Screening a neurotransmitter-receptor-related inhibitor library identifies clomipramine HCl as a potential antiviral compound against Japanese encephalitis virus.

Background: Japanese encephalitis virus (JEV) is a leading cause of viral encephalitis worldwide. JEV exhibits significant neuroinvasiveness and neurotoxicity, resulting in considerable damage to the nervous system. Japanese encephalitis is associated with high morbidity and mortality rate, seriously harming both human health and livestock production. The current lack of specific antiviral drugs means that the development of new therapeutic agents for JEV has become urgent. Methods: Anti-JEV drugs were screened from 111 inhibitors of neurotransmitter receptor-related molecules by high content technology. The antiviral effects of clomipramine HCl were evaluated through plaque assay, real-time quantitative PCR, immunofluorescence assay and western blotting assay. Bioinformatic tools were utilized to cluster the altered signaling pathway members after clomipramine HCl treatment. Finally, the anti-JEV mechanism was deeply resolved in vivo via such molecular biology and virological detection techniques. Results: In this study, we screened nine compounds with significant anti-JEV activity, of which clomipramine HCl demonstrated the most potent antiviral effect and exhibited dose-dependent activity. Mechanistically, clomipramine HCl may activate endoplasmic reticulum stress and modulate the unfolded protein response, thus inhibiting the assembly stage of JEV infection. Conclusion: This study highlights the importance of clomipramine HCl as a promising approach for JEV infection protection, which may lead to new host-directed antiviral approaches to such mosquito-borne viruses.

Japanese encephalitis virus NS1 and NS1' protein disrupts the blood-brain barrier through macrophage migration inhibitory factor-mediated autophagy.

Flaviviruses in the Japanese encephalitis virus (JEV) serogroup, such as JEV, West Nile virus, and St. Louis encephalitis virus, can cause severe neurological diseases. The nonstructural protein 1 (NS1) is a multifunctional protein of flavivirus that can be secreted by infected cells and circulate in the host bloodstream. NS1' is an additional form of NS1 protein with 52 amino acids extension at its carboxy-terminal and is produced exclusively by flaviviruses in the JEV serogroup. In this study, we demonstrated that the secreted form of both NS1 and NS1' can disrupt the blood-brain barrier (BBB) of mice, with NS1' exhibiting a stronger effect. Using the in vitro BBB model, we found that treatment of soluble recombinant JEV NS1 or NS1' protein increases the permeability of human brain microvascular endothelial cells (hBMECs) and leads to the degradation of tight junction proteins through the autophagy-lysosomal pathway. Consistently, NS1' protein exhibited a more pronounced effect compared to NS1 in these cellular processes. Further research revealed that the increased expression of macrophage migration inhibitory factor (MIF) is responsible for triggering autophagy after NS1 or NS1' treatment in hBMECs. In addition, TLR4 and NF-κB signaling was found to be involved in the activation of MIF transcription. Moreover, administering the MIF inhibitor has been shown to decrease viral loads and mitigate inflammation in the brains of mice infected with JEV. This research offers a novel perspective on the pathogenesis of JEV. In addition, the stronger effect of NS1' on disrupting the BBB compared to NS1 enhances our understanding of the mechanism by which flaviviruses in the JEV serogroup exhibit neurotropism.IMPORTANCEJapanese encephalitis (JE) is a significant viral encephalitis worldwide, caused by the JE virus (JEV). In some patients, the virus cannot be cleared in time, leading to the breach of the blood-brain barrier (BBB) and invasion of the central nervous system. This invasion may result in cognitive impairment, behavioral disturbances, and even death in both humans and animals. However, the mechanism by which JEV crosses the BBB remains unclear. Previous studies have shown that the flavivirus NS1 protein plays an important role in causing endothelial dysfunction. The NS1' protein is an elongated form of NS1 protein that is particularly produced by flaviviruses in the JEV serogroup. This study revealed that both the secreted NS1 and NS1' of JEV can disrupt the BBB by breaking down tight junction proteins through the autophagy-lysosomal pathway, and NS1' is found to have a stronger effect compared to NS1 in this process. In addition, JEV NS1 and NS1' can stimulate the expression of MIF, which triggers autophagy via the ERK signaling pathway, leading to damage to BBB. Our findings reveal a new function of JEV NS1 and NS1' in the disruption of BBB, thereby providing the potential therapeutic target for JE.

Mediation of the association between screen time and suicidality by overweight/obesity and perceived overweight: results from the youth risk behavior surveillance system of the United States.

Aim: Adolescent suicide is a major public health concern, and modifiable risk factors associated with adolescent suicide remain poorly understood. This study aimed to assess the association between screen time and overweight/obesity and self-perceived overweigh and suicidality in adolescents. Methods: Adolescents from the United States Youth Risk Behavior Surveillance System (YRBSS) between 2013 and 2019 were included in this cross-sectional study. The outcome was suicidality, including considered suicide, made a suicide plan, attempted suicide, and injurious suicide attempt. Multivariable logistic regression model was used to investigate the associations between screen time, overweight/obesity, self-perceived overweight, and suicidality, and expressed as odds ratio (OR) and 95% confidence interval (CI). Mediation analysis was used to explore the role of overweight/obesity and self-perceived overweight on the association between screen time and suicidality. Results: A total of 30,731 adolescents were included, of which 6,350 (20.65%) had suicidality, including 5,361 (17.45%) with considered suicide, 4,432 (14.42%) with made a suicide plan, 2,300 (7.45%) with attempted suicide, and 677 (2.21%) with injurious suicide attempt. Adolescents with screen time ≥3h were related to higher odds of suicidality (OR=1.35, 95%CI: 1.23-1.46), overweight/obesity (OR=1.27, 95%CI: 1.19-1.38), and self-perceived overweight (OR=1.38, 95%CI: 1.30-1.48) after adjusting confounders. Adolescents with overweight/obesity (OR=1.30, 95%CI: 1.19-1.43) and self-perceived overweight (OR=1.54, 95%CI: 1.39-1.70) were associated with higher odds of suicidality. The association between screen time and suicidality was 4.67% mediated by overweight/obesity and 9.66% mediated by self-perceived overweight. Moreover, the mediating role of overweight/obesity was observed only in females, whereas there were no sex differences in the mediating effect of self-perceived overweight. Conclusion: Both overweight/obesity and self-perceived overweight mediated the association between screen time and suicidality.

Correlation of the serum cell division cycle 42 with CD4+ T cell subsets and in-hospital mortality in Stanford type B aortic dissection patients.

Objective: Cell division cycle 42 (CDC42) regulates CD4+ T-cell differentiation and participates in vascular stiffness and atherosclerosis and is involved in the progression of Stanford type B aortic dissection (TBAD). This study aimed to explore the correlation between serum CDC42 level and CD4+ T cell subsets and in-hospital mortality in TBAD patients. Methods: Serum CDC42 and peripheral blood T-helper (Th) 1, Th2, and Th17 cells were detected in 127 TBAD patients by enzyme-linked immunosorbent assay and flow cytometry, respectively. Serum CDC42 was also quantified in 30 healthy controls. Results: Serum CDC42 was decreased in TBAD patients vs. healthy controls (median [interquartile range (IQR)]: 418.0 (228.0-761.0) pg/ml vs. 992.0 (716.3-1,445.8) pg/ml, P < 0.001). In TBAD patients, serum CDC42 was negatively correlated with Th17 cells (P = 0.001), but not Th1 (P = 0.130) or Th2 cells (P = 0.098). Seven (5.5%) patients experienced in-hospital mortality. Serum CDC42 was reduced in patients who experienced in-hospital mortality vs. those who did not (median (IQR): 191.0 (145.0-345.0) pg/ml vs. 451.5 (298.3-766.8) pg/ml, P = 0.006). By receiver operating characteristic analysis, serum CDC42 showed a good ability for estimating in-hospital mortality [area under curve = 0.809, 95% confidence interval (CI) = 0.662-0.956]. By the multivariate logistic regression analysis, elevated serum CDC42 [odd ratio (OR) = 0.994, 95% CI = 0.998-1.000, P = 0.043] was independently correlated with lower risk of in-hospital mortality, while higher age (OR = 1.157, 95% CI = 1.017-1.316, P = 0.027) was an independent factor for increased risk of in-hospital mortality. Conclusion: Serum CDC42 negatively associates with Th17 cells and is independently correlated with decreased in-hospital mortality risk in TBAD patients.

Single-cell RNA sequencing reveals the immune features and viral tropism in the central nervous system of mice infected with Japanese encephalitis virus.

Japanese encephalitis virus (JEV) is a neurotropic pathogen that causes lethal encephalitis. The high susceptibility and massive proliferation of JEV in neurons lead to extensive neuronal damage and inflammation within the central nervous system. Despite extensive research on JEV pathogenesis, the effect of JEV on the cellular composition and viral tropism towards distinct neuronal subtypes in the brain is still not well comprehended. To address these issues, we performed single-cell RNA sequencing (scRNA-seq) on cells isolated from the JEV-highly infected regions of mouse brain. We obtained 88,000 single cells and identified 34 clusters representing 10 major cell types. The scRNA-seq results revealed an increasing amount of activated microglia cells and infiltrating immune cells, including monocytes & macrophages, T cells, and natural killer cells, which were associated with the severity of symptoms. Additionally, we observed enhanced communication between individual cells and significant ligand-receptor pairs related to tight junctions, chemokines and antigen-presenting molecules upon JEV infection, suggesting an upregulation of endothelial permeability, inflammation and antiviral response. Moreover, we identified that Baiap2-positive neurons were highly susceptible to JEV. Our findings provide valuable clues for understanding the mechanism of JEV induced neuro-damage and inflammation as well as developing therapies for Japanese encephalitis.

Distribution and correlation of iron oxidizers and carbon-fixing microbial communities in natural wetlands.

Most microaerophilic Fe(II)-oxidizing bacteria (mFeOB) belonging to the family Gallionellaceae are autotrophic microorganisms that can use inorganic carbon to drive carbon sequestration in wetlands. However, the relationship between microorganisms involved in Fe and C cycling is not well understood. Here, soil samples were collected from different wetlands to explore the distribution and correlation of Gallionella-related mFeOB and carbon-fixing microorganisms containing cbbL and cbbM genes. A significant positive correlation was found between the abundances of mFeOB and the cbbL gene, as well as a highly significant positive correlation between the abundances of mFeOB and the cbbM gene, indicating the distribution of mFeOB in co-occurrence with carbon-fixing microorganisms in wetlands. The mFeOB were mainly dominated by Sideroxydans lithotrophicus ES-1 and Gallionella capsiferriformans ES-2 in all wetland soils. The structures of the carbon-fixing microbial communities were similar in these wetlands, mainly consisting of Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria. The extractable Fe(II) concentrations affected the community composition of mFeOB, resulting in a significant difference in the relative abundances of the dominant FeOB. The main factors affecting cbbL-related microbial communities were dissolved inorganic carbon and oxygen, soil redox potential, and sodium acetate-extracted Fe(II). The composition of cbbM-related microbial communities was mainly affected by acetate-extracted Fe(II) and soil redox potential. In addition, the positive correlation between these functional microorganisms suggests that they play a synergistic role in Fe(II) oxidation and carbon fixation in wetland soil ecosystems. Our results suggest a cryptic relationship between mFeOB and carbon-fixing microorganisms in wetlands and that the microbial community structure can be effectively altered by regulating their physicochemical properties, thus affecting the capacity of carbon sequestration.

Wrinkling and Strengthening Behaviors in the Two-Layer-Sheet Hot-Forming-Quenching Integrated Process for an Al-Cu-Mg-Alloy Thin-Walled Curved-Surface Shell.

The thin-walled curved-surface component is an important structural element in aerospace. Wrinkling, springback and thermal distortion occur easily when forming these components. To form thin-walled components with high precision and strength, a two-layer-sheet hot-forming-quenching integrated process was proposed, in which wrinkling is prevented by thickening the upper sheet and springback is reduced by solution and die quenching. Selecting an appropriate upper sheet is crucial to suppress wrinkling and accomplish effective die quenching. The effect of the upper sheet on the wrinkling and strengthening behaviors of an Al-Cu-Mg-alloy melon-petal shell was thus studied in detail. The anti-wrinkle mechanism was analyzed through numerical simulation. The forming quality, including forming precision, deformation uniformity and strength, were further evaluated. The wrinkle gradually decreased with the increasing thickness of the upper sheet, resulting from the depressed compressive stress at the edge of the target sheet. A defect-free specimen with a smooth surface was finally formed when the thickness of the upper sheet reached three times that of the target sheet. The profile deviation was ±0.5 mm. Excellent thickness uniformity in a specimen can be obtained with a maximum thinning rate of 6%. The full strength, ranging from 455 to 466 MPa, can be obtained in all regions of the specimen, indicating that effective strengthening can be accomplished with the two-layer-sheet die quenching. The results indicated that high forming quality and full strength can be obtained in a two-layer-sheet hot-forming-quenching integrated process. This research has great potential for engineering applications using aluminum-alloy curved-surface thin-walled components.

H3K27me3 of Rnf19a promotes neuroinflammatory response during Japanese encephalitis virus infection.

Histone methylation is an important epigenetic modification that affects various biological processes, including the inflammatory response. In this study, we found that infection with Japanese encephalitis virus (JEV) leads to an increase in H3K27me3 in BV2 microglial cell line, primary mouse microglia and mouse brain. Inhibition of H3K27me3 modification through EZH2 knockdown and treatment with EZH2 inhibitor significantly reduces the production of pro-inflammatory cytokines during JEV infection, which suggests that H3K27me3 modification plays a crucial role in the neuroinflammatory response caused by JEV infection. The chromatin immunoprecipitation-sequencing (ChIP-sequencing) assay revealed an increase in H3K27me3 modification of E3 ubiquitin ligases Rnf19a following JEV infection, which leads to downregulation of Rnf19a expression. Furthermore, the results showed that Rnf19a negatively regulates the neuroinflammatory response induced by JEV. This is achieved through the degradation of RIG-I by mediating its ubiquitination. In conclusion, our findings reveal a novel mechanism by which JEV triggers extensive neuroinflammation from an epigenetic perspective.

Biological determinants perpetuating the transmission dynamics of mosquito-borne flaviviruses.

Mosquito-borne flaviviruses present a major public health concern. Their transmission is sustained in a cycle between mosquitoes and vertebrate hosts. However, the dynamicity of the virus-mosquito-host triad has not been completely understood. Herein, we discussed determinants of viral, vertebrate host, and mosquito origins that ensure virus adaptability and transmission in the natural environment. In particular, we provided insights into how proteins and RNAs of flaviviruses, blood parameters and odours of humans, and gut microbiota, saliva, and hormones of mosquitoes coordinate with each other to perpetuate the virus transmission cycle. A better knowledge of mechanisms permitting flaviviruses dissemination in nature can provide opportunities for establishing new virus-controlling strategies and could guide future epidemic and pandemic preparedness.

The Influence of the Gut Microbiota on Alzheimer's Disease: A Narrative Review.

Alzheimer's disease (AD) is a common neurodegenerative disease that tends to occur in the elderly. The main symptom is hypomnesia. More and more older people are suffering from this disease worldwide. By 2050, 152 million people worldwide are expected to have AD. It is thought that the aggregation of amyloid-beta peptides and hyper-phosphorylated tau tangles contribute to AD. The microbiota-gut-brain (MGB) axis appears as a new concept. The MGB axis is a collection of microbial molecules produced in the gastrointestinal tract that influence the physiological function of the brain. In this review, we discuss how the gut microbiota (GM) and its metabolites affect AD in different ways. Dysregulation of the GM has been shown to be involved in various mechanisms involved in memory and learning functions. We review the current literature on the role of the entero-brain axis in the pathogenesis of AD and its potential role as a future therapeutic target in the treatment and/or prevention of AD.

The Variations of Leaf δ13C and Its Response to Environmental Changes of Arbuscular and Ectomycorrhizal Plants Depend on Life Forms.

Arbuscular mycorrhiza (AM) and ectomycorrhiza (ECM) are the two most common mycorrhizal types and are paid the most attention to, playing a vital common but differentiated function in terrestrial ecosystems. The leaf carbon isotope ratio (δ13C) is an important factor in understanding the relationship between plants and the environment. In this study, a new database was established on leaf δ13C between AM and ECM plants based on the published data set of leaf δ13C in China's C3 terrestrial plants, which involved 1163 observations. The results showed that the differences in leaf δ13C between AM and ECM plants related closely to life forms. Leaf δ13C of ECM plants was higher than that of AM plants in trees, which was mainly led by the group of evergreen trees. The responses of leaf δ13C to environmental changes were varied between AM and ECM plants. Among the four life forms, leaf δ13C of ECM plants decreased more rapidly than that of AM plants, with an increase of longitude, except for deciduous trees. In terms of the sensitivity of leaf δ13C to temperature changes, AM plants were higher than ECM plants in the other three life forms, although there was no significant difference in evergreen trees. For the response to water conditions, the leaf δ13C of ECM plants was more sensitive than that of AM plants in all life forms, except evergreen and deciduous trees. This study laid a foundation for further understanding the role of mycorrhiza in the relationship between plants and the environment.

Corresponding risk factors between cognitive impairment and type 1 diabetes mellitus: A narrative review.

Type 1 diabetes mellitus (T1DM) is a type of diabetes caused by the destruction of pancreatic ß cells and the absolute lack of insulin secretion. T1DM usually starts in adolescence or develops directly as a severe disease state of ketoacidosis. T1DM and its complications make many people suffer and have psychological problems, which make us have to pay more attention to the prevention and early control of T1DM. Cognitive impairment (CI) is one of the major complications of T1DM. It can further develop into Alzheimer's disease, which can seriously affect the quality of life of the elderly. Furthermore, the relationship between T1DM and CI is unclear. Hence, we conducted a narrative review of the existing literature through a PubMed search. We summarized some risk factors that may be associated with the cognitive changes in T1DM patients, including onset age and duration, education and gender, glycemic states, microvascular complications, glycemic control, neuropsychology and emotion, intestinal flora, dyslipidemia, sleep quality. We aimed to provide some content related to CI in T1DM, and hoped that it could play a role in early prediction and treatment to reduce the prevalence.

Excessive immunosuppression by regulatory T cells antagonizes T cell response to schistosome infection in PD-1-deficient mice.

Schistosomiasis is caused by parasitic flatworms known as schistosomes and affects over 200 million people worldwide. Prevention of T cell exhaustion by blockade of PD-1 results in clinical benefits to cancer patients and clearance of viral infections, however it remains largely unknown whether loss of PD-1 could prevent or cure schistosomiasis in susceptible mice. In this study, we found that S. japonicum infection dramatically induced PD-1 expression in T cells of the liver where the parasites chronically inhabit and elicit deadly inflammation. Even in mice infected by non-egg-producing unisex parasites, we still observed potent induction of PD-1 in liver T cells of C57BL/6 mice following S. japonicum infection. To determine the function of PD-1 in schistosomiasis, we generated PD-1-deficient mice by CRISPR/Cas9 and found that loss of PD-1 markedly increased T cell count in the liver and spleen of infected mice. IL-4 secreting Th2 cells were significantly decreased in the infected PD-1-deficient mice whereas IFN-γ secreting CD4+ and CD8+ T cells were markedly increased. Surprisingly, such beneficial changes of T cell response did not result in eradication of parasites or in lowering the pathogen burden. In further experiments, we found that loss of PD-1 resulted in both beneficial T cell responses and amplification of regulatory T cells that prevented PD-1-deficient T cells from unleashing anti-parasite activity. Moreover, such PD-1-deficient Tregs exert excessive immunosuppression and express larger amounts of adenosine receptors CD39 and CD73 that are crucial for Treg-mediated immunosuppression. Our experimental results have elucidated the function of PD-1 in schistosomiasis and provide novel insights into prevention and treatment of schistosomiasis on the basis of modulating host adaptive immunity.

Autophagy inhibitors alleviate Japanese encephalitis virus-induced cerebral inflammation in mice.

Japanese encephalitis (JE) is a zoonotic epidemic disease caused by Japanese encephalitis virus (JEV), and currently, no medicines are available to treat this disease. Autophagy modulators play an important role in the treatment of tumors, heart disease, and some viral diseases. The aim of this study was to investigate the effects of autophagy modulators on JEV infection and the host response in mice. The experimental mice were grouped as follows: DMEM (control), JEV, JEV+rapamycin (JEV+Rapa), JEV+wortmannin (JEV+Wort), JEV+chloroquine (JEV+CQ), Rapa, Wort, and CQ. The control group was treated with DMEM. The mice in other groups were infected with 105 PFU of JEV, and Rapa, Wort, and CQ were administered 2 h prior to JEV challenge and then administered daily for 10 consecutive days. All mice were monitored for neurological signs and survival. The damage of subcellular structures in the mouse brain was evaluated by transmission electron microscopy. The distribution of virus in the mouse brain was determined by RNAScope staining and immunohistochemical staining. The neuroinflammatory responses in the brain were examined via quantitative real-time PCR, and the signal pathways involved in neuroinflammation were identified by Western blot. The mice in the JEV+Wort and JEV+CQ groups showed milder neurological symptoms, less damage to the mitochondria in the brain tissue, and a higher survival rate than those in the JEV+Rapa and JEV groups. Compared with the JEV+Rapa and JEV groups, the distribution of JEV in the brain of mice in the JEV+Wort and JEV+CQ groups was lower, and the inflammatory response was weaker. No significant difference was observed in the expression of the PI3K/AKT/NF-κB pathway in mouse brain among the different groups. Our study suggests that the autophagy inhibitors Wort and CQ reduce JEV infection and weaken the inflammatory response, which does not depend on the PI3K/AKT/NF-κB pathway in mouse brain.

Arbuscular mycorrhizal fungi increase crop yields by improving biomass under rainfed condition: a meta-analysis.

BACKGROUND: Rainfed agriculture plays key role in ensuring food security and maintain ecological balance. Especially in developing areas, most grain food are produced rainfed agricultural ecosystem. Therefore, the increase of crop yields in rainfed agricultural ecosystem becomes vital as well as ensuring global food security. METHODS: The potential roles of arbuscular mycorrhizal fungi (AMF) in improving crop yields under rainfed condition were explored based on 546 pairs of observations published from 1950 to 2021. RESULTS: AMF inoculation increased 23.0% crop yields based on 13 popular crops under rainfed condition. Not only was crop biomass of shoot and root increased 24.2% and 29.6% by AMF inocula, respectively but also seed number and pod/fruit number per plant were enhanced markedly. Further, the effect of AMF on crop yields depended on different crop groups. AMF improved more yield of N-fixing crops than non-N-fixing crops. The effect of AMF changed between grain and non-grain crops with the effect size of 0.216 and 0.352, respectively. AMF inoculation enhances stress resistance and photosynthesis of host crop in rainfed agriculture. CONCLUSION: AMF increased crop yields by enhancing shoot biomass due to the improvement of plant nutrition, photosynthesis, and stress resistance in rainfed field. Our findings provide a new view for understanding the sustainable productivity in rainfed agroecosystem, which enriched the theory of AMF functional diversity. This study provided a theoretical and technical way for sustainable production under rainfed agriculture.

One-Step Microfluidic Fabrication of Multi-Responsive Liposomes for Targeted Delivery of Doxorubicin Synergism with Photothermal Effect.

INTRODUCTION: Cancer of the bladder is one of the most common and life-threatening. Compared with traditional delivery methods, intravesical administration reduces the amount of drugs required, increases the amount of drugs reaching the lesion site, and minimizes systemic exposure to therapeutic agents. To overcome the limitations of urinary voiding, low urothelium permeability, and intermittent catheterization for large dilution and irrigation of drugs in the bladder, magnetic and photothermal-responsive folate receptor-targeted thermal liposomes (FA-TMLs) were designed for the targeted delivery of doxorubicin (DOX) to bladder cancer cells. METHODS: Through a microfluidic mixer chip, the magnetic nanoparticles (MNPs), gold nanorods (GNRs) and DOX were encapsulated in folate-modified thermosensitive liposomes to form FA-TMLs@MNPs-GNRs-DOX. DLS, TEM, DSC, and magnetic hysteresis loop were used to characterize the construction of FA-TMLs@MNPs-GNRs-DOX. RESULTS: FA-TMLs@MNPs-GNRs-DOX had a size of about 230 nm and exhibited superparamagnetic properties with the saturation magnetization of 20 emu/g. The DOX loading capacity was as high as 0.57 mg/mL. Additionally, drug release of the FA-TMLs@MNPs-GNRs-DOX could be controlled by temperature change through the photothermal effect. A 980 nm laser beam was selectively irradiated on the FA-TMLs@MNPs-GNRs-DOX to trigger the structural changes of the FA-TMLs, and an average of 95% of the drug was released after 3 hours. The results of cell uptake experiments reveal indicated that FA-TMLs@MNPs-GNRs-DOX were able to specifically bind folate-receptor-positive cells and exhibited toxicity to bladder tumor cells. CONCLUSION: The present results suggest FA-TMLs@MNPs-GNRs-DOX have a promising multifunctional response and can act as an ideal multifunctional drug delivery system (DDS) for the treatment of bladder tumors.

Stoichiometry of Carbon, Nitrogen and Phosphorus in Shrub Organs Linked Closely With Mycorrhizal Strategy in Northern China.

Mycorrhizal strategies include mycorrhizal statuses and mycorrhizal types, which are important reflections of the functional characteristics of ecosystems. The stoichiometry of carbon, nitrogen, and phosphorus in plant organs is an important part of ecosystem functions, which has an important impact on the nutrient cycle of the ecosystem. The concentration of carbon, nitrogen, and phosphorus played a crucial role in ecosystem functioning and dynamics. The purpose of this study is to provide theoretical basis and data support for improving the properties of global terrestrial ecosystems by exploring the impact of mycorrhizal strategies on the stoichiometry of C, N, and P in different shrub organs. In this study, stoichiometric patterns of carbon (C), nitrogen (N) and phosphorus (P) in different shrub organs under different mycorrhizal status or types were analyzed at 725 samples across Northern China. Results showed that in different mycorrhizal status, the highest carbon concentration in shrub organs appeared in the facultatively mycorrhizal (FM) mycorrhizal status, and the highest nitrogen concentration appeared in the Non-mycorrhizal (NM) mycorrhizal status. Under different mycorrhizal types, the nitrogen concentration in the shrub organs under the arbuscular mycorrhiza (AM) mycorrhizal type was the highest, and the phosphorus concentration under the ecto-mycorrhiza (ECM) mycorrhizal type was the highest. In the OM or FM mycorrhizal status, the concentrations of C, N, and P in the stems and leaves increase with the increase of the concentrations of C, N, and P in the roots. In the NM mycorrhizal status, the N concentration in the stems and leaves increases with the increase of the N concentration in the roots. Under AM, AM+ECM, and ECM mycorrhizal type, the concentrations of C, N, and P are closely related in roots, stems and leaves. The content of plant nutrients in different organs is closely related. It turned out that mycorrhizal statuses or types are able to alter the allocation of C, N, and P in different organs, and the relationships of C, N, and P among different organs are able to present different trend with the varying of mycorrhizal statuses or types.

Evaluation of Vertebrate-Specific Replication-Defective Zika Virus, a Novel Single-Cycle Arbovirus Vaccine, in a Mouse Model.

The flavivirus Zika (ZIKV) has emerged as a global threat, making the development of a ZIKV vaccine a priority. While live-attenuated vaccines are known to induce long-term immunity but reduced safety, inactivated vaccines exhibit a weaker immune response as a trade-off for increased safety margins. To overcome the trade-off between immunogenicity and safety, the concept of a third-generation flavivirus vaccine based on single-cycle flaviviruses has been developed. These third-generation flavivirus vaccines have demonstrated extreme potency with a high level of safety in animal models. However, the production of these single-cycle, encapsidation-defective flaviviruses requires a complicated virion packaging system. Here, we investigated a new single-cycle flavivirus vaccine, a vertebrate-specific replication-defective ZIKV (VSRD-ZIKV), in a mouse model. VSRD-ZIKV replicates to high titers in insect cells but can only initiate a single-round infection in vertebrate cells. During a single round of infection, VSRD-ZIKV can express all the authentic viral antigens in vertebrate hosts. VSRD-ZIKV immunization elicited a robust cellular and humoral immune response that protected against a lethal ZIKV challenge in AG129 mice. Additionally, VSRD-ZIKV-immunized pregnant mice were protected against vertically transferring a lethal ZIKV infection to their offspring. Immunized male mice were protected and prevented viral accumulation in the testes after being challenged with lethal ZIKV. Overall, our results indicate that VSRD-ZIKV induces a potent protective immunity against ZIKV in a mouse model and represents a promising approach to develop novel single-cycle arbovirus vaccines.

Molecular Diversity and Distribution of Arbuscular Mycorrhizal Fungi at Different Elevations in Mt. Taibai of Qinling Mountain.

Arbuscular mycorrhizal fungi (AMFs) play a vital role in ecosystems, especially in ecosystem variability, diversity, and function. Understanding the AMF diversity, distribution, and their driver at different altitudinal gradients is a benefit for understanding the ecological function of AMF in mountain ecosystems. In this study, we explored the AMF molecular diversity and their distribution from 660 to 3,500 m a.s.l. in Mount Taibai of Qinling Mountains based on high-throughput sequencing technology. A total of 702 operational taxonomic units (OTUs) in 103 species of AMF are isolated from soil samples, which belong to 18 identified and 1 unidentified genus in 10 families. The fungi in the genus of Glomus is the most dominant, with the occurrence frequency of 100% and the relative abundance of 42.268% and 33.048% on the species and OTU level, respectively. The AMF colonization in root could be simulated by a cubic function with the change of altitudes with the peak and trough at a.s.l. 1,170 and 2,850 m, respectively. Further, AMF diversity indices including Sob, Shannon diversity, and Pielou evenness also showed the same cubic function change trends with increasing altitude at OTU and species levels. However, the average values of diversity indices at OTU level are always higher than these at the species level. Based on the OTU level, the highest and lowest values of Shannon and Pielou indices are observed at the altitudes of 1,400 and 2,800 m, respectively. The pattern of AMF community distribution in Mt. Taibai is driven by altitude with the characteristics of more abundance in the medium- to low-altitude than high-altitude areas. In general, abundant AMF molecular diversity and species exit in different elevations of Mt. Taibai, which indicate gradient changes with elevations.

Generation and preliminary characterization of vertebrate-specific replication-defective Zika virus.

Zika virus (ZIKV) is a mosquito-borne flavivirus that replicates in both vertebrate and insect cells, whereas insect-specific flaviviruses (ISF) replicate only in insect cells. We sought to convert ZIKV, from a dual-tropic flavivirus, into an insect-specific virus for the eventual development of a safe ZIKV vaccine. Reverse genetics was used to introduce specific mutations into the furin cleavage motif within the ZIKV pre-membrane protein (prM). Mutant clones were selected, which replicated well in C6/36 insect cells but exhibited reduced replication in non-human primate (Vero) cells. Further characterization of the furin cleavage site mutants indicated they replicated poorly in both human (HeLa, U251), and baby hamster kidney (BHK-21) cells. One clone with the induced mutation in the prM protein and at positions 291and 452 within the NS3 protein was totally and stably replication-defective in vertebrate cells (VSRD-ZIKV). Preliminary studies in ZIKV sensitive, immunodeficient mice demonstrated that VSRD-ZIKV-infected mice survived and were virus-negative. Our study indicates that a reverse genetic approach targeting the furin cleavage site in prM can be used to select an insect-specific ZIKV with the potential utility as a vaccine strain.