A correlated light and electron microscopy approach to study the subcellular localization of phosphorylated vimentin in human lung tissue.

Correlative microscopy is an important approach for bridging the resolution gap between fluorescence light and electron microscopy. Here, we describe a fast and simple method for correlative immunofluorescence and immunogold labeling on the same section to elucidate the localization of phosphorylated vimentin (P-Vim), a robust feature of pulmonary vascular remodeling in cells of human lung small arteries. The lung is a complex, soft and difficult tissue to prepare for transmission electron microscopy (TEM). Detailing the molecular composition of small pulmonary arteries (<500µm) would be of great significance for research and diagnostics. Using the classical methods of immunochemistry (either hydrophilic resin or thin cryosections), is difficult to locate small arteries for analysis by TEM. To address this problem and to observe the same structures by both light and electron microscopy, correlative microscopy is a reliable approach. Immunofluorescence enables us to know the distribution of P-Vim in cells but does not provide ultrastructural detail on its localization. Labeled structures selected by fluorescence microscope can be identified and further analyzed by TEM at high resolution. With our method, the morphology of the arteries is well preserved, enabling the localization of P-Vim inside pulmonary endothelial cells. By applying this approach, fluorescent signals can be directly correlated to the corresponding subcellular structures in areas of interest.

Exploring the neuroprotective role of artesunate in mouse models of anti-NMDAR encephalitis: insights from molecular mechanisms and transmission electron microscopy.

BACKGROUND: The pathway involving PTEN-induced putative kinase 1 (PINK1) and PARKIN plays a crucial role in mitophagy, a process activated by artesunate (ART). We propose that patients with anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis exhibit insufficient mitophagy, and ART enhances mitophagy via the PINK1/PARKIN pathway, thereby providing neuroprotection. METHODS: Adult female mice aged 8-10 weeks were selected to create a passive transfer model of anti-NMDAR encephalitis. We conducted behavioral tests on these mice within a set timeframe. Techniques such as immunohistochemistry, immunofluorescence, and western blotting were employed to assess markers including PINK1, PARKIN, LC3B, p62, caspase3, and cleaved caspase3. The TUNEL assay was utilized to detect neuronal apoptosis, while transmission electron microscopy (TEM) was used to examine mitochondrial autophagosomes. Primary hippocampal neurons were cultured, treated, and then analyzed through immunofluorescence for mtDNA, mtROS, TMRM. RESULTS: In comparison to the control group, mitophagy levels in the experimental group were not significantly altered, yet there was a notable increase in apoptotic neurons. Furthermore, markers indicative of mitochondrial leakage and damage were found to be elevated in the experimental group compared to the control group, but these markers showed improvement following ART treatment. ART was effective in activating the PINK1/PARKIN pathway, enhancing mitophagy, and diminishing neuronal apoptosis. Behavioral assessments revealed that ART ameliorated symptoms in mice with anti-NMDAR encephalitis in the passive transfer model (PTM). The knockdown of PINK1 led to a reduction in mitophagy levels, and subsequent ART intervention did not alleviate symptoms in the anti-NMDAR encephalitis PTM mice, indicating that ART's therapeutic efficacy is mediated through the activation of the PINK1/PARKIN pathway. CONCLUSIONS: At the onset of anti-NMDAR encephalitis, mitochondrial damage is observed; however, this damage is mitigated by the activation of mitophagy via the PINK1/PARKIN pathway. This regulatory feedback mechanism facilitates the removal of damaged mitochondria, prevents neuronal apoptosis, and consequently safeguards neural tissue. ART activates the PINK1/PARKIN pathway to enhance mitophagy, thereby exerting neuroprotective effects and may achieve therapeutic goals in treating anti-NMDAR encephalitis.

Thermosensitive and mucoadhesive gels containing solid lipid nanoparticles loaded with fluconazole and niosomes loaded with clindamycin for the treatment of periodontal diseases: a laboratory experiment.

BACKGROUND: Periodontal diseases may benefit more from topical treatments with nanoparticles rather than systemic treatments due to advantages such as higher stability and controlled release profile. This study investigated the preparation and characterization of thermosensitive gel formulations containing clindamycin-loaded niosomes and solid lipid nanoparticles (SLNs) loaded with fluconazole (FLZ), as well as their in vitro antibacterial and antifungal effects in the treatment of common microorganisms that cause periodontal diseases. METHODS: This study loaded niosomes and SLNs with clindamycin and FLZ, respectively, and assessed their loading efficiency, particle size, and zeta potential. The particles were characterized using a variety of methods such as differential scanning calorimetry (DSC), dynamic light scattering (DLS), and Transmission Electron Microscopy (TEM). Thermosensitive gels were formulated by combining these particles and their viscosity, gelation temperature, in-vitro release profile, as well as antibacterial and antifungal effects were evaluated. RESULTS: Both types of these nanoparticles were found to be spherical (TEM) with a mean particle size of 243.03 nm in niosomes and 171.97 nm in SLNs (DLS), and respective zeta potentials of -23.3 and -15. The loading rate was 98% in niosomes and 51% in SLNs. The release profiles of niosomal formulations were slower than those of the SLNs. Both formulations allowed the release of the drug by first-order kinetic. Additionally, the gel formulation presented a slower release of both drugs compared to niosomes and SLNs suspensions. CONCLUSION: Thermosensitive gels containing clindamycin-loaded niosomes and/or FLZ-SLNs were found to effectively fight the periodontitis-causing bacteria and fungi.

Castaways: the Leeward Antilles endemic spider genus Papiamenta (Araneae: Pholcidae).

Ninetinae is a group of small to tiny short-legged spiders largely restricted to arid habitats. Among daddy-long-legs spiders (Pholcidae) this is by far the least diverse subfamily but this may partly be a result of inadequate collecting, poor representation in collections or scientific neglect. We build on a large recent collection of the ninetine genus Papiamenta Huber, 2000 from the Leeward Antilles and use cytochrome oxidase 1 (COI ) sequences, extensive scanning electron microscopy data, transmission electron microscopy data and karyotyping to analyse this geographically isolated and poorly known island genus. COI sequences support the split between the two morphologically distinct species on Curaçao but genetic distances between these are surprisingly low (7.4-9.8%; mean 8.6%). The type species P. levii (Gertsch, 1982) may include more than one species but COI and morphology suggest conflicting clade limits. A third species, P. bonay Huber sp. nov. is newly described from Bonaire. Our data on sperm ultrastructure and karyology are puzzling as these suggest different phylogenetic affinities of Papiamenta to other genera. Males transfer sperm as individual sperm (cleistosperm), agreeing with the putative closest relatives as suggested by molecular data, the North American genera Pholcophora and Tolteca . The sex chromosome system (X 1 X 2 X 3 Y ) of P. levii , however, is as in the South American Ninetinae genera Gertschiola and Nerudia but different from the putative closest relatives. ZooBank: urn:lsid:zoobank.org:pub:7A6A2E84-3A61-4637-AF6F-0E31A9FA79A8.

Characterization of Amaranthus species: ability in nanoparticles fabrication and the antimicrobial activity against human pathogenic bacteria.

The present work aimed at differentiating five Amaranthus species from Saudi Arabia according to their morphology and the ability in nanoparticle formulation. Biogenic silver nanoparticles (AgNPs) were synthesized from leaf extracts of the five Amaranthus species and characterized by different techniques. Fourier-transform infrared spectroscopy (FT-IR) was used to identify the phyto-constituents of Amaranthus species. The nanoparticles (NPs) were characterized by UV-visible spectroscopy, dynamic light scattering (DLS), transmission electron microscopy (TEM), and energy-dispersive X-ray spectroscopy (EDX). The antibacterial activity of the synthesized NPs was tested against Staphylococcus aureus, E. coli, Klebsiella pneumoniae and Pseudomonas aeruginosa using the agar well diffusion method. Spherical NPs varying in size and functional groups from the five plant species were demonstrated by TEM, DLS and FTIR analysis, respectively. Variations in NPs characteristics could be related to the phytochemical composition of each Amaranthus species since they play a significant role in the reduction process. EDX confirmed the presence of Ag in plant fabricated AgNPs. Antibacterial activity varied among the species, possibly related to the NPs characteristics. Varied characteristics for the obtained AgNPs may reflect variations in the phytochemical composition type and concentration among Amaranthus species used for their fabrication.

How to apply the broad toolbox of correlative light and electron microscopy to address a specific biological question.

Correlative light and electron microscopy (CLEM) is an approach that combines the strength of multiple imaging techniques to obtain complementary information about a given specimen. The "toolbox" for CLEM is broad, making it sometimes difficult to choose an appropriate approach for a given biological question. In this chapter, we provide experimental details for three CLEM approaches that can help the interested reader in designing a personalized CLEM strategy for obtaining ultrastructural data by using transmission electron microscopy (TEM). First, we describe chemical fixation of cells grown on a solid support (broadest approach). Second, we apply high-pressure freezing/freeze substitution to describe cellular ultrastructure (cryo-immobilization approach). Third, we give a protocol for a ultrastructural labeling by immuno-electron microscopy (immuno-EM approach). In addition, we also describe how to overlay fluorescence and electron microscopy images, an approach that is applicable to each of the reported different CLEM strategies. Here we provide step-by step descriptions prior to discussing possible technical problems and variations of these three general schemes to suit different models or different biological questions. This chapter is written for electron microscopists that are new to CLEM and unsure how to begin. Therefore, our protocols are meant to provide basic information with further references that should help the reader get started with applying a tailored strategy for a specific CLEM experiment.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) culture and sample preparation for correlative light electron microscopy.

Correlative Light Electron Microscopy (CLEM) is a powerful technique to investigate the ultrastructure of specific cells and organelles at sub-cellular resolution. Transmission Electron Microscopy (TEM) is particularly useful to the field of virology, given the small size of the virion, which is below the limit of detection by light microscopy. Furthermore, viral infection results in the rearrangement of host organelles to form spatially defined compartments that facilitate the replication of viruses. With the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), there has been great interest to study the viral replication complex using CLEM. In this chapter we provide an exemplary workflow describing the safe preparation and processing of cells grown on coverslips and infected with SARS-CoV-2.

Some tips and tricks for a Correlative Light Electron Microscopy workflow using stable expression of fluorescent proteins.

Correlative Light Electron Microscopy (CLEM) encompasses a wide range of experimental approaches with different degrees of complexity and technical challenges where the attributes of both light and electron microscopy are combined in a single experiment. Although the biological question always determines what technology is the most appropriate, we generally set out to apply the simplest workflow possible. For 2D cell cultures expressing fluorescently tagged molecules, we report on a simple and very powerful CLEM approach by using gridded finder imaging dishes. We first determine the gross localization of the fluorescence using light microscopy and subsequently we retrace the origin/localization of the fluorescence by projecting it onto the ultrastructural reference space obtained by transmission electron microscopy (TEM). Here we describe this workflow and highlight some basic principles of the sample preparation for such a simple CLEM experiment. We will specifically focus on the steps following the resin embedding for TEM and the introduction of the sample in the electron microscope.

Investigating Epidermal Growth Factor Receptor Trafficking with High-Resolution Enzymatic Protein-Tagging and Transmission Electron Microscopy.

Enzymatic ascorbate peroxidase (APEX) tagging allows for high-resolution, three-dimensional protein distribution analyses in cells and tissues. This chapter describes the application of APEX-tagging to visualize the trafficking of the epidermal growth factor receptor (EGFR) during epidermal growth factor-mediated receptor activation. Here, we describe the preparation of cells, methods to validate the stimulation of the EGFR, and visualization of the APEX-resolved distribution of the EGFR in the transmission electron microscope.

Some Guiding Principles for a "Simple" Correlative Light Electron Microscopy Experiment.

In recent years, Correlative Multimodal Imaging (CMI) has become an "en vogue" technique and a bit of a buzzword. It entails combining information from different imaging modalities to extract more information from a sample that would otherwise not be possible from each individual technique. The best established CMI technology is correlative light and electron microscopy (CLEM), which applies light and electron microscopy on the exact same sample/structure. In general, it entails the detection of fluorescently tagged proteins or structures by light microscopy and subsequently their relative intracellular localization is determined with nanometer resolution using transmission electron microscopy (TEM). Here, we describe the different steps involved in a "simple" CLEM approach. We describe the overall workflow, instrumentation, and basic principles of sample preparation for a CLEM experiment exploiting stable expression of fluorescent proteins.

Novel Transmission Electron Microscopic Method to Quantify Protein Clustering on the Cell Surface.

The cell surface distribution patterns (clustering) of membrane proteins have been widely investigated in cell biology. Here we describe a novel transmission electron microscopic (TEM) protocol designed to improve the quality of information obtained about the protein distribution patterns detected. This novel method makes it possible to study the clustering of all transmembrane proteins on one half of the cytoplasmic membrane of a whole cell. To achieve better imaging, we combine various methods, including critical-point drying, fixation of gold beads with a carbon layer, and a newly developed chemical thinning method. In addition, in our image-processing algorithm, we implemented pair correlation and pair cross-correlation functions, providing more details and better quantitative accuracy in characterizing the size and numbers of possible protein clusters. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Sample preparation and transmission electron micrography Alternate Protocol: Direct cell labeling for transmission electron micrography Basic Protocol 2: Analysis of TEM images to detect immunogold-labeled proteins.

Photo-catalytic dye degradation potentials of Ag-Pd bimetallic nanoparticles synthesized from Vitex negundo.

Plant extracts are a great alternative to synthesizing nanoparticles of different metals and metal oxides. This green synthesis method has opened up numerous possibilities in various scientific domains. In present study, Leaf extract from Vitex negundo is a non-deciduous, long-lasting shrub from the Verbenaceae family is used as capping and reducing agents for the synthesis of silver and palladium nanoparticles. The characterization study UV-vis spectrophotometer analysis showed absorbance value around 320 nm which confirming that Ag-Pd nanoparticles have been successfully obtained. Further, SEM is used to investigate the morphology of Ag-Pd NPs, which revealing their spherical and rod-like configuration, aggregation, and the size of the particles are obtained between 50 and 100 nm. The successful synthesis of Ag-Pd NPs was further confirmed by the EDAX chart, which displayed the peak of Ag and Pd at bending energies between 0.5 and 1.5 keV. According to the quantitative study, Ag and Pd ions found about 5.24 and 13.28%, respectively. In addition, surface studies with TEM confirming that synthesized Ag-Pd NPs are predominates with spheres structure morphologies, with sizes averaging 11.20 nm and ranging from 10 to 20 nm. Further, Ag-Pd nanoparticles was applied as potential photocatalyst materials to degrade methylene blue dye and found about 85% of the degradation efficiency within 150 min of the sunlight exposure thus could be used as catalyst to removal of hazardous organic dye molecules.

An in vivo appraisal of Punica granatum peel extract's ultrastructural effect on cystic echinococcosis in mice.

In the past decade, interest has significantly increased regarding the medicinal and nutritional benefits of pomegranate (Punica granatum) peel. This study examined the effects of using pomegranate peel extract (PGE) alone and in combination with albendazole (ABZ) on ultrastructural and immunological changes in cystic echinococcosis in laboratory-infected mice. Results revealed that the smallest hydatid cyst size and weight (0.48 ± 0.47mm, 0.17 ± 0.18 gm) with the highest drug efficacy (56.2%) was detected in the PGE + ABZ group, which also exhibited marked histopathological improvement. Ultrastructural changes recorded by transmission electron microscopy including fragmentation of the nucleus, glycogen depletion, and multiple lysosomes in vacuolated cytoplasm were more often observed in PGE + ABZ group. IFN-γ levels were significantly increased in the group treated with ABZ, with a notable reduction following PGE treatment, whether administered alone or in combination with ABZ. Thus, PGE enhanced the therapeutic efficiency of ABZ, with improvement in histopathological and ultrastructural changes.

Histology, histochemistry and fine structure of the lacrimal gland in the one-humped camel (Camelus dromedarius).

Our research aimed to provide complete histological, histochemical and ultrastructural features of the lacrimal gland of the one-humped camel (Camelus dromedarius) as well as novel insights into its adaptability to the Egyptian desert. Our study was applied to 20 fresh lacrimal glands collected from 10 camels instantly after their slaughtering. The results revealed that the gland was a compound tubulo-acinar gland, and its acini were enclosed by a thick connective tissue capsule that was very rich in elastic and collagen fibres. The gland acini had irregular lumens and were composed of conical to pyramidal cells. The nuclei of secretory cells were found in the basal part, and the cytoplasm was eosinophilic and granular. The glandular tissue consisted of serous and mucous acini and seromucous secretory cells. Histochemically, there was a significant amount of neutral mucopolysaccharides in the acini in which mucous cells had a significant periodic acid-Schiff (PAS)-positive reaction, whereas seromucous cells had a mild PAS-positive reaction. Ultrastructurally, the lacrimal cells had numerous secretory vesicles with contents of moderately to highly electron-dense cytoplasm. The nuclear envelope consisted of two prominent membranes surrounding the peri-nuclear cisterna. The acinar cells had numerous electron-lucent and moderately electron-dense secretory granules, mainly situated on the apical surface, and secreted their contents into the lumen. The luminal surface of the mucous secretory cells represents the remains of secretory granules discharged by the merocrine mechanism. In conclusion, the mucous secretion is believed to aid in the washing and moistening of the eyeball, particularly in dry, hot and dusty environments.

Effect of micro- and nanoplastic particles on human macrophages.

Micro- and nanoplastics (MNPs) are ubiquitous in the environment, resulting in the uptake of MNPs by a variety of organisms, including humans, leading to particle-cell interaction. Human macrophages derived from THP-1 cell lines take up Polystyrene (PS), a widespread plastic. The question therefore arises whether primary human macrophages also take up PS micro- and nanobeads (MNBs) and how they react to this stimulation. Major aim of this study is to visualize this uptake and to validate the isolation of macrophages from peripheral blood mononuclear cells (PBMCs) to assess the impact of MNPs on human macrophages. Uptake of macrophages from THP-1 cell lines and PBMCs was examined by transmission electron microscopy (TEM), scanning electron microscopy and live cell imaging. In addition, the reaction of the macrophages was analyzed in terms of metabolic activity, cytotoxicity, production of reactive oxygen species (ROS) and macrophage polarization. This study is the first to visualize PS MNBs in primary human cells using TEM and live cell imaging. Metabolic activity was size- and concentration-dependent, necrosis and ROS were increased. The methods demonstrated in this study outline an approach to assess the influence of MNP exposure on human macrophages and help investigating the consequences of worldwide plastic pollution.

Single Extracellular Vesicle Imaging and Computational Analysis Identifies Inherent Architectural Heterogeneity.

Evaluating the heterogeneity of extracellular vesicles (EVs) is crucial for unraveling their complex actions and biodistribution. Here, we identify consistent architectural heterogeneity of EVs using cryogenic transmission electron microscopy (cryo-TEM), which has an inherent ability to image biological samples without harsh labeling methods while preserving their native conformation. Imaging EVs isolated using different methodologies from distinct sources, such as cancer cells, normal cells, immortalized cells, and body fluids, we identify a structural atlas of their dominantly consistent shapes. We identify EV architectural attributes by utilizing a segmentation neural network model. In total, 7,576 individual EVs were imaged and quantified by our computational pipeline. Across all 7,576 independent EVs, the average eccentricity was 0.5366 ± 0.2, and the average equivalent diameter was 132.43 ± 67 nm. The architectural heterogeneity was consistent across all sources of EVs, independent of purification techniques, and compromised of single spherical, rod-like or tubular, and double shapes. This study will serve as a reference foundation for high-resolution images of EVs and offer insights into their potential biological impact.

Exploring the antibiofilm and toxicity of tin oxide nanoparticles: Insights from in vitro and in vivo investigations.

BACKGROUND INFORMATION: The advancement of biological-mediated nanoscience towards higher levels and novel benchmarks is readily apparent, owing to the use of non-toxic synthesis processes and the incorporation of various additional benefits. This study aimed to synthesize stable tin oxide nanoparticles (SnO2-NPs) using S. rhizophila as a mediator. METHODS: The nanoparticles that were created by biosynthesis was examined using several analytical techniques, including Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM), X-ray diffraction (XRD), UV-visible (UV-vis) spectroscopy, and energy dispersive X-ray spectroscopy (EDS). RESULTS: The results obtained from the characterization techniques suggest that S. rhizophila effectively catalyzed the reduction of SnCl2 to SnO2-NPs duration of 90 min at ambient temperature with the ƛmax of 328 nm. The size of the nano crystallite formations was measured to be 23 nm. The present study investigates nanoscale applications' antibacterial efficacy against four bacterial strains, including Klebsiella Sp, Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. The observed zone of inhibition for the nanoparticles (NPs) varied from 10 to 25 mm. The research findings demonstrate that the nanoparticles (NPs) are effective as antibacterial, phytotoxic, and cytotoxic agents.

Exosomes from uterine fluid promote capacitation of human sperm.

Background: Extracellular vesicles (EVs) are membrane-bound vesicles containing various proteins, lipids, and nucleic acids. EVs are found in many body fluids, such as blood and urine. The release of EVs can facilitate intercellular communication through fusion with the plasma membrane or endocytosis into the recipient cell or through internalization of the contents. Recent studies have reported that EVs isolated from human endometrial epithelial cells (EECs) promote sperm fertilization ability. EVs from uterine flushing fluid more closely resemble the physiological condition of the uterus. However, it is unclear whether EVs derived directly from uterine flushing fluid have the same effect on sperm. This study aimed to research the effect of EVs from uterine flushing fluid on sperm. Methods: EVs were isolated from the uterine flushing fluid. The presence of EVs was confirmed by nanoparticle tracking analysis (NTA), Western blot, and transmission electron microscopy (TEM). EVs were incubated with human sperm for 2 h and 4 h. The effects of EVs on sperm were evaluated by analyzing acrosome reaction, sperm motility, and reactive oxygen species (ROS). Results: The EVs fractions isolated from the uterine fluid were observed in cup-shaped vesicles of different sizes by TEM. All isolated vesicles contained similar numbers of vesicles in the expected size range (30-200 nm) by NTA. CD9 and CD63 were detected in EVs by western blot. Comparing the motility of the two groups incubated sperm motility significantly differed at 4 h. The acrosome reactions were promoted by incubating with EVs significantly. ROS were increased in sperm incubated with EVs. Conclusion: Our results showed EVs present in the uterine fluid. Acrosome reactions and ROS levels increased in human sperm incubated with EVs. EVs from uterine fluid can promote the capacitation of human sperm. The increased capacitation after sperm interaction with EVs suggests a possible physiological effect during the transit of the uterus.

Evaluating the antifungal effectiveness, leaching characteristics, flexural strength, and impact strength of polymethyl methacrylate added with small-scale silver nanoparticles - An in vitro study.

AIM: (1) To assess the release of stable silver nanoparticles (AgNPs) of small scale dimension added to heat polymerized polymethyl methacrylate (PMMA) in 6 months. (2) Assessing the influence of incorporating minimal concentrations of stable AgNPs with nanoscale dimensions into heat polymerized PMMA over a 6 month period on its antifungal efficacy (AF), flexural strength (FS), and impact strength (IS). SETTINGS AND DESIGN: Incorporating nanoparticles with a very small scale may have minimal impact on mechanical properties due to their diminutive size. However, the influence of these small scaled nanoparticles on antimicrobial efficacy and potential escalation in toxicity to host cells through leaching remains unexplored. AgNPs were prepared using an Ultrasonic Probe sonicator and the addition of ammonia to obtain stabilized AgNPs (< 0.01 nm) of small scale dimension. The characterization of these AgNPs involved ultraviolet visible spectroscopy, X ray diffraction, Zetasizer, and transmission electron microscopy with energy dispersive spectroscopy (TEM). MATERIALS AND METHODS: The prepared AgNPs were then added in various percentages by weight (0%-0.5%) to fabricate 252 modified PMMA samples of sizes 10 mm × 3 mm (AF, n = 108), 65 mm × 10 mm × 3 mm (FS, n = 72), and 65 mm × 10 mm × 2.5 mm (IS, n = 72) as per ADA specification no. 12. These samples underwent testing for leaching out of AgNPs and efficacy against Candida albicans for 6 months. The effect on FS and IS was evaluated using the three point bending test and Charpy's Impact Tester, respectively. STATISTICAL ANALYSIS USED: Intergroup comparison of CFU between various concentrations of AgNP was done using the Kruskal-Wallis ANOVA test succeeded by Mann-Whitney test for pair wise comparisons. Difference in CFU of various concentrations over 6 months was seen using one way ANOVA test. Intergroup comparison of FS and IS was performed using a one way ANOVA test, followed by a post hoc Tukey's test for pair wise comparisons. RESULTS: Repeated tests showed no leaching out of AgNPs from the denture base resin into the storage medium. All concentrations of AgNPs incorporated in resin showed inhibition of Candida growth. Intergroup comparison of FS and IS revealed highly statistically significant differences (F = 15.076, P < 0.01 and F = 28.266, P < 0.01) between the groups showing a reduction in strength. CONCLUSION: The AgNPs of small scale dimension incorporated into the denture base resin imparted a strong antifungal effectiveness against C. albicans, which did not decline during the study period and did not cause any release of nanoparticles. 0.5% showed the best antifungal efficacy. This may prove to be a viable and highly effective treatment for the prevention of Candida associated denture stomatitis. However, the inclusion of these particles resulted in a decrease in both FS and IS, and this reduction was directly proportional to the percentage of added AgNPs, with 0.5% demonstrating the least IS and FS.

Redescription and molecular characterization of Loxocephalus luridus Eberhard, 1862 based on Czech populations: Implications for order Loxocephalida Jankowski, 1980.

The phylogenetic and taxonomic affinities of lineages currently assigned to the non-monophyletic ciliate order Loxocephalida Jankowski (1980) within subclass Scuticociliatia Small (1967) remain unresolved. In the current study, we redescribe the morphology of the type species, Loxocephalus luridus Eberhard (1862) based on two Czech populations and include the first scanning and transmission electron microscopy images of the species. We provide the first 18S rRNA gene sequences for L. luridus and consider its phylogenetic position. Our results support the separation of Dexiotricha from Loxocephalus; however, the former genus is recovered as non-monophyletic. The monophyly of genus Dexiotricha and that of Loxocephalus + Dexiotricha is rejected. Loxocephalus luridus, together with Dexiotricha species, nests within a fully supported clade with Conchophthirus species, long presumed to belong to the Pleuronematida. Haptophrya is recovered as sister to this clade. The monophyly of the Astomatia Schewiakoff (1896) including Haptophrya is rejected. No clear morphologic synapomorphy is identified for the fully supported clade consisting of Haptophrya, Dexiotricha, Loxocephalus, and Conchophthirus.