In the last two decades, extracellular vesicles (EVs) from the three domains of life, archae, bacteria and eukaryota, have gained increasing scientific attention. As such, the role of EVs in host-pathogen communication and immune modulation are being intensely investigated. Pivotal to EV research is the determination of how and where EVs are taken up by recipient cells and organs in vivo, which requires suitable tracking strategies including labelling. Labelling of EVs is often performed post-isolation which increases risks of non-specific labelling and the introduction of labelling artefacts. Here we exploited the inability of helminths to de novo synthesise fatty acids to enable labelling of EVs by whole organism uptake of fluorescent lipid analogues and the subsequent incorporation in EVs. We showed uptake of DOPE-Rhodamine in Anisakis spp. and Trichuris suis larvae. EVs isolated from supernatant of Anisakis spp. labelled with DOPE-Rhodamine were characterised to assess effects of labelling on size, structure and fluorescence of EVs. Fluorescent EVs were successfully taken up by the human macrophage cell line THP-1. This study therefore presents a novel staining method that can be utilized by the EV field in parasitology and potentially across multiple species.

The Encyrtidae encompass large group of insects widely used in the biocontrol programs of scale insects (Hemiptera: Coccoidea). Here we investigated antennal sensory system organization of Anagyrus vladimiri females, a species widely used as a biocontrol agent of Planococcus spp. and Pseudococcus spp. Ultrastructural investigations with scanning (SEM) and transmission (TEM) electron microscopy revealed filiform antennae made of nine segments. The scape was enlarged and paddle-like, compared to the other antennomeres. The club was mono-segmented and housed the highest number of sensilla. On the antennae, eight morphologically different types of sensilla were recorded; sensilla trichoidea I, trichoidea II, chaetica I, chaetica II, grooved peg sensilla, campaniform sensilla, multiporous plate sensilla, multiporous basiconic sensilla. The sensilla belonged to the major types sensilla types (olfactory, gustatory, thermos-hygroreceptor). The sensilla trichoidea I and multiporous plate sensilla were the most abundant types. The potential functional role of each sensillum was discussed.

Background: We assess locoregional drug targeting effectiveness of intraoperative (IO) cryoprobe-assisted injection of blue dye (BD) and cytotoxic-tracer mixture (TTM), in VX2 tumor model, and its translational value to cryo-assisted breast tumor surgery with BD alone. Methods: Under computed tomography (CT) guidance, we injected two ml TTM in five aliquots in the margin of 16 frozen or normothermic VX2 tumors. We evaluated the IO and post-operative drug targeting and therapeutic efficacy in tumor-host interface (T-HI) by CT, gross examination, and histopathology. In twenty-six T1 to T4 primary breast cancer (BRCA) we performed ultrasound-guided (US) cryoprobe-assisted tumor freezing, BD guided lymphatic mapping, and surgery. We evaluated, IO and in freshly resected specimen, BD distribution pattern in T-HI, lymph node(s), breast parenchyma, and resection cavity. Results: Fluids-impervious frozen VX2 or breast tumor transported drug(s) an arc-like pattern at T-HI regardless of freeze dose, number of freeze-thaw cycles, drug dose fractionation, tumor characteristics or dimensions. During melting, TTM spread within fifty percent VX2 tumor mirrored that of T-HI; it was massive in normothermia. In VX2 twenty percent focal margin necrosis at pathology coincided with CT gap; in both studies, BD dose-staining spread in T-HI and tumor was linear. Eighty-four patients had one to twelve stained axillary lymph nodes; sixty-nine percent and all respectively, had another quadrant and resection cavity stained. Conclusion: Intraoperative freezing-assisted drug delivery and targeting techniques during cryoablation of VX2 tumor translate successfully to locoregional BD targeting, lymphatic mapping during cryo-assisted surgery of breast cancer.

ATP-binding cassette (ABC) transporters, acting as cellular "pumps," facilitate solutes translocation through membranes via ATP hydrolysis. Their overexpression is closely tied to multidrug resistance (MDR), a major obstacle in chemotherapy and neurological disorder treatment, hampering drug accumulation and delivery. Extensive research has delved into the intricate interplay between ABC transporter structure, function, and potential inhibition for MDR reversal. Cryo-electron microscopy has been instrumental in unveiling structural details of various MDR-causing ABC transporters, encompassing ABCB1, ABCC1, ABCG2, as well as recently revealed ABCC3 and ABCC4 structures. The newly obtained structural insight has deepened our understanding of substrate and drug binding, translocation mechanisms, and inhibitor interactions. Given the growing body of structural information available for human MDR transporters and their associated mechanisms, we believe it is timely to compile a comprehensive review of these transporters and compare their functional mechanisms in the context of multidrug resistance. Therefore, this review primarily focuses on the structural aspects of clinically significant human ABC transporters linked to MDR, with the aim of providing valuable insights to enhance the effectiveness of MDR reversal strategies in clinical therapies.

Kininogens are multidomain glycoproteins found in the blood of most vertebrates. They are important in the blood coagulation cascade pathways - in intrinsic pathway activation leading to thrombin generation partially independently from tissue factor dependent extrinsic pathway, connecting blood coagulation with the kallikrein-kinin system. Nothing is known about the shape on atomic level therefore the endeavor to obtain the good-quality spatial structure of kininogens is important for a better understanding of their role in disease and treatment. Application of cryo-EM is important for solving the spatial structure of kininogens, drawing new frontiers in understanding the function and opening new pathways for drug development.

Nuclear vacuoles are specific structures present on the head of the human sperm of fertile and non-fertile men. Human sperm head vacuoles have been previously studied using motile sperm organelle morphology examination (MSOME) and their origin related to ab-normal morphology, abnormal chromatin condensation and DNA fragmentation. How-ever, other evidences argued that human sperm vacuoles are physiological structures and consequently, to date, the nature and origin of the nuclear vacuoles remains to be elucidat-ed. Here, we aim to define the incidence, position, morphology, and molecular content of the human sperm vacuoles using transmission electron microscopy (TEM) and immuno-cytochemistry techniques. Results showed that ~50% of the analyzed human sperm cells (n = 1908; 17 normozoospermic human donors) contained vacuoles mainly located (80%) in the anterior head region. A significant positive correlation was also found between the sperm vacuole and nucleus areas. Furthermore, it was confirmed that nuclear vacuoles were invaginations of nuclear envelope containing cytoskeletal proteins and a cytoplas-mic enzyme, discarding a nuclear or acrosomal origin. According to our findings, these human sperm head vacuoles are cellular structures which take the origin from nuclear invaginations and contain perinuclear theca (PT) components, allowing to define a new term of ‘nuclear invaginations’ rather than ‘nuclear vacuoles.

This study was conducted in a laboratory-controlled environment aiming at studying the physical properties and elemental composition of coal combustion particles in a brazier. Particles were sampled ~1 m above the stove using a partector, where particles were collected on gold Transmission Electron Microscopy (TEM) grids and polycarbonate filters for TEM and inductively coupled plasma mass spectrometry (ICP-MS) analysis, respectively. Particles for elemental analysis collected on a 37 µm polycarbonate filters whereby a Gillian pump was used to draw in air. During sampling, a 2.5 µm cyclone was attached to the sampling cassette to isolate larger particles. The results have shown that combustion particles emitted during the early stage of combustion where single spherical particles with a diameter of around 450 nm. As the combustion progresses, the particle diameter gradually decreases and the morphology changes to accretion chain and fluffy bead structure for the flaming and char-burning phase, respectively.

The impact of the cell community on its individual components in the prokaryotic realm is often overlooked. However, in the case of intracellular pathogens, where each infected cell can be considered as a single community, understanding how a population adapts to its environment to evolve and successfully propagate is key. Chlamydia infections are characterised by a silent propagation of the bacteria within individual hosts and the wider population. Chlamydia are strict intracellular pathogens residing within a specialised membrane-bound compartment called the inclusion. The life cycle of Chlamydia involves altering between the invasive elementary bodies (EBs) and replicative reticulate bodies (RBs). We have used cryo-soft X-ray tomography to observe individual inclusions, combining excellent resolution (40 nm) and large volume imaging (up to 16 µm). Combined with a semi-automated segmentation pipeline, we were able to consider each inclusion as an individual bacterial niche. Within the inclusion, we identified and classified different forms of the bacteria and confirmed the recent finding that RBs have a variety of volumes (small, large and abnormal). Moreover, we demonstrate that the proportions of these different RB forms depend on the bacterial concentration in the cell demonstrating the impact of the group on its individual component. We conclude that each inclusion operates as an autonomous community which regulates the characteristics of individual bacteria within the inclusion

Super-hydrophobic surfaces and coatings have stimulated a great deal of research, with the aim to achieve better wetting properties. Factors such as surface chemistry and roughness play an important role in changing the surface energy, which in turn leads to changes in the wettability. Here, we have analysed the time dependence of the oxide layer and possible surface adsorbates on the surface topography of an Al59Cu25Fe13B3 quasicrystalline material for changes to the wettability. The quasicrystalline matrix phase was 94 % of the sample volume, and covered by a very smooth, amorphous oxide layer. The AlB12 and AlFe2B2 boron-rich phases were embedded in the quasicrystalline material as a result of a 3 at.% boron addition, which makes atomisation of the material a simpler process. Under ambient conditions the sample was naturally covered by an oxide layer; therefore, it is referred to as “surfenergy”, to distinguish it from the conventional surface energy of a bare quasicrystal surface. The growth of the oxide layer with atmospheric ageing and annealing at 500° C in air for various times was investigated for both cases. The phase most prone to oxidation was the boron-rich AlFe2B2, which influenced the topography of the surface and accordingly the wetting behaviour of the specimen. We demonstrated that the surfenergy depends on the polar component, which is the most sensitive to the operating conditions. A correlation between the surfenergy components and the surface roughness was found. In addition, theoretical models to determine the wettability were included.

Soot is characterized by a multiscale structural organization and Transmission Electron Microscope (TEM) is the only diagnostic tool giving access to it. However, being a diffraction-based technique, TEM images only aromatic systems and thus, it is particularly useful to combine it with electron energy-loss spectroscopy (EELS), able to provide quantitative information about the relative abundance of sp3 and sp2 hybridized carbon. In this paper a method for the EELS spectrum analysis of carbonaceous materials recently developed for electron-irradiated graphite and glassy carbon composition analysis has been applied for the first time on soot samples, in order to test its performance in soot nanostructure study in combination with TEM and High Resolution TEM (HRTEM). Soot samples here analysed were collected in the soot inception region of premixed flames of different hydrocarbon fuels. EELS, in agreement with TEM and HRTEM, showed a quite disordered and heterogeneous structure for young soot, without any significant distinction between soot formed from methane and ethylene fuels.

This research was conducted to manufacture thermally expandable microspheres (TEMs) for vehicles’ underbody coating and to apply them on an industrial scale. TEMs heat resistance was studied depending on the ratios of a cross-linking agent and an initiator. This research focused on the content of a cross-linking agent and how it affected the results. The TEMs’ outer shell was thickened to solve the problem of the foam expansion ratio’s reduction that occurred due to the shrinkage after the maximum expansion (Tmax) was reached. After foaming, the cross-sectional thickness and surface of the sample with thickened outer shell were observed. The TEMs with the thickened shell showed the least shrinkage, which indicated excellent shrinkage stability, even after prolonged exposure to heat.

This study was conducted to improve the white index (WI) by preparing thermally expandable microspheres (TEMs) for wallpaper. The thermal properties, foam expansion ratio and WI were studied depending on the particle size of colloidal silica in the preparation of TEMs. As a result, the TEMs with small particles of colloidal silica showed the best results for whiteness and yellowing. Additionally, TGA results indicated that it was highly possible that colloidal silica with small particle sizes was physically or chemically attached to the surface of the TEMs that led to an improvement in whiteness at high temperatures.

Encapsulation of antioxidant polyphenols leads to increase solubility and bioavailability of these micronutrients in solution. The encapsulation of antioxidant resveratrol, genistein and curcumin by folic acid-chitosan nanocapsules was studied in aqueous solution, using multiple spectroscopic methods, TEM images. Structural analysis showed that polyphenol bindings are via hydrophilic, hydrophobic and H-bonding contacts with resveratrol forming more stable conjugates. As chitosan size increased, the binding efficacy and stability of polyphenol-polymer adducts were increased. Polyphenol binding induced major alterations of chitosan morphology. Chitosan nanoparticles are capable of delivery of polyphenols in vitro.

Single-particle cryo-electron microscopy (cryo-EM SPA) has recently emerged as an exceptionally well-suited technique for determining the structure of membrane proteins (MPs). Indeed, in the last years, it was observed a huge increase in the number of MPs solved by cryo-EM SPA at a resolution better than 3.0 Å in the Protein Data Bank (PDB). However, sample preparation remains a significant challenge in the field. Here, we evaluated in the MPs solved by cryo-EM SPA deposited in the PDB in the last two years at a resolution below 3.0 Å the most critical parameters for sample preparation: i) the surfactant used for protein extraction from the membrane, ii) the surfactant, amphiphiles, nanodiscs or other molecules present in the vitrification step, iii) the vitrification method employed, and iv) the type of grids used. The aim is not to provide a definitive answer on the optimal sample conditions for cryo-EM SPA of MPs, but rather assess the current trends in the MP structural biology community towards obtaining high-resolution cryo-EM structures.

A FIB/STEM interfacial study was performed on a TBC/Ti₂AlC MAX phase system, oxidized in an aggressive burner rig test (Mach 0.3 at 1300°C for 500 h). The 7YSZ TBC, a-Al₂O₃ TGO, and MAXthal 211 Ti₂AlC base were variously characterized by TEM/STEM, EDS, SADP, and HRTEM. The YSZ was a mix of ‘clean’ featureless and ‘faulted’ high contrast grains. The latter exhibited ferro-elastic domains of high Y content tetragonal t'' variants. No martensite was observed. The TGO was essentially a duplex a-Al₂O₃ structure of inner columnar plus outer equiaxed grains. It maintained a perfectly intact, clean interface with the Ti₂AlC substrate. The Ti₂AlC substrate exhibited no interfacial Al-depletion zone, but rather numerous faults along the basal plane of the hexagonal structure. These are believed to offer a means of depleting Al by forming crystallographic, low-Al planar defects, proposed as Ti_2.5AlC_1.5. These characterizations support and augment prior optical, SEM, and XRD findings that demonstrated remarkable durability for the YSZ/Ti₂AlC MAX phase system in aggressive burner tests.

Bacteriophages can provide alternative measures for the control of E. coli O157:H7 that is currently an emerging food-borne pathogen of severe public health concern. This study was aimed at characterising E. coli O157:H7 specific phages as potential biocontrol agents for these pathogens. Fifteen phages were isolated and screened against 69 environmental E. coli O157:H7. Only 3 phages displayed broad lytic spectra against environmental shiga toxin-producing E. coli O157:H7 strains. These 3 lytic phages were designated V3, V7 and V8. Subsequent characterization indicated that they displayed very high degree of similarities despite isolation from different locations. Transmission Electron microscopy (TEM) of the phages revealed that they all had isometric heads of about 73 – 77 nm in diameter and short tails ranging from 20 - 25 nm in diameter. Phages V3, V7 and V8 were assigned to the family Podoviridae based on their morphology. Pulsed field gel electrophoresis (PFGE) genome estimation of the 3 phages demonstrated identical genome sizes of ~ 69 nm. The latent periods of these phages were 20 min, 15 min, and 20 min for V3, V7 and V8 respectively while the burst sizes were 374, 349 and 419 PFU/ infected cell respectively. While all the phages were relatively stable over a wide range of salinity, temperatures and pH values, their range of infectivity or lytic profile was rather narrow on environmental E. coli O157:H7 strains isolated from cattle faeces. This study showed that the Podoviridae bacteriophages are the dominant E. coli O57:H7-infecting phages harboured in cattle faeces in the North-West Province of South Africa and due to their favourable characteristics can be exploited in the formulation of phage cocktails for the bio-control of E. coli O157:H7 in meat and other meat products.

Vibrational spectroscopies (FTIR, Raman) are exceptionally valuable tools for the identification and crystal-chemical study of fibrous minerals, and asbestos amphiboles in particular. Raman spectroscopy has been widely applied in toxicological studies and thus a large corpus of reference data on regulated species is found in the literature. However, FTIR spectroscopy has been mostly used in crystal-chemical studies and very few data are found on asbestos amphiboles. This paper is intended to fill this gap; we report new FTIR data collected on a suite of well-characterized samples of the five regulated amphibole species: anthophyllite, amosite and crocidolite, provided by the Union for International Cancer Control (UICC) Organization, and tremolite and actinolite, from two well-known occurrences. The data from these reference samples have been augmented by results from additional specimens to clarify some aspects of their spectroscopic features. We show that the FTIR spectra in both the OH-stretching region and in the lattice modes region can be effective for rapid identification of the asbestos type.

Striated muscle thick filaments are composed of myosin II and several non- myosin proteins which define the filament length and modify its function. Myosin II has a globular N-terminal motor domain comprising its catalytic and actin-binding activities and a long α-helical, coiled-coil tail that forms the dense filament backbone. Myosin alone polymerizes into filaments of irregular length, but striated muscle thick filaments have defined lengths that with the thin filament define the sarcomere structure. The motor domain structure and function are well understood but the myosin filament backbone is not. Here we report a structure of the flight muscle thick filaments from Drosophila melanogaster at 4.7 Å resolution which eliminates previous ambiguities in the non-myosin densities. The full proximal S2 region is resolved as are the connecting densities between the Ig domains of stretchin-klp. The proteins, flightin and myofilin are resolved in sufficient detail to build an atomic model based on an AlphaFold prediction. Our results suggest how flightin and myofilin cooperate to define the structure of the thick filament and explains a key myosin mutation that affects flightin incorporation. Drosophila is a genetic model organism for which our results can define strategies for functional testing.

Tick-borne encephalitis virus (TBEV) is a RNA-containing enveloped virus, a member of the Flaviviridae family. Here we describe a detailed analysis of the size and structure of inactivated TBEV (the Sofyin-Chumakov TBEV strain, used in vaccines). Four analytical methods were used to analyze individual TBEV particles—negative staining TEM, cryo-EM, AFM, and NTA. All methods confirmed that the particles were monodisperse, and their mean size was ~50 nm. Cryo-EM data were used to obtain a 3D electron density model of the virus with clearly distinguishable E-proteins. STEM-EELS analysis detected phosphorous in the particles, which was interpreted as the RNA signal. Altogether, the described analytical procedures can be valuable for the further analysis of vaccine inactivated virus samples.

With forty years of developments, bio-macromolecule cryo-electron microscopy has met its revolution of resolution and is playing a very important role in structural biology study. According to different specimen states, cryo-electron microscopy (cryo-EM) involves three specific techniques, single particle analysis (SPA), electron tomography and sub-tomogram averaging, and electron diffraction. All these three techniques have not realized their full potentials of solving structures of bio-macromolecules and therefore need to be developed in the future. In this review, the current existing bottlenecks of cryo-EM SPA are discussed with theoretical analysis, which includes air-water interface during specimen cryo-vitrification, bio-macromolecular conformational heterogeneity, focus gradient within thick specimen, and electron radiation damage. Besides, potential solutions of these bottlenecks are proposed and discussed, which are worthy of further investigations in the future.

In current work, we illustrate the effect of adding small amount of Carbon to very common Co2MnSi Heusler alloy based-glass coated microwires. A significant change in the magnetic and structure structural properties has observed for the new alloy Co2MnSiC compared to the Co2MnSi alloy. The magneto-structural investigations have performed to clarify the main phys-ical parameters i.e., structural & magnetic at a wide range of measuring temperature. The XRD analysis illustrated the well-defined crystalline structure with average grain size (Dg = 29.16 nm) and a uniform cubic structure with A2-type compared to the mixed L21 and B2 cubic structures for Co2MnSi-based glass coated microwires. The magnetic behaviour has investigated at a tem-perature range (5 to 300 K) and an external applied magnetic field (50 Oe to 20 kOe). The adding of small amount of Carbon to the Co2MnSi matrix enhance the magnetic thermal stability, where the thermomagnetic behaviour of Co2MnSiC glass-coated microwires show a perfect stable be-haviour for a temperature range from 300 K to 5 k, the differences between the coercivity value is only 0.3 Oe compared to 4 Oe for Co2MnSi-sample. In addition, M-H loops measured at tempera-ture below 50 K show unsaturated loops; meanwhile the Co2MnSi loops shows a strong antifer-romagnetic coupling for the loops measured below 50 K. By studying the field cooling (FC) and field heating (FH) magnetizations curves at a wide range of external applied magnetic field we detected a critical magnetic field (H = 1 kOe) where FC and FH curves have a stable magnetic behavior for Co2MnSiC sample, such stability does not find in Co2MnSi sample. We proposed a phenomenal expression to estimate the magnetization thermal stability, ΔM (%), of FC and FH magnetization curves and the maximum value is detected at the critical magnetic field where ΔM (%) ≈ 98 %. The promising magnetic stability of Co2MnSiC glass-coated microwires with tem-perature is due to the changing of the microstructure induced by adding Carbon, as the A2-type structure show a unique stability by variation the temperature and the external magnetic field. In addition, a unique internal mechanical stress, which induced during the fabrication process and plays on controlling magnetic behavior with temperature and external magnetic field. The ob-tained results make Co2MnSiC promising candidate for magnetic sensing devices based Heusler glass-coated microwires.

In this paper, in order to provide proper parameters for the preparation of semi-solid billets, the semi-solid annealing of hot-rolled 2A14 Al alloy was investigated. The microstructure was characterized by X-ray diffraction (XRD), scanning electron microscope (SEM) with X-ray energy dispersive spectrometer (EDS) and electron backscattered diffraction (EBSD), scanning transmission electron microscope (STEM). XRD results show that with the increase of temperature, equilibrium θ-Al2Cu gradually dissolved in the matrix. The EDS results of SEM and STEM show that there were coarse θ-Al2Cu phase, ultra-fine precipitate Al(MnFeSi) or (Mn, Fe)Al6 phase and atomic clusters in the microstructure. EBSD results show that the recrystallization mechanism was dominated by continuous static recrystallization (CSRX), the homogeneous nucleation occurred when the sample was heated to near solidus temperature, and CSRX happened in the semi-solid temperature. In the process of recrystallization, the micro-texture changed from preferred orientation to random orientation. A variety of experimental results show that static recrystallization (SRX) occurred at semi-solid temperature due to the blocking effect of atomic clusters on dislocation slip and the Zener drag effect of fine precipitates on low angle grain boundaries (LAGBs) disappeared with melting at semi-solid temperature.

Various crystallite size estimation methods were used to analyze X-ray diffractograms of spherical cerium dioxide and donut-like titanium dioxide anatase nanoparticles aiming to evaluate their reliability and limitations. The microstructural parameters were estimated from Scherrer, Monshi, Williamson-Hall, and their variants: i) uniform deformation model, ii) uniform strain deformation model, and iii) uniform deformation energy density model, and also size-strain plot, and Halder-Wagner method. For that, and improved systematic Matlab code was developed to estimate the crystallite sizes and strain, and the linear regression analysis was used to compare all the models based on the coefficient of determination, where the Halder Wagner method gave the highest value (close to 1). Therefore, being the best candidate to fit the X-ray Diffraction data of metal-oxide nanoparticles. Advanced Rietveld was introduced for comparison purposes. Refined microstructural parameters were obtained from a nanostructured 40.5 nm Lanthanum hexaboride nanoparticles and correlated with the above estimation methods and transmission electron microscopy images. In addition, electron density modelling was also studied for final refined nanostructures, and μ-Raman spectra were recorded for each material estimating the mean crystallite size and comparing by means of a phonon confinement model.

Heat stress is a serious threat to tomato production. The ultrastructure and element mapping of tomato chloroplast were investigated under different nitrogen levels to explore the regulatory effect of nitrogen on tomato chloroplast under heat stress. Experiments design two temperature treatments (25°C/15°C, as control, 40°C/30°C, as severe high-temperature, SHT ) and four nitrogen levels (0.0 kg hm-2, N1; 187.5 kg hm-2, N2; 250 kg hm-2, N3, as recommended nitrogen-application; 312.5kg hm-2 , N4) for a duration of 5 days in artificial climate chamber. The results showed that heat stress led to obvious damage to the ultrastructure of tomato chloroplast, including thylakoid stacking disorder, granum destruction, and starch grain aggregation and degradation. With the increase of nitrogen level, the ratio of long and short axes of tomato chloroplasts increased, the differentiation of chloroplasts was strengthened, the number of plastoglobuli was increased and aggregated, and the degree of damage to the ultrastructure of chloroplasts was reduced. The regulatory effect of nitrogen on the ultrastructure of tomato chloroplasts under heat stress was mainly reflected in the phosphorylation degradation process of starch granules in chloroplasts. There are obvious segmentation lines of Ca, P and Cl on both sides of the inner and outer chloroplasts, and enriched in the chloroplasts, which can outline the boundary of the chloroplasts. Element mapping analysis revealed that heat stress significantly increased the contents of calcium, phosphorus and chlorine in the chloroplast of tomato leaves, while the contents of potassium was decreased. The application of nitrogen fertilizer could significantly regulate the element distribution of tomato chloroplast under heat stress, especially the regulation of phosphorus, chlorine, calcium, magnesium. These results provide new insight into the role of nitrogen using TEM-EDX mapping in the protection of tomato chloroplast under heat stress and the mechanism of heat stress tolerance in plants.

A homogenized, supersaturated Al-Zn-Mg-Zr alloy was processed by severe plastic deformation (SPD) using high-pressure torsion (HPT) technique for different revolutions at room temperature to get ultrafine-grained (UFG) microstructure. The microstructure and mechanical properties of the UFG samples were then studied by transmission electron microscopy (TEM), differential scanning calorimetry (DSC), as well as by tensile and hardness measurements. Emphasis was placed on the effect of shear strain on the evolution of microstructure of the investigated alloy. Experimental results have shown a very interesting evolution of the decomposed microstructure in a wide range of shear strains imposed by HPT. While the global properties, such as the average grain size (~200 nm) and hardness (~2.2 GPa) appeared unchanged, the local microstructure was continuously transformed. After 1 turn of HPT, the decomposed UFG structure contains relatively large precipitates inside grains. In the sample processed by 5 turns in HPT, segregation of Zn atoms into grain boundaries (GBs) can also be observed. After 10 turns, more Zn atoms have segregated into GBs and only smaller size precipitates can be observed inside grains. The intensive segregation of solute atoms into GBs may significantly affect the ductility of the materials, leading to its ultralow-temperature superplasticity. Our findings pave the way for achieving advanced microstructural and mechanical properties in nanostructured metals and alloys by engineering their precipitation and segregation by means of applying different HPT regimes.

The effect of the melt cooling rate on the atomic ordering of austenite and, as a consequence, on the martensitic transformation of a nonstoichiometric alloy of the Ni-Mn-In system has been studied. In-situ TEM observations revealed differences in the mechanism of phase transformations of the alloy subjected to different cooling conditions. It is shown that during quenching a high density of antiphase boundaries (APB) is formed and the alloy is in the austenite-martensitic (10M and 14M) state up to a temperature of 120K. In a slowly cooled alloy, a lower APB density is observed, and a two-stage transformation L21/B2 → 10M → 14M occurs in the range of 150–120 K.

Background: in this paper, a new physical penetration technology for transdermal administration with traditional Chinese medicine (TCM) characteristics - Fu’s cupping therapy (FCT) - was established and studied by in-vitro and in-vivo experiments; the penetration effect and mechanism of FCT physical penetration technology (FCT-PPT) was preliminarily discussed. Method: Indomethacin (IM) as a model drug，by transdermal in vitro tests the establishment of the high，medium and low reference were finished as the chemical permeation system；chemical penetration enhancers and iontophoresis as a reference，the percutaneous penetration effect of FCT for IM patch was evaluated with 7 species diffusion kinetics model and in vitro drug distribution；naproxen as an internal standard，using UPLC-MS/MS technology，the IM quantitative analysis method in vivo was established，and pharmacokinetic parameters (AUC0-t，AUC0-∞，AUMC0-t，AUMC0-∞, Cmax and MRT) as indicators were used evaluate to FCT penetration role in vivo；in the same time，the group used 3K factorial design to study joint synergistic penetration effect on FCT and chemical penetration enhancers (CPEs)；by SEM and TEM，the skin micro and ultrastructural changes of the stratum corneum (SC) surface were observed, to explore pay tank penetration mechanism. Results: In vitro and in-vivo skin permeation experiments revealed that both the total cumulative percutaneous amount and in-vivo percutaneous absorption amount (AUC and AUMC) of indomethacin that permeated SD mouse skin using FCT techniques were greater than the amount observed using CPE and iontophoresis: Firstly, in contrast to the control group, the indomethacin percutaneous rate (PR) of the FCT lower group (FCTL) was 35.52%, and the enhancement ratio (ER) at 9h was 1.76X, which was roughly equivalent to the penetration enhancing effect of the CPEs and iontophoresis; secondly, the indomethacin PR of the FCT middle (FCTM) group and the FCT high intensity group (FCTH) were respectively 47.36% and 54.58%, ER at 9h were separately 3.58X and 8.39X; thirdly, pharmacokinetic studies showed that in-vivo indomethacin percutaneous absorption of the FCTs was higher than that of the control group, that of the FCTM group was slightly higher than that of the CPEs group, and that of the FCTM group was significantly higher than that of the others. Meanwhile, the variance analysis indicated that the combination of the FCT penetration enhancement method and the CPE method had beneficial effects in penetration enhancing of the skin: the significance level of the CPE method was 0.0004, which was apparently lower than the 0.001, meaning the difference was markedly significant; the significance level of the FCT was under 0.0001, its difference markedly significant; and the significance level of factor interaction A×B was lower than 0.0001, indicating that its difference of the synergism was markedly significant. Moreover, SEM and TEM images showed that the SC surfaces of SD rats treaded with FCT-PPT was damaged, and hard to observe the complete surface structure with its SC pores growing bigger and its special “brick structure” becoming looser, indicating that it broke the barrier function of skin, which revealed potentially a major route of skin penetration. Conclusion: FCT, as percutaneous penetration new technologies, has penetration effects significantly, with Chinese characteristics and highly clinical value, worth promoting development.

In this study, we introduce the concept and construction of an innovative Digital Miniature Cathode Ray Magnetometer designed for the precise detection of magnetic fields. This device addresses several limitations inherent to magnetic probes such as D.C. offset, nonlinearity, temperature drift, sensor aging, and the need for frequent recalibration, while capable of oper-ating in a wide range of magnetic fields. The core principle of this device involves the utilization of a charged particle beam as the sensitivity medium. The system leverages the interaction of an electron beam with a scintillator material, which, then, emits visible light that is captured by an imager. The emitted scintillation light is captured by a CMOS sensor. This sensor not only rec-ords the scintillation light but also accurately determines the position of the electron beam, providing invaluable spatial information crucial for magnetic field mapping. The key innova-tion lies in the combination of electron beam projection, CMOS imager scintillation-based de-tection and digital image signal processing. By employing this synergy, the magnetometer achieves remarkable accuracy, sensitivity and dynamic range. The precise position registration enabled by the CMOS sensor further enhances the device's utility in capturing complex magnetic field patterns, allowing for 2D field mapping. In this work, the optimization of the probe's per-formance is tailored for applications related to the characterization of insertion devices in light sources, including undulators.

Abstract: Due to their rapid evolution and their impact on health care, be-ta-lactamases, protein degrading beta-lactam antibiotics, are used as generic model of protein evolution. Therefore, we investigated the mutation effects in two distant beta-lactamases TEM-1 and CTX-M-15. Interestingly we found a site with a complex pattern of genetic interactions. Mutation G251W in TEM-1 is in-activating the protein’s function just as the reciprocal mutation W251G in CTXM-15. Phylogenetic analysis revealed that mutation G has been entrenched in TEM-1 background: while rarely observed throughout the phylogeny it is es-sential in TEM-1. Using a rescue experiment in TEM-1 G251W mutant, we could identify sites that alleviates the deviation from G to W. While few of these muta-tions could potentially involve local interactions, most of them were found on distant residues in the 3D structure. Many well-known mutations having an impact on protein stability, such as M182T, were recovered. Our results there-fore suggest that entrenchment of an amino acid may rely on diffuse interactions among multiple sites with a major impact on protein stability.

Corona viruses hijack human enzymes to assembly sugar coat on Spike glycoproteins. The mechanism that human antibodies may uncover the antigenic viral peptide epitopes hidden by sugar coat are unknown. In this study, we analyzed recombinant SARS-CoV-2 Spike protein secreted from BTI-Tn-5B1-4 cells, by trypsin and chymotrypsin digestion followed by mass spectrometry analysis. We acquired MS/MS spectrums for glycopeptides of all 22 predicted N-glycosylated sites. We further analyzed the surface accessibility of Spike proteins according to Cryo-EM and homolog-modeled structures, and available antibodies that bind to SARS-CoV-1. The results showed that all 22 N-glycosylated sites of SARS-CoV-2 are modified by high-mannose type of N-glycans. MS/MS fragmentation clearly established the glycopeptide identities. Electron densities of glycans cover most of the Spike receptor binding domain of SARS-CoV-2, except YQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQ, similar to a region FSPDGKPCTPPALNCYWPLNDYGFYTTTGIGYQ in SARS-CoV-1. Other surface-exposed domains included those located on Central Helix, between amino acids 967 and 1016 of SARS-CoV-1, and 985 to 1034 of SARS-CoV-2 Spike protein. As the majority of antibody paratopes bind to peptide portion with or without sugar modification, we propose a snake-catcher model that a minimal length of peptide is first clamped by a paratope, and the binding is either strengthened by sugars close to peptide, or not interfered by sugar modification.

Tick-borne encephalitis virus (TBEV) is a pathogenic, enveloped, positive-stranded RNA virus in the family Flaviviridae. Structural studies of flavivirus virions have primarily focused on mosquito-borne species with only one cryo-electron microscopy (cryo-EM) structure of a tick-borne species published. Here, we present a 3.3 Å cryo-EM structure of the TBEV virion of the Kuutsalo-14 isolate, confirming the overall organisation of the virus. We observe conformational switching of the peripheral and transmembrane helices of M protein, which can explain the quasi-equivalent packing of the viral proteins and highlights their importance in stabilizing the membrane protein arrangement in the virion. The residues responsible for the M protein inter-actions are highly conserved in TBEV but not in the structurally studied Hypr strain, nor in mosquito-borne flaviviruses. These interactions may compensate for the lower number of hydrogen bonds between E proteins in TBEV compared to the mosquito-borne flaviviruses. The structure reveals two lipids bound in the E protein, which are important for virus assembly. The lipid pockets are comparable to those recently described in mosquito-borne Zika, Spondweni, Dengue, and Usutu viruses. Our results thus advance the understanding of tick-borne flavivirus architecture and virion-stabilising interactions.

The goal of the current study was to discover a novel potential probiotic strain of Lactobacillus spp. with anti-Escherichia coli activity from locally produced yogurt in Tongi, Gazipur, Bangladesh, compare its antagonistic activity with a commercial probiotic mixture of several strains, and approve a novel method for confirming the viability and relative abundance of the microbial community in a probiotic mixer. We carried out 16S sequencing, 16S metagenomics, Transmission Electron Microscopy (TEM) analysis, and other in vitro laboratory experiments to reach this objective. The strain TY-11 was identified as Lactobacillus delbrueckii subsp. indicus (16S sequence accession number OQ652026). It was gram-positive, anaerobic, lactose fermenting, and round-ended rod that typically measured 0.7 to 1.3 µm by 2.2 to 9 µm. In addition to having seven probiotic characteristics, it also showed an antagonistic impact on six different pathogens, but what's more noteworthy is that E. coli was the pathogen it inhibited most strongly (inhibition zone diameter was 18.88±0.18 mm). The most important and ground-breaking finding of this work was determining the probiotic features of a new probiotic strain, TY-11, whose antibacterial activity was virtually as effective as that of the probiotic combination with three different strains. Furthermore, the results of 16S high throughput sequencing and the conventional plate count method demonstrated a strong correlation (0.999) at the genus level, indicating that the use of both of these approaches in combination may be a practical way to assess the relative abundance of the microbial community and their viability in commercially available probiotic blends.

Micro-nanoparticles' characteristics in geothermal fluids can be applied to detect of deep hidden geothermal resources. Observations using a nanoparticle tracking analyzer (NTA) indicated that the karst geothermal water collected in the central area of Shandong Province (Jinan and Zibo) contains many natural micro-nanoparticles with sizes primarily ranging between 100 nm and 5 μm. The micro-nanoparticles’ type, shape, crystal form, and chemical composition in the samples were analyzed using transmission electron microscopy (TEM). TEM images and energy dispersive spectroscopy showed that the micro-nanoparticles in geothermal water samples were mostly amorphous, irregular, or nearly spherical, with rough edges. The micro-nanoparticles were mainly carbonates, sulfates, and chlorine-containing Fe, Ca, Na, and Mg. The characteristics of the particles can reveal the properties of the deep, hot reservoirs and aquifers from where they originate. Therefore, we believe that natural micro-nanoparticles can be essential to detecting and studying deep, hidden geothermal resources, which is a novel approach to exploring deep, hidden geothermal resources.

There is a fast-growing interest in the use of selective laser melting (SLM) for metal/alloy additive manufacturing. Our current knowledge of SLM printed 316 stainless steel (SS316) is limited and sometimes appears sporadic, presumably due to the complex interdependent effects of a large number of process variables of the SLM processing. This is reflected from the discrepant findings in the crystallographic textures and microstructures in this investigation with those reported in the literatures, which also vary itself. The as-printed material is macroscopically asymmetric in both the structures and crystallographic textures. The <101> and <111> crystallographic directions align parallel with the SLM scanning direction (SD) and build direction (BD), respectively. Like-wise, some characteristic low angle boundary features are reported crystallographic, while this investigation unequivocally proves them non-crystallographic since they always maintain an identical alignment with the SLM laser scanning direction irrespective of the matrix material’s crystal orientation. There is also 500±200 nm columnar or cellular features, depending on the cross-section, generally found all over the sample. These columnar or cellular features are formed with walls made of dense packing of dislocations entangled with Mn, Si and O enriched amorphous inclusions. They remain stable after the ASM solution treatments at 1050 °C temperatures, and therefore, are capable of hindering boundary migration events of recrystallization and grain growth. Thus, the nanoscale structures can be retained at high temperatures. Large 2-4 µm inclusions form during the solution treatment, within which the chemical and phase distribution are heterogeneous.

Metal-organic frameworks (MOFs), composed of metal nodes and inorganic linkers are promising for a wide range of applications due to their unique periodic frameworks that can be flexibly tuned. Understanding of the structure-activity relationships can facilitate the development and application of MOFs nanomaterials. Transmission electron microscopy (TEM) is a one-of-a-kind technique capable of characterizing MOFs microstructures at the atomic scale. Besides, in-situ TEM set-ups make it possible to direct visualize the microstructural evolution of MOFs in real time, which enables monitoring transitions of MOFs during growth as well as under working conditions. However, MOFs are so sensitive to the high-energy electron beam which makes TEM-based characterizing methods challenging. In this review, we first introduce the main damage mechanisms for MOFs under electron beam irradiation and two strategies to minimize the damages: low-dose TEM and cryo-TEM. Then, we discuss the techniques combined with the two strategies to analyze the MOFs’ microstructure, including three-dimensional electron diffraction, direct detection electron counting camera, and iDPC-STEM. Groundbreaking milestones and research advances with respect to static characterization of MOFs structures are highlighted. In-situ TEM studies are exemplary reviewed to provide insights into MOFs dynamics induced by various stimuli. Additionally, perspectives on promising techniques that may further facilitate the MOFs study using TEM are analyzed.

: Poultry products in Zambia form an integral part of the human diet in many households, as they are cheap and easy to produce. The burden of poultry diseases has, however, remained a major challenge. Growing consumer demand for poultry products in Zambia has resulted in non-prudent antimicrobial use on farms, intending to prevent and treat poultry diseases for growth optimization and maximising profits. This cross-sectional study aimed to identify the different types of bacteria causing diseases in chickens in Lusaka and to detect the extended-spectrum-lactamase (ESBL)-encoding genes. We collected 215 samples from 91 diseased chickens at three post-mortem facilities and screened them for gram-negative bacteria. Of these samples, 103 tested positive for various clinically relevant Enterobacteriaceae, including Enterobacter (43/103, 41.7%), Escherichia coli (20/103, 19.4%), Salmonella (10/103, 9.7%), and Shigella (8/103, 7.8%). Other isolated bacteria included Yersinia, Morganella, Proteus, and Klebsiella, which accounted for 21.4%. E. coli, Enterobacter, Salmonella, and Shigella were subjected to antimicrobial susceptibility testing. The results revealed that E. coli, Enterobacter, and shigella were highly resistant to tetracycline, ampicillin, amoxicillin, and trimethoprim-sulfamethoxazole, while Salmonella showed complete susceptibility to all antibiotics. The observed resistance patterns correlated with antimicrobial usage estimated from sales data from a large-scale wholesale and retail company. Six (6/14, 42.9%) E. coli isolates tested positive for bla_CTX-M, whilst eight (8/14, 57.1%) Enterobacter samples tested positive for bla_TEM. Interestingly, four (4/6, 66.7%) of the E. coli isolates carrying bla_CTX-M-positive strains were also positive for bla_TEM. Sanger sequencing of the PCR products revealed that five (5/6,83.3%) of the abovementioned isolates possessed the bla_CTX-M-15 allele. The results suggest the presence of potentially pathogenic ESBL-producing Enterobacteriaceae in poultry, threatening public health. Keywords:. Enterobacteriaceae, bla_CTX-M, bla_TEM

Cryo electron microscopy (cryo-EM) instrumentation allows obtaining 3D reconstruction of the structure of biomolecular complexes in vitro (purified complexes studied by single particle analysis) and in situ (complexes studied in cells by cryo electron tomography). Standard cryo-EM approaches allow high-resolution reconstruction of only a few conformational states of a molecular complex, as they rely on data classification into a given number of classes to increase the resolution of the reconstruction from a few, most populated classes and discard all other classes. Such discrete-classification approaches result in a partial picture of the full conformational variability of the complex, due to continuous conformational transitions with many, uncountable intermediate states. In this article, we present the software with user-friendly graphical interface for running two recently introduced methods, namely MDSPACE and MDTOMO, to obtain continuous conformational landscapes of biomolecules by analyzing in vitro and in situ cryo-EM data (single particle images and subtomograms) based on molecular dynamics simulations of an available atomic model of one of the conformations. The MDSPACE and MDTOMO software is part of the open-source ContinuousFlex package, which can be run as a plugin of the Scipion software package, broadly used in the cryo-EM field.

The development of compartmentalized neuron culture systems has been invaluable in the study of neuroinvasive viruses, including the alpha herpesviruses Herpes Simplex Virus 1 (HSV-1) and Pseudorabies Virus (PRV). This chapter provides updated protocols for assembling and culturing rodent embryonic superior cervical ganglion (SCG) and dorsal root ganglion (DRG) neurons in Campenot trichamber cultures. In addition, we provide several illustrative examples of the types of experiments that are enabled by Campenot cultures: 1. Using fluorescence microscopy to investigate axonal outgrowth/extension through the chambers, and alpha herpesvirus infection, intracellular trafficking, and cell-cell spread via axons. 2. Using correlative fluorescence microscopy and cryo electron tomography to investigate the ultrastructure of virus particles trafficking in axons.

The acylation of unsymmetrical N-benzylbispidinols in aromatic solvents without external base led to formation of supramolecular gels, which possess different thickness and stability depending on the substituents in para-positions of benzylic group and nature of acylating agent as well as on the nature of the solvent used. Structural features of the native gels as well as of their dried forms were studied by complementary techniques including FT IR- and ATR-spectroscopy, AFM, TEM, SEM, SAXS. Structures of the key crystalline compounds were established by X-ray diffraction. Analysis of obtained data allowed speculating on the crucial structural and condition factors that governed the gel formation. The most important factors were: (i) absence of base, either external or internal; (ii) presence of HCl; (iii) presence of carbonyl and hydroxyl groups to allow hydrogen bonding; (iv) presence of two (hetero)aromatic rings at both sides of the molecule. The hydrogen bonding involving amide carbonyl, hydroxyl at 9th position and, very probably, ammonium N-H+ and Cl- anion appear to be responsible for the formation of infinite molecular chains required for the first step of gel formation. Subsequent lateral cooperation of molecular chains into fibers occured, presumably, due to the aromatic pi-pi-stacking interactions. sc-CO2 drying of the gels gave rise to aerogels morphology different from that of air dried samples.

Due to the ongoing global warming on the Earth, permafrost degradation has been extensively taking place, which poses a substantial threat to civil and industrial facilities and infrastructure elements, as well as to the utilization of natural resources in the Arctic and high-latitude regions. In order to prevent the negative consequences of permafrost thawing under the foundations of constructions, various geophysical techniques for monitoring permafrost have been proposed and applied so far: temperature, electrical, seismic and many others. At the same time, non-stationary electromagnetic methods seem to have significant potential and a number of important practical advantages. We propose a cross-borehole exploration system for the highest localization of target objects in the cryolithozone. A novel mathematical apparatus for three-dimensional modeling of transient electromagnetic (TEM) signals by the vector finite element method has been developed. The original combination of the latter, the Sumudu integral transform and artificial neural networks makes it possible to examine spatially heterogeneous objects of the cryolithozone with a high contrast of geoelectric parameters, significantly reducing computational costs. We have studied the sensitivity of the TEM signals to the boundary between thawed and frozen rocks depending on the inter-borehole distance and spatial orientations of the transmitters and receivers. Numerical simulation results of the TEM monitoring of industrial facilities located on permafrost are considered. The formation of a talik has been shown to significantly manifest itself in the measured electromagnetic responses, which enables timely prevention of industrial disasters and environmental catastrophes.

A homogenization of billets from Al-Cu-Li-Mg-Sc-Zr alloys should be accomplished at high annealing temperatures exceeding 500 °C. This type of aluminum alloy is susceptible to the depletion of surface layers from Li. Therefore, choosing a suitable homogenization temperature and duration is a crucial step in assuring a homogeneous distribution of alloying elements and optimal exploitation of the potential of the alloy. In-situ heating in an electron microscope was performed on a twin-roll cast Al-Cu-Li-Mg-Sc-Zr alloy to understand the peculiarities of the homogenization process. Four types of primary phase particles rich in Cu, Li, Mg, and Fe were identified in the as-cast material. They appear as coarse particles at the boundaries of eutectic cells. Their partial dissolution occurs at temperatures above 450 °C. They are almost fully dissolved at 550 °C, except for complex phases containing Fe and Cu. Small dimensions of eutectic cells in the range of 10 µm assure a homogeneous distribution of main alloying elements within the matrix after 20 min of annealing. Direct comparison with the same material prepared by mold-casting indicates that such short annealing times result in the dissolution of main primary phase particles but do not assure a homogeneous distribution of alloying elements in the whole volume of the specimen.

Macromolecular assemblies, such as protein complexes, undergo continuous structural dynamics, including global reconfigurations critical for their function. Two fast analytical methods are widely used to study these global dynamics, namely elastic network model normal mode analysis and principal component analysis of ensembles of structures. These approaches have found wide use in various computational studies, driving the development of complex pipelines in several software packages. One common one has been conformational sampling through hybrid simulations incorporating all-atom molecular dynamics and global modes of motion. However, wide functionality is only available for experienced programmers with limited capabilities for other users. We have, therefore, integrated one popular and extensively developed software for such analyses, the ProDy Python application programming interface, into the Scipion workflow engine. This enables a wider range of users to access a complete range of macromolecular dynamics pipelines beyond the core functionalities available in its command-line applications and the normal mode wizard in VMD. The new protocols and pipelines can be further expanded and integrated into larger workflows together with other software packages for cryo-electron microscopy image analysis and molecular simulations. We present the resulting plugin, Scipion-EM-ProDy, in detail, highlighting the rich functionality made available by its development.

The Golgi apparatus (GA) dysfunctions in Parkinson’s Disease (PD), neurodevelopmental disorders (NDDs), cancer, and organelle structural biology (OSB) can provide insights into therapeutic targets, gene therapy, and drug design. Primary defects and fragmentation within the GA are implicated in a wide range of neurodegenerative diseases. GA defects typically result in mislocation of proteins, accumulation of undegraded proteins, and impaired glycosylation of proteins. Inhibition of vesicular trafficking by α-synuclein (aSyn) may affect the dopamine-producing neurons and neuromodulators. GA regulates apoptosis during pathological mechanisms of neurological diseases and could provide new avenues in treatments through translation research. PD patients bearing the hereditary E46K disease mutation manifest the clinical picture of parkinsonism. How do we provide high resolution nanoimages of the GA during disease to capture dysfunction? Could we visualize the aSyn traffic jam between vesicles in the organelles ER and GA? OSB is emerging as a field as more technology advances and is more accessible. Structural studies of the GA will advance the field of neurological disease forward with an in depth understanding of dysfunction, fragmentation, and defects. Discoveries of the GA in PD, NDDs, and cancer would break new ground and provide translational medicine data of these diseases. Future research could be visualizing high angle annular dark field-STEM (HAADF-STEM) tomograms, cryogenic electron tomography (cryo-ET), multiplex correlative light and electron microscopy (cryo-CLEM), nanobody-assisted tissue immunostaining for volumetric EM (NATIVE) and using soft X-ray tomography (SXT) and computational reconstruction of the GA.

Delivery systems of the biologically active substance such as proanthocyanidins (PCAN) produced through the electrospinning method were designed using a blend of poly(L-lactide-co-glycolide) and poly[(R,S)-3-hydroxybutyrate]. The research involved the structural and thermal characteristics of the developed electrospun three-dimensional fiber matrix, as well as hydrolytic degradation tests performed on them. Additionally, the release profile of PCAN from the electrospun nonwoven was determined using UV–VIS spectroscopy. Approximately 30% of PCAN was released from the tested electrospun nonwoven during the initial 15-20 days of incubation. The chemical structure of water-soluble oligomers that were formed after the hydrolytic degradation of the developed delivery system was identified through electrospray ionization mass spectrometry. Oligomers of lactic acid and OLAGA oligocopolyester, as well as oligo-3-hydroxybutyrate terminated with hydroxyl and carboxyl end groups, were recognized as degradation products released into the water during the incubation time. It was also demonstrated that variations in the degradation rate of individual mat components influenced the degradation pattern, and the number of formed oligomers. The obtained results suggest that the incorporation of proanthocyanidins into the system slowed down the hydrolytic degradation process of the poly(L-lactide-co-glycolide)/poly[(R,S)-3-hydroxybutyrate] three-dimensional fiber matrix. In addition, in vitro cytotoxicity and antimicrobial studies advocate the use of PCAN for biomedical applications with promising antimicrobial activity.

We compare the effect on amyloid fibril formation by two homologous proteins from the family of cystatins, human stefin B (stB) and cystatin C (cysC) in presence of 3 polyphenols: curcumin, resveratrol and quercetin and 2 non-phenolic anti-oxidants: vitamin C (VitC) and N-acetyl cystein (NAC). Some of the experimental data have already been presented, here we compare, further discuss and highlight the results. The amyloid fibril formation was followed by ThT fluorescence and transmission electron microscopy. Inhibitory effects on amyloid fibrillation reaction depended on anti-oxidant class and concentration. The fact that different effect of polyphenols was observed with the two cystatins; Cur acted inhibitory on stB but not on cysC fibril formation, could be explained if the 3 polyphenols would not bind to the same binding site in the fibrils core. Other differences are pointed out and discussed. Synergistic effects of VitC and chosen polyphenols on amyloid fibrilllation of human stB have been explored and are reported here for the first time.

(1) Background: Since fullerene C60 is insoluble in water and is toxic, it is not applicable in biomedicine and cosmetics. For this reason, the first derivative FD-C60 (fullerenol) was synthesized, which solved the problem of solubility in water and partially reduced toxicity. In order to maximally reduce toxicity and increase biomedical efficiency, the second derivative SD-C60 (3HFWC- Hyper-Harmonized-Hydroxylated Fullerene Water Complex), was created. (2) Methods: In order to obtain the most reliable data, several different methods are applicable in the characterization of FD-C60 and SD-C60 with water layers (NIR, FTIR, 13C-NMR,  1H-NMR, TGA/DTA, XRD, AFM/MFM etc.). (3) Results: FD-C60 as an individual structure is about 1.3 nm in size, while SD-C60 as an individual structure is 1030 nm in size. The aforementioned methods showed that SD-C60 contains water layers, which explains the difference between those two structures in the physicochemical properties, and their effects on biomolecules. (4) Conclusions: Based on the above methods and techniques, FD-C60 and SD-C60 are two different substances in terms of size, structure, and physicochemical properties; FD-C60 at 100C has endothermic characteristics and SD-C60 at 133C has exothermic characteristics, FD-C60 possesses water molecules only from humidity, while SD-C60 has water molecules from both humidity and extra water molecules in layers, the zeta potential of FD-C60 is -25.85 mV, while it is -43.29 mV for SD-C60.

In humans, age-associated degrading changes are observed in molecular and cellular processes underly the time-dependent decline in spatial navigation, time perception, cognitive and psy-chological abilities, and memory. Cross talk of biological, cognitive, and psychological clocks provides an integrative contribution to healthy and advanced aging. At the molecular level, ge-nome, proteome, and lipidome instability are widely recognized as the primary causal factors in aging. We narrow attention to the roles of protein aging linked to prevalent amino acids chirali-ty, enzymatic and spontaneous (non-enzymatic) post-translational modifications (PTMs SP), and non-equilibrium phase transitions. The homochirality of protein synthesis, resulting in the steady-state non-equilibrium condition of protein structure, makes them prone to multiple types of enzymatic and spontaneous PTMs, including racemization and isomerization. Spontaneous racemization leads to the loss of the balanced prevalent chirality. Advanced biological aging re-lated to irreversible PTMs SP has been associated with the nontrivial interplay between poor so-matic and mental health conditions. Through stress response systems (SRS), the environmental and psychological stressors contribute to the age-associated “collapse” of protein homochirality. The role of prevalent protein chirality and entropy of protein folding in biological aging is mainly overlooked. In a more generalized context, the time-dependent shift from enzymatic to the non-enzymatic transformation of biochirality might represent an important and yet un-der-appreciated hallmark of aging.

Ribosome assembly is one of the most fundamental processes in gene expression and has served as a playground to investigate the molecular mechanisms of how protein-RNA complexes (RNPs) assemble. The bacterial ribosome is composed of around 50 ribosomal proteins several of which are co-transcriptionally assembled on a ~4,500 nucleotides long pre-rRNA transcript that is further processed and modified during transcription, the entire process taking around 2 minutes in vivo and assisted by dozens of assembly factors. How this complex molecular process works so efficiently to produce an active ribosome has been investigated over decades and has resulted in the development of a plethora of novel approaches that can also be used to study the assembly of other RNPs. Here we review biochemical, structural and biophysical methods that have been developed and integrated to provide a detailed and quantitative understanding of this complex and intricate molecular process of assembly. We also discuss emerging cutting-edge approaches that could be used in the future to study how transcription, rRNA processing, cellular factors and the native cellular environment shape ribosome assembly and RNP assembly at large.

Many studies of hypervelocity impact craters have described the characteristics of quartz grains shock-metamorphosed at high pressures of > 10 GPa. In contrast, few studies have investigated shock metamorphism at lower shock pressures. In this study, we test the hypothesis that low-pressure shock metamorphism occurs in near-surface nuclear airbursts and that this process shares essential characteristics with crater-forming impact events. To investigate low-grade shock microstructures, we compared quartz grains from Meteor Crater, a 1.2-km-wide impact crater, to those from near-surface nuclear airbursts at the Alamogordo Bombing Range, New Mexico in 1945 and Kazakhstan in 1949/1953. This investigation utilized a comprehensive analytical suite of high-resolution techniques, including transmission electron microscopy (TEM) and electron backscatter diffraction (EBSD). Meteor Crater and the nuclear test sites all exhibit quartz grains with closely-spaced, sub-micron-wide fractures that appear to have formed at low shock pressures. Significantly, these micro-fractures are closely associated with Dauphiné twins and are filled with amorphous silica (glass), widely considered a classic indicator of shock metamorphism. Thus, this study confirms that glass-filled shock fractures in quartz form during near-surface nuclear airbursts, as well as crater-forming impact events, and by extension, it suggests that they may form in any near-surface cosmic airbursts in which the shockwave is coupled to Earth’s surface. The robust characterization of such events is crucial because of their potential catastrophic effects on the Earth’s environmental and biotic systems.

Many studies of hypervelocity impact craters have described the characteristics of quartz grains shock-metamorphosed at high pressures of >10 GPa, but in contrast, few studies have investigated shock metamorphism at lower shock pressures. In this study, we test the hypothesis that low-pressure shock metamorphism occurs in near-surface nuclear airbursts and that this process shares important characteristics with impact-cratering events. To investigate low-grade shock microstructures, we compared quartz grains from Meteor Crater, a 1.2-km-wide impact crater, to those from near-surface nuclear airbursts at the Alamogordo Bombing Range, New Mexico in 1945 and Kazakhstan in 1949/1953. This investigation utilized a comprehensive analytical suite of high-resolution techniques, including transmission electron microscopy (TEM) and electron backscatter diffraction (EBSD). Meteor Crater and the nuclear test sites all exhibit metamorphosed quartz grains with closely-spaced, sub-micron-wide fractures that appear to have formed at low shock pressures. Importantly, these micro-fractures are closely associated with Dauphiné twins and are filled with amorphous silica (glass), widely considered to be a classic indicator of shock metamorphism. Thus, this study confirms that glass-filled shock fractures in quartz form during near-surface nuclear airbursts, as well as crater-forming impact events, and by extension, it suggests they also may form in near-surface cosmic airbursts.

Background. Escalating global plastic production, expected to reach 34,000 million tons by 2050, poses a significant threat to human and environmental well-being, particularly in aquatic ecosystems. Microplastics (MP) and nanoplastics (NP), which originate from the degradation of plastics, are of concern due to their potential bioaccumulation and uptake of pollutants. This study addresses identification methods and focuses on insect meal, a raw material for aquaculture feed. Methods. By using different techniques, the study was able to detect MP and NP in insect meal samples. Chemical digestion with KOH at 60°C efficiently removed organic matter without affecting the synthetic polymer polyethylene (PE). Filtration, confocal Raman microscopy, SEM and TEM were used for comprehensive analysis and integrity tests on PE films were performed using Raman and FTIR spectroscopy. The results showed the presence of PE microplastic particles in the insect meal, which was confirmed by correlative Raman and SEM mapping on a positively charged surface. In addition, the increased resolution of the Raman microscope identified submicrometric PE NP (800 nm). Transmission electron microscopy (TEM) with energy dispersive X-ray spectroscopy (EDX) confirmed plastic-like structures in the insect meal, highlighting the presence of PE plastics characterized by irregular shapes and some agglomeration. The higher carbon concentration in the EDX analysis supported the plastic nature, which was also confirmed by Raman spectroscopy. Conclusions. The study provides a robust method for the detection of MP and NP in insect meal and provides valuable insight into the possible presence of plastics in insect-based aquafeeds. The combination of different analytical methods increases the reliability of the results and sets the stage for future investigations that could focus on the quantification of NP and the assessment of their potential environmental impact.

Engineering application of additively manufactured (AM) metallic materials is quite limited by their fatigue behaviors which are very inconsistent with that of conventionally wrought or cast ones. Here, based on advanced material characterization techniques, such as focused ion beam (FIB), scanning electron microscopy (SEM) and transmission electron microscopy (TEM), the mi-crostructures underneath fracture surfaces were thoroughly investigated in an AM aluminium (AlSi10Mg) alloy with horizontal and vertical building orientation endured very-high-cycle fa-tigue (VHCF) loading under stress ratios R = –1, 0 and 0.5. Two VHCF failure specimens A and B were representatively selected to further SEM examine and TEM samples preparation via FIB milling. Specimen A was horizontally printed and failed at R = –1; specimen B was vertically printed and failed at R = 0. TEM samples A1 and B1 were lifted from locations near the crack ini-tiation sites on the fracture surfaces of specimens A and B; The locations of TEM samples A2 and B2 kept away from the crack origin sites but still within the “fish-eye” region of crack steady growth. TEM observations show that there was no characteristic microstructure induced by VHCF in different oriented specimens and under various R values.