喀什边境口岸陆运卡车及集装箱污染的新冠病毒基因特征分析

新疆维吾尔自治区喀什地区作为我国与中亚和欧洲的重要陆路货运口岸,来往货物运输频繁,引入新型冠状病毒(SARS-CoV-2)风险大,对我国新型冠状病毒肺炎(COVID-19)疫情防控造成压力.2020年11月我国新疆维吾尔自治区喀什地区发生输入SARS-CoV-2导致的本土聚集性COVID-19疫情.为明确货物运输载体携带SARS-CoV-2的基因特征以及边境快速物流系统作为SARS-CoV-2传播载体的可能性,本研究对2020年11月6日-2020年11月10日期间在喀什边境口岸货运卡车及运输的集装箱采集的35份SARS-CoV-2核酸阳性样本进行SARS-CoV-2全基因组序列测定和比对分析.结果 显示,35份样本ORFlab基因Ct值的中位数(最小值～最大值)为37.64(28.91～39.81),N基因Ct值的中位数(最小值～最大值)为36.50(26.35～39.30),Reads数匹配率的中位数(最小值～最大值)为51.95％(0.86％～99.31％),病毒载量较低;35份样本中基因组覆盖度达到70％以上的共计18份.基于Pango命名法,18条SARS-CoV-2基因组序列分别属于B.1、B.1.1、B.1.9、B.1.1.220、B.1.153和B.1.465共6个不同的基因型,其中3个基因型(B.1、B.1.1和B.1.153)在喀什边境接壤或邻近的四个国家同期采集的病例样本中也有发现.核苷酸突变位点和系统进化树分析显示,同一个地点采集的样本病毒基因组相似程度高;18条序列中的4条与喀什COVID-19疫情毒株代表序列处在同一个进化分支;其中1条序列与喀什COVID-19疫情毒株基因组存在1个或2个核苷酸突变位点差异,高度同源.本研究证实喀什COVID-19疫情期间边境货运卡车和集装箱存在境外多种基因型病毒的污染,其中存在喀什COVID-19疫情毒株的祖父代病毒,高度提示边境快速物流系统卡车及集装箱作为载体携带SARS-CoV-2病毒入境造成了本土疫情,这些数据为我国边境口岸地区的新冠防控策略制定及后续疫情溯源提供了关键的参考依据.     

呼吸道合胞病毒IgM抗体AlphaLISA快速检测方法的建立

呼吸道合胞病毒(Respiratory syncytial virus,RSV)属于副粘液病毒科,是一种下呼吸道感染最主要的RNA病毒.呼吸道合胞病毒感染是引起全球婴幼儿高致死率的呼吸道感染病原,仅次于疟疾,但用于检测RSV感染的选择相对较少.本文通过抗原抗体结合原理,建立呼吸道合胞病毒(RSV)IgM抗体AlphaLISA快速检测方法.小鼠抗人IgM单克隆抗体偶联的受体微球与临床血清样品中RSV特异性IgM抗体结合,RSV特异性IgM抗体再与生物素标记的RSV抗原结合,生物素连接偶联链霉亲和素的供体微球;受体微球和供体微球的距离被拉近小于200nm,680nm荧光激发供体微球生成单线态氧,扩散给受体微球,发射波长为520～620nm的荧光,荧光强度与血清中RSV特异性IgM抗体呈正比.结果显示,该方法批内变异系数与批间变异系数均小于10％,不与其他呼吸道病原体发生交叉反应,与间接免疫荧光法具有较好的一致性,总符合率达83.33％.该方法具有微量检测、快速省时、操作简便等优势,可快速检测RSV的IgM抗体,为早期确诊RSV感染提供高效可行方案.     

SHED aggregate exosomes shuttled miR‐26a promote angiogenesis in pulp regeneration via TGF‐β/SMAD2/3 signalling

ObjectivesPulp regeneration brings big challenges for clinicians, and vascularization is considered as its determining factor. We previously accomplished pulp regeneration with autologous stem cells from deciduous teeth (SHED) aggregates implantation in teenager patients, however, the underlying mechanism needs to be clarified for regenerating pulp in adults. Serving as an important effector of mesenchymal stem cells (MSCs), exosomes have been reported to promote angiogenesis and tissue regeneration effectively. Here, we aimed to investigate the role of SHED aggregate‐derived exosomes (SA‐Exo) in the angiogenesis of pulp regeneration.Materials and MethodsWe extracted exosomes from SHED aggregates and utilized them in the pulp regeneration animal model. The pro‐angiogenetic effects of SA‐Exo on SHED and human umbilical vein endothelial cells (HUVECs) were evaluated. The related mechanisms were further investigated.ResultsWe firstly found that SA‐Exo significantly improved pulp tissue regeneration and angiogenesis in vivo. Next, we found that SA‐Exo promoted SHED endothelial differentiation and enhanced the angiogenic ability of HUVECs, as indicated by the in vitro tube formation assay. Mechanistically, miR‐26a, which is enriched in SA‐Exo, improved angiogenesis both in SHED and HUVECs via regulating TGF‐β/SMAD2/3 signalling.ConclusionsIn summary, these data reveal that SA‐Exo shuttled miR‐26a promotes angiogenesis via TGF‐β/SMAD2/3 signalling contributing to SHED aggregate‐based pulp tissue regeneration. These novel insights into SA‐Exo may facilitate the development of new strategies for pulp regeneration.     

Extensive profiling of the expressions of tRNAs and tRNA-derived fragments (tRFs) reveals the complexities of tRNA and tRF populations in plants

Evidence is emerging that t RNA-derived fragments(t RFs) are regulatory molecules. Studies of t RFs in plants have been based on conventional small RNA sequencing, and focused on profiling of t RF-5 and t RF-3 species. A more comprehensive and quantitative analysis of the entire t RF population is highly necessary. Here, we employ t RNA-seq and YAMAT-seq, and develop a bioinformatics tool to comprehensively profile the expressions of t RNAs and t RFs in plants. We show that in Arabidopsis,approximately half of t RNA genes are extremely weakly expressed, accounting for only 1% of total t RNA abundance, while～12% of t RNA genes contribute to ～80% of t RNA abundance. Our t RNA sequencings in various plants reveal that t RNA expression profiles exhibit a cross-species conserved pattern. By characterizing the composition of a highly heterogeneous t RF population, we show that t RNA halves and previously unnoticed 10–16-nt tiny t RFs represent substantial portions. The highly accumulated 13-nt and 16-nt tiny t RFs in Arabidopsis indicate that tiny t RFs are not random t RNA degradation products. Finally,we provide a user-friendly database for displaying the dynamic spatiotemporal expressions of t RNAs and t RFs in the model plants Arabidopsis and rice.     

不稳定EGFP细胞模型的构建及其在基因编辑体系评价中的应用*

目的:为了更好地评价基因编辑效率,满足高通量筛选应用中快速、高效的检测要求,在细胞上建立一个原位检测方法具有重要的意义。通过检测荧光蛋白信号强度的变化可以评价CRISPR系统在细胞中的基因编辑情况,然而这一方法的效率受限于荧光蛋白较长的半衰期。方法:将鸟氨酸脱羧酶降解结构域(含PEST序列)与EGFP融合,通过慢病毒系统感染HEK-293T细胞,获得了表达单拷贝、EGFP-PEST报告基因的稳转细胞系。结果:与EGFP相比,EGFP-PEST在细胞内的降解速度明显加快,荧光水平在4 h内显著降低。利用该模型比较了3种商品化脂质体介导的CRISPR/Cas9基因编辑效率,能够在2~4 d实现定性和定量评价。结论:这一模型能够快速、灵敏地指示基因编辑效果,可以用于不同CRISPR系统或新递送工具的高通量筛选和评价。     

Three‐dimensional reconstruction and comparison of vacuolar membranes in response to viral infection

The vacuole is a unique plant organelle that plays an important role in maintaining cellular homeostasis under various environmental stress conditions. However, the effects of biotic stress on vacuole structure has not been examined using three‐dimensional (3D) visualization. Here, we performed 3D electron tomography to compare the ultrastructural changes in the vacuole during infection with different viruses. The 3D models revealed that vacuoles are remodeled in cells infected with cucumber mosaic virus (CMV) or tobacco necrosis virus A Chinese isolate (TNV‐A^C), resulting in the formation of spherules at the periphery of the vacuole. These spherules contain neck‐like channels that connect their interior with the cytosol. Confocal microscopy of CMV replication proteins 1a and 2a and TNV‐A^C auxiliary replication protein p23 showed that all of these proteins localize to the tonoplast. Electron microscopy revealed that the expression of these replication proteins alone is sufficient to induce spherule formation on the tonoplast, suggesting that these proteins play prominent roles in inducing vacuolar membrane remodeling. This is the first report of the 3D structures of viral replication factories built on the tonoplasts. These findings contribute to our understanding of vacuole biogenesis under normal conditions and during assembly of plant (+) RNA virus replication complexes.     

Comprehensive landscape and interference of clonal haematopoiesis mutations for liquid biopsy: A Chinese pan-cancer cohort

Tumour-derived DNA found in the plasma of cancer patients provides the probability to detect somatic mutations from circulating cell-free DNA (cfDNA) in plasma samples. However, clonal hematopoiesis (CH) mutations affect the accuracy of liquid biopsy for cancer diagnosis and treatment. Here, we integrated landscape of CH mutations in 11,725 pan-cancer patients of Chinese and explored effects of CH on liquid biopsies in real-world. We first identified 5933 CHs based on panel sequencing of matched DNA of white blood cell and cfDNA on 301 genes for 5100 patients, in which CH number of patients had positive correlation with their diagnosis age. We observed that canonical genes related to CH, including DNMT3A, TET2, ASXL1, TP53, ATM, CHEK2 and SF3B1, were dominant in the Chinese cohort and 13.29% of CH mutations only appeared in the Chinese cohort compared with the Western cohort. Analysis of CH gene distribution bias indicated that CH tended to appear in genes with functions of tyrosine kinase regulation, PI3K-Akt signalling and TP53 activity, suggesting unfavourable effects of CH mutations in cancer patients. We further confirmed effect of driver genes carried by CH on somatic mutations in liquid biopsy of cancer patients. Forty-eight actionable somatic mutations in 17 driver genes were considered CH genes in 92 patients (1.80%) of the Chinese cohort, implying potential impacts of CH on clinical decision-making. Taken together, this study exhibits strong evidence that gene mutations from CH interfere accuracy of liquid biopsies using cfDNA in cancer diagnosis and treatment in real-world.     

Numerical Study of an Ultra-Broadband and Wide-Angle Insensitive Perfect Metamaterial Absorber in the UV–NIR Region

Plasmonics - Developing a simple structure using low-cost material that enables both large-scale fabrication and broadband absorption response is highly desirable but very challenging for achieving...