Intact Particles Research Articles

Direct and Ultrasensitive Bioluminescent Detection of Intact Respiratory Viruses.

Respiratory viruses such as SARS-CoV-2, influenza, and respiratory syncytial virus (RSV) represent pressing health risks. Rapid diagnostic tests for these viruses detect single antigens or nucleic acids, which do not necessarily correlate with the amount of the intact virus. Instead, specific detection of intact respiratory virus particles may be more effective at assessing the contagiousness of a patient. Here, we report GLOVID, a modular biosensor platform to detect intact virions against a background of "free" viral proteins in solution. Our approach harnesses the multivalent display of distinct proteins on the surface of a viral particle to template the reconstitution of a split luciferase, allowing specific, single-step detection of intact influenza A and RSV virions corresponding to 0.1-0.3 fM of genomic units. The protein ligation system used to assemble GLOVID sensors is compatible with a broad range of binding domains, including nanobodies, scFv fragments, and cyclic peptides, which allows straightforward adjustment of the sensor platform to target different viruses.

Multiscale, mechanistic modeling of irradiation-enhanced silver diffusion in TRISO particles

Tristructural isotropic (TRISO) particles are under consideration for use in several proposed advanced nuclear reactor concepts. The silicon carbide (SiC) layer in TRISO acts as a barrier to prevent the release of the fission products. However, despite remarkable retention, silver (Ag) release has been observed from intact particles, which requires investigation since the Ag isotope (110m Ag) has a long half-life. Previous work focused on developing a multiscale, mechanistic model for Ag diffusion accounting for temperature and microstructure effect and has been successfully validated. In this work, we expand the previous model to account for irradiation-enhanced Ag diffusivity in SiC and improve its accuracy over a wider grain size and temperature ranges relevant for advanced reactor conditions. A temperature, grain size, and flux dependent diffusivity is therefore derived using the mesoscale code MARMOT and implemented in the fuel performance code BISON. The irradiation-enhanced Ag diffusivity in SiC is compared against experimental data and validated using BISON against Ag release measurements from the Advanced Gas Reactor Fuel Development and Qualification Program (AGR-1 and AGR-2). Herein, we quantify the impact of SiC grain size, irradiation, and temperature on Ag release. In agreement with previous studies, we find accounting for SiC grain size improves agreement between BISON predictions and experimental observations for most cases. We also find that accounting for irradiation improves agreement for cases where Ag release was underestimated, but the impact was less significant than accounting for microstructure.

Peptide ligands for the affinity purification of adenovirus from HEK293 and vero cell lysates

Adenovirus (AdVs) is the viral vector of choice in vaccines and oncolytic applications owing to its high transduction activity and inherent immunogenicity. For decades, AdV isolation has relied on ultracentrifugation and ion-exchange chromatography, which are not suitable to large-scale production and struggle to deliver sufficient purity. Immunoaffinity chromatography resins of recent introduction feature high binding capacity and selectivity, but mandate harsh elution conditions (pH 3.0), afford low yield (< 20%), and provide limited reusability. Seeking a more efficient and affordable alternative, this study introduces the first peptide affinity ligands for AdV purification. The peptides were identified via combinatorial selection and in silico design to target hexons, the most abundant proteins in the adenoviral capsid. Selected peptide ligands AEFFIWNA and TNDGPDYSSPLTGSG were conjugated on chromatographic resins and utilized to purify AdV serotype 5 from HEK293 and Vero cell lysates. The peptide-functionalized resins feature high binding capacity (> 1010 active virions per mL at the residence time of 2 min), provide high yield (> 50%) and up to 100-fold reduction of host cell proteins and DNA. Notably, the peptide ligands enable gentle elution conditions (pH 8) that prevent the “shedding” of penton and fiber proteins, thus affording intact adenovirus particles with high cell-transduction activity. The study of the peptide ligands by surface plasmon resonance and molecular docking and dynamics simulations confirmed the selective targeting of hexon proteins and elucidated the molecular-level mechanisms underlying binding and release. Collectively, these results demonstrate the strong promise of peptide ligands presented herein for the affinity purification of AdVs from cell lysates.

Tracking the movement of quartz sand particles with neural networks

Tracking the movement of particles is of paramount significance for studying the impact of particle breakage on the macroscopic mechanical behaviour of granular materials and remains a persistent challenge to date. This paper presents a novel particle tracking method that integrates Pointon and PointNetLK networks, enabling an accurate tracking of both intact and broken Leighton Buzzard sand (LBS) particles. Initially, morphological information of LBS particles was extracted from X-ray micro-tomography (CT) data collected from a miniature triaxial test. Various image processing techniques were applied to the raw CT images to achieve a realistic three-dimensional (3D) reconstruction. Subsequently, particle point cloud data was processed through sampling, Gaussian noise injection, and grouping for training and testing the Poynton and PointNetLK networks. Next, the correspondences among particles across different scans were established by PointConv, and the transformation matrix between two mutually matched particles was predicted using PointNetLK. Finally, an examination of the changes in the spatial distribution and morphological parameters of both tracked and untracked particles throughout the shearing process was conducted and followed by an analysis of particle kinematics.

Crystallographic/Morphological Changes and Performance of Lithium Iron Phosphate Deposited By Vacuum Cold Spray

Cold spray offers several advantages for material deposition, including the ability to deposit a wide range of materials without the need for high temperatures, minimizing heat-related distortion and maintaining material properties. Additionally, cold spray is a highly efficient and environmentally friendly process, as it reduces waste and energy consumption compared to traditional thermal spray methods. Lithium iron phosphate (LFP) has been gaining popularity as a cathode material for lithium-ion batteries due to its stable cycling performance, lack of oxygen generation, thermal stability, and use of environmentally friendly iron over nickel or cobalt. In this study, LFP was deposited by vacuum cold spray onto aluminum foil substrate to investigate the viability of cold spray as a possible method of manufacturing all-solid-state batteries. Results were obtained through in-situ X-ray diffraction (XRD), scanning electron microscopy (SEM), and in-situ transmission electron microscopy (TEM). Electrochemical results were obtained by assembling cold spray deposited LFP into half-cells with Li-metal anode and liquid electrolyte before being tested on electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV), and C-Rate. Due to the high impact velocities during cold spray, crystalline LFP was observed to undergo changes to an amorphous phase. Using in-situ X-ray diffraction (XRD), scanning electron microscopy (SEM), and in-situ transmission electron microscopy (TEM) the crystallographic and morphological changes to cold spray deposited LFP were investigated across various temperatures. In-situ XRD initially showed a decrease in crystallinity with crystallinity being restored with elevated temperatures, with in-situ TEM further supporting these findings. Investigation with SEM showed LFP particles undergo deformation upon impact with a mix of intact particles interspersed throughout to create a dense layer. Electrochemical impedance spectroscopy (EIS) of assembled half-cells show low resistance of 90Ω with further reduction to 60Ω upon cycling of the cell. In terms of cycling performance, initial discharge capacities showed 80 mAh/g and 20mAh/g at 0.1C and 1C respectively. These capacities dropped further with repeated cycling. Future work will be focused on improving capacity, annealing post deposition to restore crystallinity, and incorporation of solid electrolyte.

Ribosome External Electric Field Regulates Metabolic Enzyme Activity: The RAMBO Effect.

Ribosomes bind to many metabolic enzymes and change their activity. A general mechanism for ribosome-mediated amplification of metabolic enzyme activity, RAMBO, was formulated and elucidated for the glycolytic enzyme triosephosphate isomerase, TPI. The RAMBO effect results from a ribosome-dependent electric field-substrate dipole interaction energy that can increase or decrease the ground state of the reactant and product to regulate catalytic rates. NMR spectroscopy was used to determine the interaction surface of TPI binding to ribosomes and to measure the corresponding kinetic rates in the absence and presence of intact ribosome particles. Chemical cross-linking and mass spectrometry revealed potential ribosomal protein binding partners of TPI. Structural results and related changes in TPI energetics and activity show that the interaction between TPI and ribosomal protein L11 mediate the RAMBO effect.

Chemoenzymatic synthesis of sialyl-α2,3-lactoside–functionalized BSA conjugate inhibits influenza infection

Influenza remains a global public health threat, and the development of new antivirals is crucial to combat emerging drug-resistant influenza strains. In this study, we report the synthesis and evaluation of a sialyl lactosyl (TS)–bovine serum albumin (BSA) conjugate as a potential multivalent inhibitor of the influenza virus. The key trisaccharide component, TS, was efficiently prepared via a chemoenzymatic approach, followed by conjugation to dibenzocyclooctyne-modified BSA via a strain-promoted azide-alkyne cycloaddition reaction. Biophysical and biochemical assays, including surface plasmon resonance, isothermal titration calorimetry, hemagglutination inhibition, and neuraminidase inhibition, demonstrated the strong binding affinity of TS-BSA to the hemagglutinin (HA) and neuraminidase (NA) proteins of the influenza virus as well as intact virion particles. Notably, TS-BSA exhibited potent inhibitory activity against viral entry and release, preventing cytopathic effects in cell culture. This multivalent presentation strategy highlights the potential of glycocluster-based antivirals for combating influenza and other drug-resistant viral strains.

A replication-deficient gammaherpesvirus vaccine protects mice from lytic disease and reduces latency establishment

Gammaherpesviruses are oncogenic viruses that establish lifelong infections and are significant causes of morbidity and mortality. Vaccine strategies to limit gammaherpesvirus infection and disease are in development, but there are no FDA-approved vaccines for Epstein-Barr or Kaposi sarcoma herpesvirus. As a new approach to gammaherpesvirus vaccination, we developed and tested a replication-deficient virus (RDV) platform, using murine gammaherpesvirus 68 (MHV68), a well-established mouse model for gammaherpesvirus pathogenesis studies and preclinical therapeutic evaluations. We employed codon-shuffling-based complementation to generate revertant-free RDV lacking expression of the essential replication and transactivator protein encoded by ORF50 to arrest viral gene expression early after de novo infection. Inoculation with RDV-50.stop exposes the host to intact virion particles and leads to limited lytic gene expression in infected cells yet does not produce additional infectious particles. Prime-boost vaccination of mice with RDV-50.stop elicited virus-specific neutralizing antibody and effector T cell responses in the lung and spleen. In contrast to vaccination with heat-inactivated WT MHV68, vaccination with RDV-50.stop resulted in a near complete abolishment of virus replication in the lung 7 days post-challenge and reduction of latency establishment in the spleen 16 days post-challenge with WT MHV68. Ifnar1−/− mice, which lack the type I interferon receptor, exhibit severe disease and high mortality upon infection with WT MHV68. RDV-50.stop vaccination of Ifnar1−/− mice prevented wasting and mortality upon challenge with WT MHV68. These results demonstrate that prime-boost vaccination with a gammaherpesvirus that is unable to undergo lytic replication offers protection against acute replication, impairs the establishment of latency, and prevents severe disease upon the WT virus challenge. Our study also reveals that the ability of a gammaherpesvirus to persist in vivo despite potent pre-existing immunity is an obstacle to obtaining sterilizing immunity.

Pyrrhotites in asteroid 162173 Ryugu: Records of the initial changes on their surfaces with aqueous alteration

The surface chemistry of pyrrhotites from intact particles directly collected from asteroid (162173) Ryugu was investigated by micro-Raman spectroscopy. The Raman peak characteristic to pyrrhotite was observed at around 115 cm−1 in Ryugu pyrrhotites, similar to freshly cleaved surfaces of terrestrial pyrrhotites. Additional Raman bands centered at around 220, 275, and 313 cm−1 with broadened features were also detected from the Ryugu pyrrhotites. The set of Raman bands at 220 and 275 cm−1 was assigned to typical Fe-S stretching vibrations of ν2 (225 cm−1) and ν1 (275 cm−1). These bands are not clearly observed in bulk crystals of pyrrhotite but appear in its nanoparticulate phase. These bands are ordinarily seen in amorphous monosulfides that formed under low oxygen fugacity (fO2) conditions in nature, indicating that the structural alteration of pyrrhotite surfaces occurred heterogeneously on the nanoscale under low fO2 conditions. Further, the Raman band at 313 cm−1 was attributed to a characteristic tetrahedral bonding of Fe(III) in the lattice of FeII1-3xFeIII1-2xS, followed by the local breakdown of the crystal lattice structures from planar bonding with Fe(II). In addition, some areas of the Ryugu pyrrhotite grains showed corroded structures with iridescence. Furthermore, assemblages of magnetite particles were also preferentially observed on small areas of the likely-dissolved pyrrhotite crystals in phyllosilicate matrices. These characteristic features in the Raman spectra and in corroded structures of Ryugu pyrrhotites record changes in the local environmental conditions via aqueous alteration. The corrosion of pyrrhotite crystals followed by the preferential formation of magnetite particles by asteroidal water is the likely product of dissolution of Fe(II) from the pyrrhotite surface and its oxidative precipitation in microchemical environments on the Ryugu parent body.

Comparison of SERS spectra of intact and inactivated viruses via machine learning algorithms for the viral disease’s diagnosis application

In this work, Surface-enhanced Raman spectroscopy (SERS) along with machine learning algorithms (MLA) were used to detect and classify the viral particles to assess the possibility of using the spectra of inactivated influenza A viruses for MLA training and spectra database compilation for further study and diagnosis of intact forms of the virus. Viral particles inactivation was performed by formalin, ultraviolet and beta-propiolactone. Support vector method and principal component analysis allowed to classify intact and inactivated viral particles spectra with an accuracy of 80.0–96.7 %. The results obtained suggest that it is not advisable to create a spectral database and train machine learning algorithms for their further application in SERS diagnostics of intact viruses based on the spectra of the inactivated virus particles.

Impact of temperature variations on BISON predictions of Ag release in AGR-1 and AGR-2 experiments

Understanding and quantifying the release of fission products like silver (Ag) from TRistructural ISOtropic (TRISO) fuel particles is important to assess the safe operation of advanced high temperature reactors. Although the silicon carbide (SiC) layer of TRISO particles is effective as the main fission product barrier, Ag can be released from intact TRISO particles. A mechanistic model for the effective Ag diffusivity, Deff, was previously developed as a function of temperature and microstructure variables informed by atomistic modeling of Ag diffusivity. In this study, we use this model to explore how experimental temperature uncertainties impact the overall predicted Ag release. This analysis indicates that calculated temperature uncertainties have a significant impact on the overall Ag release predictions. Furthermore, we show that the time average volume average (TAVA) temperature is not an appropriate proxy for the detailed temperature histories to predict fission product release due to the Arrhenius dependence of Ag diffusivity with respect to temperature. The detailed temperature histories, therefore, provide the most accurate results and are of most importance for modeling efforts. Overall, this work shows the importance of considering the experimental uncertainty of the temperature on computational predictions of fission product transport and release and the need for more accurate temperature histories from future experiments.

A novel inhibitor of the mitochondrial respiratory complex I with uncoupling properties exerts potent antitumor activity

Cancer cells are highly dependent on bioenergetic processes to support their growth and survival. Disruption of metabolic pathways, particularly by targeting the mitochondrial electron transport chain complexes (ETC-I to V) has become an attractive therapeutic strategy. As a result, the search for clinically effective new respiratory chain inhibitors with minimized adverse effects is a major goal. Here, we characterize a new OXPHOS inhibitor compound called MS-L6, which behaves as an inhibitor of ETC-I, combining inhibition of NADH oxidation and uncoupling effect. MS-L6 is effective on both intact and sub-mitochondrial particles, indicating that its efficacy does not depend on its accumulation within the mitochondria. MS-L6 reduces ATP synthesis and induces a metabolic shift with increased glucose consumption and lactate production in cancer cell lines. MS-L6 either dose-dependently inhibits cell proliferation or induces cell death in a variety of cancer cell lines, including B-cell and T-cell lymphomas as well as pediatric sarcoma. Ectopic expression of Saccharomyces cerevisiae NADH dehydrogenase (NDI-1) partially restores the viability of B-lymphoma cells treated with MS-L6, demonstrating that the inhibition of NADH oxidation is functionally linked to its cytotoxic effect. Furthermore, MS-L6 administration induces robust inhibition of lymphoma tumor growth in two murine xenograft models without toxicity. Thus, our data present MS-L6 as an inhibitor of OXPHOS, with a dual mechanism of action on the respiratory chain and with potent antitumor properties in preclinical models, positioning it as the pioneering member of a promising drug class to be evaluated for cancer therapy.MS-L6 exerts dual mitochondrial effects: ETC-I inhibition and uncoupling of OXPHOS. In cancer cells, MS-L6 inhibited ETC-I at least 5 times more than in isolated rat hepatocytes. These mitochondrial effects lead to energy collapse in cancer cells, resulting in proliferation arrest and cell death. In contrast, hepatocytes which completely and rapidly inactivated this molecule, restored their energy status and survived exposure to MS-L6 without apparent toxicity.

Strong and efficient summertime carbon export driven by aggregation processes in a subarctic coastal ecosystem

AbstractSinking marine particles, one pathway of the biological carbon pump, transports carbon to the deep ocean from the surface, thereby modulating atmospheric carbon dioxide and supplying benthic food. Few in situ measurements exist of sinking particles in the Northern Gulf of Alaska; therefore, regional carbon flux prediction is poorly constrained. In this study, we (1) characterize the strength and efficiency of the biological carbon pump and (2) identify drivers of carbon flux in the Northern Gulf of Alaska. We deployed up to five inline drifting sediment traps in the upper 150 m to simultaneously collect bulk carbon and intact sinking particles in polyacrylamide gels and measured net primary productivity from deck‐board incubations during the summer of 2019. We found high carbon flux magnitude, low attenuation with depth, and high export efficiency. We quantitatively attributed carbon flux between 10 particle types, including various fecal pellet categories, dense detritus, and aggregates using polyacrylamide gels. The contribution of aggregates to total carbon flux (41–93%) and total carbon flux variability (95%) suggest that aggregation processes, not zooplankton repackaging, played a dominant role in carbon export. Furthermore, export efficiency correlated significantly with the proportion of chlorophyll a in the large size fraction (&gt; 20 μm), total aggregate carbon flux, and contribution of aggregates to total carbon flux. These results suggest that this stratified, small‐cell‐dominated ecosystem can have sufficient aggregation to allow for a strong and efficient biological carbon pump. This is the first integrative description of the biological carbon pump in this region.

Microstructure and viscosity of in vitro-digested rye and wheat food products

Understanding rye and wheat digestion is vital for evaluating impacts on nutrient availability and glycaemic responses. This study investigated the disintegration of processed high-fibre rye foods and refined wheat products during simulated intestinal digestion, aiming to link product characteristics with nutrient liberation. The overarching aim was to elucidate how these rye products contribute to the observed benefits in human intervention studies, particularly regarding satiety, weight loss, and metabolism.Analysis included four wholegrain rye products and three refined wheat products, spanning yeast-fermented breads and un-leavened cereal products. Microstructure examination revealed larger, partially intact digesta particles in wholegrain rye products after 120 min of digestion, alongside more aggregated and less degraded starch granules compared to refined wheat bread. Fermented rye bread exhibited greater degradation of subaleurone cell wall fragments than un-fermented rye bread. Viscosity assessments indicated lower viscosity for wheat products than for rye products, with yeast-fermented soft rye bread and rye crispbread showing notably lower viscosity than unfermented rye products. Post-digestion carbohydrate analysis uncovered higher glucose and maltose release during digestion for wheat products. PCA analysis confirmed negative correlations between glucose and maltose release and rye products, characterized by larger post-digestion bolus particles and higher dietary fibre. The elevated cell wall content in rye products acted as a protective barrier for starch granules, mitigating swelling and amylose release, explaining the observed viscosity differences between wheat and rye products and potentially influencing starch digestibility. Consequently, rye products undergo slower and less complete digestion than wheat, aligning with findings from human intervention studies.

Deposition mechanism of ceramic reinforced metal matrix composites via cold spraying

Understanding the mechanism underlying metal matrix composites (MMCs) in cold spraying is critical to fabricate innovative MMC coatings and deposits with tailored microstructures and excellent mechanical performance. In this work, we propose a systematic numerical simulation framework combined with experimental observations to reveal the co-deposition mechanism of cold-sprayed metal/ceramic MMCs. The finite element modeling results and experimental validation found some critical conclusions regarding the deposition mechanism. The modeling results show that most of alumina particles are significantly fractured in Inconel718/alumina coating in contrast to aluminum/alumina coating. This fact suggests that the critical fragmentation velocity (CFV) of ceramic particles is dependent on the hardness of the metal substrate/matrix. Ceramic particles are more prone to fracture when co-deposited with metals having high hardness. Moreover, the simulation results also indicate that the rebounded alumina particles or fragments are possibly captured by incoming metal particles and trapped into MMCs. Since intact alumina particles are barely seen in composites, especially when co-deposited with hard metals, it can be concluded that direct ceramic particle deposition significantly relies on fragmentation. In addition, secondary fragmentation plays a key role in the metal-ceramic co-deposition process. The particles that have completed the impact process may be subjected to secondary fragmentation in the form of new cracks and fragmentation due to the impact by following metal and ceramic particles.

A shedding analysis after AAV8 CNS injection revealed fragmented viral DNA without evidence of functional AAV particles in mice.

Adeno-associated viruses (AAV) are commonly used in the scientific field due to their diverse application range. However, AAV shedding, the release of virions from the host organism, can impact the safety of AAV-based approaches. An increasing number of authorities require the characterization of vector shedding in clinical trials. Recently, shedding of transduced laboratory animals has also gained attention regarding the necessary disposal measures of their waste products. However, no explicit international regulations for AAV-shedding waste exist. Generating insights into shedding dynamics becomes increasingly relevant to help authorities develop adequate regulations. To date, knowledge of AAV vector shedding in mice is very limited. Moreover, confirmation of functional shed AAV particles in mice is missing. Therefore, we examined feces, urine, and saliva of mice after CNS injection with AAV2/8. It revealed the presence of viral DNA fragments via qPCR for up to 4 days after injection. To examine AAV functionality we performed nested PCR and could not detect full-length viral genomes in any but two collected feces samples. Furthermore, a functional infection assay did not reveal evidence of intact AAV particles. Our findings are supposed to contribute murine shedding data as a foundation to help establish still lacking adequate biosafety regulations in the context of AAV shedding.

The effect of size and density on the mean retention time of particles in reindeer (Rangifer tarandus)

Particle passage from the reticulorumen (RR) depends on particle density and size. A classic way of assessing these effects is the use of plastic markers of varying density and size that are recovered in the faeces. Here, we report results of an experiment where four fistulated reindeer (Rangifer tarandus, 96 ± 12 kg) were fed two different diets (browse, voluntary dry matter intake [DMI] 70 ± 10 g/kg0.75/d; or a pelleted diet, DMI 124 ± 52 g/kg0.75/d) and dosed via fistula with 8 different particle types combining densities of 1.03, 1.22 and 1.44 g/ml and sizes of 1, 10 and 20 mm. Generally, particles that passed the digestive tract intact (not ruminated) did so relatively early after marker dosing, and therefore had shorter mean retention times (MRT) than ruminated particles. On the higher intake, the overall mean retention time (MRT) of particles was shorter, but this was not an effect of shorter MRT for either intact or ruminated particles, but due to a higher proportion of intact particles at the higher intake. This supports the concept that ruminants do not adjust chewing behaviour depending on intake, but that a lower proportion of digesta is submitted to rumination due to pressure-driven escape from the forestomach at higher gut fills. Compared to cattle (Bos primigenius taurus), muskoxen (Ovibos moschatus) and moose (Alces alces) that had received the same markers, reindeer had a lower proportion of 1 mm particles that passed intact. Our results support the concept that the critical size threshold for particles leaving the ruminant forestomach is dependent on body size. While the results likely do not represent findings peculiar for reindeer, they indicate fundamental mechanisms operating in the forestomach of ruminants.

SARS-CoV-2 presence in recreational seawater and evaluation of intestine permeability: experimental evidence of low impact on public health.

Coastal seawater pollution poses a public health risk due to the potential ingestion of contaminated water during recreational activities. Wastewater-based epidemiology has revealed the abundant presence of SARS-CoV-2 in seawater emitted from wastewater outlets. The objective of this research was to investigate the impact of seawater on SARS-CoV-2 infectivity to assess the safety of recreational activities in seawater. Wild SARS-CoV-2 was collected from oral swabs of COVID-19 affected patients and incubated for up to 90 min using the following solutions: (a) standard physiological solution (control), (b) reconstructed seawater (3.5% NaCl), and (c) authentic seawater (3.8%). Samples were then exposed to two different host systems: (a) Vero E6 cells expressing the ACE2 SARS-CoV-2 receptor and (b) 3D multi-tissue organoids reconstructing the human intestine. The presence of intracellular virus inside the host systems was determined using plaque assay, quantitative real-time PCR (qPCR), and transmission electron microscopy. Ultrastructural examination of Vero E6 cells revealed the presence of virus particles at the cell surface and in replicative compartments inside cells treated with seawater and/or reconstituted water only for samples incubated up to 2 min. After a 90-min incubation, the presence of the virus and its infectivity in Vero E6 cells was reduced by 90%. Ultrastructural analysis performed in 3D epi-intestinal tissue did not reveal intact viral particles or infection signs, despite the presence of viral nucleic acid detected by qPCR. Indeed, viral genes (Orf1ab and N) were found in the intestinal luminal epithelium but not in the enteric capillaries. These findings suggest that the intestinal tissue is not a preferential entry site for SARS-CoV-2 in the human body. Additionally, the presence of hypertonic saline solution did not increase the susceptibility of the intestinal epithelium to virus penetration; rather, it neutralized its infectivity. Our results indicate that engaging in recreational activities in a seawater environment does not pose a significant risk for COVID-19 infection, despite the possible presence of viral nucleic acid deriving from degraded and fragmented viruses.

Tracking the micro- and nanoplastics in the terrestrial-freshwater food webs. Bivalves as sentinel species

Micro- (MPs) and nanoplastics (NPs) are currently ubiquitous in the ecosystems, and freshwater biota is still insufficiently studied to understand the global fate, transport paths, and consequences of their presence. Thus, in this study, we investigated the role of bivalves and a trophic transfer of MPs and NPs in an experimental food chain. The food chain consisted of terrestrial non-selective detritivore Dendrobaena (Eisenia) sp., freshwater benthic filter feeder Unio tumidus, and freshwater benthic detritivore-collectors Asellus aquaticus or Gammarus sp. Animals were exposed to different fluorescently labeled micro- and nanoplastics (PMMA 20 μm, nanoPS 15–18 nm, and 100 nm, PS 1 μm and 20 μm, PE from cosmetics) as well as to the faeces of animals exposed to plastics to assess their influence on the environmental transportation, availability to biota, and bioaccumulation of supplied particles. Damaged and intact fluorescent particles were observed in the faeces of terrestrial detritivores and in the droppings of aquatic filter feeders, respectively. They were also present in the guts of bivalves and of crustaceans which were fed with bivalve droppings. Bivalves (Unio tumidus, and additionally Unio pictorum, and Sphaerium corneum) produced droppings containing micro- and nanoparticles filtered from suspension and deposited them onto the tank bottom, making them available for broader feeding guilds of animals (e.g. collectors, like crustaceans). Finally, the natural ageing of PS and its morphological changes, leakage of the fluorescent labelling, and agglomeration of particles were demonstrated. That supports our hypothesis of the crucial role of the characterization of physical and chemical materials in adequately understanding the mechanisms of their interaction with biota. Microscopical methods (confocal, fluorescent, scanning electron) and Raman and FT-IR spectroscopy were used to track the particles' passage in a food web and monitor structural changes of the MPs' and NPs' surface.

Structural basis for nuclear import of hepatitis B virus (HBV) nucleocapsid core.

Nuclear import of the hepatitis B virus (HBV) nucleocapsid is essential for replication that occurs in the nucleus. The ~360-angstrom HBV capsid translocates to the nuclear pore complex (NPC) as an intact particle, hijacking human importins in a reaction stimulated by host kinases. This paper describes the mechanisms of HBV capsid recognition by importins. We found that importin α1 binds a nuclear localization signal (NLS) at the far end of the HBV coat protein Cp183 carboxyl-terminal domain (CTD). This NLS is exposed to the capsid surface through a pore at the icosahedral quasi-sixfold vertex. Phosphorylation at serine-155, serine-162, and serine-170 promotes CTD compaction but does not affect the affinity for importin α1. The binding of 30 importin α1/β1 augments HBV capsid diameter to ~620 angstroms, close to the maximum size trafficable through the NPC. We propose that phosphorylation favors CTD externalization and prompts its compaction at the capsid surface, exposing the NLS to importins.