Objective:To establish a quick and convenient method for detecting human adenoviruses (Human adenoviruses, HAdV) based on immunochromatographic assay (ICA).Methods:Two antibody clones, 3C11 and 7E6 were found to bind to all tested HAdVs and then subsequently processed into ICA. The specificity and sensitivity were evaluated using representative strains of the respiratory HAdV types, including HAdV-1, 2, 3, 4, 5, 6, 7, 10 and a gastroenteric type HAdV-41 together with the original throat swabs of 10 HAdV patients confirmed by nuclear acid testing (NAT).Results:The ICA exhibited high specificity to HAdVs and its detection limitation ranged from 0.16 to 10 3 half tissue culture infectious dose (TCID 50)/ml for different types of HAdVs. All clinic samples with successful virus isolation tested by this ICA showed positive result . Conclusions:The ICA developed in the present study will be suitable for HAdVs screening in clinic setting, especially for those of respiratory types.

干扰素诱导的跨膜蛋白3 (Interferon-induced transmembrane protein 3,IFITM3)敲除小鼠(Ifitm3-/-小鼠)感染流感病毒后,表现出更为严重的病理损伤和死亡率.最近的研究发现,IFITM3蛋白表达也会影响机体针对流感病毒的适应性免疫反应效果.由于空间效应,纵膈淋巴结在机体抵抗流感病毒急性期感染的适应性免疫应答过程中发挥重要功能,本研究欲探究使用流感病毒鼠肺适应株A/PR/8/H1N1进行感染后,野生型和Ifitm3-/-小鼠的纵膈淋巴结中与免疫相关的差异表达基因.本研究对流感病毒PR8感染后第3d的纵膈淋巴结进行RNA-Seq测序.高通量测序共检测到有1 312个差异表达基因,其中上调和下调差异表达基因分别为904和408个.这些差异表达基因主要参与炎症反应与细胞凋亡信号通路、T细胞和B细胞受体信号通路、T细胞和B细胞激活与细胞因子分泌等信号通路.采用荧光定量PCR对部分免疫相关的差异表达基因进行验证,结果与RNA-Seq测序结果一致.本研究为进一步阐明IFITM3在适应性免疫应答中拮抗流感病毒的分子机制奠定了基础.

重组腺病毒是常用的基因转移载体,本文介绍一种对腺病毒基因组进行反向遗传改造的策略.拟在维持基因编码蛋白氨基酸序列不变的前提下,突变去除重组人5型腺病毒Ad5GFP基因组的PmeI酶切位点.软件分析腺病毒质粒pAd5GFP序列,选择限制性内切酶BamHI将pAd5GFP切割为大小11.7和24.6kb两个片段,24.6kb大片段自身环化形成一个质粒pAd5GB,PmeI位于其上.PmeI/AscI双酶切pAd5GB质粒,产生2.4kb和22.2kb两个片段;在引物部位引入突变的PmeI位点(由gtttaaac突变为gtttaaaT),PCR扩增得到两端各延长30bp的上述2.4kb片段,与22.2kb片段进行Gibson组装,转化E.coli TOP10感受态细胞,得到pAd5GBXP质粒.BamHI酶切pAd5GBXP,碱性磷酸酶处理,与11.7kb片段连接,还原得到腺病毒质粒pAd5GXP.PacI线性化pAd5GXP质粒,转染293细胞,拯救得到Ad5GXP病毒;酶切分析证明Ad5GXP基因组不含有PmeI位点.研究结果说明将酶切连接与DNA组装技术相结合,能够方便灵活地对腺病毒基因组进行突变改造.

Objective To study the intracellular location and characteristic of SFTSV NP protein in different phases using mini singlet oxygen generator (miniSOG) labeling technique.Metbods MiniSOG is a recently-invented genetically-encoded tag for EM.MiniSOG-fused SFTSV NP (NPSOG) gene was cloned by PCR,and inserted into pcDNA3.0 plasmid to form pTPL-NPSOG,which was used to transfect 293 cells.The transfected cells of different phases were fixed in 2.5％ glutaraldehyde in situ,stained with DAB through the photooxidation activity of miniSOG,and used to prepare ultrathin sections.Intracellular location and characteristic of SFTSV NP protein in different phases were studied by observing the sections under transmission electron microscope.Results After transfecting the plasmid with NPSOG to 293 cells,NP protein was expressed in cytoplasm and peri nucleus,and gradually aggregated,which connected with endoplasmic reticulum and Golgi apparatus to form larger volume and irregular inclusion bodies in cytoplasm.No obvious subcellular structure changes were found.Conclusions The SFTSV nucleoprotein can be expressed separately to form inclusion bodies without the assistance of other viral proteins.The formation of inclusion bodies requires the directional movement and aggregation of a certain number of NP proteins,which may involve the interaction of NP protein and host organelles during this period.