Hippo pathway deletion in adult resting cardiac fibroblasts initiates a cell state transition with spontaneous and self-sustaining fibrosis.

Cardiac fibroblasts (CFs) respond to injury by transitioning through multiple cell states, including resting CFs, activated CFs, and myofibroblasts. We report here that Hippo signaling cell-autonomously regulates CF fate transitions and proliferation, and non-cell-autonomously regulates both myeloid and CF activation in the heart. Conditional deletion of Hippo pathway kinases, Lats1 and Lats2, in uninjured CFs initiated a self-perpetuating fibrotic response in the adult heart that was exacerbated by myocardial infarction (MI). Single cell transcriptomics showed that uninjured Lats1/2 mutant CFs spontaneously transitioned to a myofibroblast cell state. Through gene regulatory network reconstruction, we found that Hippo-deficient myofibroblasts deployed a network of transcriptional regulators of endoplasmic reticulum (ER) stress, and the unfolded protein response (UPR) consistent with elevated secretory activity. We observed an expansion of myeloid cell heterogeneity in uninjured Lats1/2 CKO hearts with similarity to cells recovered from control hearts post-MI. Integrated genome-wide analysis of Yap chromatin occupancy revealed that Yap directly activates myofibroblast cell identity genes, the proto-oncogene Myc, and an array of genes encoding pro-inflammatory factors through enhancer-promoter looping. Our data indicate that Lats1/2 maintain the resting CF cell state through restricting the Yap-induced injury response.

Transcription factor NAC78 cooperates with NAC78 INTERACTING PROTEIN6 to confer drought tolerance in rice.

NAC (NAM, ATAF1/2, and CUC2) family transcription factors are involved in several cellular processes, including responses to drought, salinity, cold, and submergence. However, whether or how certain NAC proteins regulate drought tolerance in rice (Oryza sativa) remain unclear. In this study, we show that over-expression of OsNAC78 enhanced rice resistance to drought treatment, whereas Osnac78 mutant plants were susceptible to drought stress. We further characterized the OsNAC78 interacting protein, named NAC78 interacting protein6 (OsNACIP6), and found that it conferred rice drought tolerance. Our results demonstrate that OsNACIP6 enhanced transcription of OsNAC78 and promoted the expression of its downstream target OsGSTU37, encoding a glutathione reductase. The ABRE4 cis-element in the promoter region of OsNACIP675-1-127 conferred significant up-regulation of OsNACIP6 expression and initiated the OsNACIP6/OsNAC78-OsGSTU37 module that facilitates rice growth under drought conditions. Together, our results uncover a transcriptional module composed of OsNACIP6, OsNAC78, and OsGSTU37 and provide insights into the molecular mechanisms underlying the drought stress response in rice.

Coupled Poly(ethylenimine) Coreactant to Enhance Electrochemiluminescence of Polymer Dots for Array Imaging of Protein Biomarkers.

Traditional electrochemiluminescent (ECL) bioanalysis suffers from the demand for excessive external coreactants and the damage of reaction intermediates. In this work, a poly(ethylenimine) (PEI)-coupled ECL emitter was proposed by covalently coupling tertiary amine-rich PEI to polymer dots (Pdots). The coupled PEI might act as a highly efficient coreactant to enhance the ECL emission of Pdots through intramolecular electron transfer, reducing the electron transfer distance between emitter and coreactant intermediates and avoiding the disadvantages of traditional ECL systems. Through modification of the PEI-Pdots with tDNA, a sequence partially complementary to cDNA that was complementary to the aptamer of target protein biomarker (aDNA), tDNA-PEI-Pdots were obtained. The biosensors were produced using Au/indium tin oxide (ITO) with an aDNA/cDNA hybrid, and an ECL imaging biosensor array was constructed for ultrasensitive detection of protein biomarkers. Using vascular endothelial growth factor 165 (VEGF165) as a protein model, the proposed ECL imaging method containing two simple incubations with target samples and then tDNA-PEI-Pdots showed a detectable range of 1 pg mL-1 to 100 ng mL-1 and a detection limit of 0.71 pg mL-1, as well as excellent performance such as low toxicity, high sensitivity, excellent selectivity, good accuracy, and acceptable fabrication reproducibility. The PEI-coupled Pdots provide a new avenue for the design of ECL emitters and the application of ECL imaging in disease biomarker detection.

Enzyme-Triggered DNA Sensor Technology for Spatially-Controlled, Cell-Selective Molecular Imaging.

With unparalleled programmability, DNA has evolved as a powerful scaffold for engineering intricate and dynamic systems that can perform diverse tasks. By allowing serial detection of molecular targets in complex cellular milieus, increasingly sophisticated DNA sensors have not only promoted significant advances in unveiling the fundamental mechanisms of various pathophysiological processes but also provided a useful toolkit for disease diagnostics based on molecular signatures. Despite much progress, an inherent limitation of DNA-based sensors is that they often lack spatial control and cell-type selectivity for the sensing activity because of their "always active" design mechanism. Since most molecular targets of interests are not exclusive to disease cells, they are also shared by normal cells, the application of such biosensors for disease-specific imaging is limited by inadequate signal-to-background ratios due to indistinguishable signal response in both disease and normal cells. Therefore, imparting biosensors with spatial controllability remains a key issue to achieve molecular imaging with high sensitivity and cell specificity.As a biocatalyst, enzyme has been found to be closely related with the pathological conditions of numerous diseases. For example, many nucleases, protease, and kinases have been identified overexpressed in disease cells and considered as important biomarkers of cancer, inflammation, and neurological diseases. Recently, we have envisioned that such pathophysiology-associated enzymes could be leveraged as endogenous triggers to achieve spatial control over the molecular imaging activity of the DNA-based sensors with improved cell-specificity. In this Account, we outline the research efforts from our group on the development of endogenous enzyme-triggered, DNA-based sensor technology that enables spatially controlled, cell-type selective molecular imaging. With programmable DNA design and further engineering of enzymatically cleavable sites, a series of DNAzyme- and aptamer-based sensors have been developed for enzyme-controlled imaging of various molecular targets (e.g., metal ions and small molecules) in a cancer cell-selective manner. In particular, by introduction of PNA as bridge molecules to engineer DNA-based sensors with functional peptides, the conceptual design of protease-activated DNA biosensors has been established for spatioselective molecular imaging in cancer cells and extracellular tumor microenvironments. Furthermore, enzyme-triggered signal amplification approaches, such as enzymatically activated molecular beacon and catalytic hairpin assembly, have been developed for spatially selective RNA imaging in specific disease cells (e.g., inflammatory cells and cancer cells), which enables enhanced disease-site specificity and thus improved signal-to-background ratio. The signal amplification strategy is further expanded to cell-selective amplified imaging of non-RNA species through the combination with functional DNA design. Finally, the challenges and potential future directions in this burgeoning field are discussed. We hope this Account offers insights into rational design of enzymatically controlled, DNA-based sensor platforms for opening new frontiers in spatially resolved, cell-selective molecular imaging. We believe that the continuing advances in DNA-based molecular sensing technology together with the discoveries of diverse disease-associated enzymes will promise to usher a new era of diagnosis.

The impacts of basic medical insurance for urban-rural residents on the perception of social equity in China.

BACKGROUND: Based on CSS 2019 data, this article analyzes the impact of Basic Medical Insurance for Urban and Rural Residents (BMIUR) on perceived social equity of residents. METHOD: Using the CSS data of 2019, this article analyzes the influence of BMIUR on the perception of social equity of residents, on the basis of 2SLS model and mediating effect model. RESULTS: We find that BMIUR has a significantly positive impact on the perception of social equity. That is to say, BMIUR can improve residents' evaluation of social equity and further promote the level of social equity, which makes residents more happiness. The conclusion remains valid after using robustness test and propensity score matching to conduct counterfactual reasoning. The discussion of mechanism indicates that the influence of BMIUR is mediated by enhancing social capital, improving satisfaction of income distribution and reducing self-paid medical expenses. This study also finds that the influence of BMIUR is more obvious in the low-income, low-skilled and mid-west groups. CONCLUSION: The reform of the medical security system should be regarded as a foothold for improving people's well-being and promoting social equity; Expanding people's social capital through multiple channels and improving income distribution mechanisms; Strengthen vocational skills training, especially to provide more public services and social security for low-income groups, low-skilled groups, and groups in the western region of China, in order to improve the welfare and policy effectiveness of China's social security reform.

Emerging isolation and degradation technology of microplastics and nanoplastics in the environment.

Microplastics (MPs, less than 5 mm in size) are widely distributed in surroundings in various forms and ways, and threaten ecosystems security and human health. Its environmental behavior as pollutants carrier and the after-effects exposed to MPs has been extensively exploited; whereas, current knowledge on technologies for the separation and degradation of MPs is relatively limited. It is essential to isolate MPs from surroundings and/or degrade to safe levels. This in-depth review details the origin and distribution of MPs. Provides a comprehensive summary of currently available MPs separation and degradation technologies, and discusses the mechanisms, challenges, and application prospects of these technologies. Comparison of the contribution of various separation methods to the separation of NPs and MPs. Furthermore, the latest research trends and direction in bio-degradation technology are outlooked.

Solibacillus ferritrahens sp. nov., a novel siderophore-producing bacterium isolated from Wumeng Mountain National Nature Reserve in Yunnan Province.

A gram-stain-positive, aerobic, rod-shaped bacterial strain capable of producing siderophores, named YIM B08730T, was isolated from a soil sample collected from Wumeng Mountain National Nature Reserve, Zhaotong City, Yunnan Province. Growth occurred at 10-45 °C (optimum, 35-40 â„ƒ), pH 7.0-9.0 (optimum, 7.0) and in the presence of 0-5 % (w/v) NaCl (optimum, 0-1 %, w/v). A comparative analysis of the 16S rRNA gene sequence (1558 bp) of strain YIM B08730T showed the highest similarity to Solibacillus isronensis JCM 13838T (96.2 %), followed by Solibacillus silvestris DSM 12223T (96.0 %) and Solibacillus kalamii ISSFR-015T (95.4 %). The main polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine and one unidentified lipid. The main respiratory quinone of strain YIM B08730T was menaquinone 7 (MK-7). The major fatty acids were iso-C15:0 and C16:1ω7c alcohol. The digital DNA-DNA hybridization and average nucleotide identity values between strain YIM B08730T and the reference strain S. isronensis JCM 13838T were 24.8 % and 81.2 %, respectively. The G + C content of the genomic DNA was 37.1 mol%. The genome of the novel strain contained genes associated with the production of siderophores, and it also revealed other functional gene clusters involved in plant growth promotion and soil bioremediation. Based on these phenotypic, chemotaxonomic and phylogenetic analyses, strain YIM B08730T is considered to be a novel species of the genus Solibacillus, for which the name Solibacillus ferritrahens sp. nov. is proposed. The type strain is YIM B08730T (= NBRC 116268T = CGMCC 1.60169T).

Dual PI3K/mTOR Inhibitor BEZ235 combined with BMS-1166 Promoting Apoptosis in Colorectal Cancer.

Background: BMS-1166, a PD-1/PD-L1 inhibitor, inhibits the binding of PD-L1 to PD-1, restores T cell function, and enhances tumor immune response. However, mutations in the tumor suppressor or impaired cellular signaling pathways may also lead to cellular transformation. In this study, the SW480 and SW480R cell lines were used as the model to elucidate the treatment with BMS-1166, BEZ235, and their combination. Methods: MTT and colony-formation assays were used to evaluate cell proliferation. Wound-healing assay was used to assess cell migration. Cell cycle and apoptosis were analyzed by flow cytometry. The phosphorylation level of the key kinases in the PI3K/Akt/mTOR and MAPK pathways, PD-L1, and the protein levels related to the proliferation, migration, and apoptosis were assessed using western blotting. Results: BEZ235 enhanced BMS-1166-mediated cell proliferation and migration inhibition in SW480 and SW480R cells and promoted apoptosis. Interestingly, the downregulation of the negative regulator PTEN raised the PD-L1 level, which was abolished by the inhibition of Akt. BMS-1166 promoted PI3K, Akt, mTOR, and Erk phosphorylation. However, the combination of BEZ235 with BMS-1166 suppressed the expression of PI3K, p-Akt, p-mTOR, and p-Erk in SW480 and SW480R cells compared to BMS-1166 or BEZ235 single treatment by inhibiting the binding of PD1 to PD-L1. Conclusions: PD-1 binds to PD-L1 and activates the PI3K/mTOR and MAPK pathways, which might be the molecular mechanism of acquired resistance of CRC to BMS-1166. The combination of the two drugs inhibited the phosphorylation of PI3K, Akt, and Erk in the PI3K/mTOR and MAPK pathway, i.e., BEZ235 enhanced the BMS-1166 treatment effect by blocking the PI3K/mTOR pathway and interfering with the crosstalk of the MAPK pathway. Therefore, these findings provide a theoretical basis for BMS-1166 combined with BEZ235 in the trial treatment of colorectal cancer.

Psychromicrobium Xiongbiense sp. nov., an Actinobacterium Isolated from Forest Soil.

A novel Gram-stain-positive, oval-shaped, and non-flagellated bacterial strain YIM S02556T was isolated from forest soil in Xiongbi Town, Shizong County, Qujing City, Yunnan Province, southwestern China. The strain exhibited high pairwise 16 S rRNA gene sequence similarity with Psychromicrobium lacuslunae (97.3%) and Psychromicrobium silvestre (96.3%). Strain YIM S02556T exhibited an average nucleotide identity (ANI) of 72.5% with P. lacuslunae IHBB 11,108T and 72.8% ANI with P. silvestre AK 20-18T. The digital DNA-DNA hybridization (dDDH) value between strain YIM S02556T and P. lacuslunae IHBB 11,108T was 20.2%, while with P. silvestre AK 20-18T, the dDDH value was 20.8%. Strain YIM S02556T exhibited optimal growth at 28 °C, pH 7.0, without NaCl. Growth occurred within 10-37 ℃, pH 5.0-8.0, and in the presence of up to 5% w/v NaCl concentration. The genome size was 3.1 Mbp with 64.2% G + C content. The predominant menaquinone was MK-8(H4). The major cellular fatty acid was anteiso-C15:0. Based on the polyphasic analysis, strain YIM S02556T (= KCTC 49,805T = CCTCC AB2020166T) represents a novel Psychromicrobium species in which the name Psychromicrobium xiongbiense sp.nov. was proposed.

Identification of the absorbed constituents and metabolites of Huangqi Guizhi Wuwu decoction by ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

Huangqi Guizhi Wuwu decoction (HGWWD) is a widely used traditional Chinese medicine (TCM) preparation for the treatment of ischemic stroke and diabetes peripheral neuropathy. However, the material basis for the efficacy of HGWWD remains unclear. In this study, a rapid, sensitive and selective ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) method was developed to separate and identify the absorbed components and metabolites of HGWWD in rat plasma after oral administration for the first time. By comparing the retention time, high-resolution mass spectrometry primary and secondary mass spectrometry data of blank plasma and drug-containing plasma, a total of 42 constituents, including 24 prototype compounds and 18 metabolites, were identified or tentatively characterized. The results indicated that monoterpenes, flavonoids, organic acids, amino acids, gingerols and alkaloids were main prototype compounds in rat plasma, and flavonoid-related metabolites, organic acid-related metabolites and gingerol-related metabolites were major metabolites. It is concluded the developed UHPLC-Q-TOF-MS method with high sensitivity and resolution is suitable for identifying and characterizing the absorbed components and metabolites of HGWWD, and the results will provide important data for further study on the relationship between the chemical constituents and pharmacological activities of HGWWD.

Enzyme and MicroRNA Dual-Regulated Photodynamic Molecular Beacons for Cell-Selective Amplification of Antitumor Efficacy.

Photodynamic molecular beacons (PMBs) are highly appealing for activatable photodynamic therapy (PDT), but their applications are hindered by limited therapeutic efficacy. Here, by molecular engineering of enzyme-responsive units in the loop region of DNA-based PMBs, we present for the first time the modular design of an enzyme/microRNA dual-regulated PMB (D-PMB) to achieve cancer-cell-selective amplification of PDT efficacy. In the design, the "inert" photosensitizers in D-PMB could be repeatedly activated in the presence of both tumor-specific enzyme and miRNA, leading to amplified generation of cytotoxic singlet oxygen species and therefore enhanced PDT efficacy in vitro and in vivo. By contrast, low photodynamic activity could be observed in healthy cells, as D-PMB activation has been largely avoided by the dual-regulatable design. This work presents a cooperatively activated PDT strategy, which enables enhanced therapeutic efficacy with improved tumor-specificity and thus conceptualizes an approach to expand the repertoire of designing smart tumor treatment modality.

DNA Nanotechnology Targeting Mitochondria: From Subcellular Molecular Imaging to Tailor-Made Therapeutics.

Mitochondria, one of the most important organelles, represent a crucial subcellular target for fundamental research and biomedical applications. Despite significant advances in the design of DNA nanotechnologies for a variety of bio-applications, the dearth of strategies that enable mitochondria targeting for subcellular molecular imaging and therapy remains an outstanding challenge in this field. In this Minireview, we summarize the recent progresses on the emerging design and application of DNA nanotechnology for mitochondria-targeted molecular imaging and tumor treatment. We first highlight the engineering of mitochondria-localized DNA nanosensors for in situ detection and imaging of diverse key molecules that are essential to maintain mitochondrial functions, including mitochondrial DNA and microRNA, enzymes, small molecules, and metal ions. Then, we compile the developments of DNA nanotechnologies for mitochondria-targeted anti-tumor therapy, including modularly designed DNA nanodevices for subcellular delivery of therapeutic agents, and programmed DNA assembly for mitochondrial interference. We will place an emphasis on clarification of the chemical principles of how DNA nanobiotechnology can be designed to target mitochondria for various biomedical applications. Finally, the remaining challenges and future directions in this emerging field will be discussed, hoping to inspire further development of advanced DNA toolkits for both academic and clinical research regarding mitochondria.

Modular Weaving DNAzyme in Skeleton of DNA Nanocages for Photoactivatable Catalytic Activity Regulation.

DNAzymes exhibit tremendous application potentials in the field of biosensing and gene regulation due to its unique catalytic function. However, spatiotemporally controlled regulation of DNAzyme activity remains a daunting challenge, which may cause nonspecific signal leakage or gene silencing of the catalytic systems. Here, we report a photochemical approach via modular weaving active DNAzyme into the skeleton of tetrahedral DNA nanocages (TDN) for light-triggered on-demand liberation of DNAzyme and thus conditional control of gene regulation activity. We demonstrate that the direct encoding of DNAzyme in TDN could improve the biostability of DNAzyme and ensure the delivery efficiency, comparing with the conventional surface anchoring strategy. Furthermore, the molecular weaving of the DNA nanostructures allows remote control of DNAzyme-mediated gene regulation with high spatiotemporal precision of light. In addition, we demonstrate that the approach is applicable for controlled regulation of the gene editing functions of other functional nucleic acids.

Scalable Precise Nanofilm Coating and Gradient Al Doping Enable Stable Battery Cycling of LiCoO2 at 4.7†V.

The quest for smart electronics with higher energy densities has intensified the development of high-voltage LiCoO2 (LCO). Despite their potential, LCO materials operating at 4.7 V faces critical challenges, including interface degradation and structural collapse. Herein, we propose a collective surface architecture through precise nanofilm coating and doping that combines an ultra-thin LiAlO2 coating layer and gradient doping of Al. This architecture not only mitigates side reactions, but also improves the Li+ migration kinetics on the LCO surface. Meanwhile, gradient doping of Al inhibited the severe lattice distortion caused by the irreversible phase transition of O3-H1-3-O1, thereby enhanced the electrochemical stability of LCO during 4.7 V cycling. DFT calculations further revealed that our approach significantly boosts the electronic conductivity. As a result, the modified LCO exhibited an outstanding reversible capacity of 230 mAh g-1 at 4.7 V, which is approximately 28 % higher than the conventional capacity at 4.5 V. To demonstrate their practical application, our cathode structure shows improved stability in full pouch cell configuration under high operating voltage. LCO exhibited an excellent cycling stability, retaining 82.33 % after 1000 cycles at 4.5 V. This multifunctional surface modification strategy offers a viable pathway for the practical application of LCO materials, setting a new standard for the development of high-energy-density and long-lasting electrode materials.

Lycopene inhibits endothelial-to-mesenchymal transition of choroidal vascular endothelial cells in laser-induced mouse choroidal neovascularization.

Choroidal neovascularization (CNV), is a major cause of irreversible blindness among the elderly population in developed countries, which is resulted from subretinal fibrosis without effective therapeutic strategies. Endothelial-to-mesenchymal transition (EndMT) of choroidal vascular endothelial cells (CVECs) contributes to subretinal fibrosis. Lycopene (LYC), a non-pro-vitamin A carotenoid, plays an anti-fibrotic role. Herein, we explored the effect and mechanism of LYC on the EndMT of CVECs during CNV. Firstly, LYC inhibited EndMT in hypoxic human choroidal endothelial cells (HCVECs). Meanwhile, LYC inhibited proliferation, androgen receptor (AR) expression and nuclear localization in hypoxic HCVECs. Then LYC-inhibited AR promotes the activation of microphthalmia-associated transcription factor (MITF) in hypoxic HCVECs. In addition, LYC down-regulated AR and induced MITF up-regulated pigment epithelium-derived factor (PEDF) transcription and expression in hypoxic HCVECs. Moreover, LYC-induced PEDF bound to laminin receptor (LR), inhibiting EndMT of hypoxic HCVECs via down-regulating protein kinase B (AKT)/ß-catenin pathway. In vivo, LYC alleviated mouse laser-induced subretinal fibrosis secondary to CNV via up-regulating PEDF without any ocular or systemic toxicity. These results indicate that LYC inhibits EndMT of CVECs via modulating AR/MITF/PEDF/LR/AKT/ß-catenin pathway, showing LYC is a promising therapeutic agent for CNV.

Protease-Triggered, Spatially Controlled DNA Assembly in Apoptotic Cells for Early Evaluation of Therapeutic Efficacy.

Despite numerous advances in the use of DNA as building blocks to assemble complex structures, the dearth of strategies that allow for protease-controlled in situ DNA assembly in living cells remains a bottleneck in this field. Here, we present a modular engineering approach to achieve protease-triggered self-assembly of DNA in apoptotic cells for early evaluation of tumor response to drug treatment. In the design, peptide nucleic acid is introduced as a building bridge to engineer DNA building blocks with peptides and thus to suppress their self-assembly activity, while caspase-3 (Casp-3) protease-mediated enzymatic cleavage of the peptide substrate enables the activation of the DNA assembly, generating fluorescence signal output for real-time monitoring of Casp-3 activity. Furthermore, the specific protease triggering imparts DNA assembly with spatial selectivity to apoptotic cells in vivo, allowing for early evaluation of tumor therapeutic efficacy. Moreover, the strategy is extended to probe the activity of MMP-2 for lymph node metastasis imaging, demonstrating the universality of this approach. This work highlights protease-controlled DNA assembly in ways that are simple and versatile, with the potential to expand the repertoire of DNA nanotechnology for diverse biomedical applications.

Modular Engineering of DNAzyme-Based Sensors for Spatioselective Imaging of Metal Ions in Mitochondria.

DNAzyme-based sensors remain at the forefront of metal-ion imaging efforts, but most lack the subcellular precision necessary to their applications in specific organelles. Here, we seek to overcome this limitation by presenting a DNAzyme-based biosensor technology for spatiotemporally controlled imaging of metal ions in mitochondria. A DNA nanodevice was constructed by integrating an optically activatable DNAzyme sensor and an upconversion nanoparticle with an organelle-targeting signal. We exemplify that this approach allows for mitochondria-specific imaging of Zn2+ in living cells in a near-infrared light-controlled manner. Based on this, the system is used for the monitoring of mitochondrial Zn2+ during drug treatment in a cellular model of ischemia insult. Furthermore, the DNA nanodevice is employed to assess dynamic Zn2+ change and pharmacological interventions in an injury cell model of Zn2+ toxicity. This method paves the way for engineering of DNAzyme sensors to investigate the pathophysiological roles of metal ions at the subcellular level.

Regulation of Target-activated CRISPR/Cas12a on Surface Binding of Polymer Dots for Sensitive Electrochemiluminescence DNA Analysis.

Polymer dots (Pdots) have emerged as a type of attractive electrochemiluminescence (ECL) emitter. However, the low ECL efficiency severely limits their practicability. In this work, we develop a sensitive ECL biosensing strategy for the detection of human papilloma virus subtype (HPV-16) DNA by using target-activated CRISPR/Cas12a to regulate the binding of Pdots-DNA to biosensor and local surface plasmon resonance (LSPR) effect of electrochemically deposited Au nanoparticles (depAuNPs) to enhance the ECL emission of Pdots bound on biosensor. The biosensor is prepared by simply assembling hairpin DNA on depAuNPs modified electrode. In the presence of target DNA, the designed specific CRISPR/Cas12a can be activated to digest single-stranded assistant DNA, which decreases the amount of hairpin DNA opened by assistant DNA to bind Pdots-DNA on the biosensor surface, thus reduces the ECL emission. The integration of target DNA-triggered catalysis and the LSPR effect of depAuNPs greatly improves the sensitivity of ECL analysis. Using HPV-16 DNA as a target model, the proposed method shows a limit of detection (LOD) of 3.2 fM at a signal-to-noise ratio of 3 and a detectable concentration range of 5.0 fM to 50 pM. The high sensitivity, excellent selectivity, good testing stability, and acceptable fabrication reproducibility of the designed ECL biosensing strategy demonstrate its potential application in DNA bioanalysis.

Pseudomonas subflava sp. nov., a new Gram-negative bacterium isolated from Guishan in Yunnan province, south-west China.

A new Gram-negative, rod-shaped, flagellated bacterium was isolated from soil in the Guishan, Xinping County, Yuxi City, Yunnan Province, China, and named YIM B01952T. Growth occurred at 10-40 °C (optimum, 30 °C), pH 6.0-9.0 (optimum, pH 7.5) and with up to ≤ 5.0% (w/v) NaCl on Tryptic Soy Broth Agar (TSA) plates. Phylogenetic analysis based on the 16S rRNA gene and draft-genome sequence showed that strain YIM B01952T belonged to the genus Pseudomonas, and was closely related to the type strain of Pseudomonas alcaligenes (sequence similarity was 98.8%). The digital DNA-DNA hybridization (dDDH) value between strain YIM B01952T and the parallel strain P. alcaligenes ATCC 14909T was 49.0% based on the draft genome sequence. The predominant menaquinone was Q-9. The major fatty acids were summed feature 8 (C18:1 ω6c and/or C18:1 ω7c), summed feature 3 (C16:1 ω6c and/or C16:1 ω7c) and C16:0. The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, and phosphatidylglycerol. The genome size of strain YIM B01952T was 4.341 Mb, comprising 4156 predicted genes with a DNA G + C content of 66.4 mol%. In addition, we detected that strain YIM B01952T had some traditional functional genes (plant growth promotion and multidrug resistance), unique genes through genome comparison and analysis with similar strains. Based on genetic analyses and biochemical characterization, the strain YIM B01952T was identified as a novel species in the genus Pseudomonas, for which the name Pseudomonas subflava sp. nov. is proposed. The type strain is YIM B01952T (=CCTCC AB 2021498T = KCTC 92073T).

Characterization of chemical constituents in Huangqi Guizhi Wuwu decoction using ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

Huangqi Guizhi Wuwu decoction (HGWWD) is a classic traditional Chinese medicine prescription for the treatment of ischemic stroke, etc. However, the material basis of its efficacy remains unclear, seriously affecting drug development and clinical applications. In the present study, an ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry method was developed to separate and identify the chemical components of HGWWD. A total of 81 compounds were identified and tentatively characterized. Eight compounds were accurately identified by comparing the retention time and mass spectrometry data with those of reference substances, the remaining compounds were characterized by comparing the mass spectrometry data and reference information. Based on the results of compound attribution, 35 compounds were from Astragali Radix, six compounds were from Cinnamomi Ramulus, 23 compounds were from Paeoniae Radix Alba, eight compounds were from Zingiberis Rhizoma Recens and nine compounds were from Jujubae Fructus. The results showed that monoterpenoids, flavonoids, organic acids, triterpenes, amino acids, gingerols, alkaloids, and glycosides were the main chemical components of HGWWD. This analytical method is suitable for characterizing the chemical constituents of HGWWD, and the results provide important information for elucidating its pharmacodynamic material basis and mechanism of action.