DNA Framework-Programmed Nanoscale Enzyme Assemblies.

Multienzyme assemblies mediated by multivalent interaction play a crucial role in cellular processes. However, the three-dimensional (3D) programming of an enzyme complex with defined enzyme activity in vitro remains unexplored, primarily owing to limitations in precisely controlling the spatial topological configuration. Herein, we introduce a nanoscale 3D enzyme assembly using a tetrahedral DNA framework (TDF), enabling the replication of spatial topological configuration and maintenance of an identical edge-to-edge distance akin to natural enzymes. Our results demonstrate that 3D nanoscale enzyme assemblies in both two-enzyme systems (glucose oxidase (GOx)/horseradish peroxidase (HRP)) and three-enzyme systems (amylglucosidase (AGO)/GOx/HRP) lead to enhanced cascade catalytic activity compared to the low-dimensional structure, resulting in ∼5.9- and ∼7.7-fold enhancements over homogeneous diffusional mixtures of free enzymes, respectively. Furthermore, we demonstrate the enzyme assemblies for the detection of the metabolism biomarkers creatinine and creatine, achieving a low limit of detection, high sensitivity, and broad detection range.

Identification of genes associated with sperm storage capacity in hens at different times after insemination by RNA-seq and Ribo-seq.

BACKGROUND: Sperm storage capacity (SSC) determines the duration of fertility in hens and is an important reproduction trait that cannot be ignored in production. Currently, the genetic mechanism of SSC is still unclear in hens. Therefore, to explore the genetic basis of SSC, we analyzed the uterus-vagina junction (UVJ) of hens with different SSC at different times after insemination by RNA-seq and Ribo-seq. RESULTS: Our results showed that 589, 596, and 527 differentially expressed genes (DEGs), 730, 783, and 324 differentially translated genes (DTGs), and 804, 625, and 467 differential translation efficiency genes (DTEGs) were detected on the 5th, 10th, and 15th days after insemination, respectively. In transcription levels, we found that the differences of SSC at different times after insemination were mainly reflected in the transmission of information between cells, the composition of intercellular adhesion complexes, the regulation of ion channels, the regulation of cellular physiological activities, the composition of cells, and the composition of cell membranes. In translation efficiency (TE) levels, the differences of SSC were mainly related to the physiological and metabolic activities in the cell, the composition of the organelle membrane, the physiological activities of oxidation, cell components, and cell growth processes. According to pathway analysis, SSC was related to neuroactive ligand-receptor interaction, histidine metabolism, and PPAR signaling pathway at the transcriptional level and glutathione metabolism, oxidative phosphorylation, calcium signaling pathway, cell adhesion molecules, galactose metabolism, and Wnt signaling pathway at the TE level. We screened candidate genes affecting SSC at transcriptional levels (COL4A4, MUC6, MCHR2, TACR1, AVPR1A, COL1A1, HK2, RB1, VIPR2, HMGCS2) and TE levels(COL4A4, MUC6, CYCS, NDUFA13, CYTB, RRM2, CAMK4, HRH2, LCT, GCK, GALT). Among them, COL4A4 and MUC6 were the key candidate genes differing in transcription, translation, and translation efficiency. CONCLUSIONS: Our study used the combined analysis of RNA-seq and Ribo-seq for the first time to investigate the SSC and reveal the physiological processes associated with SSC. The key candidate genes affecting SSC were screened, and the theoretical basis was provided for the analysis of the molecular regulation mechanism of SSC.

Iron-Doped Polymer Dots with Enhanced Fluorescence and Dual Enzyme Activity for Versatile Bioassays.

Nanozymes with multiple functionalities endow biochemical sensing with more sensitive and efficient analytical performance by widening the sensing modes. Meanwhile, the target-oriented design of multifunctional nanozymes for certain biosensing remains challenging. Herein, a constructive strategy of doping iron into polymer dots (PDs) to achieve nanozymes with excellent oxidase-mimicking and peroxidase-mimicking activity is proposed. Compared with the Fe-free PDs prepared under the same mild condition, the Fe-doped PDs (Fe-PDs) exhibit greatly boosted fluorescence at 500 nm. While applying 3,3',5,5'-tetramethylbenzidine (TMB) as a chromogenic substrate, the fluorescence of the Fe-PDs can be further quenched by oxTMB due to the inner filter effect (IFE). Inspired by this, a simple but efficient colorimetric and fluorometric dual-mode sensing platform is developed for monitoring the reducing substances ascorbic acid (AA), α-glucosidase (α-Glu), and its inhibitors (AGIs). We believe that such multifunctional enzyme-mimic materials will provoke the exploration of multimode sensing strategy with strong practicality to serve as a versatile tool in biochemical sensing.

Combination of Ternary Electrochemiluminescence System of BNQDs/AgMOG-K2S2O8 and Electrochemiluminescence Resonance Energy Transfer Strategy for Ultrasensitive Immunoassay of Amyloid-ß Protein.

In this work, based on boron nitride quantum dots (BNQDs) as energy donors and MnO2@MWCNTs-COOH as energy receptors, we designed an efficient electrochemiluminescence resonance energy transfer (ECL-RET) immunosensor for the detection of amyloid-ß (Aß42) protein, a biomarker of Alzheimer's disease (AD). First, the signal amplification of a ternary ECL system composed of BNQDs (as the ECL emitter), K2S2O8 (as the coreactant), and silver metal-organic gels (AgMOG, as the coreaction accelerator) was realized, and PDDA as stabilizer was added, a strong and stable initial ECL signal was obtained. AgMOG could not only support a large amount of BNQDs and Aß42 capture antibody (Ab1) through Ag-N bond but also exhibit excellent ECL catalytic performance and enhance the luminescent intensity of BNQDs@PDDA-K2S2O8 system. In addition, due to the broad absorption spectrum of MnO2@MWCNTs-COOH and the extensive overlap with the ECL emission spectrum of BNQDs, the quenching probe Ab2-MnO2@MWCNTs-COOH could be introduced into the ternary system through a sandwich immune response. On this basis, the signal on-off ECL immunosensor was constructed to achieve the ultrasensitive detection of Aß42 through signal transformation. Under the optimal conditions, the prepared ECL biosensor manifested a wide linear range (10 fg/mL-100 ng/mL) with a detection limit of 2.89 fg/mL and showed excellent stability, selectivity, and repeatability, which provided an effective strategy for the ultrasensitive detection of biomarkers in clinical analysis.

Encapsulation of Enzymes into Hydrophilic and Biocompatible Metal Azolate Framework: Improved Functions of Biocatalyst in Cascade Reactions and its Sensing Applications.

Multiple enzyme-triggered cascade biocatalytic reactions are vital in vivo or vitro, considering the basic biofunction preservation in living organisms and signals transduction for biosensing platforms. Encapsulation of such enzymes into carrier endows a sheltering effect and can boost catalytic performance, although the selection and preparation of an appropriate carrier is still a concern. Herein, focusing on MAF-7, a category of metal azolate framework (MAF) with superiority against the topologically identical ZIF-8, this enzyme@MAF system can ameliorate the sustainability of encapsulating natural enzymes into carriers. The proposed biocatalyst composite AChE@ChOx@MAF-7/hemin is constructed via one-pot in situ coprecipitation method. Subsequently, MAF-7 is demonstrated to exhibit an excellent capacity of the carrier and protection against external factors in the counterpart of ZIF-8 through encapsulated and free enzymes. In addition, detections for specific substrates or inhibitors with favorable sensitivity are accomplished, indicating that the properties above expectation of different aspects of the established platform are successfully realized. This biofunctional composite based on MAF-7 can definitely provide a potential approach for optimization of cascade reaction and enzyme encapsulation.

Comparing the Catalytic Effect of Metals for Energetic Materials: Machine Learning Prediction of Adsorption Energies on Metals.

Energetic materials (EMs) and metals are the important components of solid propellants, and a strong catalysis of metals on EMs could further enhance the combustion performance of solid propellants. Accordingly, the study on the adsorption of EMs such as octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX), hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), and ammonium dinitramide (ADN) on metals (Ti, Zr, Fe, Ni, Cu, and Al) was carried out by density functional theory (DFT) to reveal the catalytic effect of metals. The deep dissociation of EMs on Ti and Zr represents a stronger interaction and corresponds to the rapid thermal decomposition behavior of the EMs/metal composite in the experiment. It is expected that DFT calculation can be selected instead of experiments to compare the catalytic effect of metals and preliminarily screen out potential high-performance metals. Based on the data set of the calculated adsorption energy, further machine learning (ML) was used to predict the adsorption energy of EMs on metals for a convenient comparison of the catalytic effect of metals, since a quite high adsorption energy value represents a more thorough dissociation. The kernel ridge regression (KRR) method shows the best performance on predicting adsorption energy and helps to choose the metals for efficiently catalyzing ammonium nitrate (AN) and hexanitrohexaazaisowurtzitane (CL-20). Such adsorption computation and ML not only reveal the decomposition mechanism of EMs on metals but also provide a simple underlying method to predict the catalytic effect of metals.

Enhanced Detection of Novel Low-Frequency Gene Fusions via High-Yield Ligation and Multiplexed Enrichment Sequencing.

Panel-based methods are commonly employed for the analysis of novel gene fusions in precision diagnostics and new drug development in cancer. However, these methods are constrained by limitations in ligation yield and the enrichment of novel gene fusions with low variant allele frequencies. In this study, we conducted a pioneering investigation into the stability of double-stranded adapter DNA, resulting in improved ligation yield and enhanced conversion efficiency. Additionally, we implemented blocker displacement amplification, achieving a remarkable 7-fold enrichment of novel gene fusions. Leveraging the pre-enrichment achieved with this approach, we successfully applied it to Nanopore sequencing, enabling ultra-fast analysis of novel gene fusions within one hour with high sensitivity. This method offers a robust and remarkably sensitive mean of analyzing novel gene fusions, promising the discovery of pivotal biomarkers that can significantly improve cancer diagnostics and the development of new therapeutic strategies.

Transcriptome profiling analysis of uterus during chicken laying periods.

The avian eggshell is formed in the uterus. Changes in uterine function may have a significant effect on eggshell quality. To identify the vital genes impacting uterine functional maintenance in the chicken, uteri in three different periods (22W, 31W, 51W) were selected for RNA sequencing and bioinformatics analysis. In our study, 520, 706 and 736 differentially expressed genes (DEGs) were respectively detected in the W31 vs W22 group, W51 vs W31 group and W51 vs W22 group. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated DEGs were enriched in the extracellular matrix, extracellular region part, extracellular region, extracellular matrix structural constituent, ECM receptor interaction, collagen-containing extracellular matrix and collagen trimer in the uterus (P < 0.05). Protein-protein interaction analysis revealed that FN1, LOX, THBS2, COL1A1, COL1A2, COL5A1, COL5A2, POSTN, MMP13, VANGL2, RAD54B, SPP1, SDC1, BTC, ANGPTL3 might be key candidate genes for uterine functional maintenance in chicken. This study discovered dominant genes and pathways which enhanced our knowledge of chicken uterine functional maintenance.

Room-Temperature Synthesized Iron/Cobalt Metal-Organic Framework Nanosheets with Highly Efficient Catalytic Activity toward Luminol Chemiluminescence Reaction.

Two-dimensional (2D) iron/cobalt metal-organic framework nanosheets (Fe/Co-MOF NSs) were synthesized via the cooperative self-assembly reaction of Fe3+/Co2+ and terephthalic acid at room temperature. The as-prepared 2D Fe/Co-MOF NSs display superior performance in catalysis of the chemiluminescence (CL) reaction between luminol and H2O2. The CL spectrum, UV-vis absorption spectroscopy, radical scavenger experiments, and electron spin resonance (ESR) spectroscopy are utilized to research the possible CL mechanism of the luminol-H2O2-Fe/Co-MOF NSs system. All results indicate that Fe/Co-MOF NSs present outstanding peroxidase-like activity and could catalyze H2O2 to produce 1O2, O2·-, and ·OH, which could react rapidly with the luminol anion radical and result in strong CL. With the highly efficient CL of the luminol-H2O2-Fe/Co-MOF NSs system, a sensitive sensor for the detection of dopamine (DA) is developed based on the inhibitory effect of DA on the CL intensity. Good linearity over the range of 50-800 nM is achieved with a limit of detection of 20.88 nM (S/N = 3). This research demonstrates that 2D Fe/Co-MOF NSs is a highly effective catalyst for luminol CL reaction and has great application potential in the CL field.

PacBio full-length transcriptome analysis provides new insights into transcription of chloroplast genomes.

Chloroplast and mitochondrial DNA (cpDNA and mtDNA) are apart from nuclear DNA (nuDNA) in a eukaryotic cell. The transcription system of chloroplasts differs from those of mitochondria and eukaryotes. In contrast to nuDNA and animal mtDNA, the transcription of cpDNA is still not well understood, primarily due to the unresolved identification of transcription initiation sites (TISs) and transcription termination sites (TTSs) on the genome scale. In the present study, we characterized the transcription of chloroplast (cp) genes with greater accuracy and comprehensive information using PacBio full-length transcriptome data from Arabidopsis thaliana. The major findings included the discovery of four types of artifacts, the validation and correction of cp gene annotations, the exact identification of TISs that start with G, and the discovery of polyA-like sites as TTSs. Notably, we proposed a new model to explain cp transcription initiation and termination at the whole-genome level. Four types of artifacts, degraded RNAs and splicing intermediates deserve the attention from researchers working with PacBio full-length transcriptome data, as these contaminant sequences can lead to incorrect downstream analysis. Cp transcription initiates at multiple promoters and terminates at polyA-like sites. Our study provides new insights into cp transcription and new clues to study the evolution of promoters, TISs, TTSs and polyA tails of eukaryotic genes.

Piriformospora indica Increases Resistance to Fusarium pseudograminearum in Wheat by Inducing Phenylpropanoid Pathway.

Fusarium crown rot (FCR), mainly caused by Fusarium pseudograminearum, not only seriously threatens the yield and quality of wheat, but also endangers the health and safety of humans and livestock. Piriformospora indica is a root endophytic fungus that colonizes plant roots extensively and can effectively promote plant growth and improve plant resistance to biotic and abiotic stresses. In this study, the mechanism of FCR resistance mediated by P. indica in wheat was revealed from the phenylpropanoid metabolic pathway. The results showed that the colonization of P. indica significantly reduced the progression of wheat disease, the amount of F. pseudograminearum colonization, and the content of deoxynivalenol (DON) in wheat roots. RNA-seq suggested that P. indica colonization could reduce the number of differentially expressed genes (DEGs) in the transcriptome caused by F. pseudograminearum infection. The DEGs induced by the colonization of P. indica were partially enriched in phenylpropanoid biosynthesis. Transcriptome sequencing and qPCR indicated that the colonization of P. indica up-regulated the expression of genes involved in the phenylpropanoid biosynthesis pathway. The metabolome analysis indicated that the colonization of P. indica increased the metabolites' accumulation in the phenylpropanoid biosynthesis. Consistent with transcriptome and metabolomic analysis, microscopic observations showed enhanced lignin accumulation in the roots of the Piri and Piri+Fp lines, most likely contributing to the arrested infection by F. pseudograminearum. These results suggested that P. indica increased resistance to F. pseudograminearum in wheat by inducing the phenylpropanoid pathway.

Comparison of Transcriptome between Tolerant and Susceptible Rice Cultivar Reveals Positive and Negative Regulators of Response to Rhizoctonia solani in Rice.

Rice (Oryza sativa L.) is one of the world's most crucial food crops, as it currently supports more than half of the world's population. However, the presence of sheath blight (SB) caused by Rhizoctonia solani has become a significant issue for rice agriculture. This disease is responsible for causing severe yield losses each year and is a threat to global food security. The breeding of SB-resistant rice varieties requires a thorough understanding of the molecular mechanisms involved and the exploration of immune genes in rice. To this end, we conducted a screening of rice cultivars for resistance to SB and compared the transcriptome based on RNA-seq between the most tolerant and susceptible cultivars. Our study revealed significant transcriptomic differences between the tolerant cultivar ZhengDao 22 (ZD) and the most susceptible cultivar XinZhi No.1 (XZ) in response to R. solani invasion. Specifically, the tolerant cultivar showed 7066 differentially expressed genes (DEGs), while the susceptible cultivar showed only 60 DEGs. In further analysis, we observed clear differences in gene category between up- and down-regulated expression of genes (uDEGs and dDEGs) based on Gene Ontology (GO) classes in response to infection in the tolerant cultivar ZD, and then identified uDEGs related to cell surface pattern recognition receptors, the Ca2+ ion signaling pathway, and the Mitogen-Activated Protein Kinase (MAPK) cascade that play a positive role against R. solani. In addition, DEGs of the jasmonic acid and ethylene signaling pathways were mainly positively regulated, whereas DEGs of the auxin signaling pathway were mainly negatively regulated. Transcription factors were involved in the immune response as either positive or negative regulators of the response to this pathogen. Furthermore, our results showed that chloroplasts play a crucial role and that reduced photosynthetic capacity is a critical feature of this response. The results of this research have important implications for better characterization of the molecular mechanism of SB resistance and for the development of resistant cultivars through molecular breeding methods.

Grade follicles transcriptional profiling analysis in different laying stages in chicken.

During follicular development, a series of key events such as follicular recruitment and selection are crucially governed by strict complex regulation. However, its molecular mechanisms remain obscure. To identify the dominant genes controlling chicken follicular development, the small white follicle (SWF), the small yellow follicle (SYF), and the large yellow follicle (LYF) in different laying stages (W22, W31, W51) were collected for RNA sequencing and bioinformatics analysis. There were 1866, 1211, and 1515 differentially expressed genes (DEGs) between SWF and SYF in W22, W31, and W51, respectively. 4021, 2295, and 2902 DEGs were respectively identified between SYF and LYF in W22, W31, and W51. 5618, 4016, and 4809 DEGs were respectively identified between SWF and LYF in W22, W31, and W51. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that extracellular matrix, extracellular region, extracellular region part, ECM-receptor interaction, collagen extracellular matrix, and collagen trimer were significantly enriched (P < 0.05). Protein-protein interaction analysis revealed that COL4A2, COL1A2, COL4A1, COL5A2, COL12A1, ELN, ALB, and MMP10 might be key candidate genes for follicular development in chicken. The current study identified dominant genes and pathways contributing to our understanding of chicken follicular development.

Ferrocene-Functionalized Covalent Organic Frameworks and Target Catalyzed Hairpin Assembly Strategy for Amplified Electrochemical Determination of MicroRNAs.

MicroRNAs (miRNAs) are considered as significant biomarkers in early diagnosis and treatment of diseases. Herein, an electrochemical biosensor that uses ferrocene (Fc)-functionalized covalent organic frameworks (COFs), a DNA tetrahedron nanostructure (DTN) biosensing interface, and a target catalyzed hairpin assembly (CHA) strategy has been fabricated and successfully developed for the sensitive and specific determination of microRNA-21 (miR-21). The COF served as a linked substrate for immobilization of gold nanoparticles (AuNPs), Fc-COOH, and complementary DNA probe L1 to prepare the electrochemical signal probe COF/Au/Fc/L1, which has a large surface area, extraordinary catalytic properties, and superior biocompatibility to amplify the current signal. The DTN containing a hairpin sequence H1 at one vertex was elaborately designed to construct the biosensing interface; thus, the CHA could be implemented on the electrode surface. In the presence of miR-21, the CHA reaction between H1 and the hairpin H2 was triggered to produce a great number of duplex DNA (H1/H2) with sticky ends. Then, the signal probe COF/Au/Fc/L1 was modified on the electrode surface through the hybridization between L1 and the sticky end of H1/H2, thereby obtaining an amplified Fc current signal. Under optimal conditions, the biosensor showed a wide linear response ranging from 1 fM to 10 nM miR-21, with a low detection limit of 0.33 fM (S/N = 3). Meanwhile, the method showed acceptable accuracy and precision for the determination of miR-21 in human serum.

Electrochemiluminescence Cascade Amplification Platform for Detection of Dual-microRNA and Operation of Concatenated Logic Circuit.

The detection of multiple biomarkers is of great significance to the accurate diagnosis of diseases. Herein, in this work, we constructed an electrochemiluminescence (ECL) cascade amplification platform for dual acute myocardial infarction (AMI)-related microRNA detection. The Zn2+-dependent DNAzyme digestion reaction initiated by miR-133a and the duplex-specific nuclease (DSN) cleavage circuit initiated by miR-499 were carried out independently to form a fuel hairpin DNA and active initiator strand, respectively, to trigger a hybridization chain reaction, which constituted a two-input-regulated "AND" logic circuit based on single ECL signal output. The use of single signal probe (Ru(bpy)32+) avoided the time-consuming and costly process of multiple signal molecule labeling or modification. The independent operation of the DNAzyme digestion reaction and DSN-assisted target recycling improved the detection efficiency of the system. In addition, the detection of each miRNA had undergone a cascade amplification process, which improved the detection sensitivity for each target. Furthermore, benefitting from the strong complexation of EDTA with Zn2+ and the flexible design of DNA sequences, the two-input "AND" logic gate was extended to a four-input "INHIBIT-AND-INHIBIT" concatenated logic circuit, which broadens the application of the ECL method in logic gates. We anticipate that this cascading amplification strategy can be widely applied in accurate diagnosis of AMI and the construction of ECL-based logic devices.

Versatile Electrochemiluminescence Biosensing Platform Based on DNA Nanostructures and Catalytic Hairpin Assembly Signal Amplification.

Achieving rapid and highly sensitive detection of biomarkers is crucial for disease diagnosis and treatment. Here, a highly sensitive and versatile dual-amplification electrochemiluminescence (ECL) biosensing platform was constructed for target detection based on DNA nanostructures and catalyzed hairpin assembly (CHA). Specifically, when the target DNA was present, it would hybridize with the auxiliary strands (D1 and D2) to form an I-shaped nanostructure, which in turn triggered the subsequent catalytic hairpin assembly reaction to generate plenty of double-stranded DNA complexes (H1-H2). The resulting double-stranded complex could be trapped on the electrode surface and adsorbed the ECL signal probe Ru(phen)32+.We found that the I-shaped nanostructure-triggered CHA reaction had higher amplification efficiency compared with traditional CHA amplification. Thus, a sensitive "signal-on" ECL biosensor was constructed for target DNA detection with a detection limit of 1.09 fM. Additionally, by combining the binding properties of C-Ag+-C with an elaborately designed "Ag+-helper" probe, the proposed strategy could be immediately utilized for the highly sensitive and selective detection of silver ions, demonstrating the versatility of the developed biosensing platform. This strategy provided a new approach with potential applications in disease diagnosis and environmental monitoring.

Colorimetric Assay for Acetylcholinesterase Activity and Inhibitor Screening Based on Metal-Organic Framework Nanosheets.

Alzheimer's disease (AD) is a common chronic neurodegenerative disease that manifests as cognitive impairment and behavioral deficits and severely threatens the health of the elderly. Acetylcholinesterase (AChE) plays a vital role in biological signaling and is an essential target for the early diagnosis and treatment of AD. Herein, 2D Zn-TCPP(Fe) nanosheets (NSs) employing Zn2+ and Fe-bound tetrakis(4-carboxyphenyl)porphyrin ligands were prepared through a surfactant-assisted synthetic method. The ultrathin two-dimensional (2D) metal-organic framework structures exhibited high peroxidase-like activity, which allowed the catalysis of the H2O2-initiated oxidation of colorless 3,3',5,5'-tetramethylbenzidine (TMB) to blue oxidized TMB (ox-TMB). Such catalytic performance inspired us to develop a convenient, rapid, and sensitive acetylcholinesterase activity assay, during which AChE can catalyze the substrate acetylthiocholine (ATCh) to produce thiocholine (TCh), and TCh could especially enable the degradation of 2D Zn-TCPP(Fe) NSs accompanied by the reduction of ox-TMB production. Our proposed sensing system exhibited favorable selectivity and sensitivity (LOD of 0.029 mU/mL) and has excellent potential to evaluate AChE activity in human serum samples and to screen AChE inhibitors. This colorimetric assay could provide an alternative pathway for early diagnosis and drug screening of AD, facilitating the development of AD therapy.

Protective Effects of Three Flavonoids from Dendrobium huoshanense Flowers on Alcohol-Induced Hepatocyte Injury via Activating Nrf2 and Inhibiting NF-κB Pathways.

Dendrobium huoshanense flowers have been widely used for liver protection in China. This work was aimed to discover the natural products with activity of mitigating alcoholic hepatocyte injury from Dendrobium huoshanense flowers via bioactivity-guided isolation, and to clarify the underlying mechanisms of these natural products. As a result, three flavonoids, 3'-O-methylquercetin-3-O-ß-D-galactopyranoside (1), 3'-O-methylquercetin-3-O-ß-D-glucopyranoside (2) and quercetin-3-O-ß-D-glucopyranoside (3), were firstly isolated from D. huoshanense flowers. Results exhibited that flavonoids 1-3 could enhance the cell viability, decrease the expression of ALT and AST, inhibit the cell apoptosis, alleviate the oxidative stress, and mitigate the inflammatory response of alcohol-induced L02 cells. Mechanism study exhibited that flavonoids 1-3 could increase the expression of Nrf2 as well as its downstream antioxidation genes of alcohol-induced L02 cells, while ML-385 (Nrf2 inhibitor) could abolish the inhibitory effects of 1-3 on alcohol-induced hepatocyte injury. Flavonoids 1-3 could also reduce the phosphorylation levels of IκBα and NF-κB p65 of alcohol-induced L02 cells, while SC75741 (NF-κB inhibitor) could not enhance the inhibitory effects of 1-3 on alcohol-induced L02 cells injury. The data above indicated that flavonoids 1-3 could inhibit alcohol-induced hepatocyte injury, which might be attributed to alleviating oxidative stress and mitigating inflammatory response by activating Nrf2 and inhibiting NF-κB pathways.

First report of Leaf Spot Caused by Paramyrothecium foliicola on Tomato (Solanum lycopersicum) in China.

Tomato (Solanum lycopersicum) is a staple vegetable across the world. In October 2019, leaf spots were observed on tomato (cv. Tianmi) in a greenhouse in JiZhou District Tianjin, China(117°10 'E; 39°55 'N). Symptoms initially appeared as small brown spots, which gradually expanded and turned into circular, oval or irregular spots (some spots with distinct concentric zones). In severe cases, some spots coalesced and eventually covered the whole leaf. Disease incidence ranged between 12 and 18%. Twenty symptomatic leaves from five plants were collected and cut into small pieces, surface disinfested in 2% NaClO for 60 s, rinsed three times in sterile water, and subsequently plated on potato dextrose agar (PDA). Plates were incubated at 25°C in the dark for 7 days. A total of 102 isolates were obtained and 92 isolates had the same morphology. Colonies were initially white with abundant aerial mycelia and formed sporodochia with conidial masses in olivaceous green concentric rings. All isolates formed single-celled, hyaline, and rod-shaped conidia were 4.91 to 7.43 (avg. 6.53±0.72) × 1.41 to 2.45 （avg. 2.11±0.30）µm with rounded ends (n=50). Conidiophores were highly branched. These characteristics resembled a Paramyrothecium-like fungus (Lombard et al. 2016). The genomic DNA of three representative single-spored isolates TJJXPF1-3 were extracted and the internal transcribed spacer (ITS) region, ß-tubulin (tub2), large subunit ribosomal RNA (LSU), calmodulin (cmdA) and translation elongation factor 1-alpha (tef1) genes were amplified and sequenced using the primer pairs ITS4/ITS5 (White et al. 1990), Bt2a/Bt2b (Glass and Donaldson 1995), LR0R/LR5 (Rehner and Samuels 1995; Vilgalys and Hester 1990), CAL-228F/CAL2Rd (Carbone and Kohn 1999; Groenewald et al. 2013) and EF1-728F/EF2 (O'Donnell et al. 1998), respectively. All sequences were deposited in GenBank (ITS: MW463444, OM368178, OM368179; tub2: MW269542，OM714930，OM714931; LSU: OM349050, OM397398, OM390582; cmdA: MW280443, OM350474, OM350476; tef1: MW560083, OM350475, OM350477). BLASTN analysis showed 99.3-100% similarity with reference isolate QB1 of P. foliicola (MK335967, MT415353, MT415362, MT415356 and MT415359). Multilocus phylogenetic analysis showed that TJJXPF1-3 best grouped with the P. foliicola clade, which was identified by morphological characteristics and phylogenetic analysis. To fulfill Koch's postulates, pathogenicity tests were conducted by spray-inoculation with a conidial suspension of isolate TJJXPF1 prepared with distilled water (1×105 conidia/mL) on five 45-day old tomato plants. Three healthy plants were sprayed with sterile water as control. All treatments were incubated in an artificial climate chamber (25°C, 80% RH, 12h light/12h dark ). After two weeks, leaf spots were observed on all inoculated plants, which were similar to those in the greenhouse of JiZhou District, while control plants remained asymptomatic. Additionally, the pathogens were reisolated from symptomatic leaves and three representative isolates TJJXPF4-6 were identified as P. foliicola. The pathogenicity tests were repeated thrice. To our knowledge, this is the first report of leaf spot caused by P. foliicola on tomato in China. This disease could be a serious threat to tomato production in the future. Our findings will help to differentiate this disease from other leaf spot-like diseases and develop disease control strategies.

Transcriptome landscapes of differentially expressed genes related to fat deposits in Nandan-Yao chicken.

Nandan-Yao chicken is a Chinese native chicken with lower fat deposition and better meat quality. Fat deposition is a quite complex and important economic trait. However, its molecular mechanism is still unknown in chickens. In the current study, Nandan-Yao chicken was divided into two groups based on the rate of abdominal fat at 120 days old, namely the high-fat group and low-fat group. The total RNAs were isolated and sequenced by RNA sequencing (RNA-seq). After quality control, we gained 1222, 902, 784, 624, and 736 differentially expressed genes (DEGs) in abdominal fat, back skin, liver, pectoral muscle, and leg muscle, respectively. Analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) showed that significantly enriched GO term and KEGG signaling pathway mainly involved cytosolic ribosome, growth development, PPAR signaling pathway, Wnt signaling pathway, and linoleic acid metabolism in abdominal fat, back skin, and liver. While in pectoral muscle and leg muscle, it is mainly enriched in phosphatidylinositol signaling system, adrenergic signaling in cardiomyocytes, cytosolic ribosome, and cytosolic part. Sixteen genes were differentially expressed in all five tissues. Among them, PLA2G4A and RPS4Y1 might be the key regulators for fat deposition in Nandan-Yao chicken. The protein-protein interaction (PPI) network analysis of DEGs showed that PCK1 was the most notable genes. The findings in the current study will help to understand the regulation mechanism of abdominal fat and intramuscular fat in Nandan-Yao chicken and provide a theoretical basis for Chinese local chicken breeding.