烟草NbbZIP28突变体的创建及其对病毒侵染胁迫的响应

【目的】bZIP类转录因子参与植物的生长发育、激素信号、抗病性及抗逆性等多种生物胁迫过程。前期研究表明黄瓜花叶病毒(Cucumber mosaic virus,CMV)或烟草花叶病毒(Tobacco mosaic virus,TMV)侵染本氏烟(Nicotiana benthamiana)上调内质网应激（endoplasmic reticulum stress,ERs）因子NbbZIP28;NbbZIP28沉默导致病毒积累量上升,本研究旨在验证NbbZIP28对病毒侵染胁迫的响应机制。【方法】通过CRISPR/Cas9基因编辑技术和烟草遗传转染创建NbbZIP28基因突变植株,以野生型植株为对照,分别浸润接种侵染性克隆TMV-GFP、摩擦接种TMV-GFP或CMV接种液,检测突变体植株对病毒侵染胁迫的敏感性变化,接种CMV后0-48 h,采用qRT-PCR检测内质网应激相关的未折叠蛋白反应（unfolded protein response,UPR)基因的表达。本氏烟接种TMV 24 h、接种CMV 48 h后,在接种叶上浸润融合蛋白NbbZIP28-GFP,瞬时表达48 h后,采用Western blot检测病毒诱导NbbZIP28蛋白的水解激活;采用在线搜索工具PlantCARE分析NbbZIP28启动子区域中参与防卫和应激反应的顺式作用元件。【结果】CRISPR/Cas9定点敲除NbbZIP28后,目的基因靶位点缺失了10个碱基,导致翻译错误、蛋白功能变化。在正常生长条件下,转基因阳性植株与野生型无显著的表型差异。植株接种TMV-GFP后4-8 d,突变体中的病毒浸润斑亮度或初侵染点数目均显著高于野生型,扩展至新叶的速度较快。接种CMV后12-48 h,突变体中UPR相关基因BiP、PDI、CAM及NbbZIP28下游基因NF-YC2的表达量显著低于野生型;5-7 d突变体中的病毒外壳蛋白（coat protein,CP）基因表达量显著高于野生型,花叶及皱缩症状更显著。蛋白质序列比对分析显示NbbZIP28具有与拟南芥AtbZIP28相同的S1P和S2P蛋白酶水解位点。Western blot检测发现,与接种清水对照相比,TMV或CMV侵染显著促进了全长的融合蛋白NbbZIP28-GFP在S1P和S2P位点发生裂解。NbbZIP28启动子序列中包含5个参与热应激反应的顺式作用元件（heat stress response element,HSE）、3个参与低温反应的顺式作用元件（low temperature response element,LTR）、1个参与防卫和应激反应的顺式作用元件（TC-rich repeats）。【结论】病毒侵染促进NbbZIP28的水解激活并上调相关的UPR基因;NbbZIP28敲除导致植株对病毒的敏感性上升,病毒诱导的UPR基因表达被抑制。NbbZIP28为病毒侵染胁迫下的UPR调控因子,在病毒侵染早期通过上调UPR信号和提高寄主基础防卫反应而延缓病毒的侵染和增殖。     

温度对锦鲤疱疹病毒体外培养和致病性的影响

【目的】证实温度对锦鲤疱疹病毒CyHV-3体外增殖和致病性的影响。【方法】选用CyHV-3-EGFP病毒株为研究对象,采用间接免疫荧光染色法测定病毒滴度,使用荧光显微镜观察病毒体外增殖,研究病毒的热稳定性、以及温度对病毒的体外培养和体内致病性的影响。【结果】CyHV-3在4~36℃条件下具有很好的稳定性,温度超过36℃后,病毒活力随着温度升高而降低,50℃时病毒活力完全丧失。感染温度对CyHV-3病毒活力影响不大,高温(37℃)和低温(4℃)时病毒都具有感染能力。但是,孵育温度明显影响病毒活力,温度高于30℃时,CyHV-3的增殖能力减退甚至消失。鲤鱼感染CyHV-3病毒后,低温组(15℃)的死亡率为26.67%,常温组(25℃)的死亡率为73.33%,高温组(30℃)和对照组的死亡率均为0。【结论】证实了CyHV-3病毒在低温下能够增殖并且具有致病性,解释了近年来锦鲤疱疹病毒(KHVD)的暴发及流行不再受限于春秋季节的现象。     

额尔齐斯河湖拟鲤寄生虫季节动态及其优势虫种的寄生情况调查研究

按照鱼类寄生虫的常规调查及研究方法,对额尔齐斯河(中国段)湖拟鲤(Rutilus rutilus lacustris Pallas)寄生虫的种类、感染情况、季节动态及优势寄生虫种的寄生情况进行调查。结果共发现6种寄生虫,分别为维氏指环虫(Dactylogyrus vistulae Prost)、西伯利亚副双身虫(Paradiplozoon homoion homoion Bychowsky et Nagibina)、双穴吸虫幼虫(Diplostomum sp.)、绦虫幼虫(Cestode sp.larva)、对盲囊线虫(Contracaecum sp.)和椭圆尾鲺(Ar-gulus ellipticaudatus Wang),隶属于3门,5纲,6目,6科,6属。春、夏、秋三季,均发现双穴吸虫幼虫,夏季及秋季的感染率高于春季;维氏指环虫春季的感染率高于夏季,而秋季没有检出;西伯利亚副双身虫、对盲囊线虫、绦虫幼虫和椭圆尾鲺仅在夏季发现,且感染率及感染强度均不高。     

广西园林菟丝子发生危害情况调查与分析

【目的】明确广西园林植物种类及菟丝子寄主、种类和分布,掌握其发生危害规律,为园林菟丝子防控提供科学依据。【方法】2011-2013年,每年4-10月,以南宁市为中心选择16个有代表性市（县）进行园林树木及地被植物感染菟丝子实地调查,记录不同树木感染菟丝子种类、生育期、危害的植物学特征及程度,计算感染率和危害指数。【结果】调查鉴定广西园林树木88种,其中菟丝子寄主62种,发生率达70.45%,62种寄主分属39科,51属,其中桑科最多有6种,其次是豆科、大戟科各5种。广西分布有3种菟丝子,分别为日本菟丝子、中国菟丝子和大花菟丝子,还有一种无根藤,它们分别分布在16、9、7和5个市（县）,寄主分别达到38、20、15、11种,分别占寄主的61.29%、32.25%、24.19%和17.74%。南宁市和防城港市菟丝子种类分布最多,3种菟丝子和无根藤均有感染。中国菟丝子主要以种子繁殖危害,每年3月下旬开始萌芽侵染为害,日本菟丝子、大花菟丝子及无根藤主要以藤茎繁殖危害,4月初开始生长,11月果实成熟。寄主危害指数较高的园林树木依次是八月桂、福建茶、龙船花、黄素梅、小叶榕、非洲茉莉,危害指数达4.01-5.48,属于易感树木。【结论】调查鉴定广西园林树木88种,其中菟丝子及无根藤寄主共62种,分属39科,51属;广西分布有3种菟丝子,分别为日本菟丝子、中国菟丝子、大花菟丝子,还有一种无根藤,其中日本菟丝子为优势种类,为重点防控对象。每年4-10月是菟丝子及无根藤主要侵染危害期,也是防除的重点时期。     

坏死梭杆菌FN(A)p2001株小鼠感染模型的建立

将鹿源坏死梭杆菌FN(A)p2001分离株以3×108、3×107、3×106、3×105、3×104、3×103个/只等不同菌量,分别接种于6组小鼠,每组10只,逐日观察小鼠感染情况.结果表明,3×106、3×107、3×108个/只菌量感染组小鼠,在接种后3 d～8 d先后死亡,5 d后死亡小鼠的病理变化明显,并从死亡小鼠内脏及脓汁中均检到长丝状及小杆状等多形态典型的坏死梭杆菌;3×104个/只和3×105个/只菌量感染仅表现体重减轻,3×103个/只菌量感染无明显临床表现.由此表明,坏死梭杆菌FN(A)P2001分离株具有很强的感染毒力;对小鼠的最小致死量为106个/只菌量;腹腔接种是适宜的感染途径.从而建立了坏死梭杆菌分离株小鼠感染模型.     

Gene Discovery and Characterization from Non-sequenced Medicinal Plants

Plant-derived secondary metabolites still constitute important resources for currently prescribed drugs and for the discovery of active principles which are effective in new indication areas. Promising compounds are often exclusively found in wild medicinal plants and the classical plant-based pharmaceutical production approach has been hampered by low yields, lack of reproducibility and the high complexity of plant extracts （Rischer and Oksman-Caldentey, 2005）. Influencing the accumulation of target compounds by breeding or genetic engineering is greatly impeded because in most cases the general biosynthetic routes are incompletely understood.     

A role for Toll-like receptor 4 in the host response to the lung infection of Yersinia pseudotuberculosis in mice

Although a Yersinia pseudotuberculosis (Yptb) lung infection model has been developed to study Y. pestis pathogenesis, it is still necessary to establish a new animal model to mimic the pathophysiological features induced by Y. pestis infection. Here, we provide a new lung infection model using the Yptb strain, IP2777, which displayed rapid spread of bacteria to the liver, spleen, and blood. In addition, we examined whether TLR4 is involved in Yptb-induced pathogenesis in the lung infection model of mice we generated. Following lung infection of WT and TLR4-deficient mice with the Yptb strain IP2777, the survival rate, bacterial colonization, histopathology, and level of cytokines and chemokines in the lung, spleen, liver, and blood were analyzed. TLR4-deficient mice had a lower survival rate than WT mice in response to Yptb lung infection. Although the bacterial colonization and pathology of the lung were comparable between WT and TLR4-deficient mice, those of the spleen and liver were more severe in TLR4-deficient mice. In addition, the levels of TNF-α and CXCL2 in the liver and IL-6 and CXCL2 in the blood were higher in TLR4-deficient mice than in WT mice. Our results demonstrate that TLR4 is necessary for optimal host protection against Yptb lung infection and TLR4-deficient mice may serve as a better genetic model of Yptb infection for mimicking Y. pestis infection.     

宁夏农民中专学历教育探析

结合工作实际,从宁夏农民接受教育的现状入手,分析了农民学历教育存在的一些问题,探索了解决问题的办法。     

景泰川灌区次生盐渍化及其防治

景泰川灌区位于甘肃省北部,景泰县内。1974年景泰电力提水工程建成,最大提水高度406.2米,设计流量10.5米/秒,有效灌溉面积30万余亩。本区为干旱区大陆性气候,年平均降水量186毫米,蒸发量3600毫米,且风沙多。过去由于没有灌溉条件,大面积土地长期荒芜。景电工程的建成,彻底改变了本区的自然面貌和生产条件,短短的十四年间,这里建成了荒漠中的大片绿洲。但是,随着土壤灌溉,灌区的次生盐渍化已经露头。根据第二次土壤普查统计,己经盐化面积30482亩,占总耕地面积的10.16%。因此,充分发挥提黄河水灌溉的效益,并防治土     

Genetic diversity and mixed infections of begomoviruses infecting tomato, pepper and cucurbit crops in Nicaragua

Begomoviruses were detected in Nicaraguan fields of tomato ( Lycopersicon esculentum ) and adjacently growing plants of pepper ( Capsicum annuum ), chilli pepper ( C . baccatum ), cushaw ( Cucurbita argyrosperma ) and Mexican fireplant ( Euphorbia heterophylla ) using polymerase chain reaction (PCR) and universal begomovirus primers. All tomato and Mexican fireplant plants showing symptoms were infected with begomoviruses, while only 30–46% of the pepper, chilli pepper and cushaw plants showing symptoms tested virus-positive. No begomoviruses were found in potato. The virus species were provisionally identified by sequencing 533 bp of the viral coat protein gene ( AV1 ). Tomato severe leaf curl virus (ToSLCV), Tomato leaf curl Sinaloa virus (ToLCSinV) and Pepper golden mosaic virus (PepGMV) were found to infect both tomato and pepper. A new provisional species designated Tomato leaf curl Las Playitas virus (ToLCLPV) was detected in a tomato plant. Squash yellow mottle virus (SYMoV) and PepGMV were found in cucurbits, the latter for the first time in this host. Euphorbia mosaic virus (EuMV) was detected in Mexican fireplant. Sequencing of a larger number of PCR-amplified clones from selected plants revealed intraspecific viral sequence variability, and also multiple begomovirus infections which could represent up to three species in a single tomato or cushaw plant. Phylogenetic grouping of virus sequences did not correlate with the host of origin.