-associated transposition/evolution and dissemination. Shower/bath and sluice/mop-associated sites were more likely to be KPC-E-positive (Adjusted Odds Ratio [95% CI]: 2.69 [1.44-5.01], p=0.0019 and 2.60 [1.04-6.52], p=0.0410, respectively). Different strains had different transmission and blaKPC dissemination dynamics.InterpretationThere may be substantial KPC-E colonisation of wastewater sites and patients in KPC-E-endemic healthcare settings. Niche-specific factors, and different strains with different transmission dynamics influence carbapenemase gene dissemination. New transmission models incorporating complex, multi-level dynamics are needed to better quantify CPE dissemination to inform interventions and reduce transmission.FundingThis study was supported by the National Institute for Health Research, UK.RESEARCH IN CONTEXTEvidence before this studyWe searched PubMed to identify previous studies using whole genome sequencing (WGS) to characterise in-hospital dissemination and persistence of carbapenemase-producing Enterobacterales (CPEs) or the presence of CPEs in hospital wastewater reservoirs. In our search (01/Jan/1996-30/Sep/2021) we used the terms ((((CPE) OR (KPC)) AND (healthcare-associated OR nosocomial OR hospital-associated)) AND (genom*) AND ((environment OR patient))). We had no restrictions on language. We found sixty-six studies, but these were limited by: (i) evaluating single species or lineages only (n=34), (ii) including patients or environmental outbreaks only, and not both (n=23), (iii) sampling only small numbers (nex vivo or in vivo evolution), two studies did not use WGS, and one study was not primary research.Added value of this studyWe provide a comprehensive assessment of healthcare-associated CPE dissemination and persistence in hospital wastewater reservoirs through systematic sampling of a large number of environmental sites (n=349 sites, n=4,488 samples) and patients (n=4,425) over 6-12 months in a single hospital. For all CPE-positive environmental samples (n=319) and a subset of CPE-positive patient samples (n=97/399 [24.3%] samples from 76/183 [41.5%] CPE-positive patients), we sampled up to five CPE isolates per sample to capture within-niche diversity, and used short-read Illumina WGS (n=1,732 isolates successfully sequenced) to characterise within-niche diversity, changes in colonisation over time, and genetic overlap between patient and environmental niches.Implications of all the available evidenceThrough a detailed and resource-intensive study we captured the dynamics of seeding and dissemination of important carbapenemase genes amongst patients and environmental reservoirs within a hospital. We found differential colonisation and dissemination dynamics for different species and lineages within and between niches. Improved approaches incorporating variable within-niche diversity, accessory distances and horizontal gene transfer in transmission evaluations are required to better understand CPE dissemination and persistence, in order to direct interventions limiting transmission.

The evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus leads to new variants that warrant timely epidemiological characterization. Here we use the dense genomic surveillance datagenerated by the COVID-19 Genomics UK Consortium to reconstruct the dynamics of 71 different lineages in each of 315 English local authorities between September 2020 and June 2021. This analysis reveals a series of subepidemics that peaked in early autumn 2020, followed by a jump in transmissibility of the B.1.1.7/Alpha lineage. The Alpha variant grew when other lineages declined during the second national lockdown and regionally tiered restrictions between November and December 2020. A third more stringent national lockdown suppressed the Alpha variant and eliminated nearly all other lineages in early 2021. Yet a series of variants (most of which contained the spike E484K mutation) defied these trends and persisted at moderately increasing proportions. However, by accounting for sustained introductions, we found that the transmissibility of these variants is unlikely to have exceeded the transmissibility of the Alpha variant. Finally, B.1.617.2/Delta was repeatedly introduced in England and grew rapidly in early summer 2021, constituting approximately 98% of sampled SARS-CoV-2 genomes on 26 June 2021.

The vast scale of SARS-CoV-2 sequencing data has made it increasingly challenging to comprehensively analyze all available data using existing tools and file formats. To address this, we present a database of SARS-CoV-2 phylogenetic trees inferred with unrestricted public sequences, which we update daily to incorporate new sequences. Our database uses the recently-proposed mutation-annotated tree (MAT) format to efficiently encode the tree with branches labeled with parsimony-inferred mutations as well as Nextstrain clade and Pango lineage labels at clade roots. As of June 9, 2021, our SARS-CoV-2 MAT consists of 834,521 sequences and provides a comprehensive view of the virus’ evolutionary history using public data. We also present matUtils – a command-line utility for rapidly querying, interpreting and manipulating the MATs. Our daily-updated SARS-CoV-2 MAT database and matUtils software are available at http://hgdownload.soe.ucsc.edu/goldenPath/wuhCor1/UShER_SARS-CoV-2/ and https://github.com/yatisht/usher, respectively.

Tuberculosis (TB) is a difficult-to-treat infection because of multidrug regimen requirements based on drug susceptibility profiles and treatment observance issues. TB cure is defined by mycobacterial sterilization, technically complex to systematically assess. We hypothesized that microbiological outcome was associated with stage-specific immune changes in peripheral whole blood during TB treatment. The T-cell phenotypes of treated TB patients were prospectively characterized in a blinded fashion using mass cytometry after Mycobacterium tuberculosis (Mtb) antigen stimulation, and then correlated to sputum culture status. At two months of treatment, cytotoxic and terminally differentiated CD8+ T-cells were under-represented and naïve CD4+ T-cells were over-represented in positive-versus negative-sputum culture patients, regardless of Mtb drug susceptibility. At treatment completion, an antigen-driven T-cell immune shift towards differentiated subpopulations was associated with TB cure. Overall, we identified specific T-cell profiles associated with slow sputum converters, which brings new insights in TB prognostic biomarker research designed for clinical application.SummaryIn patients treated for pulmonary TB, high-dimensional immune profiling with mass cytometry revealed that Mycobacterium tuberculosis culture conversion is associated with newly characterized peripheral CD8+ T-cell phenotypes. This paves the way for new immune biomarkers associated with mycobacterial sterilization.

Funder: Alfred P. Sloan Foundation; funder-id: http://dx.doi.org/10.13039/100000879, Funder: European Molecular Biology Laboratory (EMBL), The SARS-CoV-2 pandemic has led to unprecedented, nearly real-time genetic tracing due to the rapid community sequencing response. Researchers immediately leveraged these data to infer the evolutionary relationships among viral samples and to study key biological questions, including whether host viral genome editing and recombination are features of SARS-CoV-2 evolution. This global sequencing effort is inherently decentralized and must rely on data collected by many labs using a wide variety of molecular and bioinformatic techniques. There is thus a strong possibility that systematic errors associated with lab-or protocol-specific practices affect some sequences in the repositories. We find that some recurrent mutations in reported SARS-CoV-2 genome sequences have been observed predominantly or exclusively by single labs, co-localize with commonly used primer binding sites and are more likely to affect the protein-coding sequences than other similarly recurrent mutations. We show that their inclusion can affect phylogenetic inference on scales relevant to local lineage tracing, and make it appear as though there has been an excess of recurrent mutation or recombination among viral lineages. We suggest how samples can be screened and problematic variants removed, and we plan to regularly inform the scientific community with our updated results as more SARS-CoV-2 genome sequences are shared (https://virological.org/t/issues-with-sars-cov-2-sequencing-data/473 and https://virological.org/t/masking-strategies-for-sars-cov-2-alignments/480). We also develop tools for comparing and visualizing differences among very large phylogenies and we show that consistent clade- and tree-based comparisons can be made between phylogenies produced by different groups. These will facilitate evolutionary inferences and comparisons among phylogenies produced for a wide array of purposes. Building on the SARS-CoV-2 Genome Browser at UCSC, we present a toolkit to compare, analyze and combine SARS-CoV-2 phylogenies, find and remove potential sequencing errors and establish a widely shared, stable clade structure for a more accurate scientific inference and discourse.

In Sulfolobus solfataricus, Sso, the ADP-ribosylating thermozyme is known to carry both auto- and heteromodification of target proteins via short chains of ADP-ribose. Here, we provide evidence that this thermoprotein is a multifunctional enzyme, also showing ATPase activity. Electrophoretic and kinetic analyses were performed using NAD+ and ATP as substrates. The results showed that ATP is acting as a negative effector on the NAD+-dependent reaction, and is also responsible for inducing the dimerization of the thermozyme. These findings enabled us to further investigate the kinetic of ADP-ribosylation activity in the presence of ATP, and to also assay its ability to work as a substrate. Moreover, since the heteroacceptor of ADP-ribose is the sulfolobal Sso7 protein, known as an ATPase, some reconstitution experiments were set up to study the reciprocal influence of the ADP-ribosylating thermozyme and the Sso7 protein on their activities, considering also the possibility of direct enzyme/Sso7 protein interactions. This study provides new insights into the ATP-ase activity of the ADP-ribosylating thermozyme, which is able to establish stable complexes with Sso7 protein.

In the context of climatic change, more severe and long-lasting droughts will modify the fitness of plants, with potentially worse consequences on the relict trees. We have investigated the leaf phenotypic (anatomical, physiological and biochemical) plasticity in well-watered, drought-stressed and re-watered plants of two populations of Platanus orientalis, an endangered species in the west of the Mediterranean area. The two populations originated in contrasting climate (drier and warmer, Italy (IT) population, more humid and colder, Bulgaria (BG) population). The IT control plants had thicker leaves, enabling them to maintain higher leaf water content in the dry environment, and more spongy parenchyma, which could improve water conductivity of these plants and may result in easier CO2 diffusion than in BG plants. Control BG plants were also characterized by higher photorespiration and leaf antioxidants compared to IT plants. BG plants responded to drought with greater leaf thickness shrinkage. Drought also caused substantial reduction in photosynthetic parameters of both IT and BG plants. After re-watering, photosynthesis did not fully recover in either of the two populations. However, IT leaves became thicker, while photorespiration in BG plants further increased, perhaps indicating sustained activation of defensive mechanisms. Overall, our hypothesis, that plants with a fragmented habitat (i.e., the IT population) lose phenotypic plasticity but acquire traits allowing better resistance to the climate where they became adapted, remains confirmed.

Phytocannabinoids (pCBs) are a large family of meroterpenoids isolated from the plant Cannabis sativa. &Delta, 9-Tetrahydrocannabinol (THC) and cannabidiol (CBD) are the best investigated phytocannabinoids due to their relative abundance and interesting bioactivity profiles. In addition to various targets, THC and CBD are also well-known agonists of peroxisome proliferator-activated receptor gamma (PPAR&gamma, ), a nuclear receptor involved in energy homeostasis and lipid metabolism. In the search of new pCBs potentially acting as PPAR&gamma, agonists, we identified cannabimovone (CBM), a structurally unique abeo-menthane pCB, as a novel PPAR&gamma, modulator via a combined computational and experimental approach. The ability of CBM to act as dual PPAR&gamma, /&alpha, agonist was also evaluated. Computational studies suggested a different binding mode toward the two isoforms, with the compound able to recapitulate the pattern of H-bonds of a canonical agonist only in the case of PPAR&gamma, Luciferase assays confirmed the computational results, showing a selective activation of PPAR&gamma, by CBM in the low micromolar range. CBM promoted the expression of PPAR&gamma, target genes regulating the adipocyte differentiation and prevented palmitate-induced insulin signaling impairment. Altogether, these results candidate CBM as a novel bioactive compound potentially useful for the treatment of insulin resistance-related disorders.

Mediterranean tomato landraces adapted to arid environments represent an option to counteract drought, and to address the complexity of responses to water deficit and recovery, which is a crucial component of plant adaptation mechanisms. We investigated physiological, biochemical and molecular responses of two Mediterranean tomato landraces, "Locale di Salina" (Lc) and "Pizzutello di Sciacca" (Pz) under two dehydration periods and intermediate rehydration in greenhouse pot-experiments. Relationship between CO2 assimilation and stomatal conductance under severe water stress (gs < 0.05 mol m-2 s-1 ) indicated the occurrence of stomatal and non-stomatal limitations of photosynthesis. Gas-exchanges promptly recovered within 2-3-day re-hydration. ABA and gs showed a strict exponential relationship. Both leaf ABA and proline peaked under severe water stress. Lc showed higher accumulation of ABA and higher induction of the expression of both NCED and P5CS genes than Pz. PARP increased during imposition of stress, mainly in Lc, and decreased under severe water stress. The two landraces hardly differed in their physiological performance. Under severe water stress, stomatal conductance showed low sensitivity to ABA, which instead controlled stomatal closure under moderate water stress (gs > 0.15 mol m-2 s-1 ). The prompt recovery after re-dehydration of both landraces confirmed their drought tolerant behaviour. Differences between the two landraces were instead observed at biochemical and molecular levels. This article is protected by copyright. All rights reserved.

DING proteins represent a new group of 40 kDa-related members, ubiquitous in living organisms. The family also include the DING protein from Sulfolobus solfataricus, functionally related to poly(ADP-ribose) polymerases. Here, the archaeal protein has been compared with the human Phosphate-Binding Protein and the Pseudomonas fluorescence DING enzyme, by enzyme assays and immune cross-reactivity. Surprisingly, as the Sulfolobus enzyme, the Human and Pseudomonas proteins display poly(ADP-ribose) polymerase activity, whereas a phosphatase activity was only present in Sulfolobus and human protein, despite the conserved phosphate-binding site residues in Pseudomonas DING. All proteins were positive to anti-DING antibodies and gave a comparable pattern of anti-poly(ADP-ribose) polymerase immunoreactivity with two bands, at around 40 kDa and roughly at the double of this molecular mass. The latter signal was present in all Sulfolobus enzyme preparations and proved not due to either a contaminant or a precursor protein, but likely being a dimeric form of the 40 kDa polypeptide. The common immunological and partly enzymatic behavior linking human, Pseudomonas and Sulfolobus DING proteins, makes the archaeal protein an important model system to investigate DING protein function and evolution within the cell.

Accurate and timely detection of recombinant lineages is crucial for interpreting genetic variation, reconstructing epidemic spread, identifying selection and variants of interest, and accurately performing phylogenetic analyses. During the SARS-CoV-2 pandemic, genomic data generation has exceeded the capacities of existing analysis platforms, thereby crippling real-time analysis of viral recombination. Low SARS-CoV-2 mutation rates make detecting recombination difficult. Here, we develop and apply a novel phylogenomic method to exhaustively search a nearly comprehensive SARS-CoV-2 phylogeny for recombinant lineages. We investigate a 1.6M sample tree, and identify 606 recombination events. Approximately 2.7% of sequenced SARS-CoV-2 genomes have recombinant ancestry. Recombination breakpoints occur disproportionately in the Spike protein region. Our method empowers comprehensive real time tracking of viral recombination during the SARS-CoV-2 pandemic and beyond.

Objectives: To describe a rapid workflow based on the direct detection of Escherichia coli (Ec) and Klebsiella pneumoniae (Kp) producing CTX-M extended-spectrum β-lactamase (ESBL) and/or carbapenemases (eg, KPC, VIM) from blood cultures (BCs) and the infectious disease (ID) consulting for timely appropriate antimicrobial therapy. Methods: This observational, retrospective study included adult patients with a first episode of Ec or Kp bloodstream infection (BSI) in a large Italian university hospital, where an inpatient ID consultation team (IDCT) has been operational. Results from the BCs tested for detecting blaCTX-M, blaKPC, blaNDM, blaOXA-48-like, and blaVIM genes by the eazyplex® SuperBug CRE assay in Ec and Kp organisms had been notified for antimicrobial therapy consulting. Results: In 321 BSI episodes studied, we found that 151 (47.0%) of Ec or Kp organisms harbored blaCTX-M and/or blaKPC and/or blaVIM (meantime from BC collection: 18.5 h). Empirical antimicrobial treatment was appropriate in 21.8% (33/151) of BSIs, namely 5.9% (3/51) of BSIs caused by KPC/VIM producers and 30.0% (30/100) of BSIs caused by CTX-M producers. After notification of results, the IDCT modified antimicrobial therapy (mean time from BC collection: 20 h) such that the proportion of appropriate treatments increased to 84.8% (128/151) of BSIs, namely 70.6% (36/51) of BSIs caused by KPC/VIM producers and 92.0% (92/100) of BSIs caused by CTX-M producers. Conclusion: Our study shows that a rapid diagnostic-driven clinical strategy allowed for early prescription of potentially effective antimicrobial therapy in BSIs caused by CTX-M ESBL- and/or KPC/VIM carbapenemase-producing Ec and Kp organisms.

Tuberculosis (TB) remains a global threat and there is an urgent need for improved drugs and treatments, particularly against the drug-resistant strains of Mycobacterium tuberculosis (Mtb). Graphene oxide (GO) is an innovative bi-dimensional nanomaterial that when administered in vivo accumulates in the lungs. Further, GO is readily degraded by peroxidases and has a high drug loading capacity and antibacterial properties. In this study, we first evaluated the GO anti-mycobacterial properties using Mycobacterium smegmatis (Ms) as a model. We observed that GO, when administered with the bacteria, was able to trap Ms in a dose-dependent manner, reducing entry of bacilli into macrophages. However, GO did not show any anti-mycobacterial activity when used to treat infected cells or when macrophages were pre-treated before infection. Similar results were obtained when the virulent Mtb strain was used, showing that GO was able to trap Mtb and prevent entry into microphages. These results indicate that GO can be a promising tool to design improved therapies against TB.

Background Older aged adults and those with pre-existing conditions are at highest risk for severe COVID-19 associated outcomes. Methods Using a large dataset of genome-wide RNA-seq profiles derived from human dermal fibroblasts (GSE113957) we investigated whether age affects the expression of pattern recognition receptor (PRR) genes and ACE2, the receptor for SARS-CoV-2. Results Extremes of age are associated with increased expression of selected PRR genes, ACE2 and four genes that encode proteins that have been shown to interact with SAR2-CoV-2 proteins. Conclusions Assessment of PRR expression might provide a strategy for stratifying the risk of severe COVID-19 disease at both the individual and population levels.

Objectives Genotyping of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been instrumental in monitoring viral evolution and transmission during the pandemic. The quality of the sequence data obtained from these genotyping efforts depends on several factors, including the quantity/integrity of the input material, the technology, and laboratory-specific implementation. The current lack of guidelines for SARS-CoV-2 genotyping leads to inclusion of error-containing genome sequences in genomic epidemiology studies. We aimed to establish clear and broadly applicable recommendations for reliable virus genotyping. Methods We established and used a sequencing data analysis workflow that reliably identifies and removes technical artefacts; such artefacts can result in miscalls when using alternative pipelines to process clinical samples and synthetic viral genomes with an amplicon-based genotyping approach. We evaluated the impact of experimental factors, including viral load and sequencing depth, on correct sequence determination. Results We found that at least 1000 viral genomes are necessary to confidently detect variants in the SARS-CoV-2 genome at frequencies of ≥10%. The broad applicability of our recommendations was validated in over 200 clinical samples from six independent laboratories. The genotypes we determined for clinical isolates with sufficient quality cluster by sampling location and period. Our analysis also supports the rise in frequencies of 20A.EU1 and 20A.EU2, two recently reported European strains whose dissemination was facilitated by travel during the summer of 2020. Conclusions We present much-needed recommendations for the reliable determination of SARS-CoV-2 genome sequences and demonstrate their broad applicability in a large cohort of clinical samples.

In the absence of effective treatment, COVID-19 is likely to remain a global disease burden. Compounding this threat is the near certainty that novel coronaviruses with pandemic potential will emerge in years to come. Pan-coronavirus drugs – agents active against both SARS-CoV-2 and other coronaviruses – would address both threats. A strategy to develop such broad-spectrum inhibitors is to pharmacologically target binding sites on SARS-CoV-2 proteins that are highly conserved in other known coronaviruses, the assumption being that any selective pressure to keep a site conserved across past viruses will apply to future ones. Here, we systematically mapped druggable binding pockets on the experimental structure of fifteen SARS-CoV-2 proteins and analyzed their variation across twenty-seven α- and β-coronaviruses and across thousands of SARS-CoV-2 samples from COVID-19 patients. We find that the two most conserved druggable sites are a pocket overlapping the RNA binding site of the helicase nsp13, and the catalytic site of the RNA-dependent RNA polymerase nsp12, both components of the viral replication-transcription complex. We present the data on a public web portal (https://www.thesgc.org/SARSCoV2_pocketome/) where users can interactively navigate individual protein structures and view the genetic variability of drug binding pockets in 3D.

Sepsis is a life-threatening condition that arises from a poorly regulated inflammatory response to pathogenic organisms. Current treatments are limited to antibiotics, fluid resuscitation, and other supportive therapies. New targets for monitoring disease progression and therapeutic interventions are therefore critically needed. We previously reported that lipocalin-2 (Lcn2), a bacteriostatic mediator with potent pro-apoptotic activities, was robustly induced in sepsis. Other studies showed that Lcn2 was a predictor of mortality in septic patients. However, how Lcn2 is regulated during sepsis is poorly understood. We evaluated how IkBζ, an inducer of Lcn2, was regulated in sepsis using both the cecal ligation and puncture (CLP) and endotoxemia (LPS) animal models. We show that Nfkbiz, the gene encoding for IkBζ, was rapidly stimulated but, unlike Lcn2, whose expression persists during sepsis, mRNA levels of Nfkbiz decline to near basal levels several hours after its induction. In contrast, we observed that IkBζ expression remained highly elevated in septic animals following CLP but not LPS, indicating the occurrence of a CLP-specific mechanism that extends IkBζ half-life. By using aninhibitor of IkBζ, we determined that the expression of Lcn2 was largely controlled by IkBζ. Altogether, these data indicate that the high IkBζ expression in tissues likely contributes to the elevated expression of Lcn2 in sepsis. Since IkBζ is also capable of promoting or repressing other inflammatory genes, it might exert a central role in sepsis.

Several studies evaluated the presence of SARS-CoV-2 in the environment. Saliva and nasopharyngeal droplets can land on objects and surfaces, creating fomites., Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) environmental contamination occurs through droplets and biological fluids released in the surroundings from patients or asymptomatic carriers. Surfaces and objects contaminated by saliva or nose secretions represent a risk for indirect transmission of coronavirus disease 2019 (COVID-19). We assayed surfaces from hospital and living spaces to identify the presence of viral RNA and the spread of fomites in the environment. Anthropic contamination by droplets and biological fluids was monitored by detecting the microbiota signature using multiplex quantitative real-time PCR (qPCR) on selected species and massive sequencing on 16S amplicons. A total of 92 samples (flocked swabs) were collected from critical areas during the pandemic, including indoor (three hospitals and three public buildings) and outdoor surfaces exposed to anthropic contamination (handles and handrails, playgrounds). Traces of biological fluids were frequently detected in spaces open to the public and on objects that are touched with the hands (>80%). However, viral RNA was not detected in hospital wards or other indoor and outdoor surfaces either in the air system of a COVID hospital but only in the surroundings of an infected patient, in consistent association with droplet traces and fomites. Handled objects accumulated the highest level of multiple contaminations by saliva, nose secretions, and fecal traces, further supporting the priority role of handwashing in prevention. In conclusion, anthropic contamination by droplets and biological fluids is widespread in spaces open to the public and can be traced by qPCR. Monitoring fomites can support evaluation of indirect transmission risks for coronavirus or other flu-like viruses in the environment. IMPORTANCE Several studies have evaluated the presence of SARS-CoV-2 in the environment. Saliva and nasopharyngeal droplets can land on objects and surfaces, creating fomites. A suitable indicator would allow the detection of droplets or biofluids carrying the virus. Therefore, we searched for viral RNA and droplets and fomites on at risk surfaces. We monitored by qPCR or next generation sequencing (NGS) droplets through their microbiota. Although the study was performed during the pandemic, SARS-CoV-2 was not significantly found on surfaces, with the only exception of environmental areas near infectious patients. Conversely, anthropic contamination was frequent, suggesting a role for biofluids as putative markers of indirect transmission and risk assessment. Moreover, all SARS-CoV-2-contaminated surfaces showed droplets’ microbiota. Fomite monitoring by qPCR may have an impact on public health strategies, supporting prevention of indirect transmission similarly to what is done for other communicable diseases (e.g., influenza and influenza-like infections).

F-type plasmids are diverse and of great clinical significance, often carrying genes conferring antimicrobial resistance (AMR) such as extended-spectrum β-lactamases, particularly in Enterobacterales. Organising this plasmid diversity is challenging, and current knowledge is largely based on plasmids from clinical settings. Here, we present a network community analysis of a large survey of F-type plasmids from environmental (influent, effluent and upstream/downstream waterways surrounding wastewater treatment works) and livestock settings. We use a tractable and scalable methodology to examine the relationship between plasmid metadata and network communities. This reveals how niche (sampling compartment and host genera) partition and shape plasmid diversity. We also perform pangenome-style analyses on network communities. We show that such communities define unique combinations of core genes, with limited overlap. Building plasmid phylogenies based on alignments of these core genes, we demonstrate that plasmid accessory function is closely linked to core gene content. Taken together, our results suggest that stable F-type plasmid backbone structures can persist in environmental settings while allowing dramatic variation in accessory gene content that may be linked to niche adaptation. The association of F-type plasmids with AMR may reflect their suitability for rapid niche adaptation.

Studies have shown that HCV subtype 3a had likely been circulating in South Asia before its global spread. However, the time and route of this dissemination remain unclear. For the first time, we generated host and virus genome-wide data for more than 500 patients infected with HCV subtype 3a from the UK, North America, Australia and New Zealand. We used the host genomic data to infer the ancestry of the patients and used this information to investigate the epidemic history of HCV subtype 3a. We observed that viruses from hosts of South Asian ancestry clustered together near the root of the tree, irrespective of the sampling country and that they were more diverse than viruses from other host ancestries. We also inferred that three independent transmission events resulted in the spread of the virus from South Asia to the UK, North America and the Australian continent. This initial spread happened during or soon after the end of the second world war. This was followed by an exponential growth in the effective population size of HCV subtype 3a worldwide and many independent transmissions between the UK, North America and Australian continent. Using both host and virus genomic information can be highly informative in studying the virus epidemic history especially in the context of chronic infections.

Weather and the susceptibility of children to severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection is still a debated question and currently a hot topic, particularly in view of important decisions regarding opening schools. Therefore, we performed this prospective analysis of anti‐SARS‐CoV‐2 immunoglobulin G (IgG) antibodies in children with known household exposure to SARS‐CoV‐2 and compared their IgG status with the other adults exposed to the index case in the same household. A total of 30 families with a documented COVID‐19 index case were included. A total of 44 out of 80 household contacts (55%) of index patients had anti SARS‐CoV‐2 IgG antibodies. In particular, 16/27 (59,3%) adult partners had IgG antibodies compared with 28/53 (52,3%) of pediatric contacts (p > .05). Among the pediatric population, children ≥5 years of age had a similar probability of having SARS‐CoV‐2 IgG antibodies (21/39, 53.8%) compared to those less than 5 years old (7/14, 50%) (p > .05). Adult partners and children also had a similar probability of having SARS‐CoV‐2 IgG antibodies. Interestingly, 10/28 (35.7%) of children and 5/27 (18.5%) of adults with SARS‐CoV‐2 IgG antibodies were previously diagnosed as COVID‐19 cases. Our study shows evidence of a high rate of IgG antibodies in children exposed to SARS‐CoV‐2. This report has public health implications, highlighting the need to establish appropriate guidelines for school openings and other social activities related to childhood.

Among molecular assays currently developed for detection and identification of pathogens (and their antimicrobial resistance genes) in positive blood cultures (BCs) (1), the BioFire FilmArray blood culture identification (BCID) panel (bioMerieux, Marcy l'Etoile, France)-a multiplex PCR assay with less than 2 minutes of hands-on time and a ∼1-hour turnaround time-allows syndromic diagnosis of bloodstream infection (BSI) (2, 3). Previously, the panel could identify 24 etiological agents of BSI (11 Gram-negative bacteria, 8 Gram-positive bacteria, and 5 yeast species), as well as three antimicrobial resistance genes (mecA, vanA/B, and bla KPC, which encodes Klebsiella pneumoniae carbapenemase). Now, the BioFire FilmArray BCID2 panel encompasses 43 molecular targets associated with BSI, including 15 Gram-negative bacteria, 11 Gram-positive bacteria, 7 yeast species, and 10 antimicrobial resistance genes (https://www.biomerieux-diagnostics.com/biofire-bcid-panel). The last targets include genes encoding for carbapenemases (IMP, KPC, OXA-48-like, NDM, and VIM), colistin resistance (mcr-1), ESBL (CTX-M), methicillin-resistance (mecA/C and, specifically for methicillin-resistant Staphylococcus aureus [MRSA], mecA/C and MREJ [mec right-extremity junction]), or vancomycin resistance (vanA/B). Unlike BCID, no published studies to date reported on the BCID2 performance. This study evaluated and compared the accuracy of BCID2 with that of BCID to identify bacterial species and relative antimicrobial resistance genes directly from positive BCs.

The COVID-19 pandemic has seen an unprecedented response from the sequencing community. Leveraging the sequence data from more than 140,000 SARS-CoV-2 genomes, we study mutation rates and selective pressures affecting the virus. Understanding the processes and effects of mutation and selection has profound implications for the study of viral evolution, for vaccine design, and for the tracking of viral spread. We highlight and address some common genome sequence analysis pitfalls that can lead to inaccurate inference of mutation rates and selection, such as ignoring skews in the genetic code, not accounting for recurrent mutations, and assuming evolutionary equilibrium. We find that two particular mutation rates, G →U and C →U, are similarly elevated and considerably higher than all other mutation rates, causing the majority of mutations in the SARS-CoV-2 genome, and are possibly the result of APOBEC and ROS activity. These mutations also tend to occur many times at the same genome positions along the global SARS-CoV-2 phylogeny (i.e., they are very homoplasic). We observe an effect of genomic context on mutation rates, but the effect of the context is overall limited. Although previous studies have suggested selection acting to decrease U content at synonymous sites, we bring forward evidence suggesting the opposite.

The present study reports data on a 20 months campaign monitoring enteric viruses (hepatitis A, norovirus, rotavirus, astrovirus, sapovirus, and aichivirus) and bacteria (Salmonella spp.) in seawater. The aim of this work was to assess the potential correlation among the presence of viruses/bacteria and different environmental factors like seasonality, water discharge sources (treated and untreated wastewater, mixed waters and raw water) as well as influence of the Italian lockdown measure against COVID-19 pandemic. Results showed different prevalence of the investigated viruses with values equal to 16 % for norovirus GI, 15.1 % for norovirus GII, followed by 13.8 % for astrovirus, and 13.3 % for sapovirus. Rotavirus was detected in the 8.4 % of samples and aichivirus was detected with the lowest prevalence of 3.5 %. Hepatitis A virus was never identified in the monitoring campaign. Salmonella spp. was detected with a prevalence of 36.6 %. Statistical analysis displayed a high correlation for the two noroviruses simultaneous detection (NGI and NGII) while a lower correlation was found for co-presence of noroviruses with astrovirus, sapovirus or Salmonella spp. A significant decrease of enteric pathogens in seawater was observed during the restrictions period. Results on seasonality highlighted a higher viral prevalence correlated to the wet season for all the pathogens but rotavirus and aichivirus, which instead showed an opposite trend and a higher incidence in the dry season. With respect to discharge typology, some viruses displayed a higher prevalence in treated waters (astrovirus, rotavirus, sapovirus and aichivirus) while the other investigated pathogens (noroviruses and Salmonella spp.) showed a higher prevalence in mixed waters. The main observations of this work were used to define a potential monitoring strategy that could be useful for sanitary Authorities to implement surveillance plans aimed at preventing possible sanitary outbreaks and/or environmental quality deterioration.

Anthropometrics and sex influence balance performances, and visual information can change anthropometrics’ relation and the postural sway. Therefore, the aim of the present study was to evaluate the effect of anthropometric characteristics, sex, and visual biofeedback and/or their interaction on a computerized wobble board. Twenty-seven (14 females, 13 males) young adults performed three 30-s double leg stance trials on a wobble board during two conditions: with visual and without visual biofeedback. Visual biofeedback improved (p = 0.010) balance on a wobble board with respect to the condition without visual biofeedback. Regardless of sex, no differences between conditions were found (p = 0.088). When investigating the effect of anthropometrics variables, sex, and their interactions on conditions, a significant main effect of the lower limb/height ratio, sex, and their interaction on the condition without visual biofeedback was found (p = 0.0008, R2 = 0.57). For the visual biofeedback condition, significant effects for sex and body mass (p = 0.0012, R2 = 0.43) and sex and whole-body moment of inertia (p = 0.0030, R2 = 0.39) were found. Results from the present study showed (1) visual biofeedback improved wobble board balance performance, (2) a significant main effect of lower limb/height ratio, sex, and their interaction on the wobble board performances without visual biofeedback emerged, (3) significant effects were found for sex and body mass and sex and moment of inertia in the visual biofeedback condition. Findings from the present study could have an impact on training and evaluations protocols, especially when several populations such as children, athletes, older adults and people with balance disorders are involved.

In this chapter, we provide a methodological description of the process to perform gastrointestinal (GIT) microbiota profiling on human stool samples. The process includes: (i) collection of feces, (ii) isolation of DNA from fecal community bacteria, (iii) selection of both 16S rDNA sequencing target and next-generation sequencing platform, and (iv) analysis and interpretation of sequence data. The process culminates into a comprehensive report on the GIT microbiota composition and structure that may translate into clinically actionable results.

The aim of this study was to characterize COVID-19 (SARS-CoV-2-infected) patients who develop bloodstream infection (BSI) and to assess risk factors associated with in-hospital mortality. We conducted a retrospective observational study of adult patients admitted for ≥48 h to a large Central Italy hospital for COVID-19 (1 March to 31 May 2020) who had or had not survived at discharge. We included only patients having blood cultures drawn or other inclusion criteria satisfied. Kaplan–Meier survival or Cox regression analyses were performed of 293 COVID-19 patients studied, 46 patients (15.7%) had a hospital-acquired clinically relevant BSI secondary to SARS-CoV-2 infection, accounting for 58 episodes (49 monomicrobial and 9 polymicrobial) in total. Twelve episodes (20.7%) occurred at day 3 of hospital admission. Sixty-nine species were isolated, including Staphylococcusaureus (32.8%), Enterobacterales (20.7%), Enterococcusfaecalis (17.2%), Candida (13.8%) and Pseudomonas aeruginosa (10.3%). Of 69 isolates, 27 (39.1%) were multidrug-resistant organisms. Twelve (54.5%) of 22 patients for whom empirical antimicrobial therapy was inappropriate were infected by a multidrug-resistant organism. Of 46 patients, 26 (56.5%) survived and 20 (43.5%) died. Exploring variables for association with in-hospital mortality identified > 75-year age (HR 2.97, 95% CI 1.15–7.68, p = 0.02), septic shock (HR 6.55, 95% CI 2.36–18.23, p < 0.001) and BSI onset ≤ 3 days (HR 4.68, 95% CI 1.40–15.63, p = 0.01) as risk factors independently associated with death. In our hospital, mortality among COVID-19 patients with BSI was high. While continued vigilance against these infections is essential, identification of risk factors for mortality may help to reduce fatal outcomes in patients with COVID-19.

Background The severe acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2) has a tropism for the gastrointestinal tract and several studies have shown an alteration of the gut microbiota in hospitalized infected patients. However, long-term data on microbiota changes after recovery are lacking. Methods We enrolled 30 patients hospitalized for SARS‑CoV‑2-related pneumonia. Their gut microbiota was analyzed within 48 h from the admission and compared with (1) that of other patients admitted for suspected bacterial pneumonia (control group) (2) that obtained from the same subject 6 months after nasopharyngeal swab negativization. Results Gut microbiota alpha-diversity increased 6 months after the resolution of SARS-CoV-2 infection. Bacteroidetes relative abundance was higher (≈ 36.8%) in patients with SARS-CoV-2, and declined to 18.7% when SARS-CoV-2 infection resolved (p = 0.004). Conversely, Firmicutes were prevalent (≈ 75%) in controls and in samples collected after SARS-CoV-2 infection resolution (p = 0.001). Ruminococcaceae, Lachnospiraceae and Blautia increased after SARS-CoV-2 infection resolution, rebalancing the gut microbiota composition. Conclusion SARS-CoV-2 infection is associated with changes in the gut microbiome, which tend to be reversed in long-term period.

Isoniazid (INH) is the cornerstone of the anti-tuberculosis regimens and emergence of Mycobacterium tuberculosis (Mtb) resistant strains is a major threat to our ability to control tuberculosis (TB) at global level. Mutations in the gene coding the catalase KatG confer resistance to high level of INH. In this paper, we describe for the first time a complete deletion of the genomic region containing the katG gene in an Mtb clinical strain isolated in Italy in a patient with HIV infection that previously completed INH preventive therapy. We genotypically characterized the Mtb strain and showed that katG deletion confers high-level resistance to INH (MIC > 25.6 μg/mL). The katG deletion did not impact significantly on Mtb fitness as we did not detect enhanced susceptibility to H2O2 compared to the wild type Mtb strains nor impaired growth in in vitro infection models. These findings highlight the ability of Mtb to acquire resistance to INH while maintaining fitness and pathogenic potential.

Ti3C2TX nano-sheets (MXenes) with excellent light-conversion capacity have gained importance in treating infectious diseases due to their limited bacterial resistance. In this study, we exploit this property to design photothermal antibacterial therapy using few- (FX) and multi-layer (MX) Ti3C2Tx nano-sheets. We demonstrate that FX have a higher cytocompatibility and conversion of light to heat, but MX show a better efficacy in inhibiting growth of S. aureus and E. coli due to MXenes’ reversible bacteria trapping. For MX (25 µg/mL), □37% of E. coli and □23% of S. aureus cells survived, while the effect was less pronounced for FX with □72% of E. coli and □46% of S. aureus viable cells after treatment. After using 100 µg/mL of MX, □11% of E. coli and □4% of S. aureus survived, while FX had only a mild effect on both species. The NIR laser treatment increased the efficacy of both materials: 100 µg/mL of MX combined with 5 min laser treatment at 5.7 W cm − 2 completely killed both species. For FX, the treatment with 3 W cm − 2 and the highest concentration (100 µg/mL) induced an effect comparable to MX (87% on E. coli, 95% on S.aureus). The combined NIR-MXene treatment causes an irreversible cell death linked to the loss of cell integrity (DNA release quantification and bacteria debris observation).

PE_PGRS proteins are surface antigens ofMycobacterium tuberculosis(Mtb) and a few other pathogenic mycobacteria. The PE_PGRS33 protein is among the most studied PE_PGRSs. It is known that the PE domain of PE_PGRS33 is required for the protein translocation through the mycobacterial cell wall, where the PGRS domain remains available for interaction with host receptors. Interaction with Toll like receptor 2 (TLR2) promotes secretion of inflammatory chemokines and cytokines, which are key in the immunopathogenesis of tuberculosis (TB). In this review, we briefly address some key challenges in the development of a TB vaccine and attempt to provide a rationale for the development of new vaccines aimed at fostering a humoral response againstMtb. Using PE_PGRS33 as a model for a surface-exposed antigen, we exploit the availability of current structural data using homology modeling to gather insights on the PGRS domain features. Our study suggests that the PGRS domain of PE_PGRS33 exposes four PGII sandwiches on the outer surface, which, we propose, are directly involved through their loops in the interactions with the host receptors and, as such, are promising targets for a vaccination strategy aimed at inducing a humoral response.

The 70 kDa heat shock proteins (Hsp70) are prone to self-assembly under thermal stress conditions, forming supramolecular assemblies (SMA), what may have detrimental consequences for cellular viability. In mitochondria, the cochaperone Hsp70-escort protein 1 (Hep1) maintains mitochondrial Hsp70 (mtHsp70) in a soluble and functional state, contributing to preserving proteostasis. Here we investigated the interaction between human Hep1 (hHep1) and HSPA9 (human mtHsp70) or HSPA1A (Hsp70-1A) in monomeric and thermic SMA states to unveil further information about the involved mechanisms. hHep1 was capable of blocking the formation of HSPA SMAs under a thermic treatment and stimulated HSPA ATPase activity in both monomeric and preformed SMA. The interaction of hHep1 with both monomeric and SMA HSPAs displayed a stoichiometric ratio close to 1, suggesting that hHep1 has access to most protomers within the SMA. Interestingly, hHep1 remodeled HSPA9 and HSPA1A SMAs into smaller forms. Furthermore, hHep1 was detected in the mitochondria and nucleus of cells transfected with the respective coding DNA and interacted with liposomes resembling mitochondrial membranes. Altogether, these new features reinforce that hHep1 act as a "chaperone for a chaperone", which may play a critical role in cellular proteostasis.

Ocean Acidification (OA) is likely to affect macroalgal diversity in the future with species-specific responses shaping macroalgal communities. In this framework, it is important to focus research on the photosynthetic response of habitat-forming species which have an important structural and functional role in coastal ecosystems. Most of the studies on the impacts of OA involve short-term laboratory or micro/mesocosm experiments. It is more challenging to assess the adaptive responses of macroalgal community to decreasing ocean pH over long-term periods, as they represent the basis of trophic dynamics in marine environments. This work aims to study the physiological traits of a population of Sargassum vulgare that lives naturally in the high pCO 2 vents system in Ischia (Italy), in order to predict the species behaviour in a possible OA future scenario. With this purpose, the photosynthetic performance of S. vulgare was studied in a wild, natural population living at low pH (6.7) as well as in a population transplanted from native (6.7) to ambient pH (8.1) for three weeks. The main results show that the photochemical activity and Rubisco expression decreased by 30% after transplanting, whereas the non-photochemical dissipation mechanisms and the photosynthetic pigment content increased by 50% and 40% respectively, in order to compensate for the decrease in photochemical efficiency at low pH. Our data indicated a stress condition for the S. vulgare population induced by pH variation, and therefore a reduced acclimation capability at different pH conditions. The decline of the PS II maximum quantum yield (F v /F m ) and the increase of PARP enzyme activity in transplanted thalli further supported this hypothesis. The absence of the species at ambient pH conditions close to the vent system, as well as the differences in physiological traits, suggest a local adaptation of S. vulgare at pH 6.7, through optimization of photosynthetic performance.

The ageing process in living organisms is characterised by the accumulation of several deleterious changes occurring in cells and tissues. The increase of reactive oxygen species with the advancement of age is responsible for the oxidative damage to proteins, lipids and DNA, enhancing the risk of diseases. The antioxidant response and the activation of the poly(ADP-ribosyl)ation process represent the first defences activated by organisms at all life stages to counteract damage to cell structures and genomic material. The regulation of poly(ADP ribosyl)ation with age is little known in plants, especially in combination with antioxidant defences modulation. In this study, the relationships between poly (ADP-ribose) polymerase (PARP) activity and enzymatic and non-enzymatic antioxidant pool have been studied together with the photosynthetic apparatus efficiency in the Mediterranean species Cistus incanus L., examining leaves at different developmental stages: young, mature and senescent. The photosynthetic performance was evaluated by chlorophyll a fluorescence measurement, the total soluble and fat-soluble antioxidant capacity, as well as the activities of enzymes superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and glutathione-S-transferase (GST), were determined by spectrophotometer, PARP activity was assessed by radioactive labelling. The highest photochemical activity was observed in young leaves, together with the highest GST activity. With the progress of the ageing process, the non-enzymatic antioxidant pool (namely ascorbic acid, &alpha, tocopherol) declined, reaching the lowest value in senescent leaves, whereas PARP activity rose significantly. The overall results indicate that the decline of photosynthetic apparatus efficiency during senescence is due to the reduction of specific defences against oxidative damages, which increase the damages to DNA, as demonstrated by PARP activity rise.