Human induced pluripotent stem cells are resistant to human cytomegalovirus infection primarily at the attachment level due to the reduced expression of cell-surface heparan sulfate.

Cytomegalovirus (CMV), a type of herpes virus, is the predominant cause of congenital anomalies due to intrauterine infections in humans. Adverse outcomes related to intrauterine infections with human cytomegalovirus (HCMV) vary widely, depending on factors such as fetal infection timing, infection route, and viral virulence. The precise mechanism underlying HCMV susceptibility remains unclear. In this study, we compared the susceptibility of neonatal human dermal fibroblast cells (NHDFCs) and human induced pluripotent stem cells (hiPSCs) derived from NHDFCs, which are genetically identical to HCMV, using immunostaining, microarray, in situ hybridization, quantitative PCR, and scanning electron microscopy. These cells were previously used to compare CMV susceptibility, but the underlying mechanisms were not fully elucidated. HCMV susceptibility of hiPSCs was significantly lower in the earliest phase. No shared gene ontologies were observed immediately post-infection between the two cell types using microarray analysis. Early-stage expression of HCMV antigens and the HCMV genome was minimal in immunostaining and in in situ hybridization in hiPSCs. This strongly suggests that HCMV does not readily bind to hiPSC surfaces. Scanning electron microscopy performed using the NanoSuit method confirmed the scarcity of HCMV particles on hiPSC surfaces. The zeta potential and charge mapping of the charged surface in NHDFCs and hiPSCs exhibited minimal differences when assessed using zeta potential analyzer and scanning ion conductance microscopy; however, the expression of heparan sulfate (HS) was significantly lower in hiPSCs compared with that in NHDFCs. Thus, HS expression could be a primary determinant of HCMV resistance in hiPSCs at the attachment level. IMPORTANCE: Numerous factors such as attachment, virus particle entry, transcription, and virus particle egress can affect viral susceptibility. Since 1984, pluripotent cells are known to be CMV resistant; however, the exact mechanism underlying this resistance remains elusive. Some researchers suggest inhibition in the initial phase of HCMV binding, while others have suggested the possibility of a sufficient amount of HCMV entering the cells to establish latency. This study demonstrates that HCMV particles rarely attach to the surfaces of hiPSCs. This is not due to limitations in the electrostatic interactions between the surface of hiPSCs and HCMV particles, but due to HS expression. Therefore, HS expression should be recognized as a key factor in determining the susceptibility of HCMV in congenital infection in vitro and in vivo. In the future, drugs targeting HS may become crucial for the treatment of congenital CMV infections. Thus, further research in this area is warranted.

Identifying Active Progeny Virus Particles in Formalin-Fixed, Paraffin-Embedded Sections Using Correlative Light and Scanning Electron Microscopy.

Immunohistochemical analysis of formalin-fixed paraffin-embedded (FFPE) tissue blocks is routinely used to identify virus-infected cells. However, detecting virus particles in FFPE sections using light microscopy is difficult because of the light diffraction resolution limitations of an optical microscope. In this study, light microscopy and field emission scanning electron microscopy were performed to observe 3-dimensional virus particles in FFPE sections in a nondestructive manner using NanoSuit or osmium conductive treatment methods. The virus particles in FFPE sections were immunostained with specific antibodies against the surface antigens of the viral particles and stained with 3,3'-diaminobenzidine. A metal solution (0.2% gold chloride or 2% osmium tetroxide) was applied to enhance the 3,3'-diaminobenzidine-stained area. This procedure is nondestructive for FFPE sections and is a simpler method than transmission electron microscopy. To validate the applicability of this technique, we performed 3-dimensional imaging of the virus particles of different sizes, such as human papillomavirus, cytomegalovirus, and varicella-zoster virus. Furthermore, ultrathin sections from the FFPE sections that were observed to harbor viral particles using field emission scanning electron microscopy were prepared and assessed using transmission electron microscopy. In the correlative areas, transmission electron microscopy confirmed the presence of large numbers of virus particles. These results indicated that the combination of marking viral particles with 3,3'-diaminobenzidine/metal staining and conductive treatment can identify active progeny virus particles in FFPE sections using scanning electron microscopy. This easy correlative imaging of field emission scanning electron microscopy of the identical area of FFPE in light microscopy may help elucidate new pathological mechanisms of virus-related diseases.

Primary Cilia Are Frequently Present in Small Cell Lung Carcinomas but Not in Non-Small Cell Lung Carcinomas or Lung Carcinoids.

Most human malignant neoplasms show loss of primary cilia (PC). However, PC are known to be retained and involved in tumorigenesis in some types of neoplasms. The PC status in lung carcinomas remains largely uninvestigated. In this study, we comprehensively assessed the PC status in lung carcinomas. A total of 492 lung carcinomas, consisting of adenocarcinomas (ACs) (n = 319), squamous cell carcinomas (SCCs) (n = 152), and small cell lung carcinomas (SCLCs) (n = 21), were examined by immunohistochemical analysis using an antibody against ARL13B, a marker of PC. The PC-positive rate was markedly higher in SCLCs (81.0%) than in ACs (1.6%) and SCCs (7.9%). We subsequently performed analyses to characterize the PC-positive lung carcinomas further. PC-positive lung carcinomas were more numerous and had longer PC than normal cells. The presence of PC in these cells was not associated with the phase of the cell cycle. We also found that the PC were retained even in metastases from PC-positive lung carcinomas. Furthermore, the hedgehog signaling pathway was activated in PC-positive lung carcinomas. Because ARL13B immunohistochemistry of lung carcinoids (n = 10) also showed a statistically significantly lower rate (10.0%) of PC positivity than SCLCs, we searched for a gene(s) that might be upregulated in PC-positive SCLCs compared with lung carcinoids, but not in PC-negative carcinomas. This search, and further cell culture experiments, identified HYLS1 as a gene possessing the ability to regulate ciliogenesis in PC-positive lung carcinomas. In conclusion, our findings indicate that PC are frequently present in SCLCs but not in non-SCLCs (ACs and SCCs) or lung carcinoids, and their PC exhibit various specific pathobiological characteristics. This suggests an important link between lung carcinogenesis and PC.

Identification and characterization of primary cilia-positive salivary gland tumours exhibiting basaloid/myoepithelial differentiation.

Primary cilia (PC) are non-motile, antenna-like structures on the cell surface. Many types of neoplasms exhibit PC loss, whereas in some neoplasms PC are retained and involved in tumourigenesis. To elucidate the PC status and characteristics of major salivary gland tumours (SGTs), we examined 100 major SGTs encompassing eight histopathological types by immunohistochemical analysis. PC were present in all (100%) of the pleomorphic adenomas (PAs), basal cell adenomas (BCAs), adenoid cystic carcinomas (AdCCs), and basal cell adenocarcinomas (BCAcs) examined, but absent in all (0%) of the Warthin tumours, salivary duct carcinomas, mucoepidermoid carcinomas, and acinic cell carcinomas examined. PC were also detected by electron-microscopic analysis using the NanoSuit method. It is worthy of note that the former category and latter category of tumours contained and did not contain a basaloid/myoepithelial differentiation component, respectively. The four types of PC-positive SGTs showed longer PC than normal and exhibited a characteristic distribution pattern of the PC in the ductal and basaloid/neoplastic myoepithelial components. Two PC-positive carcinomas (AdCC and BCAc) still possessed PC in their recurrent/metastatic sites. Interestingly, activation of the Hedgehog signalling pathway, shown by predominantly nuclear GLI1 expression, was significantly more frequently observed in PC-positive SGTs. Finally, we identified tau tubulin kinase 2 (TTBK2) as being possibly involved in the production of PC in SGTs. Taken together, our findings indicate that SGTs that exhibit basaloid/myoepithelial differentiation (PA, BCA, AdCC, and BCAc) are ciliated, and their PC exhibit tumour-specific characteristics, are involved in activation of the Hedgehog pathway, and are associated with TTBK2 upregulation, providing a significant and important link between SGT tumourigenesis and PC. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Exosomes from dental pulp cells attenuate bone loss in mouse experimental periodontitis.

BACKGROUND AND OBJECTIVE: Exosomes are small vesicles secreted from many cell types. Their biological effects largely depend on their cellular origin and the physiological state of the originating cells. Exosomes secreted by mesenchymal stem cells exert therapeutic effects against multiple diseases and may serve as potential alternatives to stem cell therapies. We previously established and characterized human leukocyte antigen (HLA) haplotype homo (HHH) dental pulp cell (DPC) lines from human wisdom teeth. In this study, we aimed to investigate the effect of local administration of HHH-DPC exosomes in a mouse model of periodontitis. METHODS: Exosomes purified from HHH-DPCs were subjected to particle size analysis, and expression of exosome markers was confirmed by western blotting. We also confirmed the effect of exosomes on the migration of both HHH-DPCs and mouse osteoblastic MC3T3-E1 cells. A mouse experimental periodontitis model was used to evaluate the effect of exosomes in vivo. The morphology of alveolar bone was assessed by micro-computed tomography (µCT) and histological analysis. The effect of exosomes on osteoclastogenesis was evaluated using a co-culture system. RESULTS: The exosomes purified from HHH-DPCs were homogeneous and had a spherical membrane structure. HHH-DPC exosomes promoted the migration of both human DPCs and mouse osteoblastic cells. The MTT assay showed a positive effect on the proliferation of human DPCs, but not on mouse osteoblastic cells. Treatment with HHH-DPC exosomes did not alter the differentiation of osteoblastic cells. Imaging with µCT revealed that the exosomes suppressed alveolar bone resorption in the mouse model of periodontitis. Although no change was apparent in the dominance of TRAP-positive osteoclast-like cells in decalcified tissue sections upon exosome treatment, HHH-DPC exosomes significantly suppressed osteoclast formation in vitro. CONCLUSIONS: HHH-DPC exosomes stimulated the migration of human DPCs and mouse osteoblastic cells and effectively attenuated bone loss due to periodontitis.

Sky radiance and spectral gradient are orienting cues for the sandhopper Talitrus saltator (Crustacea, Amphipoda).

The sandhopper Talitrus saltator relies on both the sun and the moon compasses to return to the belt of damp sand on the beach in which it lives buried during the day. In addition to the sun, the gradient of radiance and the spectral distribution across the sky could provide directional information that T. saltator can potentially use to orient itself during the day even when the sun is not visible (e.g. cloudy sky). The scope of this work was (1) to determine the intensity levels of sky radiance that the sandhoppers use in their zonal recovery and (2) to investigate whether this species relies on the celestial spectral gradient in its zonal recovery. Sandhoppers were tested in the laboratory under artificial radiance or spectral gradients. Our results show that under an artificial sky simulating the natural radiance gradient on a cloudless day, sandhoppers orientated toward the correct seaward direction of their home beach; however, individuals lost their ability to use the intensity gradient as an orientation cue when the radiance was attenuated by at least 40%. Sandhoppers were also able to head in the correct seaward direction of their home beach at any time of the day by using the spectral gradient as their only source of visual orientation reference.

Visual adaptation of opsin genes to the aquatic environment in sea snakes.

BACKGROUND: Evolutionary transitions from terrestrial to aquatic life history cause drastic changes in sensory systems. Indeed, the drastic changes in vision have been reported in many aquatic amniotes, convergently. Recently, the opsin genes of the full-aquatic sea snakes have been reported. However, those of the amphibious sea snakes have not been examined in detail. RESULTS: Here, we investigated opsin genes and visual pigments of sea snakes. We determined the sequences of SWS1, LWS, and RH1 genes from one terrestrial, three amphibious and four fully-aquatic elapids. Amino acid replacements at four and one spectra-tuning positions were found in LWS and RH1, respectively. We measured or predicted absorption of LWS and RH1 pigments with A1-derived retinal. During their evolution, blue shifts of LWS pigments have occurred stepwise in amphibious sea snakes and convergently in both amphibious and fully-aquatic species. CONCLUSIONS: Blue shifted LWS pigments may have adapted to deep water or open water environments dominated by blue light. The evolution of opsins differs between marine mammals (cetaceans and pinnipeds) and sea snakes in two fundamental ways: (1) pseudogenization of opsins in marine mammals; and (2) large blue shifts of LWS pigments in sea snakes. It may be possible to explain these two differences at the level of photoreceptor cell composition given that cone and rod cells both exist in mammals whereas only cone cells exist in fully-aquatic sea snakes. We hypothesize that the differences in photoreceptor cell compositions may have differentially affected the evolution of opsins in divergent amniote lineages.

The NanoSuit method: a novel histological approach for examining paraffin sections in a nondestructive manner by correlative light and electron microscopy.

Histological examination using the light microscopy is currently the gold standard for life science research and diagnostics. However, magnified observations are limited because of the limitations intrinsic to light microscopy. Thus, a dual approach, known as correlative light and electron microscopy (CLEM), has emerged, although several technical challenges remain in terms of observing myriad stored paraffin sections. Previously, we developed the NanoSuit method, which enabled us to keep multicellular organisms alive/wet in the high vacuum of a scanning electron microscope by encasing the sample in a thin, vacuum-proof membrane. The approach uses the native extracellular substance (ECS) or an ECS-mimicking substance to polymerize a membrane by plasma or electron beam irradiation. Since the resulting NanoSuit is flexible and dense enough to prevent a living organism's bodily gas and liquids from evaporating (which we refer to as the "surface shield enhancer" (SSE) effect), it works like a miniature spacesuit with sufficient electron conductivity for an SEM observation. Here, we apply the NanoSuit method to CLEM analysis of paraffin sections. Accordingly, the NanoSuit method permits the study of paraffin sections using CLEM at low and high magnification, with the following features: (i) the integrity of the glass slide is maintained, (ii) three-dimensional microstructures of tissue and pathogens are visualized, (iii) nuclei and 3,3'-diaminobenzidine-stained areas are distinct because of gold chloride usage, (iv) immunohistochemical staining is quantitative, and (v) contained elements can be analyzed. Removal of the SSE solution after observation is a further advantage, as this allows slides to be restained and stored. Thus, the NanoSuit method represents a novel approach that will advance the field of histology.

Specializations in the compound eye of Talitrus saltator (Crustacea, Amphipoda).

We investigated the eye regionalization in Talitrus saltator by morphological, electrophysiological and behavioural experiments. Each ommatidium possesses five radially arranged retinular cells producing a square fused rhabdom by R1-R4 cells; the smaller R5 exists between R1 and R4. The size of R5 rhabdomere is larger in the dorsal part and becomes smaller in the median and ventral parts of the eye. Spectral-sensitivity by electroretinograms were recorded from dorsal or ventral parts of the eye. The dorsal part possesses maxima at green and UV-blue region. The main response region in the ventral part is only from UV (390 nm) to blue (430 nm) decreasing at longer wavelengths. To evaluate the sandhoppers' celestial orientation, their eyes were painted black either in the dorsal or ventral part, under the natural sky or a blue filter with or without the vision of the sun. Sandhoppers with the dorsal region of the eyes painted and tested under the screened sun were more dispersed and their directions varied more than in other groups of individuals. Sandhoppers with this area of the eye obscured display considerable difficulties to head in a specific direction. This work suggests the existence of regional specializations in the eye of T. saltator.

Liquid Marbles in Nature: Craft of Aphids for Survival.

Some aphids that live in the leaf galls of the host plant are known to fabricate liquid marbles consisting of honeydew and wax particles as an inner liquid and a stabilizer, respectively. In this study, the liquid marbles fabricated by the galling aphids, Eriosoma moriokense, were extensively characterized with respect to size and size distribution, shape, nanomorphology, liquid/solid weight ratio, and chemical compositions. The stereo microscopy studies confirmed that the liquid marbles have a near-spherical morphology and that the number-average diameter was 368 ± 152 µm, which is 1 order of magnitude smaller than the capillary length of the honeydew. The field emission scanning electron microscopy studies indicated that micrometer-sized wax particles with fiber- and dumpling-like shapes coated the honeydew droplets, which rendered the liquid marbles hydrophobic and nonwetting. Furthermore, the highly magnified scanning electron microscopy images confirmed that the wax particles were formed with assemblage of submicrometer-sized daughter fibers. The contact angle measurements indicated that the wax was intrinsically hydrophobic and that the liquid marbles were stabilized by the wax particles in the Cassie-Baxter model. The weight ratio of the honeydew and the wax particles was determined to be 96/4, and the honeydew consisted of 19 wt % nonvolatile components and 81 wt % water. The 1H nuclear magnetic resonance, Fourier transform infrared spectroscopy, and mass spectroscopy studies confirmed that the wax mainly consisted of triglycerides and that the honeydew mainly consisted of saccharides (glucose and fructose) and ribitol. The atomic force microscopy studies confirmed that honeydew is sticky in nature.

An RNAi Screen for Genes Involved in Nanoscale Protrusion Formation on Corneal Lens in Drosophila melanogaster.

The "moth-eye" structure, which is observed on the surface of corneal lens in several insects, supports anti-reflective and self-cleaning functions due to nanoscale protrusions known as corneal nipples. Although the morphology and function of the "moth-eye" structure, are relatively well studied, the mechanism of protrusion formation from cell-secreted substances is unknown. In Drosophila melanogaster, a compound eye consists of approximately 800 facets, the surface of which is formed by the corneal lens with nanoscale protrusions. In the present study, we sought to identify genes involved in "moth-eye" structure, formation in order to elucidate the developmental mechanism of the protrusions in Drosophila. We re-examined the aberrant patterns in classical glossy-eye mutants by scanning electron microscope and classified the aberrant patterns into groups. Next, we screened genes encoding putative structural cuticular proteins and genes involved in cuticular formation using eye specific RNAi silencing methods combined with the Gal4/UAS expression system. We identified 12 of 100 candidate genes, such as cuticular proteins family genes (Cuticular protein 23B and Cuticular protein 49Ah), cuticle secretion-related genes (Syntaxin 1A and Sec61 ßß subunit), ecdysone signaling and biosynthesis-related genes (Ecdysone receptor, Blimp-1, and shroud), and genes involved in cell polarity/cell architecture (Actin 5C, shotgun, armadillo, discs large1, and coracle). Although some of the genes we identified may affect corneal protrusion formation indirectly through general patterning defects in eye formation, these initial findings have encouraged us to more systematically explore the precise mechanisms underlying the formation of nanoscale protrusions in Drosophila.

A thin polymer membrane, nano-suit, enhancing survival across the continuum between air and high vacuum.

Most multicellular organisms can only survive under atmospheric pressure. The reduced pressure of a high vacuum usually leads to rapid dehydration and death. Here we show that a simple surface modification can render multicellular organisms strongly tolerant to high vacuum. Animals that collapsed under high vacuum continued to move following exposure of their natural extracellular surface layer (or that of an artificial coat-like polysorbitan monolaurate) to an electron beam or plasma ionization (i.e., conditions known to enhance polymer formation). Transmission electron microscopic observations revealed the existence of a thin polymerized extra layer on the surface of the animal. The layer acts as a flexible "nano-suit" barrier to the passage of gases and liquids and thus protects the organism. Furthermore, the biocompatible molecule, the component of the nano-suit, was fabricated into a "biomimetic" free-standing membrane. This concept will allow biology-related fields especially to use these membranes for several applications.

A 'NanoSuit' surface shield successfully protects organisms in high vacuum: observations on living organisms in an FE-SEM.

Although extremely useful for a wide range of investigations, the field emission scanning electron microscope (FE-SEM) has not allowed researchers to observe living organisms. However, we have recently reported that a simple surface modification consisting of a thin extra layer, termed 'NanoSuit', can keep organisms alive in the high vacuum (10(-5) to 10(-7) Pa) of the SEM. This paper further explores the protective properties of the NanoSuit surface-shield. We found that a NanoSuit formed with the optimum concentration of Tween 20 faithfully preserves the integrity of an organism's surface without interfering with SEM imaging. We also found that electrostatic charging was absent as long as the organisms were alive, even if they had not been coated with electrically conducting materials. This result suggests that living organisms possess their own electrical conductors and/or rely on certain properties of the surface to inhibit charging. The NanoSuit seems to prolong the charge-free condition and increase survival time under vacuum. These findings should encourage the development of more sophisticated observation methods for studying living organisms in an FE-SEM.

The use of polarized light in the zonal orientation of the sandhopper Talitrus saltator (Montagu).

It is well known that the celestial polarization is used as a compass cue by many species of insects and crustaceans. Although it has been shown that the sandhopper Talitrus saltator perceives polarized light and possesses an arrangement of the rhabdomeres that could allow e-vector interpretation and utilization, T. saltator does not use the e-vector of the skylight polarization as a compass cue when making excursions along the sea-land axis of sandy shores. We performed tests in confined conditions to clarify if skylight polarization is somehow involved in the zonal recovery of T. saltator. We observed the directional responses of sandhoppers in a transparent bowl under an artificial sky (an opaline Plexiglas dome). The bowl was covered by a blue gelatin filter with a grey filter (control condition) and a linear polarizing filter (experimental conditions) positioned under the blue one in such a way as to occupy half of the upper surface of the Plexiglas bowl so as to create a linear polarization gradient. Our experiments confirm that T. saltator perceives polarized light and highlight that this visual capability determines the perception, or perhaps the increase, of the radiance and/or spectral gradient and their use as compass cues in the zonal orientation. Moreover, our findings confirm that the radiance gradient is used as a chronometric compass orienting reference in the absence of other celestial orienting cues.

Route of pesticide spread on the body surface of Blattella germanica (Linnaeus): a NanoSuit-energy dispersive X-ray spectroscopy analysis.

Numerous studies have focussed on the mechanisms of entry of pesticides into insect body parts such as oral intake, penetration through the integument of the body wall, and inhalation through spiracles. However, little is known about how insecticides spread to the points of entry or the paths on the body surface that are used to reach the target sites. In this study, elemental signals of pesticide-mimicking test solutions were tracked and their routes of spreading in experimental insects (Blattella germanica L.) were investigated using NanoSuit (a method of surface modification) and energy dispersive X-ray spectroscopy, combined with high-resolution scanning electron microscopy. When the test solution initially adhered to the dorsal and/or ventral body surface, it tended to spread horizontally to reach lateral plates. Whereas, when the solution directly adhered to the anterior side of the lateral plates, it spread to posterior segments. In this case, however, spreading in the opposite direction (i.e., the solution directly adhered to the posterior side of the lateral plates) was interrupted at a boundary erected by different groups of fine structures; each protrusion was large, and the arrangement was rather dense in the posterior segments. Morphological features of these fine structures and chemical characteristics of the hydrophobic surface substances potentially regulate the strength of the capillary force, which determines pesticide spreading.

Association between human papillomavirus particle production and the severity of recurrent respiratory papillomatosis.

Recurrent respiratory papillomatosis (RRP) has a wide range of severity. We investigate the relationship between human papillomavirus (HPV) particle production and severity of RRP. From September 2005 to June 2021, 68 RRP samples (from 29 patients) were included. HPV type was determined. HPV viral load, physical status, and demographic and clinical characteristics were assessed. Immunohistochemistry (IHC) was performed for p16, Ki-67, L1, and E4. We used NanoSuit-CLEM (correlative light and electron microscopy) and transmission electron microscopy (TEM) to examine the samples. The total number of surgeries in HPV-positive and HPV-negative cases were 3.78 (n = 55/68, range: 1-16) and 1.30 (n = 13/68, range: 1-3), respectively (p = 0.02). IHC showed that L1 and E4 were correlated and expressed on the tumour surface. NanoSuit-CLEM and TEM revealed HPV particles in L1-positive nuclei. L1 IHC-positive cases had a shorter surgical interval (p < 0.01) and more frequent surgeries (p = 0.04). P16 IHC, viral load, and physical status were not associated with disease severity. This study visualised HPV particle production in RRP for the first time. Persistent HPV particle infection was associated with severity. We suggest L1 IHC for evaluating RRP severity in addition to the Derkay score.

Highly Sensitive and Quantitative Diagnosis of SARS-CoV-2 Using a Gold/Platinum Particle-Based Lateral Flow Assay and a Desktop Scanning Electron Microscope.

The gold standard test for identifying SARS-CoV-2, the causative agent of COVID-19, is polymerase chain reaction (PCR). Despite their limited sensitivity, SARS-CoV-2 antigen rapid diagnostic tests are vital tools in the fight against viral spread. Owing to its simplicity and low cost, the lateral flow assay (LFA) is the most extensively used point-of-care diagnostic test. Here, we report a newly designed LFA-NanoSuit method (LNSM) that works in conjunction with desktop scanning electron microscopy (SEM) to detect SARS-CoV-2. LNSM requires no standard SEM treatment, avoids cellulose and residual buffer deformation, and enables the capture of high-resolution images of antibody-labeled gold/platinum particles reacting with SARS-CoV-2 antigens. To assess its applicability, we compared clinical SARS-CoV-2 samples via visual detection of LFA, LSNM detection of LFA, and real-time reverse transcription-PCR (qRT-PCR). Compared to qRT-PCR, LNSM showed 86.7% sensitivity (26/30; 95% confidence interval (CI): 69.28-96.24%) and 93.3% specificity (14/15; 95% CI: 68.05-99.83%) for SARS-CoV-2. In samples with a relatively low SARS-CoV-2 RNA copy number (30 < Ct ≤ 40), the sensitivity of LNSM was greater (73.3%) than that of visual detection (0%). A simple, sensitive, and quantitative LNSM can be used to diagnose SARS-CoV-2.

Microscopy and biomimetics: the NanoSuit® method and image retrieval platform.

This review aims to clarify a suitable method towards achieving next-generation sustainability. As represented by the term 'Anthropocene', the Earth, including humans, is entering a critical era; therefore, science has a great responsibility to solve it. Biomimetics, the emulation of the models, systems and elements of nature, especially biological science, is a powerful tool to approach sustainability problems. Microscopy has made great progress with the technology of observing biological and artificial materials and its techniques have been continuously improved, most recently through the NanoSuit® method. As one of the most important tools across many facets of research and development, microscopy has produced a large amount of accumulated digital data. However, it is difficult to extract useful data for making things as biomimetic ideas despite a large amount of biological data. Here, we would like to find a way to organically connect the indispensable microscopic data with the new biomimetics to solve complex human problems.

Combination of the NanoSuit method and gold/platinum particle-based lateral flow assay for quantitative and highly sensitive diagnosis using a desktop scanning electron microscope.

Owing to its simplicity and low cost, the lateral flow assay (LFA) is one of the most commonly used point-of-care diagnostic techniques, despite its low sensitivity and poor quantification. Here, we report a newly developed LFA-NanoSuit method (LNSM) combined with a desktop scanning electron microscope (SEM) for the direct observation of immunocomplexes labeled with a colloidal metal instead of signal enhancement strategies, such as using color, electrochemical signals, silver enhancement, magnetic properties, luminescent, and surface-enhanced Raman spectroscopy (SERS). The proposed LNSM suppresses cellulose deformity, thereby allowing the acquisition of high-resolution images of gold/platinum-labeled immunocomplexed pathogens such as influenza A, without conductive treatment as in conventional SEM. Electron microscopy-based diagnosis of influenza A exhibited 94 % clinical sensitivity (29/31; 95 % confidence interval [CI]: 79.3-98.2 %) and 100 % clinical specificity (95 % CI: 98.1-100 %), which was more sensitive (71.4 %) than visual detection (14.3 %), especially in the lower influenza A-RNA copy number group. The detection ability of our method was nearly comparable to that of real-time reverse transcription-PCR. This is the first report on the diagnosis of clinical diseases using LFA equipped with a desktop SEM. This simple and highly sensitive quantitative analysis method involving LFA can be used to diagnose various diseases in humans and livestock, including highly infectious diseases such as COVID-19.

Synergistic Effects of the Red Light and Blue Traps on Control of Thrips palmi (Thysanoptera: Thripidae).

The melon thrips (Thrips palmi) is a serious insect pest of vegetables and ornamental plants. To control and monitor this thrips, blue traps are widely used. We previously reported that irradiation of cucumber plants with red light-emitting diodes (660 nm) causes T. palmi to avoid the plants. Here, we evaluated the responses of T. palmi adults to a cucumber plant in an experimental arena with either a transparent sticky trap or a blue sticky trap, without or with illumination by a red LED panel. When T. palmi were introduced into the arena, the number of thrips attracted to the plant was lowest in treatments using the red light and blue trap. On the other hand, when T. palmi were inoculated on plants first, most thrips stayed put on the plants. Placing both red LEDs and blue sticky traps in greenhouses before the thrips invade the greenhouse from the outside might effectively control T. palmi.