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A recent study on forest harvest in the EU (Ceccherini et al. 2020) reported a strong increase in clear-cut harvested area in recent years, based on remote sensing information. This triggered a heated debate and many critical comments. Apart from several fair and constructive criticisms, which were welcome, we found that some comments have been either not based on evidence or affected by serious misunderstandings. Here we clarify some technical aspects that were omitted or misrepresented in the public debate. Overall, the original study used in a scientifically correct way the best information available at that time. After the study was published, a previously undocumented inconsistency in the time series emerged in the original dataset used. After correcting for this inconsistency, updated results confirm an increase in clear-cut harvested area, but not as abrupt as originally reported. Contrary to what many critics say, this information should be seen as complementing and not necessarily contradicting country statistics, because the latter typically refer to total harvest (including thinning, etc.) and not clear-cut only. Finally, it should not be overlooked that the main aim of the original study was to offer a vision for integrating satellite data into the monitoring of forest resources. This was achieved: the JRC study showed the potential (and limitations) for high-resolution satellite maps to track the temporal evolution of clear-cut forest harvest in EU.

Ennio Polilli,1 Valeria Cento,2 Umberto Restelli,3,4 Francesca Ceccherini-Silberstein,2 Marianna Aragri,2 Velia Chiara Di Maio,2 Antonina Sciacca,1 Fiorenzo Santoleri,5 Paolo Fazii,6 Alberto Costantini,5 Carlo Federico Perno,2 Giustino Parruti1 1Infectious Diseases Unit, Pescara General Hospital, Pescara, 2Department of Experimental Medicine and Surgery, University of Rome “Tor Vergata”, Rome, 3CREMS - Centre for Research on Health Economics, Social and Health Care Management, Carlo Cattaneo – LIUC University, Castellanza, Italy; 4School of Public Health, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; 5Hospital Pharmacy, Pescara General Hospital, 6Microbiology and Virology Unit, Pescara General Hospital, Pescara, Italy Abstract: Available commercial assays may yield inaccurate hepatitis C virus (HCV) genotype assignment in up to 10% of cases. We investigated the cost-effectiveness of re-evaluating HCV genotype by population sequencing, prior to choosing a direct acting antiviral (DAA) regimen. Between March and September 2015, HCV sequence analysis was performed in order to confirm commercial LiPA-HCV genotype (Versant® HCV Genotype 2.0) in patients eligible for treatment with DAAs. Out of 134 consecutive patients enrolled, sequencing yielded 21 (15.7%) cases of discordant results. For three cases of wrong genotype assignment, the putative reduction in efficacy was gauged between 15% and 40%. Among the eight cases for whom G1b was assigned by commercial assays instead of G1a, potentially suboptimal treatments would have been prescribed. Finally, for five patients with G1 and indeterminate subtype, the choice of regimens would have targeted the worst option, with a remarkable increase in costs, as in the case of the four mixed HCV infections for whom pan-genotypic regimens would have been mandatory. Precise assignment of HCV genotype and subtype by sequencing may, therefore, be more beneficial than expected, until more potent pan-genotypic regimens are available for all patients. Keywords: HCV, HCV sequence analysis, HCV genotype, direct acting antiviral, treatment costs

Atherosclerosis is an inflammatory disease wherein neutrophils play a key role in plaque evolution. We observed that neutrophil CD11b was associated with a higher necrotic core volume in coronary plaques. Since platelets modulate neutrophil function, we explored the influence of neutrophil–platelet conjugates on the cytokine-modulated neutrophil complex CD11b/CD18 and CCR5 receptor expression. In 55 patients [68.53 ± 7.95 years old (mean ± SD); 71% male], neutrophil positivity for CD11b, CD18 and CCR5 was expressed as Relative Fluorescence Intensity (RFI) and taken as a dependent variable. Cytokines and chemokines were assessed by ELISA. Following log-10-based logarithmic transformation, they were used as independent variables in Model 1 of multiple regression together with Body Mass Index and albumin. Model 1 was expanded with the RFI of neutrophil CD41a+ (model 2). The RFI of neutrophil CD41a+ correlated positively and significantly with CD11b, CD18, and CCR5. In Model 2, CCR5 correlated positively only with the RFI of neutrophil CD41a+. Albumin maintained its positive effect on CD11b in both models. These observations indicate the complexity of neutrophil phenotypic modulation in stable CAD. Despite limitations, these findings suggest there is a role played by neutrophil–platelet interaction on the neutrophil cytokine-modulated expression of adhesive and chemotactic receptors.

Congenital central hypoventilation syndrome (CCHS), a rare genetic disease caused by heterozygous PHOX2B mutations, is characterized by life-threatening breathing deficiencies. PHOX2B is a transcription factor required for the specification of the autonomic nervous system, which contains, in particular, brainstem respiratory centers. In CCHS, PHOX2B mutations lead to cytoplasmic PHOX2B protein aggregations, thus compromising its transcriptional capability. Currently, the only available treatment for CCHS is assisted mechanical ventilation. Therefore, identifying molecules with alleviating effects on CCHS-related breathing impairments is of primary importance. A transcriptomic analysis of cells transfected with different PHOX2B constructs was used to identify compounds of interest with the CMap tool. Using fluorescence microscopy and luciferase assay, the selected molecules were further tested in vitro for their ability to restore the nuclear location and function of PHOX2B. Finally, an electrophysiological approach was used to investigate ex vivo the effects of the most promising molecule on respiratory activities of PHOX2B-mutant mouse isolated brainstem. The histone deacetylase inhibitor SAHA was found to have low toxicity in vitro, to restore the proper location and function of PHOX2B protein, and to improve respiratory rhythm-related parameters ex vivo. Thus, our results identify SAHA as a promising agent to treat CCHS-associated breathing deficiencies.

Microplastics (MPs) are recognized as a major environmental problem due to their ubiquitous presence in ecosystems and bioaccumulation in food chains. Not only humans are continuously exposed to these pollutants through ingestion and inhalation, but recent findings suggest they may trigger vascular inflammation and potentially worsen the clinical conditions of cardiovascular patients. Here we combine headspace analysis by needle trap microextraction-gas chromatography-mass spectrometry (HS-NTME-GC-MS) and biological assays to evaluate the effects of polystyrene, high- and low-density polyethylene MPs on phenotype, metabolic activity, and pro-inflammatory status of Vascular Smooth Muscle Cells (VSMCs) the most prominent cells in vascular walls. Virgin and artificially aged MPs (4 weeks at 40 °C and 750 W/m2 simulated solar irradiation) were comparatively tested at 1 mg/mL to simulate a realistic exposure scenario. Our results clearly show the activation of oxidative stress and inflammatory processes when VSMCs were cultured with aged polymers, with significant overexpression of IL-6 and TNF-α. In addition, volatile organic compounds (VOCs), including pentane, acrolein, propanal, and hexanal as the main components, were released by VSMCs into the headspace. Type-specific VOC response profiles were induced on vascular cells from different MPs.

• Human immunodeficiency virus (HIV) drug resistance has implications for antiretroviral treatment strategies and for containing the HIV pandemic because the development of HIV drug resistance leads to the requirement for antiretroviral drugs that may be less effective, less well-tolerated, and more expensive than those used in first-line regimens.  • HIV drug resistance studies are designed to determine which HIV mutations are selected by antiretroviral drugs and, in turn, how these mutations affect antiretroviral drug susceptibility and response to future antiretroviral treatment regimens.  • Such studies collectively form a vital knowledge base essential for monitoring global HIV drug resistance trends, interpreting HIV genotypic tests, and updating HIV treatment guidelines.  • Although HIV drug resistance data are collected in many studies, such data are often not publicly shared, prompting the need to recommend best practices to encourage and standardize HIV drug resistance data sharing.  • In contrast to other viruses, sharing HIV sequences from phylogenetic studies of transmission dynamics requires additional precautions as HIV transmission is criminalized in many countries and regions.  • Our recommendations are designed to ensure that the data that contribute to HIV drug resistance knowledge will be available without undue hardship to those publishing HIV drug resistance studies and without risk to people living with HIV.

Numerous efforts have been made at Tri-Alpha Energy (TAE) to theoretically explore the physics of microwave electron heating in field-reversed configuration (FRC) plasmas. For the fixed 2D profiles of plasma density and temperature for both electrons and thermal ions and equilibrium field of the C-2U machine, simulations with GENRAY-C ray-tracing code have been conducted for the ratios of ω/ωci[D] in the range of 6 - 20. Launch angles and antenna radial and axial positions have been optimized in order to simultaneously achieve good wave penetration into the core of FRC plasmas and efficient power damping on electrons. It is found that in an optimal regime, single pass absorption efficiency is 100% and most of the power is deposited inside the separatrix of FRC plasmas, with power damping efficiency of about 72% on electrons and less than 19% on ions. Calculations have clearly demonstrated that substantial power absorption on electrons is mainly attributed to high beta enhancement of magnetic pumping; complete power damping occurs before Landau damping has a significant effect on power absorption.

The 2022 edition of the IAS-USA drug resistance mutations list updates the Figure last published in September 2019. The mutations listed are those that have been identified by specific criteria for evidence and drugs described. The Figure is designed to assist practitioners to identify key mutations associated with resistance to antiretroviral drugs, and therefore, in making clinical decisions regarding antiretroviral therapy.

Objectives: To investigate whether SARS-CoV-2 messenger RNA (mRNA) vaccination has an impact on HIV-related viro-immunological parameters. Methods: People with HIV (PWH) in the VAXICONA-ORCHESTRA cohort who received one or more doses of SARS-CoV-2 mRNA vaccine and for whom paired measures of immuno-virological markers (viral load, clusters of differentiation [CD]4, and CD8 count 1 month before and after a vaccine dose [VD]) were available were included. Paired t-test and generalized estimating equation linear regression analyses were used to study changes over ± 1 month around the VD. Subgroup analyses were performed. Results: A total of 510 PWH were enrolled: the median age was 55 years (interquartile range 46-60 years), the CD4 and CD8 count were 489 (287-719) and 790 (59-1104) cells/mm3, respectively, and 81% received three VDs. After a median of 28 (3-53) days from VD, CD4 count increased by +15 cells/mm3 (SD ± 129.7, P = 0.001) and CD8 by +12 (±250.5, P = 0.199) and the viral load decreased by −0.11 log10 (±0.88, P = 0.001). Similar results were observed after restricting the analysis to viro-suppressed PWH, with CD4 ≤200/mm3, more than 6 months of antiretroviral therapy before VD and after excluding previous COVID-19. Conclusions: A small significant increase in CD4 count and a negligible drop in HIV RNA were observed. Our findings are consistent with the hypothesis that SARS-CoV-2 mRNA vaccine can prime CD4 T spike-specific cells, even in the more immuno-compromised PWH.

CLN8 is an endoplasmic reticulum cargo receptor and a regulator of lysosome biogenesis whose loss of function leads to neuronal ceroid lipofuscinosis. CLN8 has been linked to autophagy and lipid metabolism, but much remains to be learned, and there are no therapies acting on the molecular signatures in this disorder. The present study aims to characterize the molecular pathways involved in CLN8 disease and, by pinpointing altered ones, to identify potential therapies. To bridge the gap between cell and mammalian models, we generated a new zebrafish model of CLN8 deficiency, which recapitulates the pathological features of the disease. We observed, for the first time, that CLN8 dysfunction impairs autophagy. Using autophagy modulators, we showed that trehalose and SG2 are able to attenuate the pathological phenotype in mutant larvae, confirming autophagy impairment as a secondary event in disease progression. Overall, our successful modeling of CLN8 defects in zebrafish highlights this novel in vivo model's strong potential as an instrument for exploring the role of CLN8 dysfunction in cellular pathways, with a view to identifying small molecules to treat this rare disease.

Objectives: Heavily treatment-experienced (HTE) people living with HIV (PLWH) pose unique challenges due to limited antiretroviral treatment (ART) options. Our study aimed to investigate the prevalence and features of HTE individuals followed up in the Italian Cohort Naïve Antiretrovirals (ICONA) cohort as of December 31, 2021. Methods: HTE were defined based on meeting specific conditions concerning their current ART and their ART history up to December 31, 2021. Descriptive statistics were performed by HTE status. Regression analyses explored factors associated with becoming HTE based on pre-ART patients' characteristics. Cluster dendrogram analysis provided insights into subgroups with inadequate responses based on clusters of differentiation (CD4) counts and viral load (VL) trajectories. Results: Among the 8758 PLWH actively followed in our cohort, 163 individuals (1.9%), mainly female, younger, Italian, and infected through heterosexual contact, met the HTE criteria. A lower CD4 count at ART initiation (odds ratio [OR] 1.60 per 100 cells/mmc lower CD4, 95% confidence interval [CI] 1.06-2.41, P = 0.03) and hepatitis C virus antibody positivity (OR 1.90, 95% CI 1.16-3.11, P = 0.01) were associated with higher HTE risk. Thirty PLWH exhibited ongoing immune-virological failure (18% of the HTE subgroup and 0.003% of the total population). Thirty PLWH exhibited ongoing immune-virological failure (i.e., with a current CD4 count 200 copies/mL). A cluster analysis identified 13 (43%) with a current CD4 count

IntroductionAutosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a rare incurable neurodegenerative disease caused by mutations in the SACS gene, which codes for sacsin, a large protein involved in protein homeostasis, mitochondrial function, cytoskeletal dynamics, autophagy, cell adhesion and vesicle trafficking. However, the pathogenic mechanisms underlying sacsin dysfunction are still largely uncharacterized, and so attempts to develop therapies are still in the early stages.MethodsTo achieve further understanding of how processes are altered by loss of sacsin, we used untargeted proteomics to compare protein profiles in ARSACS fibroblasts versus controls.ResultsOur analyses confirmed the involvement of known biological pathways and also implicated calcium and lipid homeostasis in ARSACS skin fibroblasts, a finding further verified in SH-SY5Y SACS–/– cells. Validation through mass spectrometry-based analysis and comparative quantification of lipids by LC-MS in fibroblasts revealed increased levels of ceramides coupled with a reduction of diacylglycerols.DiscussionIn addition to confirming aberrant Ca2+ homeostasis in ARSACS, this study described abnormal lipid levels associated with loss of sacsin.

IntroductionComplement factor H (FH) is a major regulator of the complement alternative pathway, its mutations predispose to an uncontrolled activation in the kidney and on blood cells and to secondary C3 deficiency. Plasma exchange has been used to correct for FH deficiency and although the therapeutic potential of purified FH has been suggested by in vivo experiments in animal models, a clinical approved FH concentrate is not yet available. We aimed to develop a purification process of FH from a waste fraction rather than whole plasma allowing a more efficient and ethical use of blood and plasma donations.MethodsWaste fractions from industrial plasma fractionation (pooled human plasma) were analyzed for FH content by ELISA. FH was purified from unused fraction III and its decay acceleration, cofactor, and C3 binding capacity were characterized in vitro. Biodistribution was assessed by high-resolution dynamic PET imaging. Finally, the efficacy of the purified FH preparation was tested in the mouse model of C3 glomerulopathy (Cfh−/− mice).ResultsOur purification method resulted in a high yield of highly purified (92,07%), pathogen-safe FH. FH concentrate is intact and fully functional as demonstrated by in vitro functional assays. The biodistribution revealed lower renal and liver clearance of human FH in Cfh-/- mice than in wt mice. Treatment of Cfh-/- mice documented its efficacy in limiting C3 activation and promoting the clearance of C3 glomerular deposits.ConclusionWe developed an efficient and economical system for purifying intact and functional FH, starting from waste material of industrial plasma fractionation. The FH concentrate could therefore constitute possible treatments options of patients with C3 glomerulopathy, particularly for those with FH deficiency, but also for patients with other diseases associated with alternative pathway activation.

Objectives: Here we investigate Hepatitis D virus (HDV)-prevalence in Italy and its fluctuations over time and we provide an extensive characterization of HDV-infected patients. Methods: The rate of HDV seroprevalence and HDV chronicity was assessed in 1579 hepatitis B surface antigen (HBsAg)+ patients collected from 2005 to 2022 in Central Italy. Results: In total, 45.3% of HBsAg+ patients received HDV screening with an increasing temporal trend: 15.6% (2005-2010), 45.0% (2011-2014), 49.4% (2015-2018), 71.8% (2019-2022). By multivariable model, factors correlated with the lack of HDV screening were alanine-aminotransferase (ALT) less than two times of upper limit of normality (

Mortality in children accounts for 15% of all AIDS-related deaths globally, with a higher burden among Cameroonian children (25%), likely driven by poor virological response. We sought to evaluate viral suppression (VS) and its determinants in a nationally representative paediatric and young adult population receiving antiretroviral therapy (ART). A cross-sectional and multicentric study was conducted among Cameroonian children (p < 0.001. VS was 85.1% on a DTG-based regimen versus 80.0% on efavirenz/nevirapine and 65.6% on lopinavir/ritonavir or atazanavir/ritonavir. VS was higher in females versus males (85.8% versus 78.2%, p < 0.001). The VS rate remained stable around 85% at 12 and 24 months but dropped to about 80% at 36 months after ART initiation, p < 0.009. Independent predictors of non-VS were younger age, longer ART duration (>36 months), backbone drug (non-TDF/3TC) and anchor drug (non-DTG based). In this Cameroonian paediatric population with varying levels of transition to DTG, overall VS remains below the 95% targets. Predictors of non-VS are younger age, non-TDF/3TC- and non-DTG-based regimens. Thus, efforts toward eliminating paediatric AIDS should prioritise the transition to a DTG-based regimen in this new ART era.

Background: Next-generation sequencing (NGS) is gradually replacing Sanger sequencing for HIV genotypic drug resistance testing (GRT). This work evaluated the concordance among different NGS-GRT interpretation tools in a real-life setting. Methods: Routine NGS-GRT data were generated from viral RNA at 11 Italian laboratories with the AD4SEQ HIV-1 Solution v2 commercial kit. NGS results were interpreted by the SmartVir system provided by the kit and by two online tools (HyDRA Web and Stanford HIVdb). NGS-GRT was considered valid when the coverage was >100 reads (100×) at each PR/RT/IN resistance-associated position listed in the HIVdb 9.5.1 algorithm. Results: Among 629 NGS-GRT, 75.2%, 74.2%, and 70.9% were valid according to SmartVir, HyDRA Web, and HIVdb. Considering at least two interpretation tools, 463 (73.6%) NGS-GRT had a valid coverage for resistance analyses. The proportion of valid samples was affected by viremia 10% showed fair concordance among different interpretation tools. Conclusion: This Italian survey on NGS resistance testing suggests that viremia levels and HIV subtype affect NGS-GRT coverage. Within the current routine method for NGS-GRT, only mutations with frequency >10% seem reliably detected across different interpretation tools.

Post COVID-19 condition (PCC) is defined as ongoing symptoms at ≥1 month after acute COVID-19. We investigated the risk of PCC in an international cohort according to viral variants. We included 7699 hospitalized patients in six centers (January 2020–June 2023); a subset of participants with ≥1 visit over the year after clinical recovery were analyzed. Variants were observed or estimated using Global Data Science Initiative (GISAID) data. Because patients returning for a post COVID-19 visit may have a higher PCC risk, and because the variant could be associated with the probability of returning, we used weighted logistic regressions. We estimated the proportion of the effect of wild-type (WT) virus vs. Omicron on PCC, which was mediated by Intensive Care Unit (ICU) admission, through a mediation analysis. In total, 1317 patients returned for a post COVID visit at a median of 2.6 (IQR 1.84–3.97) months after clinical recovery. WT was present in 69.6% of participants, followed by the Alpha (14.4%), Delta (8.9%), Gamma (3.9%) and Omicron strains (3.3%). Among patients with PCC, the most common manifestations were fatigue (51.7%), brain fog (32.7%) and respiratory symptoms (37.2%). Omicron vs. WT was associated with a reduced risk of PCC and PCC clusters; conversely, we observed a higher risk with the Delta and Alpha variants vs. WT. In total, 42% of the WT effect vs. Omicron on PCC risk appeared to be mediated by ICU admission. A reduced PCC risk was observed after Omicron infection, suggesting a possible reduction in the PCC burden over time. A non-negligible proportion of the variant effect on PCC risk seems mediated by increased disease severity during the acute disease.