Pathological characteristics of enterotoxigenic Escherichia coli K88 and screening of its probiotics

在获得绿色荧光蛋白(GFP)标记的菌株K88-GFP并证明其与K88具有遗传同质性的基础上,以K88-GFP为致病菌,腹腔注射侵染小鼠,在不同时间进行眼球采血,测定血液生理生化指标,并采取不同器官或组织,培养后利用紫外光激发K88-GFP的绿色荧光,观察计数这种产肠毒素性大肠杆菌(ETEC)在小鼠体内的分布。同时通过体外抑制和活体饲喂试验,进行了益生菌的筛选。结果证实,ETEC致病菌具有较强的侵袭性,它可以侵袭小鼠肝、肾、心、肺及脑、肌肉等器官和组织,尤其可对肝、肾造成严重的损伤;筛选得到益生菌株PB JK-2,在体内外均对K88具有较好的抑制作用。Healthy Kunming mice were infected with Escherichia coli strain K88-GFP,which was marked with green fluorescence protein(GFP) gene and shared homology with enterotoxigenic E.coli(ETEC) K88,by intraperitoneal injection.Eyeball blood of the infected mice was sampled and the standard blood biochemical indexes were determined at different moment post-injection.Different organs and tissues of the mice-killed at different time points post-injection were sampled and cultured.Then,the distribution of the ETEC in the mice was observed by ultraviolet radiating of the green florescence of K88-GFP clones.Probiotics were screened by using inhibition test in vitro and mice feeding test in vivo.The results showed that the ETEC was invasive,and could infect murine liver,kidney,heart,lung,brain and muscles but damage seriously the liver and kidney.Probiotics strain PB JK-2 was selected and proved to suppress the pathogeny K88 well in vivo and in vitro.福建省青年科技人才创新计划项目(2005J064

Tissue Culture Technique of Acacia mangium Elite Trees

作者简介: 黄烈健( 1971—) ，男,博士,副研究员,研究方向为林木遗传育种与森林培育，电话: 020-87033463,E-mail: hlj@ ritf.ac.cnTaking Acacia mangium elite trees as explants，the tissue culture technique system for dormant bud of 3-5 year-old elite trees was established． The system includes the germination-inducing of the dormant buds，the multiplying of shoots，the rooting of adventitious shoots，and the pre-treating and transplanting of seedlings． The medium MS + Sucrose 30 g·L－1 + BA 0.5 mg·L－1 + NAA 0.1 mg·L－1 was used for inducing the dormant buds; the medium MS + BA 1.0 mg·L－1 + NAA 0.05 mg·L－1 was used for multiplying; while the medium 1 /2 MS +IBA 2.0 mg·L－1 + NAA 0.5 mg·L－1 was used for rooting.The results revealed that it was viable to producing field seedlings with micropropagation.Although the branches with dormant buds harbored many kinds of microbes and the adventitious shoots were not easy to root，20% to 30% healthy germination could be yielded and the rooting rate of adventitious shoots could be higher than 85%.Pretreated with ABT powder ( rooting hormone) ，both the rooting rate and survival rate of adventitious shoots were nearly 100%．国家“十一五”科技支撑计划项目(2006BAD01A15-5,2006BAD32B010-3); 农业科技成果转化项目(2009GB24320482

Study on the Self-incompatibility of Cauliflower(Brassica oleracea L.var.botrytis L.)by Molecular Markers

运用随机扩增多态性DNA(RAPD)和inter简单重复序列(ISSR)等分子标记技术,分别对5组花椰菜自交不亲和系和对应的自交亲和系的基因组进行指纹差异分析.采用10个10mer随机引物和5个ISSR引物进行PCR扩增,结果表明,扩增片段分子量在0.3～5kb之间,指纹图谱的稳定性和重复性较好.经过3次以上重复发现,ISSR4引物扩增图谱中,自交亲和系比不亲和系多扩增出3800bp片段;ISSR6引物扩增图谱中,除第3组外,不亲和系比亲和系多扩增出1100bp片段.结果表明,花椰菜自交不亲和系与自交系基因组DNA之间存在差异,扩增出的差异片段与花椰菜自交不亲和性相关,并可作为自交不亲和系和自交亲和系育种筛选的分子标记.初步探讨了花椰菜自交不亲和性的遗传机制及RAPD和ISSR技术在自交不亲和系选育过程中的应用前景.Genetic differences between self-incompatible and correspondent self-compatible among five lines of cauliflowers were analyzed by using random amplified polymorphism DNA (RAPD) and inter-simple sequence repeats (ISSR).Ten selective primers were used in RAPD analysis and five selective primers were used in ISSR analysis.The results show that the molecular weight of amplified fragments were between 0.3 kb and 5 kb,and the genetic fingerprint could be reproductive for more than 3 times.It was found that the self-compatible lines amplified a 3 800 bp fragment with ISSR4 primer but self-incompatible lines can not;the self-incompatible lines amplified a 1 100 bp fragment with ISSR6 primer but self-incompatible lines can not except one group.The different bands were related to cauliflower self-incompatibility.The genetic mechanism of cauliflower self-incompatibility and the application of RAPD and ISSR in breeding of self-incompatibility cauliflower were preliminary discussed.教育部科技重点项目(104105);; 福建省青年科技人才创新项目(2001J033)资

Review on Achievement to Fusarium Wilt Disease in Agricultural Crops

对中国农作物枯萎病的研究进行综述.主要包括枯萎病的发生与危害、寄主范围、致病力分化、病原菌鉴定、生理小种、生理研究、生化防治等.The present paper dealt with the review of achievement to Fusarium wilt disease in agricultural crops in China. The discussion topics included the damage of Fusarium wilt disease to crops,host ranges,identification of the disease,differentiation of pathogenicities,separation of races and forms,physiological research,chemical and biological control etc.138 references were cited in the paper.国家863计划项目(2002AA244031 2);; 福建省计委项目(闽计农经[2002]48号)资

Study on the Transformation and Expression of Green Fluorescent Protein Gene in the Enterotoxigenic Escherichia coli

通过电击转化法,将带有gfp标记基因的pGLO质粒成功转化到肠毒素型大肠杆菌K88、K99中,经过紫外检测及荧光显微镜检测均发现转化菌株发出绿色的荧光,表明gfp基因得到稳定、高效的表达.进一步通过PCR分子鉴定、血清型鉴定、微生物学特性分析均表明,转化菌株与原始菌株是一致的.该转化菌株将为研究肠毒性大肠杆菌的致病机理及益生素的筛选提供有效的手段.The plasmid pGLO with green fluorescent protein gene(gfp gene) was transformed successfully to Enterotoxigenic Escherichia coli strains K88 and K99 by the method of electroshock.It was observed that the gfp gene expressed stably and efficiently in transformed stains by the detection of ultraviolet radiating and fluorescent microscope.By detection of PCR amplification of a structural gene in flagellum,serological identification and growth curve analysis,the transformed stains and the origin strains were proved to have the same genetic background.Transformed strains could provide a new way for the study of pathogenic mechanism of Enterotoxigenic Escherichia coli and the selection of probiotics.福建省发展和改革委员会重大产业技术开发专项(2004-4-7)资

Effect of Bacillus cereus on Virulence of Ralstonia solanacearum for Tomato Bacterial Wilt

青枯雷尔氏菌(Ralstonia solanacearum)从其致病性上可分为能使植株发病的强致病力菌株和不能使植株发病的弱致病力菌株.利用蜡状芽孢杆菌(Bacillus cereus)与致病性稳定的青枯雷尔氏菌强致病力菌株进行共培养,处理后的青枯雷尔氏菌在TTC平板上表现为弱毒株的菌落特征,出现了致弱现象.经过16S rDNA基因序列测定,证明了用蜡状芽孢杆菌处理前后的菌株为青枯雷尔氏菌,并非是其它的污染菌株.通过回接番茄盆栽苗发现致弱前的青枯雷尔氏菌能有效引起番茄发生青枯病,而致弱后的菌株丧失致病力,不能引起番茄发病.Ralstonia solanacearum is divided into two strains from its pathogenicity,one is virulent strain which can induce plants disease,and the other is avirulent strain which can not.After cocultivated with Bacillus cereus,the virulent strain of Ralstonia solanacearum was attenuated and showed the characters of weak pathogenicity strain on the TTC medium.It is improved that the strains before and after cocultivated with Bacillus cereus are Ralstonia solanacearum,not other pollutional bacteria,by sequencing the 16S rDNA.The results showd that the original Ralstonia solanacearum could induce bacterial wilt when inoculated to tomato and the attenuate strain can not.福建省科技重点项目(2005N045);; 细胞生物学与肿瘤细胞工程教育部重点实验室开放基金资

Moleculor Clone and Functional Analysis of Betaine Aldehyde Dehydrogenase Gene of Avicennia marina

甜菜碱在植物体内以胆碱为底物,经两步酶催化反应合成,其中催化第二步反应的是甜菜碱醛脱氢酶(Betaine A l-dehyde Dehydrogenase,BADH).为研究甜菜碱醛脱氢酶基因在酵母中是否具有生物学功能,通过PCR-RACE技术从红树植物白骨壤(Avicennia marina)中分离得到BADH基因,将其转入酿酒酵母AH109中.通过对重组酵母生长曲线的测定,发现重组酵母对NaC l的耐受度由转化前的9%提高到14%,表明白骨壤BADH基因在酵母中能得到有效表达出具生物学活性的蛋白质.The synthesis of betaine in plants is two-step oxidation reaction of choline,and the the second step is catalyzed by betaine aldehyde dehydrogenase(BADH).In order to study the function of BADH gene in yeast,the BADH in Avicennia marina was obtained with PCR technique and transformed into Saccharomyces cerevisiae AH109.The salt tolerance of recombinant yeast increased from 9% to 14% by reorganizing the recombinant yeast′s growth curve.The results showed that the BADH gene can be effectively produce protein with biological activity in the recombinant yeast.国家自然科学基金(30771304);; 福建省自然科学基金(B0510002);; 国家基础科学人才培养基金项目(J0630649)资

Avirulent Strain of Ralstonia solanacearum Inducing Plant Systemic Acquired Resistance in Tomato

通讯作者: htzhou@ xmu.edu.cn; saiqunzh@163. com[中文文摘]青枯雷尔氏菌无致病力突变株RS-F523接种后能显著提高番茄对青枯病的抗性,但RS-F523在体外对强致病力菌株RS-F052没有直接的抑制作用.RS-F523接种后植株中与系统获得性抗性(systemic acquired resistance,SAR)相关的过氧化物酶(POD)活性、过氧化氢酶(CAT)活性,H2O2含量以及植株可溶性总蛋白都发生了波动变化,其变化方式和特点与RS-F052接种植株后的变化完全不同,提示RS-F523的生防机制可能与其接种后导致植物的SAR有关.[英文文摘]Tomato resistance ability to Ralstonia solanacearum RS-F052 improves after being inoculated with avirulent strain RS-F523,although RS-F523 cannot inhibit the growing of RS-F052 in vitro.The activities of POD and CAT,the amount of H2O2 and the soluble total proteins,which are all believed to associate with systemic acquired resistance,are tested in the plant which is inoculated with RS-F523.The results show that each of the indexes above fluctuates and differs from that of the plant inoculated with RS-F052,indicating that the biocon trolmechanism of RS-F523 is associated with SAR.国家自然科学基金(30771304);福建省自然科学基金(B0510002);福建省科技计划项目(2005N045);国家基础科学人才培养基金项目(J0630649)资

Effects of Ultrasonic Treatment on Pathogenicity of Ralstonia solanacearum Strain F.1.3-010703-01

用频率为40kHz,功率为250W的超声波处理野生型番茄强致病力青枯雷尔氏菌F.1.3 010703 01,结果表明:在5、10和20min的处理出现3.24%～5.54%的抑制率,而在15和25min的处理出现-1.04%～-2.82%的抑制率;强致病力菌株的弱化指数在0.4461～0.5376之间,强致病力菌株所占的比例在95.83%～96.14%之间;超声波处理不同时间的菌株回接番茄组培苗,回接发病率皆为100%;处理组与对照之间无极显著性差异(P≥0.01).表明处理菌株保持着强致病力菌株的特性.作为一种机械能量形式,超声波对番茄青枯雷尔氏菌F.1.3 010703 01的生长和致病力都没有影响.作为一种物理致弱因子,超声波对青枯雷尔氏菌无致弱作用.Effects of ultrasonic treatment on the virulent strain,Ralstonia solanacearum F.1.3-010703-01 isolated from tomato were conducted.The result showed that number of the bacteria was slightly reduced to 3.24%～5.54% under 5,10 and 20 min treatments,somewhat increased 1.04%～2.82% under 15 and 25 min treatments.After the ultrasonic treatment,the attenuation index and number ratio of the virulent bacteria were 0.4461～0.5376 and 95.83%～96.14% respectively.The moralities for all in the bioassays infecting the tomato tissue culture plantlet with R.solanacearum treated under ultrasonic were 100%.F text illustrated that there was no significant difference between treatments and control.It was concluded that,as a kind of mechanic energy, ultrasonic had no effect on the pathogenicity of R.solanacearum.All results showed that,as a kind of physical factor,ultrasonic had no attenuation ability to the virulent R.solanacearum.国家863计划项目(2002AA244031 2);; 福建省科技厅重大项目(2000Z031);; 福建省计委项目(闽计农经[2002]48号

Study on the Growing Competition Relationship Between Virulent and Avirulent Strains of Ralstonia solanacearum Isolated from Tomato

在单独和混合培养条件下,研究了番茄青枯雷尔氏菌的强致病力(RV)与无致病力菌株(RA)的生长能力的差异.单独培养时,24h前无致病力菌株的菌体生长量超过强致病力菌株,24h后强致病力菌株菌体生长量超过无致病力菌株,12h时无致病力菌株的生长速率是强致病力菌株的5.6倍,60h时无致病力菌株的菌量仅为强致病力菌株的0.37倍.混合培养时,混合比例RV RA1 and reduced when RVRA=1 or <1;the revised increased ration of the virulent in five mixture treatments were -119.57%～-33.57%.All the amounts of the avirulent strain in five mixture treatments increased,the revised increased ration were 32.03%～346.09%.It was concluded that the avirulent had higher growth ability than the virulent in short time under sole culture condition,and could inhibit the growth of the virulent under mixture culture condition,which could be used as an biocontrol agent for the short-term control efficiency of ABPS to bacterial wilt disease of tomato in the field.国家863计划项目(2002AA244031 2);; 福建省科技厅重大项目(2000Z031);; 福建省计委项目(闽计农经[2002]48号)资