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Abstract Patients with rare diseases often experience prolonged diagnostic delays. Ordering appropriate genetic tests is crucial yet challenging, especially for general pediatricians without genetic expertise. Recent American College of Medical Genetics (ACMG) guidelines embrace early use of exome sequencing (ES) or genome sequencing (GS) for conditions like congenital anomalies or developmental delays while still recommend gene panels for patients exhibiting strong manifestations of a specific disease. Recognizing the difficulty in navigating these options, we developed a machine learning model trained on 1005 patient records from Columbia University Irving Medical Center to recommend appropriate genetic tests based on the phenotype information. The model achieved a remarkable performance with an AUROC of 0.823 and AUPRC of 0.918, aligning closely with decisions made by genetic specialists, and demonstrated strong generalizability (AUROC:0.77, AUPRC: 0.816) in an external cohort, indicating its potential value for general pediatricians to expedite rare disease diagnosis by enhancing genetic test ordering.

Abstract Background Our previous study revealed that feeding the antimicrobial peptide (AMP) product Scy-hepc significantly enhances the growth of mariculture fish through the activation of the GH-Jak2-STAT5-IGF1 axis. However, the contribution of gut microbiota to this growth enhancement remains unclear. This study aimed to elucidate the potential mechanism involved in intestinal absorption and modulation of gut microbiota in Epinephelus akaara following Scy-hepc feeding. Results The results showed that a 35 day regimen of Scy-hpec markedly promoted the growth of E. akaara compared to groups supplemented with either florfenicol, B. subtilis, or a vector. The growth enhancement is likely attributed to alterations in microbiota colonization in the foregut and midgut, characterized by an increasing abundance of potential probiotics (Rhizobiaceae and Lysobacter) and a decreased abundance of opportunistic pathogens (Psychrobacter and Brevundimonas) as determined by 16S rRNA analysis. Additionally, similar to the effect of florfenicol feeding, Scy-hepc significantly improved host survival rate by over 20% in response to a lethal dose challenge with Edwardsiella tarda. Further investigations demonstrated that Scy-hepc is absorbed by the fish foregut (20–40 min) and midgut (20–30 min) as confirmed by Western blot, ELISA, and Immunofluorescence. The absorption of Scy-hepc affected the swimming, swarming and surfing motility of Vibrio harveyi and Bacillus thuringiensis isolated from E. akaara’s gut. Moreover, Scy-hepc induced the downregulation of 40 assembly genes and the upregulation expression of 5, with the most significant divergence in gene expression between opportunistic pathogens and probiotics concentrated in their motility genes (PomA/B, MotA/B). Conclusions In summary, this study shows that feeding AMP Scy-hepc can promote growth and bolster immunity in E. akaara. These beneficial effects are likely due to the absorption of Scy-hepc in the fish’s foregut and midgut, which modulates the colonization and motility of commensal bacteria, leading to favorable changes in the composition of the foregut and midgut microbiota. Therefore, a profound understanding of the mechanisms by which antimicrobial peptides affect host gut microbiota will contribute to a comprehensive assessment of their advantages and potential application prospects as substitutes for antibiotics in fish health and improving aquaculture practices.

Abstract BackgroundThe association between social media usage and the risk of depressive symptoms has attracted increasing attention. WeChat is a popular social media software in China. The impact of using WeChat and posting WeChat moments on the risk of developing depressive symptoms among community-based middle-aged and older adults in China is unknown. ObjectiveThe objective was to assess the association between using WeChat and posting WeChat moments and the risk of depressive symptoms among middle-aged and older adults in China. MethodsA prospective national cohort study was designed based on the data obtained from the fourth and fifth waves of the China Health and Retirement Longitudinal Study (CHARLS). The strength of association between using WeChat and posting WeChat moments and the risk of depressive symptoms was estimated by modified Poisson regressions. Depressive symptoms were determined using the 10-item Center for Epidemiologic Studies Depression Scale. Meanwhile, the heterogeneity of the associations was explored through multiple subgroup analyses. Moreover, multiple sensitivity analyses were performed to verify the robustness of the associations between the exposures and depressive symptoms. ResultsA total of 9670 eligible participants were included in the cohort study, and the incidence rate of depressive symptoms was 19.08% (1845/9670, 95% CI 19.07%‐19.09%) from the fourth to fifth waves of the CHARLS. Using WeChat (adjusted relative risk [aRR] 0.691, 95% CI 0.582‐0.520) and posting WeChat moments (aRR 0.673, 95% CI 0.552‐0.821) reduced the risk of depressive symptoms among middle-aged and older Chinese adults. The association between the exposures and depressive symptoms was robust, proved through multiple sensitivity analyses (all P ConclusionsUsing WeChat and especially posting WeChat moments can mitigate the risk of depressive symptoms among community-based middle-aged and older Chinese adults. However, there is likely a need for a longer follow-up period to explore the impact of the exposures on the risk of long-term depressive outcomes.

Abstract Vertigo is a common symptom of various diseases that affects a large number of people worldwide. Current leading treatments for intractable peripheral vertigo are to intratympanically inject ototoxic drugs such as gentamicin to attenuate the semicircular canal function but inevitably cause hearing injury. Photodynamic therapy (PDT) is a noninvasive therapeutic approach by precisely targeting the diseased tissue. Here, we developed a PDT‐based method for treating intractable peripheral vertigo in a mouse model using a polymer‐coated photosensitizer chlorin e6 excited by red light. We found that a high dose of PDT attenuated the function of both semicircular canals and otolith organs and damaged their hair cells. Conversely, the PDT exerted no effect on hearing function or cochlear hair‐cell viability. These results suggest the therapeutic potential of PDT for treating intractable peripheral vertigo without hurting hearing. Besides, the attenuation level and affected area can be precisely controlled by adjusting the light exposure time. Furthermore, we demonstrated the potential of this therapeutic approach to be minimally invasive with light irradiation through bone results. Thus, our PDT‐based approach for attenuating the function of the semicircular canals offers a basis for developing a less‐invasive and targeted therapeutic option for treating vertigo.

Abstract Background The gut microbiota significantly influences the health and growth of red-spotted grouper (Epinephelus akaara), a well-known commercial marine fish from Fujian Province in southern China. However, variations in survival strategies and seasons can impact the stability of gut microbiota data, rendering it inaccurate in reflecting the state of gut microbiota. Which impedes the effective enhancement of aquaculture health through a nuanced understanding of gut microbiota. Inspired by this, we conducted a comprehensive analysis of the gut microbiota of wild and captive E. akaara in four seasons. Results Seventy-two E. akaara samples were collected from wild and captive populations in Dongshan city, during four different seasons. Four sections of the gut were collected to obtain comprehensive information on the gut microbial composition and sequenced using 16S rRNA next-generation Illumina MiSeq. We observed the highest gut microbial diversity in both captive and wild E. akaara during the winter season, and identified strong correlations with water temperature using Mantel analysis. Compared to wild E. akaara, we found a more complex microbial network in captive E. akaara, as evidenced by increased abundance of Bacillaceae, Moraxellaceae and Enterobacteriaceae. In contrast, Vibrionaceae, Clostridiaceae, Flavobacteriaceae and Rhodobacteraceae were found to be more active in wild E. akaara. However, some core microorganisms, such as Firmicutes and Photobacterium, showed similar distribution patterns in both wild and captive groups. Moreover, we found the common community composition and distribution characteristics of top 10 core microbes from foregut to hindgut in E. akaara. Conclusions Collectively, the study provides relatively more comprehensive description of the gut microbiota in E. akaara, taking into account survival strategies and temporal dimensions, which yields valuable insights into the gut microbiota of E. akaara and provides a valuable reference to its aquaculture.

Abstract A scalable, reusable, and broad-coverage unified material knowledge representation shows its importance and will bring great benefits to data sharing among materials communities. A knowledge graph (KG) for materials terminology, which is a formal collection of term entities and relationships, is conceptually important to achieve this goal. In this work, we propose a KG for materials terminology, named Materials Genome Engineering Database Knowledge Graph (MGED-KG), which is automatically constructed from text corpus via natural language processing. MGED-KG is the most comprehensive KG for materials terminology in both Chinese and English languages, consisting of 8,660 terms and their explanations. It encompasses 11 principal categories, such as Metals, Composites, Nanomaterials, each with two or three levels of subcategories, resulting in a total of 235 distinct category labels. For further application, a knowledge web system based on MGED-KG is developed and shows its great power in improving data sharing efficiency from the aspects of query expansion, term, and data recommendation.

Abstract In the process of coping with energy and environmental protection issues, technologies such as energy materials, energy devices, and energy systems have made great progress. With excellent performance, film capacitors play an increasingly important role in energy‐related fields. With the increase of application scenarios and the continuous development of film material technology, it is urgent to establish a better theoretical connection from films to capacitors. First, the main components of the capacitor including the film and the positional relationships among them are given. Then, from the two perspectives of indirect calculation according to the volume and the direct calculation according to the winding process, the equation between the dielectric constant of films and the corresponding capacitance of capacitors is established. Further, the measurement data and error analysis results of the built test platform prove the accuracy and great potential of the proposed calculation methods. In addition, error sources, including film thickness uniformity, are listed. Finally, the challenges faced by the proposed calculation methods and the paths that can be referenced for future research are summarised and discussed.

Osteosarcoma (OS) is the most common primary bone cancer in children and young adults, patient survival rates have not improved in recent decades. To further understand the interrelationship between different cell types in the tumor microenvironment of osteosarcoma, we comprehensively analyzed single-cell sequencing data from six patients with untreated osteosarcoma. Nine major cell types were identified from a total of 46,046 cells based on unbiased clustering of gene expression profiles and canonical markers. Osteosarcoma from different patients display heterogeneity in cellular composition. Myeloid cells were the most commonly represented cell type, followed by osteoblastic and TILs. Copy number variation (CNV) results identified amplifications and deletions in malignant osteoblastic cells and fibroblasts. Trajectory analysis based on RNA velocity showed that osteoclasts in the OS microenvironment could be differentiated from myeloid cells. Furthermore, we explored the intercellular communications in OS microenvironment and identified multiple ligand-receptor pairs between myeloid cells, osteoblastic cells and their cells, including 21 ligand-receptor pair genes that significantly associated with survival outcomes. Importantly, we found chemotherapy may have an effect on cellular communication in the OS microenvironment by analyzing single-cell sequencing data from seven primary osteosarcoma patients who received chemotherapy. We believe these observations will improve our understanding of potential mechanisms of microenvironment contributions to OS progression and help identify potential targets for new treatment development in the future.

In recent years, new emerging pathogenic microorganisms have frequently appeared in animals, including marine fish, possibly due to climate change, anthropogenic activities, and even cross-species transmission of pathogenic microorganisms among animals or between animals and humans, which poses a serious issue for preventive medicine. In this study, a bacterium was clearly characterized among 64 isolates from the gills of diseased large yellow croaker Larimichthys crocea that were raised in marine aquaculture. This strain was identified as K. kristinae by biochemical tests with a VITEK 2.0 analysis system and 16S rRNA sequencing and named K. kristinae_LC. The potential genes that might encode virulence-factors were widely screened through sequence analysis of the whole genome of K. kristinae_LC. Many genes involved in the two-component system and drug-resistance were also annotated. In addition, 104 unique genes in K. kristinae_LC were identified by pan genome analysis with the genomes of this strain from five different origins (woodpecker, medical resource, environment, and marine sponge reef) and the analysis results demonstrated that their predicted functions might be associated with adaptation to living conditions such as higher salinity, complex marine biomes, and low temperature. A significant difference in genomic organization was found among the K. kristinae strains that might be related to their hosts living in different environments. The animal regression test for this new bacterial isolate was carried out using L. crocea, and the results showed that this bacterium could cause the death of L. crocea and that the fish mortality was dose-dependent within 5 days post infection, indicating the pathogenicity of K. kristinae_LC to marine fish. Since K. kristinae has been reported as a pathogen for humans and bovines, in our study, we revealed a new isolate of K. kristinae_LC from marine fish for the first time, suggesting the potentiality of cross-species transmission among animals or from marine animals to humans, from which we would gain insight to help in future public prevention strategies for new emerging pathogens.

The gut microbiota plays an important role in animal health and behavior. In marine fish, the composition of the gut microbiota is affected by many complex factors, such as diet, species, and regional factors. Since more than one hundred fish species have been cultured in fish farms along with the 3,324 km coastline of Fujian Province in South China, we chose this region to study the gut microbiota composition of marine commercial fishes because sufficient different species, diets, and regional factors were observed. We investigated the distribution characteristics of the gut microbiota of seven cultured species (Epinephelus akaara, Epinephelus coioides, Epinephelus lanceolatus ♂ × Epinephelus fuscoguttatus ♀, Siganus fuscescens, Pagrus major, Lateolabrax japonicus, and Acanthopagrus schlegelii) living in the same aquatic region and one species (E. akaara) living separately in five regions separated by latitude. The impacts of diet, region, and species factors on fish gut microbiota were also evaluated. Diversity and multivariate analyses showed that the patterns of the microbiota were significantly different in different fish species within the same habitat and E. akaara with five latitude regions. Mantel analysis showed that AN, SiO32–, DO, and NO2– were the principal factors affecting the microbial community of E. akaara in the five habitats. Additionally, similar distribution characteristics occurred in different gut parts of different fishes, with an increasing trend of Proteobacteria and Vibrionaceae abundance and a decreasing trend of Firmicutes and Bacillaceae abundance from the foregut to the hindgut. Vibrionaceae was the most abundant family in the content. This study highlights that a persistent core microbiota was established in marine commercial fishes spanning multiple scales. The factors with the greatest effect on fish gut microbiota may be (i) host genetics and (ii) geographic factors rather than the microbiota in the diet and water environment. These core microbes regularly colonized from the foregut to the hindgut, which was driven by their underlying functions, and they were well adapted to the gut environment. Moreover, the microbiota in the content may have contributed more to the gut microbial communities than previously reported. This study could complement basic data on the composition of marine commercial fishes and facilitate relatively complete investigations, which would be beneficial for the healthy and sustainable development of aquaculture.

Antimicrobial peptides (AMPs) may be the most promising substitute for antibiotics due to their effective bactericidal activity and multiple antimicrobial modes against pathogenic bacteria. In this study, a new functional gene named Spgillcin was identified in Scylla paramamosain, which encoded 216 amino acids of mature peptide. In vivo, Spgillcin was dominantly expressed in the gills of male and female crabs, offering the highest expression level among all tested organs or tissues. The expression pattern of Spgillcin was significantly altered when challenged by Staphylococcus aureus, indicating a positive immune response. In vitro, a functional truncated peptide Spgillcin177–189 derived from the amino acid sequence of Spgillcin was synthesized and showed a broad-spectrum and potent antibacterial activity against several bacterial strains, including the clinical isolates of multidrug-resistant (MDR) strains, with a range of minimum inhibitory concentrations from 1.5 to 48 μM. Spgillcin177–189 also showed rapid bactericidal kinetics for S. aureus and Pseudomonas aeruginosa but did not display any cytotoxicity to mammalian cells and maintained its antimicrobial activity in different conditions. Mechanistic studies indicated that Spgillcin177–189 was mainly involved in the disruption of cell membrane integrity where the membrane components lipoteichoic acid and lipopolysaccharide could significantly inhibit the antimicrobial activity in a dose-dependent manner. In addition, Spgillcin177–189 could change the membrane permeability and cause the accumulation of intracellular reactive oxygen species. No resistance was generated to Spgillcin177–189 when the clinical isolates of methicillin-resistant S. aureus and MDR P. aeruginosa were treated with Spgillcin177–189 and then subjected to a long term of continuous culturing for 50 days. In addition, Spgillcin177–189 exerted a strong anti-biofilm activity by inhibiting biofilm formation and was also effective at killing extracellular S. aureus in the cultural supernatant of RAW 264.7 cells. Taken together, Spgillcin177–189 has strong potential as a substitute for antibiotics in future aquaculture and medical applications.

Crustins are the most abundant class of antimicrobial peptides in crustaceans and are essential for protecting animals from infection. Among them, type II crustins usually exhibit potent antimicrobial activity. Interestingly, in this study, a newly identified type II crustin gene homolog (named SpCrus8) from mud crab Scylla paramamosain, the recombinant proteins of which (rSpCrus8 and rTrx-SpCrus8) showed no obvious antibacterial effects, but could significantly reduce the bacterial load in crab hemolymph and improve the survival rate of crabs infected with Vibrio alginolyticus. The immune-related function of SpCrus8 and the underlying mechanism deserve further investigation. It was found that the SpCrus8 gene was widely distributed in various tissues of adult crabs. In the hepatopancreas of crabs infected with V. alginolyticus or Staphylococcus aureus, transcripts of the SpCrus8 gene were remarkably induced, indicating that the SpCrus8 gene was involved in the immune response to bacterial infection in vivo. In addition, rSpCrus8 and rTrx-SpCrus8 had strong binding activity not only to microbial surface components (lipopolysaccharide, lipoteichoic acid, peptidoglycan, and glucan), but also to the tested bacteria (S. aureus, Pseudomonas aeruginosa and V. alginolyticus). Notably, rSpCrus8 and rTrx-SpCrus8 could significantly promote hemocyte phagocytosis. After rSpCrus8 and rTrx-SpCrus8 treatment, a large number of fluorescent microspheres were observed to aggregate into clusters and be phagocytosed by multiple hemocytes, while hemocytes in the control group phagocytosed only individual microspheres, indicating that SpCrus8 played an important role in opsonization. When the SpCrus8 gene was knocked down, the expression levels of the key phagocytosis-related genes SpRab5 and SpRab7 were significantly downregulated, as well as the IMD signaling pathway genes SpIKKβ and SpRelish, and another crustin gene SpCrus5. Correspondingly, all the SpIKKβ, SpRelish and SpCrus5 genes were significantly upregulated after rSpCrus8 treatment, suggesting that SpCrus8 might be involved in the immunomodulation of S. paramamosain. Taken together, this study revealed the immune-related functions of the SpCrus8 gene in opsonization and regulation, which will help us further understand the role of the crustin gene family in the immune system of mud crabs and provide new insights into the function of type II crutins.

This paper presents the results of experimental investigations on the durability of glass-fiber-reinforced polymer (GFRP) pultruded profiles made of unsaturated polyester (UP) and vinylester (VE) resins commonly used in civil engineering. The water absorption, tensile properties, and microstructures of GFRP profiles exposed to several typical accelerated aging environments (e.g., deionized water, salt water, salt fog, and combined hygrothermal cycles) for 12 months were investigated. Moreover, a sustained loading factor was included in the test to reflect the behavior of the GFRP profiles in real structures. A normalization approach based on the controlled specimens was used to assess the effectiveness and relevance of the accelerated exposure. The results indicated that the maximum moisture absorption of both UP and VE GFRP profiles was immersed in deionized water, where the masses increased by 1.03 and 0.53%, respectively, leading to the maximum degradation of tensile strength (24.03%) of UP GFRP profile immersion in deionized water after 360 days of aging. However, the tensile modulus was more sensitive to high temperatures and has the maximum degradation (47.03%) after hygrothermal cycles. Moreover, VE GFRP profiles show superior humidity and temperature endurance. Furthermore, the sustained loading exacerbated the degradation of tensile properties slightly under the same conditions. Finally, by incorporating the cumulative damage caused by the sustained loading and a time-dependent factor into a residual strength model, a revised model was proposed to describe the tensile strength loss of pultruded UP GFRP profiles.

Most cases of acquired hearing loss are due to degeneration and subsequent loss of cochlear hair cells. Whereas mammalian hair cells are not replaced when lost, in zebrafish, they constantly renew and regenerate after injury. However, the molecular mechanism among this difference remains unknown. Dual-specificity phosphatase 14 (DUSP14) is an important negative modulator of mitogen-activated protein kinase (MAPK) signaling pathways. Our study was to investigate the effects of DUSP14 on supporting cell development and hair cell regeneration and explore the potential mechanism. Our results showed that dusp14 gene is highly expressed in zebrafish developing neuromasts and otic vesicles. Behavior analysis showed that dusp14 deficiency resulted in hearing defects in zebrafish larvae, which were reversed by dusp14 mRNA treatment. Moreover, knockdown of dusp14 gene caused a significant decrease in the number of neuromasts and hair cells in both neuromast and otic vesicle, mainly due to the inhibition of the proliferation of supporting cells, which results in a decrease in the number of supporting cells and ultimately in the regeneration of hair cells. We further found significant changes in a series of MAPK pathway genes through transcriptome sequencing analysis of dusp14-deficient zebrafish, especially mapk12b gene in p38 signaling. Additionally, inhibiting p38 signaling effectively rescued all phenotypes caused by dusp14 deficiency, including hair cell and supporting cell reduction. These results suggest that DUSP14 might be a key gene to regulate supporting cell development and hair cell regeneration and is a potential target for the treatment of hearing loss.

Hearing loss is a serious illness affecting people’s normal life enormously. The acoustic properties of a tympanic membrane play an important role in hearing, and highly depend on its geometry, composition, microstructure and connection to the surrounding annulus. While the conical geometry of the tympanic membrane is critical to the sound propagation in the auditory system, it presents significant challenges to the study of the 3D microstructure of the tympanic membrane using traditional 2D imaging techniques. To date, most of our knowledge about the 3D microstructure and composition of tympanic membranes is built from 2D microscopic studies, which precludes an accurate understanding of the 3D microstructure, acoustic behaviors and biology of the tissue. Although the tympanic membrane has been reported to contain elastic fibers, the morphological characteristic of the elastic fibers and the spatial arrangement of the elastic fibers with the predominant collagen fibers have not been shown in images. We have developed a 3D imaging technique for the three-dimensional examination of the microstructure of the full thickness of the tympanic membranes in mice without requiring tissue dehydration and stain. We have also used this imaging technique to study the 3D arrangement of the collagen and elastic fibrillar network with the capillaries and cells in the pars tensa-annulus unit at a status close to the native. The most striking findings in the study are the discovery of the 3D form of the elastic and collagen network, and the close spatial relationships between the elastic fibers and the elongated fibroblasts in the tympanic membranes. The 3D imaging technique has enabled to show the 3D waveform contour of the collagen and elastic scaffold in the conical tympanic membrane. Given the close relationship among the acoustic properties, composition, 3D microstructure and geometry of tympanic membranes, the findings may advance the understanding of the structure—acoustic functionality of the tympanic membrane. The knowledge will also be very helpful in the development of advanced cellular therapeutic technologies and 3D printing techniques to restore damaged tympanic membranes to a status close to the native.

The emergence of multidrug-resistant (MDR) pathogens has become a global public health crisis. Among them, MDR Pseudomonas aeruginosa is the main cause of nosocomial infections and deaths. Antimicrobial peptides (AMPs) are considered as competitive drug candidates to address this threat. In the study, we characterized two AMPs (AS-hepc3(41-71) and AS-hepc3(48-56)) that had potent activity against 5 new clinical isolates of MDR P. aeruginosa. Both AMPs destroyed the integrity of the cell membrane, induced leakage of intracellular components, and ultimately led to cell death. A long-term comparative study on the bacterial resistance treated with AS-hepc3(41-71), AS-hepc3(48-56) and 12 commonly used antibiotics showed that P. aeruginosa quickly developed resistance to the nine antibiotics tested (including aztreonam, ceftazidime, cefepime, imipenem, meropenem, ciprofloxacin, levofloxacin, gentamicin, and piperacillin) as early as 12 days after 150 days of successive culture generations. The initial effective concentration of 9 antibiotics against P. aeruginosa was greatly increased to a different high level at 150 days, however, both AS-hepc3(41-71) and AS-hepc3(48-56) maintained their initial MIC unchangeable through 150 days, indicating that P. aeruginosa did not produce any significant resistance to both AMPs. Furthermore, AS-hepc3(48-56) did not show any toxic effect on mammalian cells in vitro and mice in vivo. AS-hepc3(48-56) had a therapeutic effect on MDR P. aeruginosa infection using a mouse lung infection model and could effectively increase the survival rate of mice by inhibiting bacterial proliferation and attenuating lung inflammation. Taken together, the short peptide AS-hepc3(48-56) would be a promising agent for clinical treatment of MDR P. aeruginosa infections.

Due to the increasing prevalence of drug-resistant fungi and the limitations of current treatment strategies to fungal infections, exploration and development of new antifungal drugs or substituents are necessary. In the study, a novel antimicrobial peptide, named Sparamosin, was identified in the mud crab Scylla paramamosain, which contains a signal peptide of 22 amino acids and a mature peptide of 54 amino acids. The antimicrobial activity of its synthetic mature peptide and two truncated peptides (Sparamosin1–25 and Sparamosin26–54) were determined. The results showed that Sparamosin26–54 had the strongest activity against a variety of Gram-negative bacteria, Gram-positive bacteria and fungi, in particular had rapid fungicidal kinetics (killed 99% Cryptococcus neoformans within 10 min) and had potent anti-biofilm activity against C. neoformans, but had no cytotoxic effect on mammalian cells. The RNA-seq results showed that after Sparamosin26–54 treatment, the expression of genes involved in cell wall component biosynthesis, cell wall integrity signaling pathway, anti-oxidative stress, apoptosis and DNA repair were significantly up-regulated, indicating that Sparamosin26–54 might disrupt the cell wall of C. neoformans, causing oxidative stress, DNA damage and cell apoptosis. The underlying mechanism was further confirmed. Sparamosin26–54 could bind to several phospholipids in the cell membrane and effectively killed C. neoformans through disrupting the integrity of the cell wall and cell membrane observed by electron microscope and staining assay. In addition, it was found that the accumulation of reactive oxygen species (ROS) increased, the mitochondrial membrane potential (MMP) was disrupted, and DNA fragmentation was induced after Sparamosin26–54 treatment, which are all hallmarks of apoptosis. Taken together, Sparamosin26–54 has a good application prospect as an effective antimicrobial agent, especially for C. neoformans infections.

Abstract Background Inner ear supporting cells (SCs) in the neonatal mouse cochlea are a potential source for hair cell (HC) regeneration, but several studies have shown that the regeneration ability of SCs decreases dramatically as mice age and that lost HCs cannot be regenerated in adult mice. To better understand how SCs might be better used to regenerate HCs, it is important to understand how the gene expression profile changes in SCs at different ages. Methods Here, we used Sox2 GFP/+ mice to isolate the Sox2+ SCs at postnatal day (P)3, P7, P14, and P30 via flow cytometry. Next, we used RNA-seq to determine the transcriptome expression profiles of P3, P7, P14, and P30 SCs. To further analyze the relationships between these age-related and differentially expressed genes in Sox2+ SCs, we performed gene ontology (GO) analysis. Results Consistent with previous reports, we also found that the proliferation and HC regeneration ability of isolated Sox2+ SCs significantly decreased as mice aged. We identified numerous genes that are enriched and differentially expressed in Sox2+ SCs at four different postnatal ages, including cell cycle genes, signaling pathway genes, and transcription factors that might be involved in regulating the proliferation and HC differentiation ability of SCs. We thus present a set of genes that might regulate the proliferation and HC regeneration ability of SCs, and these might serve as potential new therapeutic targets for HC regeneration. Conclusions In our research, we found several genes that might play an important role in regulating the proliferation and HC regeneration ability of SCs. These datasets are expected to serve as a resource to provide potential new therapeutic targets for regulating the ability of SCs to regenerate HCs in postnatal mammals.

Hepcidin is widely present in many kinds of fish and is an important innate immune factor. A variety of HAMP2-type hepcidins have strong antimicrobial activity and immunomodulatory functions and are expected to be developed as substitutes for antibiotics. In this study, the antimicrobial activity of Hepc2 from Japanese seabass (Lateolabrax japonicus) (designated as LJ-hep2) was investigated using its recombinant precursor protein (rLJ-hep2) expressed in Pichia pastoris and a chemically synthesized mature peptide (LJ-hep2(66–86)). The results showed that both rLJ-hep2 and synthetic LJ-hep2(66–86) displayed broad antimicrobial spectrum with potent activity against gram-negative and gram-positive bacteria, and fungi. Especially, LJ-hep2(66–86) had stronger antimicrobial activity and exhibited potent activity against several clinically isolated multidrug-resistant bacteria, including Acinetobacter baumannii, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae and Enterococcus faecium. Moreover, LJ-hep2(66–86) exerted rapid bactericidal kinetic (killed tested bacteria within 2 h), induced significant morphological changes and promoted agglutination of E. coli, P. aeruginosa and Aeromonas hydrophila. The activity of LJ-hep2(66–86) against E. coli, P. aeruginosa and A. hydrophila was stable and remained active when heated for 30 min. In addition, LJ-hep2(66–86) exhibited no cytotoxicity to the mammalian cell line HEK293T and fish cell lines (EPC and ZF4). In vivo study showed that LJ-hep2(66–86) could improve the survival rate of marine medaka (Oryzias melastigma) by about 40% under the challenge of A. hydrophila, indicating its immunoprotective function. Taken together, both rLJ-hep2 and LJ-hep2(66–86) have good prospects to be used as potential antimicrobial agents in aquaculture and medicine in the future.

C-type lectin (CTL), a well-known immune-related molecule, has received more and more attention due to its diverse functions, especially its important role in development and host defense of vertebrate and invertebrate. Since the research on crab CTLs is still lack, we screened a new CTL homolog, named SpCTL6 from mud crab Scylla paramamosain. The full-length cDNA sequence of SpCTL6 was 738 bp with a 486 bp of ORF, and the deduced amino acids were 161 aa. SpCTL6 was predicted to have a 17 aa signal peptide and its mature peptide was 144 aa (MW 16.7 kDa) with pI value of 5.22. It had typical CTL structural characteristics, such as a single C-type lectin-like domain, 4 conserved cysteines, similar tertiary structure to that of vertebrate CTLs and a mutated Ca2+ binding motif Gln-Pro-Thr (QPT), clustering into the same branch as the crustacean CTLs. SpCTL6 was highly expressed in the entire zoeal larval stages and widely distributed in adult crab tissues with the highest transcription level in testis. During the molting process of juvenile crabs, the expression level of SpCTL6 was remarkably increased after molting. SpCTL6 could be significantly upregulated in two larval stages (Z1 and megalopa) and adult crab testis under immune challenges. Recombinant SpCTL6 (rSpCTL6) was successfully obtained from eukaryotic expression system. rSpCTL6 exhibited binding activity with PAMPs (LPS, lipoteichoic acid, peptidoglycan, and glucan) and had a broad spectrum bacterial agglutination activity in a Ca2+-dependent manner. In addition, rSpCTL6 could enhance the encapsulation activity of hemocytes and has no cytotoxic effect on hemocytes. Although rSpCTL6 had no bactericidal activity on Vibrio alginolyticus, rSpCTL6 treatment could significantly reduce the bacterial endotoxin level in vitro and greatly improved the survival of S. paramamosain under V. alginolyticus infection in vivo. The immunoprotective effect of rSpCTL6 might be due to the regulatory role of rSpCTL6 in immune-related genes and immunological parameters. Our study provides new information for understanding the immune defense of mud crabs and would facilitate the development of effective strategies for mud crab aquaculture disease control.

Hair cells in the inner ear and lateral lines are mechanosensitive receptor cells whose development and function are tightly regulated. Several transcription factors as well as splicing factors have been identified to play important roles in hair cell development, whereas the role of RNA stability in this process is poorly understood. In the present work, we report that RNA-binding motif protein 24a (Rbm24a) is indispensable for hair cell development in zebrafish. Rbm24a expression is detected in the inner ear as well as lateral line neuromasts. Albeit rbm24a deficient zebrafish do not survive beyond 9 days post fertilization (dpf) due to effects outside of the inner ear, rbm24a deficiency does not affect the early development of inner ear except for delayed otolith formation and semicircular canal fusion. However, hair cell development is severely affected and hair bundle is disorganized in rbm24a mutants. As a result, the auditory and vestibular function of rbm24a mutants are compromised. RNAseq analyses identified several Rbm24a-target mRNAs that are directly bound by Rbm24a and are dysregulated in rbm24a mutants. Among the identified Rbm24a-target genes, lrrc23, dfna5b, and smpx are particularly interesting as their dysregulation might contribute to the inner ear phenotypes in rbm24a mutants. In conclusion, our data suggest that Rbm24a affects hair cell development in zebrafish through regulating mRNA stability.

Overuse or abuse of antibiotics has undoubtedly accelerated the increasing prevalence of global antibiotic resistance crisis, and thus, people have been trying to explore approaches to decrease dosage of antibiotics or find new antibacterial agents for many years. Antimicrobial peptides (AMPs) are the ideal candidates that could kill pathogens and multidrug-resistant bacteria either alone or in combination with conventional antibiotics. In the study, the antimicrobial efficacy of mud crab Scylla paramamosain AMPs Sphistin and Sph12−38 in combination with eight selected antibiotics was evaluated using a clinical pathogen, Pseudomonas aeruginosa. It was interesting to note that the in vitro combination of rifampicin and azithromycin with Sphistin and Sph12−38 showed significant synergistic activity against P. aeruginosa. Moreover, an in vivo study was carried out using a mouse model challenged with P. aeruginosa, and the result showed that the combination of Sph12−38 with either rifampicin or azithromycin could significantly promote the healing of wounds and had the healing time shortened to 4–5 days compared with 7–8 days in control. The underlying mechanism might be due to the binding of Sphistin and Sph12−38 with P. aeruginosa lipopolysaccharides (LPS) and subsequent promotion of the intracellular uptake of rifampicin and azithromycin. Taken together, the significant synergistic antibacterial effect on P. aeruginosa in vitro and in vivo conferred by the combination of low dose of Sphistin and Sph12−38 with low dose of rifampicin and azithromycin would be beneficial for the control of antibiotic resistance and effective treatment of P. aeruginosa-infected diseases in the future.

Natural antimicrobial peptides (AMPs) are potential antibiotic alternatives. Marine crustaceans are thought to generate more powerful and various AMPs to protect themselves from infections caused by pathogenic microorganisms in their complex aquatic habitat, thus becoming one of the most promising sources of AMPs or other bioactive substances. In the study, a novel protein was identified as an interacting partner of male-specific AMP SCY2 in Scylla paramamosain and named scyreprocin. The recombinant product of scyreprocin (rScyreprocin) was successfully expressed in Escherichia coli. rScyreprocin exerted potent, broad-spectrum antifungal, antibacterial, and anti-biofilm activity (minimum inhibitory concentrations from 0.5 to 32 μM) through differential modes of action, including disruption of cell membrane integrity and induction of cell apoptosis, and has rapid bactericidal (in 0.5–2 h) and fungicidal (in 8–10 h) kinetics. In addition to its fungicidal activity against planktonic fungi, rScyreprocin also prevented the adhesion of fungal cells, inhibited biofilm formation, and eradicated the mature biofilms. Moreover, rScyreprocin showed a profound inhibitory effect on spore germination of Aspergillus spp. (minimum inhibitory concentrations from 4 to 8 μM). This peptide was not cytotoxic to murine and mammalian cells and could increase the survival rate of Oryzias melastigma under the challenge of Vibrio harveyi. Taken together, the novel AMP scyreprocin would be a promising alternative to antibiotics used in aquaculture and medicine.

Abstract Background Herpes simplex virus (HSV) has various presentations, depending on the patient’s immune status, age, and the route of transmission. In adults, HSV type 1 is found predominantly in the oral area, and HSV type 2 (HSV-2) is commonly found in the genital area. HSV-2 infection without genital lesions is uncommon. Herein we report a unique case of pharyngotonsillitis as an initial manifestation of disseminated HSV-2 infection without genital involvement. Case presentation A 46-year-old male was admitted to our hospital with a 1-week history of fever and sore throat. His past medical history included hypereosinophilic syndrome diagnosed at age 45 years. Physical examination revealed throat congestion, bilaterally enlarged tonsils with exudates, tender cervical lymphadenopathy in the left posterior triangle, and mild epigastric tenderness. The laboratory data demonstrated bicytopenia, elevated liver enzyme levels, and hyperferritinemia. A bone marrow smear showed hypocellular marrow with histiocytes and hemophagocytosis. The diagnosis of HLH was confirmed, and the patient was treated with methylprednisolone pulse therapy on days 1–3. On day 5, despite initial improvement of the fever and sore throat, multiple, new, small bullae developed on the patient’s face, trunk, and extremities. Additional testing showed that he was positive for HSV-specific immunoglobulin M and immunoglobulin G. Disseminated HSV infection was suspected, and intravenous acyclovir (10 mg/kg every 8 h) was begun. A subsequent direct antigen test of a bulla sample was positive for HSV-2. Moreover, tonsillar and esophageal biopsies revealed viral inclusion bodies. Immunohistochemical staining and a quantitative real-time polymerase chain reaction (PCR) assay confirmed the presence of HSV-2. Disseminated HSV-2 infection with multiple bullae, tonsillitis, esophagitis, and suspected hepatic involvement was diagnosed. After a 2-week course of intravenous acyclovir, his hematological status and liver function normalized, and his cutaneous skin lesions resolved. He was discharged on day 22 in good general health and continued taking oral valacyclovir for viral suppression due to his immunosuppressed status. Conclusion Disseminated HSV-2 infection should be considered as one of the differential diagnoses in patients with pharyngotonsillitis and impaired liver function of unknown etiology even if there are no genital lesions.

The utricle is one of the five sensory organs in the mammalian vestibular system, and while the utricle has a limited ability to repair itself, this is not sufficient for the recovery of vestibular function after hair cell (HC) loss induced by ototoxic drugs. In order to further explore the possible self-recovery mechanism of the adult mouse vestibular system, we established a reliable utricle epithelium injury model for studying the regeneration of HCs and examined the toxic effects of 3,3′-iminodiproprionitrile (IDPN) on the utricle in vivo in C57BL/6J mice, which is one of the most commonly used strains in inner ear research. This work focused on the epithelial cell loss, vestibular dysfunction, and spontaneous cell regeneration after IDPN administration. HC loss and supporting cell (SC) loss after IDPN treatment was dose-dependent and resulted in dysfunction of the vestibular system, as indicated by the swim test and the rotating vestibular ocular reflex (VOR) test. EdU-positive SCs were observed only in severely injured utricles wherein above 47% SCs were dead. No EdU-positive HCs were observed in either control or injured utricles. RT-qPCR showed transient upregulation of Hes5 and Hey1 and fluctuating upregulation of Axin2 and β-catenin after IDPN administration. We conclude that a single intraperitoneal injection of IDPN is a practical way to establish an injured utricle model in adult C57BL/6J mice in vivo. We observed activation of Notch and Wnt signaling during the limited spontaneous HC regeneration after vestibular sensory epithelium damage, and such signaling might act as the promoting factors for tissue self-repair in the inner ear.

Mutation patterns of DNA adducts, such as mutational spectra and signatures, are useful tools for diagnostic and prognostic purposes. Mutational spectra of carcinogens derive from three sources: adduct formation, replication bypass, and repair. Here, we consider the repair aspect of 1,N6-ethenoadenine (εA) by the 2-oxoglutarate/Fe(II)-dependent AlkB family enzymes. Specifically, we investigated εA repair across 16 possible sequence contexts (5′/3′ flanking base to εA varied as G/A/T/C). The results revealed that repair efficiency is altered according to sequence, enzyme, and strand context (ss- versus ds-DNA). The methods can be used to study other aspects of mutational spectra or other pathways of repair.

Zebrafish (Danio rerio) larvae are widely used to study otic functions because they possess all five typical vertebrate senses including hearing and balance. Powerful genetic tools and the transparent body of the embryo and larva also make zebrafish a unique vertebrate model to study otic development. Due to its small larval size and moisture requirement during experiments, accurately acquiring the vestibulo-ocular reflex (VOR) of zebrafish larva is challenging. In this report, a new VOR testing device has been developed for quantifying linear VOR (LVOR) in zebrafish larva, evoked by the head motion about the earth horizontal axis. The system has a newly designed larva-shaped chamber, by which live fish can be steadily held without anesthesia, and the system is more compact and easier to use than its predecessors. To demonstrate the efficacy of the system, the LVORs in wild-type (WT), dlx3b and dlx4b morphant zebrafish larvae were measured and the results showed that LVOR amplitudes were consistent with the morphological changes of otoliths induced by morpholino oligonucleotides (MO). Our study represents an important advance to obtain VOR and predict the vestibular conditions in zebrafish.

The discovery that an apparent forward-propagating otoacoustic emission (OAE) induced basilar membrane vibration has created a serious debate in the field of cochlear mechanics. The traditional theory predicts that OAE will propagate to the ear canal via a backward traveling wave on the basilar membrane, while the opponent theory proposed that the OAE will reach the ear canal via a compression wave. Although accepted by most people, the basic phenomenon of the backward traveling wave theory has not been experimentally demonstrated. In this study, for the first time, we showed the backward traveling wave by measuring the phase spectra of the basilar membrane vibration at multiple longitudinal locations of the basal turn of the cochlea. A local vibration source with a unique and precise location on the cochlear partition was created to avoid the ambiguity of the vibration source in most previous studies. We also measured the vibration pattern at different places of a mechanical cochlear model. A slow backward traveling wave pattern was demonstrated by the time-domain sequence of the measured data. In addition to the wave propagation study, a transmission line mathematical model was used to interpret why no tonotopicity was observed in the backward traveling wave.

Lgr5+ supporting cells (SCs) are enriched hair cell (HC) progenitors in the cochlea. Both in vitro and in vivo studies have shown that HC injury can spontaneously activate Lgr5+ progenitors to regenerate HCs in the neonatal mouse cochlea. Promoting HC regeneration requires the understanding of the mechanism of HC regeneration, and this requires knowledge of the key genes involved in HC injury-induced self-repair responses that promote the proliferation and differentiation of Lgr5+ progenitors. Here, as expected, we found that neomycin-treated Lgr5+ progenitors (NLPs) had significantly greater HC regeneration ability, and greater but not significant proliferation ability compared to untreated Lgr5+ progenitors (ULPs) in response to neomycin exposure. Next, we used RNA-seq analysis to determine the differences in the gene-expression profiles between the transcriptomes of NLPs and ULPs from the neonatal mouse cochlea. We first analyzed the genes that were enriched and differentially expressed in NLPs and ULPs and then analyzed the cell cycle genes, the transcription factors, and the signaling pathway genes that might regulate the proliferation and differentiation of Lgr5+ progenitors. We found 9 cell cycle genes, 88 transcription factors, 8 microRNAs, and 16 cell-signaling pathway genes that were significantly upregulated or downregulated after neomycin injury in NLPs. Lastly, we constructed a protein-protein interaction network to show the interaction and connections of genes that are differentially expressed in NLPs and ULPs. This study has identified the genes that might regulate the proliferation and HC regeneration of Lgr5+ progenitors after neomycin injury, and investigations into the roles and mechanisms of these genes in the cochlea should be performed in the future to identify potential therapeutic targets for HC regeneration.

Cochlear supporting cells (SCs) have been shown to be a promising resource for hair cell (HC) regeneration in the neonatal mouse cochlea. Previous studies have reported that Lgr5+ SCs can regenerate HCs both in vitro and in vivo and thus are considered to be inner ear progenitor cells. Lgr5+ progenitors are able to regenerate more HCs than Lgr5- SCs, and it is important to understand the mechanism behind the proliferation and HC regeneration of these progenitors. Here, we isolated Lgr5+ progenitors and Lgr5- SCs from Lgr5-EGFP-CreERT2/Sox2-CreERT2/Rosa26-tdTomato mice via flow cytometry. As expected, we found that Lgr5+ progenitors had significantly higher proliferation and HC regeneration ability than Lgr5- SCs. Next, we performed RNA-Seq to determine the gene expression profiles of Lgr5+ progenitors and Lgr5- SCs. We analyzed the genes that were enriched and differentially expressed in Lgr5+ progenitors and Lgr5- SCs, and we found 8 cell cycle genes, 9 transcription factors, and 24 cell signaling pathway genes that were uniquely expressed in one population but not the other. Last, we made a protein–protein interaction network to further analyze the role of these differentially expressed genes. In conclusion, we present a set of genes that might regulate the proliferation and HC regeneration ability of Lgr5+ progenitors, and these might serve as potential new therapeutic targets for HC regeneration.

Bulky organic carcinogens are activated in vivo and subsequently react with nucleobases of cellular DNA to produce adducts. Some of these DNA adducts exist in multiple conformations that are slowly interconverted to one another. Different conformations have been implicated in different mutagenic and repair outcomes. However, studies on the conformation-specific inhibition of replication, which is more relevant to cell survival, are scarce, presumably due to the structural dynamics of DNA lesions at the replication fork. It is difficult to capture the exact nature of replication inhibition by existing end-point assays, which usually detect either the ensemble of consequences of all the conformers or the culmination of all cellular behaviors, such as mutagenicity or survival rate. We previously reported very unusual sequence-dependent conformational heterogeneities involving FABP-modified DNA under different sequence contexts (TG1*G2T [67%B:33%S] and TG1G2*T [100%B], G*, N-(2′-deoxyguanosin-8-yl)-4′-fluoro-4-aminobiphenyl) (Cai et al. Nucleic Acids Research, 46, 6356–6370 (2018)). In the present study, we attempted to correlate the in vitro inhibition of polymerase activity to different conformations from a single FABP-modified DNA lesion. We utilized a combination of surface plasmon resonance (SPR) and HPLC-based steady-state kinetics to reveal the differences in terms of binding affinity and inhibition with polymerase between these two conformers (67%B:33%S and 100%B).

Ocean Acidification (OA) is known to affect various aspects of physiological performances of diatoms, but little is known about the underlining molecular mechanisms involved. Here, we show that in the model diatom Phaeodactylum tricornutum, the expression of key genes associated with photosynthetic light harvesting as well as those encoding Rubisco, carbonic anhydrase, NADH dehydrogenase and nitrite reductase, are modulated by OA (1000 μatm, pHnbs 7.83). Growth and photosynthetic carbon fixation were enhanced by elevated CO2. OA treatment decreased the expression of β-carbonic anhydrase (β-ca), which functions in balancing intracellular carbonate chemistry and the CO2 concentrating mechanism (CCM). The expression of the genes encoding fucoxanthin chlorophyll a/c protein (lhcf type (fcp)), mitochondrial ATP synthase (mtATP), ribulose-1, 5-bisphosphate carboxylase/oxygenase large subunit gene (rbcl) and NADH dehydrogenase subunit 2 (ndh2), were down-regulated during the first four days (< 8 generations) after the cells were transferred from LC (cells grown under ambient air condition; 390 μatm; pHnbs 8.19) to OA conditions, with no significant difference between LC and HC treatments with the time elapsed. The expression of nitrite reductase (nir) was up-regulated by the OA treatment. Additionally, the genes for these proteins (NiR, FCP, mtATP synthase, β-CA) showed diel expression patterns. It appeared that the enhanced photosynthetic and growth rates under OA could be attributed to stimulated nitrogen assimilation, increased CO2 availability or saved energy from down-regulation of the CCM and consequently lowered cost of protein synthesis versus that of non-nitrogenous cell components.

The zebrafish has become an established model organism for the study of hearing and balance systems in the past two decades. The classical approach to examine hair cells is to use dye to conduct selective staining, which shows the number and morphology of hair cells but does not reveal their function. Startle response is a behavior closely related to the auditory function of hair cells; therefore it can be used to measure the function of hair cells. In this study, we developed a device to measure the startle response of zebrafish larvae. By applying various levels of stimulus, it showed that the system can discern a 10 dB difference. The hair cell in zebrafish can regenerate after damage due to noise exposure or drug treatment. With this device, we measured the startle response of zebrafish larvae during and after drug treatment. The results show a similar trend to the classical hair cell staining method. The startle response was reduced with drug treatment and recovered after removal of the drug. Together it demonstrated the capability of this behavioral assay in evaluating the hair cell functions of fish larvae and its potential as a high-throughput screening tool for auditory-related gene and drug discovery.

Goal. To establish a reliable instrumental system for synchronized reactivation of a unilaterally paralyzed vocal fold and evaluate its functional feasibility. Methods. Unilateral vocal fold paralysis model was induced by destruction of the left recurrent laryngeal nerve (RLN) in anesthetized dogs. With a micro controller-based electronic system, electromyography (EMG) signals from cricothyroid (CT) muscle on the ipsilateral side were recorded and used to trigger pacing of paralyzed vocalis muscles. The dynamic movement of vocal folds was continuously monitored using an endoscope, and the opening and closing of the glottis were quantified with customized imaging processing software. Results. The recorded video images showed that left side vocal fold was obviously paralyzed after destructing the RLN. Using the pacing system with feedback triggering EMG signals from the ipsilateral CT muscle, the paralyzed vocal fold was successfully reactivated, and its movement was shown to be synchronized with the healthy side. Significance. The developed unilateral laryngeal pacing system triggered by EMG from the ipsilateral side CT muscle could be successfully used in unilateral vocal fold paralysis with the advantage of avoiding disturbance to the healthy side muscles.

SpHyastatin was first identified as a new cationic antimicrobial peptide (AMP) in hemocytes of the mud crab Scylla paramamosain. Based on the amino acid sequences deduced, it was predicted that this peptide was composed of two different functional domains, a proline-rich domain (PRD) and a cysteine-rich domain (CRD). The recombinant product of SpHyastatin displayed potent antimicrobial activities against the human pathogen Staphylococcus aureus and the aquatic animal pathogens Aeromonas hydrophila and Pseudomonas fluorescens. Compared with the CRD of SpHyastatin, the PRD presented better antimicrobial and chitin binding activities, but both regions were essential for allowing SpHyastatin complete antimicrobial activity. The binding properties of SpHyastatin to different microbial surface molecules suggested that this might be an initial and crucial step for performing its antimicrobial activities. Evaluated using propidium iodide uptake assays and scanning electron microscopy images, the antimicrobial mechanism of SpHyastatin was found to be prone to disrupt cell membrane integrity. Interestingly, SpHyastatin exerted its role specifically on the surface of S. aureus and Pichia pastoris whereas it directly killed P. fluorescens through simultaneous targeting the membrane and the cytoplasm, indicating that SpHyastatin could use different antimicrobial mechanisms to kill different species of microbes. As expected, the recombinant SpHyastatin increased the survival rate of crabs challenged with Vibrio parahaemolyticus. In addition, SpHyastatin could modulate some V. parahaemolyticus-responsive genes in S. paramamosain.

A phase-sensitive fluidic biosensor based on a spectral-domain low-coherence interferometer is presented in this paper. With a fiber optic probe employing the common-path interferometric configuration, subnanometric changes in thickness of the molecular layers can be detected through phase analysis of the acquired interference signal from the sensor surface. Advantages of this biosensor include its picometer-scale thickness sensitivity, 13.9-ms time response, and tolerance to the fluctuation in concentration of the target solution. The capabilities of this biosensor in monitoring specific molecular binding and recognizing specific molecular was successfully demonstrated by using the reactions between the molecules of protein A and IgG. The calculated minimum detectable concentration of IgG is 0.11 µg/mL.

The resting membrane potential (RP) of vascular smooth muscle cells (VSMCs) is a major determinant of cytosolic calcium concentration and vascular tone. The heterogeneity of RPs and its underlying mechanism among different vascular beds remain poorly understood. We compared the RPs and vasomotion properties between the guinea pig spiral modiolar artery (SMA), brain arterioles (BA) and mesenteric arteries (MA). We found: 1) RPs showed a robust bimodal distribution peaked at -76 and -40 mV evenly in the SMA, unevenly at -77 and -51 mV in the BA and ~-71 and -52 mV in the MA. Ba(2+) 0.1 mM eliminated their high RP peaks ~-75 mV. 2) Cells with low RP (~-45 mV) hyperpolarized in response to 10 mM extracellular K(+), while cells with a high RP depolarized, and cells with intermediate RP (~-58 mV) displayed an initial hyperpolarization followed by prolonged depolarization. Moderate high K(+) typically induced dilation, constriction and a dilation followed by constriction in the SMA, MA and BA, respectively. 3) Boltzmann-fit analysis of the Ba(2+)-sensitive inward rectifier K(+) (Kir) whole-cell current showed that the maximum Kir conductance density significantly differed among the vessels, and the half-activation voltage was significantly more negative in the MA. 4) Corresponding to the whole-cell data, computational modeling simulated the three RP distribution patterns and the dynamics of RP changes obtained experimentally, including the regenerative swift shifts between the two RP levels after reaching a threshold. 5) Molecular works revealed strong Kir2.1 and Kir2.2 transcripts and Kir2.1 immunolabeling in all 3 vessels, while Kir2.3 and Kir2.4 transcript levels varied. We conclude that a dense expression of functional Kir2.X channels underlies the more negative RPs in endothelial cells and a subset of VSMC in these arterioles, and the heterogeneous Kir function is primarily responsible for the distinct bimodal RPs among these arterioles. The fast Kir-based regenerative shifts between two RP states could form a critical mechanism for conduction/spread of vasomotion along the arteriole axis.

Pathogens can enter their host cells by way of endocytosis in which the membrane lipid raft gene flotillins are probably involved in the invasion process and this is an important way to cause infection. In this study, a new gene SpFLT-1 was identified in Scylla paramamosain, which shared high identity with the flotillin-1 of other species. The SpFLT-1 gene was widely distributed in tissues and showed the highest level of mRNA transcripts in the hemocytes. This gene might be a maternal gene based on the evident results that it was highly expressed in maternal ovaries and in the early developmental stages of the zygote and early embryo stage whereas it gradually decreased in zoea 1. SpFLT-1 positively responded to the challenge of Vibrio alginolyticus with a significantly increased level of mRNA expression in the hemocytes and gills at 3 hours post infection (hpi). The SpFLT-1 protein was detected densely in the same fraction layer where the Vibrio protein was most present in the hemocytes and gills at 3 hpi. Furthermore, it was found that the expression of SpFLT-1 decreased to the base level following disappearance of the Vibrio protein at 6 hpi in the gills. Silencing SpFLT-1 inhibited the endocytosis rate of V. alginolyticus but overexpression of the gene could facilitate bacterial entry into the epithelioma papulosum cyprinid cells. Our study indicated that SpFLT-1 may act as a key protein involved in the process of bacterial infection and this sheds light on clarifying the pathogenesis of pathogens infecting S. paramamosain.

Mammalian hearing is refined by amplification of the sound-evoked vibration of the cochlear partition. This amplification is at least partly due to forces produced by protein motors residing in the cylindrical body of the outer hair cell. To transmit power to the cochlear partition, it is required that the outer hair cells dynamically change their length, in addition to generating force. These length changes, which have not previously been measured in vivo, must be correctly timed with the acoustic stimulus to produce amplification.Using in vivo optical coherence tomography, we demonstrate that outer hair cells in living guinea pigs have length changes with unexpected timing and magnitudes that depend on the stimulus level in the sensitive cochlea.The level-dependent length change is a necessary condition for directly validating that power is expended by the active process presumed to underlie normal hearing.