Morphological and pathogenic investigation of the emerging fungal threat Emergomyces africanus.

Emergomyces africanus is a highly fatal fungal pathogen affecting individuals with advanced HIV disease. Molecular patterns and ultrastructural aspects of E. africanus are unknown, and pathogenic models have not been investigated in detail. Since the cell wall of fungi is a determinant for interaction with the host and antifungal development, we characterized the ultrastructural aspects of E. africanus and the general properties of cell wall components under different conditions of growth in vitro and in vivo. We also tested the pathogenic potential of E. africanus in a Galleria mellonella model of infection. Transmission electron microscopy revealed the common intracellular, ultrastructural features of fungi in association with a thick cell wall. Scanning electron microscopy revealed a smooth cell surface, with no apparent decorative structures. Yeast cultures of E. africanus showed the distribution of chitin, chitooligomers, and mannoproteins commonly observed in fungi. However, in mixed microenvironments containing yeast and filamenting forms of E. africanus, the detection of chitooligomers was increased in comparison with isolated yeast cells, while the detection of these components in filamenting forms was markedly reduced. These observations were suggestive of the ability of E. africanus to change its cell wall composition in response to different microenvironments. Although E. africanus was unable to kill G. mellonella, this infection model allowed us to isolate infected hemocytes for further analysis of mannoproteins, chitin, and chitooligomers. Once again, the detection of E. africanus chitooligomers was markedly increased. These results reveal previously unknown ultrastructural features of E. africanus and suggest a high plasticity in the cell wall of this lethal pathogen. IMPORTANCE: The epidemiology of fungal infections is very dynamic, and novel health emergencies are hard to predict. New fungal pathogens have been continuously emerging for the last few decades, and Emergomyces africanus is one of these threats to human health. This complex scenario points to the need for generating knowledge about emerging pathogens so that new therapeutic strategies can be designed. In this study, we characterized the general cellular and pathogenic properties of the emerging fungal pathogen E. africanus. Our results reveal that E. africanus manifests some of the typical properties of fungal cells but also exhibits some unique characteristics that might be helpful for the future development of therapeutic strategies.

Antimicrobial Composites Based on Methacrylic Acid-Methyl Methacrylate Electrospun Fibers Stabilized with Copper(II).

This study presents fibers based on methacrylic acid-methyl methacrylate (Eudragit L100) as Cu(II) adsorbents, resulting in antimicrobial complexes. Eudragit L100, an anionic copolymer synthesized by radical polymerization, was electrospun in dimethylformamide (DMF) and ethanol (EtOH). The electrospinning process was optimized through a 22-factorial design, with independent variables (copolymer concentration and EtOH/DMF volume ratio) and three repetitions at the central point. The smallest average fiber diameter (259 ± 53 nm) was obtained at 14% w/v Eudragit L100 and 80/20 EtOH/DMF volume ratio. The fibers were characterized using scanning electron microscopy (SEM), infrared spectroscopy in attenuated total reflectance mode (FTIR-ATR), and differential scanning calorimetry (DSC). The pseudo-second-order mechanism explained the kinetic adsorption toward Cu(II). The fibers exhibited a maximum adsorption capacity (qe) of 43.70 mg/g. The DSC analysis confirmed the Cu(II) absorption, indicating complexation between metallic ions and copolymer networks. The complexed fibers showed a lower degree of swelling than the non-complexed fibers. The complexed fibers exhibited bacteriostatic activity against Gram-negative (Pseudomonas aeruginosa) and Gram-positive (Staphylococcus aureus) bacteria. This study successfully optimized the electrospinning process to produce thin fibers based on Eudragit L100 for potential applications as adsorbents for Cu(II) ions in aqueous media and for controlling bacterial growth.

Receptor transfer between immune cells by autoantibody-enhanced, CD32-driven trogocytosis is hijacked by HIV-1 to infect resting CD4 T cells.

Immune cell phenotyping frequently detects lineage-unrelated receptors. Here, we report that surface receptors can be transferred from primary macrophages to CD4 T cells and identify the Fcγ receptor CD32 as driver and cargo of this trogocytotic transfer. Filamentous CD32+ nanoprotrusions deposit distinct plasma membrane patches onto target T cells. Transferred receptors confer cell migration and adhesion properties, and macrophage-derived membrane patches render resting CD4 T cells susceptible to infection by serving as hotspots for HIV-1 binding. Antibodies that recognize T cell epitopes enhance CD32-mediated trogocytosis. Such autoreactive anti-HIV-1 envelope antibodies can be found in the blood of HIV-1 patients and, consistently, the percentage of CD32+ CD4 T cells is increased in their blood. This CD32-mediated, antigen-independent cell communication mode transiently expands the receptor repertoire and functionality of immune cells. HIV-1 hijacks this mechanism by triggering the generation of trogocytosis-promoting autoantibodies to gain access to immune cells critical to its persistence.

The influence of cation exchange on the possible mechanism of erionite toxicity: A synchrotron-based micro-X-ray fluorescence study on THP-1-derived macrophages exposed to erionite-Na.

Fibrous erionite is the only zeolite classified as Group 1 carcinogen by the International Agency for Research on Cancer (IARC). Carcinogenesis induced by erionite is thought to involve several factors as biopersistence, the iron role and cation exchange processes. To better understand these mechanisms, a detailed investigation at the micro scale was performed, collecting elemental information on iron and cation release and their distribution in biological systems by synchrotron micro-X-ray fluorescence mapping (SR-micro-XRF) and synchrotron micro-X-ray absorption spectroscopy (SR-micro-XANES) at the TwinMic beamline (Elettra synchrotron) and at the ID21 beamline of the European Synchrotron Radiation Facility (ESRF). By microscopy and chemical mapping, highly detailed maps of the chemical and morphological interaction of biological systems with fibres could be produced. In detail, THP-1 cell line derived macrophages, used as in vitro model, were analysed during erionite-Na phagocytosis at different time intervals, after single dose exposure. For comparison, cellular fluorescent probes were also used to evaluate the intracellular free sodium and calcium concentrations. Synchrotron analyses visualised the spatial distribution of both fibre and mineral particle associated metals during the phagocytosis, describing the mechanism of internalisation of erionite-Na and its accessory mineral phases. The intracellular distribution of metals and other cations was mapped to evaluate metal release, speciation changes and/or cation exchange during phagocytosis. The fluorescent probes complemented microchemical data clarifying, and confirming, the cation distribution observed in the SR-micro-XRF maps. The significant cytoplasmic calcium decrease, and the concomitant sodium increase, after the fibre phagocytosis seemed due to activation of plasma membrane cations exchangers triggered by the internalisation while, surprisingly, the ion-exchange capacity of erionite-Na could play a minor role in the disruption of the two cations intracellular homeostasis. These results help to elucidate the role of cations in the toxicity of erionite-treated THP-1 macrophages and add knowledge to its carcinogenicity process.

Thermal shakedown in granular materials with irregular particle shapes.

Granular materials with irregular particle shapes undergo a myriad of temperature variations in natural and engineered systems. However, the impacts of cyclic temperature variations on the mechanics of granular materials remain poorly understood. Specifically, little is known about the response of granular materials to cyclic temperature variations as a function of the following central variables: particle shape, applied stress level, relative density, and temperature amplitude. This paper presents advanced laboratory experiments to explore the impacts of cyclic temperature variations on the mechanics of granular materials, with a focus on sands. The results show that cyclic temperature variations applied to sands induce thermal shakedown: the accumulation of irreversible bulk deformations due to microstructural rearrangements caused by thermal expansions and contractions of the constituting particles. The deformation of sands caused by thermal shakedown strongly depends on particle shape, stress level, relative density, and temperature amplitude. This deformation is limited for individual thermal cycles but accumulates and becomes significant for multiple thermal cycles, leading to substantial compaction in sands and other granular materials, which can affect various natural and engineered systems.

Iron nuclearity in mineral fibres: Unravelling the catalytic activity for predictive modelling of toxicity.

Chronic inflammation induced in vivo by mineral fibres, such as asbestos, is sustained by the cyclic formation of cytotoxic/genotoxic oxidant species that are catalysed by iron. High catalytic activity is observed when iron atoms are isolated in the crystal lattice (nuclearity=1), whereas the catalytic activity is expected to be reduced or null when iron forms clusters of higher nuclearity. This study presents a novel approach for systematically measuring iron nuclearity across a large range of iron-containing standards and mineral fibres of social and economic importance, and for quantitatively assessing the relation between nuclearity and toxicity. The multivariate curve resolution (MCR) empirical approach and density functional theory (DFT) calculations were applied to the analysis of UV-Vis spectra to obtain information on the nature of iron and nuclearity. This approach led to the determination of the nuclearity of selected mineral fibres which was subsequently used to calculate a toxicity-related index. High nuclearity-related toxicity was estimated for chrysotile samples, fibrous glaucophane, asbestos tremolite, and fibrous wollastonite. Intermediate values of toxicity, corresponding to a mean nuclearity of 2, were assigned to actinolite asbestos, amosite, and crocidolite. Finally, a low nuclearity-related toxicity parameter, corresponding to an iron-cluster with a lower catalytic power to produce oxidants, was assigned to asbestos anthophyllite.

Revisiting anthelmintic resistance in sheep flocks from São Paulo State, Brazil.

Haemonchus contortus and Trichostrongylus colubriformis are the most important gastrointestinal nematodes causing serious losses in sheep production of tropical and subtropical regions. Prophylaxis of gastrointestinal nematode infections is based on anthelmintics use, but their frequent administration selects multiple-resistant parasites. To evaluate how the situation has changed over the last decades, the anthelmintic resistance status of gastrointestinal nematodes in sheep flocks was assessed in the current study and compared to previous surveys. In each one of the 15 flocks evaluated, animals (n ≥ 7) were allocated into at least five groups and treated as follows: 1) untreated control; 2) albendazole; 3) levamisole; 4) ivermectin; and 5) monepantel. If more animals were available, two additional groups were included: 6) closantel, and 7) moxidectin. The faecal egg count reduction test (FECRT) was carried out to evaluate the pre- and post-treatment using the SHINY tool. Haemonchus spp. was the most prevalent nematode from faecal cultures. The mean efficacy of albendazole was 40%. Only in two farms, levamisole presented a relatively high percentage of reduction in the FECRT about 90%, while ivermectin and moxidectin presented the worst mean efficacy of 34% and 21% among all farms, respectively. Like other anthelmintics, closantel demonstrated low efficacy (63%) across all farms evaluated. Monepantel presented an overall mean efficacy of 79%, but it was the only anthelmintic that presented efficacy ≥95%, in five farms. The results revealed that gastrointestinal nematodes with multiple anthelmintic resistance were prevalent in all 15 sheep herds. The research suggests that nematodes are becoming more and more resistant to various anthelmintic compounds, which has made the problem worse. This circumstance highlights the necessity to put into practice sustainable and long-lasting methods to prevent gastrointestinal nematode infections in sheep husbandry.

Mechanisms of action of mineral fibres in a placental syncytiotrophoblast model: An in vitro toxicology study.

Asbestos has been widely used due to its unique characteristics. It is known that exposure to asbestos causes serious damage to health but one species, chrysolite, is still used because it is considered less toxic and not biopersistent in some countries. The aim of our study was to investigate if cellular process underlying the proliferation, differentiation and cell death of placental tissues could be modify in presence of asbestos fibres (50 µg/ml final concentration), long chrysolite fibres (CHR-L) and short chrysolite fibres (CHR-S), using BeWo cell line, an in vitro model that mimics the syncytiotrophoblast (STB), the outer layer of placental villi. Our data demonstrated that none of the fibres analysed alter syncytiotrophoblast formation but all of them induce ROS formation and reduced cell proliferation. Moreover, we showed that only CHR-L fibre induced was able to induce irreversible DNA alterations that carried cells to apoptosis. In fact, BeWo cells exposed to CHR-L fibre showed a significant increase in cleaved CASP3 protein, a marker of apoptosis. These data suggest that CHR-L may induce death of the placental villi leading to impaired placental development. The impairment of placental development is the basis of many gestational pathologies such as preeclampsia and intrauterine growth retardation. Since these pathologies are very dangerous for foetal and maternal life, we suggest to the gynaecologists to carefully evaluate the area of maternal residence, the working environment, the food used, and the materials used daily to avoid contact with these fibres as much as possible.

Upper and lower limb tremor in inflammatory neuropathies.

OBJECTIVE: The mechanisms underlying neuropathic tremor remain incompletely understood and a distinction has not been drawn between proximal and distal neuropathies. Lower limb tremor contributes to imbalance in chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), but this is unexplored in other neuropathies. We characterized upper and lower limb tremor in chronic immune sensory polyradiculopathy (CISP) and distal acquired demyelinating neuropathy with anti-MAG antibodies (DADS-MAG), contrasted to CIDP. METHODS: This was a cross-sectional study of 38 patients (CIDP [n = 25], CISP [n = 7], DADS-MAG [n = 6]). Clinical assessment, tremor study recordings, nerve conduction studies, and somatosensory evoked potentials were performed. Balance was measured by force platform. RESULTS: Upper limb tremor was prevalent (CIDP 66%, CISP 70%, DADS-MAG 100%). Peak frequencies followed a gradient along the upper limb, unchanged by weight-loading. Lower limb tremor was also present (CIDP 32%, CISP 29%, DADS-MAG 66%) and associated with imbalance. Nerve conduction parameters correlated with upper limb tremor in DADS-MAG and CISP, and imbalance in CISP. CONCLUSIONS: Upper limb tremor is mediated by peripheral and central mechanisms regardless of distal or proximal pathology. Lower limb tremor correlates with peripheral nerve function and contributes to imbalance. SIGNIFICANCE: This study contributes to the understanding of neuropathic tremor. Addressing lower limb tremor may be of therapeutic importance for neuropathy-associated imbalance.

Upper and lower limb tremor in Charcot-Marie-Tooth neuropathy type 1A and the implications for standing balance.

BACKGROUND: Neuropathic tremor occurs in Charcot-Marie-Tooth neuropathy type 1A (CMT1A; hereditary motor and sensory neuropathy, HMSN), although the pathophysiological mechanisms remain to be elucidated. Separately, lower limb tremor has not been explored in CMT1A and could be associated with imbalance as in other neuropathies. The present study aimed to determine tremor characteristics in the upper and lower limbs in CMT1A and relate these findings to clinical disability, particularly imbalance. METHODS: Tremor and posturography studies were undertaken in phenotyped and genotyped CMT1A patients. Participants underwent detailed clinical assessment, tremor study recordings, and nerve conduction studies. Tremor stability index was calculated for upper limb tremor and compared to essential tremor. RESULTS: Seventeen patients were enrolled. Postural and kinetic upper limb tremors were evident in 65%, while postural and orthostatic lower limb tremors were seen in 35% of CMT1A patients. Peak upper limb frequencies were lower distally (~ 6 Hz) and higher proximally (~ 9 Hz), were unchanged by weight-loading, and not impacted by fatigue. The tremor stability index was significantly higher in CMT1A than in essential tremor. A 5-6 Hz lower limb tremor was recorded which did not vary along the limb and was unaffected by fatigue. Balance was impaired in patients with postural lower limb tremor. A high frequency peak on posturography was associated with 'good' balance. CONCLUSIONS: Tremor is a common clinical feature in CMT1A, distinct from essential tremor, mediated by a complex interaction between peripheral and central mechanisms. Postural lower limb tremor is associated with imbalance; strategies aimed at tremor modulation could be of therapeutic utility.

Zinc oxide nanoparticle-reinforced pectin/starch functionalized films: A sustainable solution for biodegradable packaging.

Environmental pollution caused by non-biodegradable plastic pollutants adversely affects various ecosystems. This study proposes the development of novel functional and biodegradable films based on corn starch (CST) and pectin (PEC) containing zinc oxide nanoparticles (ZnONPs) from the casting method. The films exhibited processability, transparency, low water vapor permeation, and desirable mechanical properties for food packaging and coating applications. The ZnONPs acted as a plasticizer, enhancing the film elongation at the break, increasing the pec25-1 (PEC 25 wt% and ZnONPs 1 wt%) elongation from 79.85 to 162.32 %. The improved film elasticity supported by ZnONPs reduced the material stiffness. However, the films still demonstrated an average tensile strength (0.69 MPa) 17-fold higher than the tensile strength (0.04 MPa) of the non-biodegradable commercial film based on poly(vinyl chloride). Furthermore, the ZnONPs enhanced the UV-blocking capabilities of the films, leading to wettable materials with water contact angles lower than 90°. The films showed high biodegradation rates under natural disposal conditions. The results indicated that the pec25-1/ZnONPs film is a promising eco-friendly coating in food preservation due to its biodegradability, suitable mechanical properties, low water vapor permeability, and UV-blocking properties.

Expanding the Scope of an Amphoteric Condensed Tannin, Tanfloc, for Antibacterial Coatings.

Bacterial infections are a common mode of failure for medical implants. This study aims to develop antibacterial polyelectrolyte multilayer (PEM) coatings that contain a plant-derived condensed tannin polymer (Tanfloc, TAN) with inherent antimicrobial activity. Tanfloc is amphoteric, and herein we show that it can be used as either a polyanion or a polycation in PEMs, thereby expanding the possibility of its use in PEM coatings. PEMs are ordinarily formed using a polycation and a polyanion, in which the functional (ionic) groups of the two polymers are complexed to each other. However, using the amphoteric polymer Tanfloc with weakly basic amine and weakly acidic catechol and pyrogallol groups enables PEM formation using only one or the other of its functional groups, leaving the other functional group available to impart antibacterial activity. This work demonstrates Tanfloc-containing PEMs using multiple counter-polyelectrolytes including three polyanionic glycosaminoglycans of varying charge density, and the polycations N,N,N-trimethyl chitosan and polyethyleneimine. The layer-by-layer (LbL) assembly of PEMs was monitored using in situ Fourier-transform surface plasmon resonance (FT-SPR), confirming a stable LbL assembly. X-ray photoelectron spectroscopy (XPS) was used to evaluate surface chemistry, and atomic force microscopy (AFM) was used to determine the surface roughness. The LDH release levels from cells cultured on the Tanfloc-containing PEMs were not statistically different from those on the negative control (p > 0.05), confirming their non-cytotoxicity, while exhibiting remarkable antiadhesive and bactericidal properties against Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus), respectively. The antibacterial effects were attributed to electrostatic interactions and Tanfloc's polyphenolic nature. This work underscores the potential of Tanfloc as a versatile biomaterial for combating infections on surfaces.

Vaccine development for pathogenic fungi: current status and future directions.

INTRODUCTION: Fungal infections are caused by a broad range of pathogenic fungi that are found worldwide with different geographic distributions, incidences, and mortality rates. Considering that there are relatively few approved medications available for combating fungal diseases and no vaccine formulation commercially available, multiple groups are searching for new antifungal drugs, examining drugs for repurposing and developing antifungal vaccines, in order to control deaths, sequels, and the spread of these complex infections. AREAS COVERED: This review provides a summary of advances in fungal vaccine studies and the different approaches under development, such as subunit vaccines, whole organism vaccines, and DNA vaccines, as well as studies that optimize the use of adjuvants. We conducted a literature search of the PubMed with terms: fungal vaccines and genus of fungal pathogens (Cryptococcus spp. Candida spp. Coccidioides spp. Aspergillus spp. Sporothrix spp. Histoplasma spp. Paracoccidioides spp. Pneumocystis spp. and the Mucorales order), a total of 177 articles were collected from database. EXPERT OPINION: Problems regarding the immune response development in an immunocompromised organism, the similarity between fungal and mammalian cells, and the lack of attention by health organizations to fungal infections are closely related to the fact that, at present, there are no fungal vaccines available for clinical use.

Traversing the Cell Wall: The Chitinolytic Activity of Histoplasma capsulatum Extracellular Vesicles Facilitates Their Release.

Histoplasma capsulatum is the causative agent of histoplasmosis. Treating this fungal infection conventionally has significant limitations, prompting the search for alternative therapies. In this context, fungal extracellular vesicles (EVs) hold relevant potential as both therapeutic agents and targets for the treatment of fungal infections. To explore this further, we conducted a study using pharmacological inhibitors of chitinase (methylxanthines) to investigate their potential to reduce EV release and its subsequent impact on fungal virulence in an in vivo invertebrate model. Our findings revealed that a subinhibitory concentration of the methylxanthine, caffeine, effectively reduces EV release, leading to a modulation of H. capsulatum virulence. To the best of our knowledge, this is the first reported instance of a pharmacological inhibitor that reduces fungal EV release without any observed fungicidal effects.

Abomasal RNA-seq reveals a strong local cellular response in suckling lambs with resistance against Haemonchus contortus.

Santa Ines (SI) and Ile de France (IF) sheep are known to be resistant and susceptible to Haemonchus contortus infection, respectively. Several studies have shown some genes as potential biological markers for sheep resistance against gastrointestinal nematodes using molecular tools, including transcriptomic analysis. In this study, we sequenced the polyadenylated RNA of the abomasal tissue of SI and IF suckling lambs to identify mucosa-specific transcript alterations between breeds artificially infected with H. contortus. Naïve SI (n = 4) and IF (n = 4) lambs were artificially infected every other day, over a period of 52 days, from 14 to 66 days old, with a total of 5,400 H. contortus infective larvae. Fundic abomasal tissue samples were collected at 68 days old, and submitted to high-throughput RNA sequencing (RNA-seq). Differential expression analysis (P value < 0.001 and False Discovery Rate (FDR) < 0.05) between SI and IF samples identified 292 genes, most of which showed greater expression in SI lambs. To help annotate and assign possible function to differentially expressed genes (DEGs), we used previously available single-cell RNA-seq (scRNA-seq) data from ovine abomasal mucosa to putatively identify cell types and possible mechanisms involved in resistance to H. contortus. In particular, genes associated with endothelial and tuft cells showed the greatest increases in expression in SI relative to IF lambs. SI lambs had higher percentages of tuft cells than IF lambs in the fundic abomasal mucosa. Although we found innate immunity (cell-mediated in mucosa) acting as a protagonist in impairing H. contortus infection, a stronger acquired immune response was being modulated at an earlier stage by SI lambs. We suggest that the complex connection between innate and adaptive immunity is via cellular antigen processing and presentation (APP). Based on comparison with scRNA-seq data, SI lambs showed a robust APP mechanism characterized mainly by greater T cell APP, macrophage differentiation, and cytokine signalling. We identified potential mechanisms and markers to advance knowledge for selection of H. contortus resistance at a very early age, in SI as well as in other commercial sheep breeds.

κ-Carrageenan/poly(vinyl alcohol) functionalized films with gallic acid and stabilized with metallic ions.

Given the environmental issues caused by the extensive use of conventional petroleum-based packaging, this work proposes functional films based on commercial κ-carrageenan (κc), poly(vinyl alcohol) (PVA), and gallic acid (GA) prepared by the "casting" method. Metallic ions in the κc composition stabilized the films, supporting processability and suitable mechanical properties. However, the incorporated GA amount (6.25 and 10 wt%) in the films created from an aqueous κc solution at 3.0 % wt/v (κc3) prevented crystalline domains in the resulting materials. The κc3/GA6.25 and κc3/GA10 films had less tensile strength (8.50 ± 0.61 and 10.28 ± 0.65 MPa) and high elongation at break (2.36 ± 0.16 and 1.19 ± 0.17 %) compared to the other samples, respectively. Low κc contents (κc2.5/GA6.25 and κc2.5/GA10) promoted stiff films and less permeability to water vapor (5.36 ± 0.51 and 3.76 ± 0.02 [×10-12 g(Pa × m × s)-1], respectively. The κc/GA weight ratio also influenced the film wettability, indicating water contact angles (WCAs) between 55 and 74°. The surface wettability implies a low oil permeability and high water swelling capacity of up to 1600 %. The κc/GA also played an essential role in the film's antimicrobial action against Staphylococcus aureus and Escherichia coli. Thus, the κc3/GA10 film showed suitable physical, chemical, and biological properties, having the potential to be applied as food coatings.

There is plenty of asbestos at the bottom. The case of magnesite raw material contaminated with asbestos fibres.

Although all six asbestos minerals (the layer silicate chrysotile and five chain silicate species actinolite asbestos, amosite, anthophyllite asbestos, crocidolite and tremolite asbestos) are classified as carcinogenic, chrysotile is still mined and used in many countries worldwide. Other countries, like Italy, impose zero tolerance for all asbestos species, but conflicting views repress the development of globally uniform treaties controlling international trade of asbestos-containing materials. Hence, countries with more severe legislations against the use of these hazardous materials lack of an international safety net against importation of non-compliant products. This research reports the first discovery of commercial magnesite raw materials contaminated with white asbestos (chrysotile). X-ray powder diffraction and thermogravimetric/thermodifferential measurements showed the presence of serpentine group minerals in both the semi-processed (powder) and quarried material. The univocal identification of chrysotile in the powders was confirmed by its peculiar Raman bands of the OH stretching vibrations between 3500 and 3800 cm-1, with an intense peak at ∼3695 cm-1 and a weak contribution at ∼3647 cm-1. Transmission electron microscope showed that chrysotile forms fibres up to a few microns long and up to 80 nm thick with a nanotube structure characterized by inner channels as large as 30-40 nm. Fibres size analysis obtained by scanning electron microscopy indicates mean length and diameter of 5.95 and 0.109 µm with medians of 2.62 and 0.096 µm, respectively; some among the fibres analysed exhibit the so-called "Stanton size" (i.e., asbestos fibres longer than 8 µm and thinner than 0.25 µm that are strongly carcinogenic). Quantitative analysis showed a chrysotile content around 0.01 wt% not allowed by current regulations in Italy and many other countries. More generally, our findings demonstrate that without shared policies aimed at regulating asbestos circulation on the global market, "asbestos-free" national policies will inevitably fail.