菌根真菌(mycorrhizal fungi,MF)对植物土传病害有一定的拮抗或抑制作用,能提高植物对土传病害的抗/耐病性。MF菌剂尤其是丛枝菌根真菌(arbuscular mycorrhizal fungi,AMF)及外生菌根真菌(ectomycorrhizal fungi,ECMF)菌剂作为一种生物防治剂在生产实践中的应用也是人们关心和感兴趣的问题。本文对MF菌剂防治土传病害的可能性、MF生态生理学、MF菌剂的生产、MF菌剂应用方法以及今后努力的方向进行了系统分析,并对MF菌剂防治土传病害的潜力和应用范围进行了评价。

从中国南宁柑桔叶片中分离到对柑桔溃疡病菌有拮抗作用的细菌菌株Bc51。根据16S rDNA序列同源性、形态学特征和生理生化反应,将该菌株鉴定为洋葱伯克霍尔德氏菌(Burkholderia cepacia)。温度、pH值和培养基对洋葱伯克霍尔德氏菌抑制柑桔溃疡病菌生长的能力有显著影响。在温室测试中,观察到60.3%柑桔溃疡病斑的形成受到洋葱伯克霍尔德氏菌抑制。连续8次在人工培养基上转代培养,洋葱伯克霍尔德氏菌对柑桔溃疡病菌生长的抑制力没有发生明显改变。洋葱伯克霍尔德氏菌对植物病原菌有较宽的抑菌谱,表明除柑桔溃疡病外,该菌对其它作物病害的防治也具有潜在的应用价值。

2003~2005年,从云南省德宏州新兴冬作马铃薯主产区采集的马铃薯晚疫病病样中分离纯化到86株致病疫霉菌,对其代表性菌株的表型特征进行了较系统的研究。交配型实验结果表明:所测定的62个菌株都属于A1交配型,未发现A2交配型或自育型菌株。毒性基因鉴定、监测及生理小种组成分析显示:德宏州存在所有已知可鉴定的毒性基因,而且生理小种组成非常复杂;所测定的66个菌株可区分出36个生理小种,其中,出现频率最高的小种为1,2,3,4,5,6,7,8,9,10,11,占被测菌株的18.18%,在主产区潞西县和盈江县均有分布。对60个菌株的甲霜灵抗性测定结果表明:56.67%的菌株表现为敏感,25.00%的菌株中抗,18.33%的菌株对甲霜灵有抗性。本文对此实验研究结果作了初步的分析和讨论。

对4种柑橘衰退病毒源及其31个单蚜传毒分离株的CP基因做了限制性片段长度多态性(RFLP)和单链构象多态性(SSCP)分析,并对其中8个单蚜传毒分离株的CP基因进行了序列分析。实验明确了所研究柑橘衰退病毒(Citrus tristeza virus,CTV)的CPG/Hinf I RFLP组群和CPG/SSCP模式,二者能较好地对应并有效验证了单蚜传毒对CTV毒源的分离纯化作用;CP基因序列分析结果表明,相同CPG/Hinf I RFLP组群的单蚜传毒分离株间具有高度的序列相似性,而不同CPG/Hinf I RFLP组群单蚜传毒分离株间则存在较大差异;通过与已知生物学特性CTV分离株比较,初步建立了上述CP基因分子特征与病毒生物学特性之间的联系。

本研究以广东省农业科学院果树研究所隔离网室内通过嫁接分别感染CTV、CEVd和CTLV的柑橘树皮为实验材料,建立了以oligo(dT)为反转录引物的RT-PCR检测体系,成功检测到在病毒RNA 3'末端带有ploy_(A)加尾的CTV和CTLV。实验过程中,发现不具有ploy_(A)加尾的环状类病毒CEVd,同样可以用oligo(dT)反转录的RT-PCR检测出来,通过测序分析,其同源性在96%以上,并发现在CEVd序列中有一个富含A的区段。在此基础上,进一步研究了同时检测CTV、CEVd和CTLV的多重RT-PCR检测体系,该方法能为这3种病害的检测简化步骤、节省时间、降低成本。

应用等位基因特异性PCR(allele-specific PCR,AS-PCR)技术,建立了大豆猝死综合症(sudden death syndrome of soybean,SDS)病原菌北美种Fusarium virguliforme与其近似种Fusarium phaseoli(菜豆根腐病菌)的鉴别方法。针对翻译延长因子(EF-1α)基因上的3个SNP(单核苷酸多态性)位点,参照引物设计原则和等位基因特异引物(allele-specific primer,ASP)的要求设计了1对引物Fsg-α-1/2,能特异地扩增F.virguliforme,产生327 bp的电泳条带。另外,根据ITS区序列设计了1对通用引物Fu1/2,能扩增F.virguliforme和F.phaseoli,产生452 bp的电泳条带。本实验在传统的AS-PCR基础上进行改进,引入Fu1/2作为阳性质控引物,将ASP的特异性与双重PCR的严谨性相结合,建立了简便可靠的SDS病原菌鉴定方法。

水稻和稻瘟病菌互作是研究植物与病原菌互作的模式体系。本文利用3个水稻抗稻瘟病近等基因品系(CO39、C101LAC和C101A51)和2个特异性菌株(M209和M210)构成不同亲和程度的互作关系,研究了稻瘟病菌对3个水稻信号传导途径关键酶合成基因、5个防御反应病程相关蛋白基因和1个防御反应转录因子调控基因诱导表达的作用。结果表明:稻瘟病菌诱导了各种互作关系中水稻OsLOX、OsAOS和OsPAL酶合成基因的表达,水稻启动了茉莉酸和水杨酸防御反应信号传导途径。在不亲和的互作反应中,稻瘟病菌能不同程度地诱导水稻OsPR1a、OsPR2、OsPR3-1、OsPR3-2和OsPR4基因的表达,从而有效激活了防御反应系统,使水稻植株表现为抗病;而在亲和的互作反应中,多数OsPR基因的表达水平低、时间短或没有表达,水稻植株表现为感病。OsMyb基因在各种互作关系中有不同的诱导表达。说明这些防御相关基因的诱导表达可能与水稻抗稻瘟病性相关。

利用抑制差减杂交技术构建了条锈菌(Puccinia striiformis)诱导的小麦叶片cDNA文库。文库质量检测表明:差减杂交效率较高,质量较好。用DNAStar 5.0对172条高质量的ESTs进行聚类,共获得120个contigs。对功能已知的contigs分类,能量和初级代谢类占32.5%,其中与光合作用相关的基因出现频率最高;参与膜及转运、抗病与防御的基因分别位于第二、三位;次生代谢、蛋白质合成加工和细胞结构建成的基因较少。通过分析抗病相关基因,初步推测HR和SAR可能为小麦抗条锈病过程中的2种抗性形式。最后利用RT-PCR对4条感兴趣基因进行分析,明确其在抗病过程中的表达模式。

电镜检查表明,接种烟草花叶病毒(TMV)表现系统侵染的烟草植株叶肉细胞和筛管内有大量病毒粒子和病毒结晶体,细胞质中出现髓鞘样结构、多泡体和次级囊泡。叶片薄壁细胞内叶绿体、线粒体等细胞器变形解体,膜结构增生。经壳寡糖(50 μg/mL)诱导处理的烟草,系统症状减轻,叶肉细胞和筛管中病毒粒子显著减少,偶见病毒结晶体;叶肉细胞中细胞器基本完好,但叶绿体内出现较大的淀粉粒;液泡内和细胞间隙出现大量电子致密物质。在未接毒和未经壳寡糖处理的植株中未见此种电子致密物,其产生可能与诱导抗病性表达有关。

本文研究了过氧化氢(H₂O₂)、一氧化氮(NO)和Ca²⁺在棉疫病菌激发子PB90诱导烟草气孔运动中的作用。1 nmol/L激发子PB90可诱导野生型烟草Bel-W3气孔关闭,且在相同条件下该激发子诱导反义抑制抗坏血酸过氧化物酶anti-APX烟草气孔关闭的孔径比野生型的更小;进一步药理学证明,抗氧化剂DTT和NADPH氧化酶抑制剂DPI(diphenylene iodonium)、一氧化氮合酶(NOS)抑制剂L-NAME、Ca²⁺螯合剂EGTA和Ca²⁺信号途径中CaMPK Ⅱ的竞争抑制剂KN93与磷脂酶C抑制剂U73122都能抵消PB90诱导气孔关闭的效应。推测该激发子PB90通过NADPH氧化酶途径形成H₂O₂、NOS途径形成NO和Ca²⁺信号途径进而诱导气孔关闭。表明H₂O₂、NO、Ca²⁺在激发子PB90诱导气孔关闭信号传递中作为第二信使起着重要的作用。

本研究克隆了来源于马铃薯Y病毒坏死株系(PVY^N)外壳蛋白(CP)基因(PVY^N CP)5'端和3'端的反向重复cDNA序列,并构建了植物表达载体pROKⅡ-5'IR和pROKⅡ-3'IR,利用农杆菌介导的叶盘法转化烟草NC89,分别获得166和126株转基因烟草。抗病性试验表明,转化PVY^N CP基因3'端茎50 bp(环50 bp) hpRNA(hairpin RNA)的烟草抗病率达69%,而转化PVY^N CP基因5'端相同片段长度的hpRNA的烟草却全部发病。Southern blot结果表明,转基因植株的抗病性与转基因的拷贝数之间无明显的相关性;Northern blot结果表明,目的片段转录产物的积累量与植株的抗病性呈负相关,证明所获得的抗病性是RNA介导的抗病性。研究结果证明PVY^N CP基因的3'端比5'端更能有效地诱发基因沉默,并且hpRNA茎部的长度可减少到50 bp。该结果为探索利用CP基因的最短长度和有效部位的基因片段来培育多抗病毒转基因植物提供了依据。

小麦条锈病是我国小麦生产中的重要病害,对该病防治的关键是做好监测工作。本研究利用ASD手持野外光谱仪[ASD FieldSpec HandHeld FR(325~1 075 nm)]和热气球在近地与高空研究了发病小麦冠层的高光谱遥感数据特征,获得了近地和对应高空2个不同平台光谱数据。经比较分析,发现高空数据与近地数据之间存在明显而有规律的变异关系,即高空获得的光谱反射率在可见光谱区域明显大于近地获得的光谱反射率。进一步对差异最显著的绿峰580 nm和黄边610 nm处数据进行回归分析,获得了高空光谱反射率值与近地光谱反射率值之间的回归模型,为进一步研究利用高平台遥感监测小麦条锈病奠定了一定的理论基础。

病毒与宿主相互作用中,存在宿主主动或被动摆脱病毒的机制。本研究用光照诱导-单孢分离、原生质体再生和菌丝尖端分离3种方法对携带低毒病毒的板栗疫病菌(Cryphonectria parasitica)的EP721、EP713和Euro7三个菌株的脱毒效率进行了比较。结果表明:经光照诱导-单孢分离后所有菌株均可获得脱毒菌株,而原生质体再生对EP721的脱毒效率最高,是继光照诱导-单孢分离脱毒法之后另一种新的有效的脱毒方法,特别适用于不产孢的真菌脱毒。

观测了枯草芽孢杆菌BS、GF1和荧光假单胞菌LX1、BCA1 4株生防菌对7株来源不同的辣椒疫病病菌的拮抗作用和防治效果。结果表明:BS对辣椒疫病病菌菌丝的拮抗作用显著高于其它3株生防菌,抑制率达到33.2%~59.4%;温室防病试验中,BS喷雾处理和BCA1灌根处理对辣椒疫病具有明显的防治效果,防效分别达到56.83%和57.81%。BS抗菌粗提物对病菌菌丝生长具有明显的抑制作用,抑菌作用与抗菌粗提物稀释倍数呈负相关,稀释10倍时抑制率达到100%。BS抗菌粗提物处理使辣椒疫病病菌菌丝分枝增多,分枝间距明显缩短,分枝顶端原生质消解;同时可以显著抑制辣椒疫病病菌游动孢子的萌发速度和萌发率。对绿色荧光蛋白标记的BS菌株(gfp-BS)在辣椒幼苗各部位定殖情况的检测表明:该菌可以通过种子细菌化成功定殖于辣椒植株体内,出苗60 d后仍能检测到荧光标记的菌落,其定殖量维持在10³ cfu/g以上。

Effect of essential oil from Asarum heterotropoides Fr. Schmidt var. mandshuricum (Maxim.) Kitag on hypha growth of sixteen plant pathogenic fungi was tested at concentration of 500 μg/mL. The inhibi-tion rates to fourteen plant pathogenic fungi were above 50%, which showed the essential oil had broad-spec-trum antifungal activity. According to the toxicity curve of essential oil to Alternaria humicola, the EC₅₀ to mycelium growth and spores germination were 326.86 μg/mL and 406.24 μg/mL respectively. Furthermore, the effect of essential oil on cell membrane potential of A. humicola was tested by the conductivity method. The results showed the action of essential oil was changing osmosis of cell membrane of fungi.

Rose leaves with virus symptoms (mosaic, ring spot and line pattern) were detected by DAS-ELISA and RT-PCR. The results showed Prunus necrotic ring spot virus (PNRSV) infection in the leaves. CP gene of PNRSV isolated from Yunnan province was cloned and sequenced. Analysis of sequence showed that CP gene of PNRSV-yunnan contained 683 nucleotides encoding 165 aminoacids (EMBL accession num-ber:AY684271). Nucleotide similarity of CP gene was more than 98% between PNRSV-yunnan and group I isolates, and less than 95% between PNRSV-yunnan and group Ⅱ and Ⅲ isolates. According to the results, PNRSV-yunnan was identified as a member of group I.

Endophytic bacteria B20-006, an antagonistic microbe, was applied to coat maize seeds and resul-ted in 67.9% control effect on maize sheath blight caused by Rhizoctonia solani. Moreover, B20-006 was en-sured the ability to penetrate roots, and in turn move along vascular elements of maize plants. For further study of biocontrol mechanism of the antagonistic agent, the changes of major defensive enzymes (PAL, POD, SOD and EST) in specific activity were assayed in plants due to interaction between B20-006 and host maize.

Fifty six endophytic bacteria strains were isolated from the roots of wheat, and one strain named G-32 was screened with antagonism on pathogenic fungi of Gaeumannomyces graminis var. tritici. Preliminary characterization indicated that G-32 was belonged to Bacillus cereus. The inhibitory action to the fungi was as-sayed in lab by using the fermentable extracts of antagonistic endophytic bacteria. The results showed that my-celium of G. graminis var. tritici growed slowly and the biomass reduced 89% compared with the control 6 d after inoculation. The hyphae were deformative which broke into pieces. Colonization was studied with dual-resistant label which suggested that the endophytic bacteria strain could colonize in the root systems of wheat. The control efficacy with G-32 strain against the disease of the wheat take-all in pot tests was better than that of common used chemical fungicide Diniconazole.

The differential genes expression of adult plant resistance gene of TcLr35 was analysed by using cDNA-AFLP technique. The results showed that 33 of 843 amplified bands were differential ones in different growth stages of TcLr35. A polymorphic band amplified by primer pair E-TG/M-CTG was found at different leaf-growing stages from the second to the sixth leaf of TcLr35 and absent in Thatcher and the seedling stage of TcLr35 on TcLr35 without inoculated with Puccinia recondita. Reverse Northern blotting test suggested that band E-TG/M-CTG-192 was a piece of special gene fragment of TcLr35. The band was cloned and sequenced subsequently. The fragment of 192 bp produced by E-TG/M-CTG contained an open reading frame (ORF), but did not show homologous to the published wheat gene sequences.

植物寄生线虫种类繁多、危害严重,给世界农业生产造成巨大经济损失。目前防治线虫通常采用轮作、杀线虫药剂、生物防治和应用抗性品种等措施,但存在一定局限性。随着植物与寄生线虫之间相互作用机制的深入研究以及分子遗传操作技术的逐渐成熟,利用基因工程技术构建环保、方便、有效的线虫防治策略逐渐成为研究热点。本文从植物抗线虫基因、抑制线虫的外源活性蛋白、特异表达启动子,以及RNAi介导的抗线虫基因工程策略等方面,简要概述了国内外近年来植物抗线虫基因工程新途径研究进展以及在分子抗病育种中的应用。

应用营养亲和性方法研究了尖孢镰刀菌菌株抗氯酸盐突变体和nit突变体的诱发规律及分布特性,以及菌株营养体亲和群(VCG)的划分。研究表明,不同寄主(黄瓜、甜瓜和西瓜)分离的尖孢镰刀菌菌株形成的抗氯酸盐突变体数目差异不显著,平均为每个接种点产生0.89~0.98个;但寄主不同部位(根部、茎基部和茎中部)分离的菌株间差异显著,形成的数目分别为1.27、0.75及0.76个。菌株产生的nit1突变体比例(75.40%)显著高于nitM突变体比例(13.17%);nit1突变体数目会因菌株的寄主及菌株寄主部位的不同而有差异,寄主为黄瓜、甜瓜和西瓜的菌株产生的比例依次为67.73%、83.71%和77.50%,根部、茎基部及茎中部分离菌株产生的比例依次为81.82%、78.48%和68.64%,而在致病菌株与非致病菌株间无显著差异,分别为74.43%和79.63%;nitM突变体数目受菌株寄主影响较小,所占比例在11.17%~13.92%之间;而在寄主不同部位分离的菌株及致病菌株与非致病菌株间差异显著,分离自茎基部的菌株所占比例最高为15.97%,茎中部菌株所占比例最低为9.87%,致病菌株与非致病菌株所占比例分别为14.08%和9.26%。供试菌株分为7个VCGs,其特点为来源于不同寄主的尖孢镰刀菌菌株互不亲和,同一寄主的致病菌株与非致病菌株均不亲和,同一寄主不同部位分离的菌株可亲和。

柑橘衰退病毒(Citrus tristeza virus,CTV)存在复杂的株系分化现象,运用弱毒株交叉保护防治柑橘衰退病时需了解不同类型柑橘对CTV构成的影响。作者将CTV分离株TR-L514嫁接接种到6类柑橘植物上获得了18个亚分离株,并对这18个亚分离株和TR-L514进行了指示植物鉴定,p25基因的限制性片段长度多态性(RFLP)和单链构象多态性(SSCP)分析,p23基因的序列比较。结果表明,CTV株系对不同柑橘类植株的适应性存在差异。TR-L514嫁接到不同柑橘类植株其构成会发生变化,在甜橙上可以同时检测出p25//HinfⅠ RFLP第1和6组群,并具有比在其它4类柑橘上更为复杂的SSCP谱型构成,因此甜橙可能更适宜于CTV的增殖。TR-L514及18个亚分离株的p23基因序列差异可能与不同类柑橘植株的适应性有关。

从云南砚山刺茄(Solanum aculeatissimum)上分离到病毒分离物Y322,全序列测定表明,Y322 DNA-A全长2 730个核苷酸,共编码6个ORF。基因组比较发现,Y322 DNA-A与中国番茄黄化曲叶病毒(TYLCCNV)各分离物的同源性最高(88.3%~99.2%),而与其它双生病毒的同源性均在79.6%以下,表明Y322是TYLCCNV的一个分离物。利用DNAβ的特异性引物在Y322中扩增到DNAβ分子(Y322 β),序列分析表明,其全长1 331个核苷酸,与TYLCCNV伴随的DNAβ的同源性最高,达75.1%~93.1%,而与已报道的其它种类的DNAβ的同源性均低于55.4%。这是首次在刺茄中检测到中国番茄黄化曲叶病毒。

将稻瘟病菌10个两性可育菌株,包括GUY11、KA3、KA9、KA7、6023、2539W、8773R-19、8773R-27、8113R-2和8113R-10相互交配,结果表明,10个菌株育性好,交配型明确;2539W已丧失雌性功能,不再是两性菌株。原先由6023/2539W测为可育的24个福建省田间稻瘟病菌菌株分别与另外8个两性菌株交配,结果得出不同两性菌株测得同一组稻瘟病菌菌株的育性差异很大,这可能是由于两性菌株的交配能力强弱差异以及两性菌株与待测菌株之间的亲和性不同引起的。分析认为,GUY11和KA3最适于测定稻瘟病菌群体的育性和交配型,并用其测定了227个来自福建省田间稻瘟病菌菌株的育性。结果表明,福建省稻瘟病菌群体可育菌株率为81.1%,其中81.0%为交配型MAT1-2,19.0%为MAT1-1。研究结果将有助于进一步认识稻瘟病菌的群体结构特点及其演化规律。

明尼苏达被毛孢(Hirsutella minnesotensis)是中国大豆胞囊线虫重要的内寄生菌,本研究采取固体和液体2种培养方法测定了6种天然培养基、20种碳源、19种氮源和9种维生素对其生长、产孢和孢子萌发的影响。结果表明:H. minnesotensis在酵母葡萄糖琼脂(Yeast dextrose agar,YDA)培养基上生长最快,在麦芽浸膏琼脂(Malt extract agar,MEA)培养基上产孢最好,胰化大豆琼脂(Tryptic soy agar,TSA)培养基既不适合生长,也不适合产孢。在碳氮源研究中,Melibiose在固体培养基上显著促进H. minnesotensis气生菌丝生长,Glycogen是液体培养和孢子萌发最好的碳源。H. minnesotensis在以Casein为氮源的培养基上气生菌丝最多,Peptone是H. minnesotensis液体培养最好的氮源,大多数的氮源可以促进H. minnesotensis孢子萌发,但L-Cystine抑制孢子萌发,且在固体培养中不能被H. minnesotensis利用。维生素对H. minnesotensis的生长、产孢和孢子萌发有一定的促进作用。

以克隆载体pGEMT-PVY-NHC为模板,用PCR方法获得HC-Pro基因,构建表达载体pPIC9K-HC,将重组质粒经Sal I单酶切后电转化Pachia pichia GS115菌株,筛选出对G418有高抗性和在MM培养基上生长缓慢的转化子。经摇瓶发酵和1%甲醇诱导后,SDS-PAGE检测发酵液上清,在66 kD处有1个特异条带表达,目的条带蛋白占上清液总蛋白的40%。Western blot鉴定表明,表达蛋白可以和HC抗体发生结合反应。

黄瓜花叶病毒M株系在白肋烟上具有典型的症状恢复现象,本研究用提纯病毒和DAS-ELISA建立了CMV-M病毒定量检测方法。研究发现,症状严重程度与叶片中的病毒浓度呈正相关:最早发病的黄化叶每克病组织中病毒可高达790 μg,而恢复叶片上部再发病的花叶症状病叶中每克病组织中病毒也可高达508 μg,恢复叶片中病毒含量很低,每克叶片中最高也没有超过6 μg,仅为发病叶片病毒浓度的1/85~1/135,远低于根和茎中的病毒浓度。RT-PCR和生物学检测结果表明恢复叶片中确实存在具侵染活性的病毒,而且病毒在长达半个月以上一直保持很低浓度。结果表明恢复叶中可能存在有效的病毒防御机制,其具体机理有待进一步研究。

2004年春季参试的湖北、安徽2省11个芜菁花叶病毒(Turnip mosaic virus,TuMV)分离物感染4个油菜品种,病情指数幅度为26.2~76.0;秋季参试5个TuMV分离物感染14个油菜品种,病情指数幅度为38.3~55.9,均值方差分析表明,致病力差异分别达到极显著和显著水平。壳蛋白(CP)基因序列分析表明,来源于2省油菜、白菜、红菜薹、芝麻和萝卜的17个TuMV分离物与浙江分离物ZJB3序列同源性在97%以上,同属于MB类群;而另一个萝卜分离物WRS1与ZJR1、CH1和CH2分离物序列同源性在95.4%~98.7%之间,属于MR类群。类群间分离物序列同源性仅为88.0%~92.2%。遗传进化树分析表明,萝卜分离物WRS2在MB类群中单独构成一个分支,可能是MR类群和MB类群发生重组的后代。

小麦数量抗病品种Aquileja(AQ)、Libellula(LB)、Luke(LK)、Nugaines(NG)和咸农4号(XN)具有持久抗条锈病特点,本文对这5个品种的抗性组分及它们之间的遗传距离进行了研究,以铭贤169(MX)为感病对照品种。苗期室内抗性组分试验表明:LB、XN、AQ 3个品种与LK、NG 2个品种相比,前者的气孔下泡囊密度低于后者;前者的菌落长度短于后者。其中AQ和XN的吸器密度低于LK和NG而高于LB;LK和NG的气孔下泡囊密度甚至高于感病对照品种MX。田间抗性组分试验表明:5个数量抗病品种AQ、LB、LK、NG和XN的反应型(IT)、严重度(DS)、病情发展曲线下面积(AUDPC)和病斑长度(LL)值均低于感病对照品种MX;数量抗病品种AQ、LB、LK、NG、XN之间相比,AQ的IT值最高而LB最低;NG的DS和AUDPC值最高而LB最低;AQ、NG和XN的LL值高于LB和LK;AQ的病斑密度值低于NG和LK而高于XN和LB。由此可见,LK和NG没有抗侵入能力而在成株期具有抗扩展能力,LB、AQ和XN除了具有抗扩展能力外,还具有降低侵入频率能力。基于SSR DNA指纹数据的聚类分析,AQ与LB为同一类,NG与XN为同一类,LK单独为一类。由此推测,AQ或LB与其它3个数量抗病品种之间杂交的后代中可能会出现性状超亲分离,本实验室报道的AQ/NG杂交组合的结果证明了这一点。本文为进一步的抗病遗传育种工作提供了有用信息。

经过9年在塔里木盆地边缘干旱区对伽师县等3个相互隔离的甜瓜霜霉病流行系统的实地调查研究,提出了甜瓜霜霉病远距离传播的三级空间结构。一个甜瓜霜霉病流行系统单元是在一个绿洲内,围绕可提供初侵染病原的黄瓜温室区而形成的,其空间结构分为区系(包含若干种植区)、种植区(包含若干田块)和田块三级结构。伽师甜瓜霜霉病流行区系分为3个种植区,甜瓜霜霉病每年春季最先从位于种植一区发生霜霉病的黄瓜温室附近的甜瓜开始发生,之后甜瓜霜霉病由近向远逐步传播。种植二区和种植三区甜瓜霜霉病的初发病时间(初发病中心出现的时间)比种植一区的分别平均晚20和45d,发病中心与初菌源地的平均距离分别是56.4和106.5km,表明此地甜瓜霜霉病菌远距离传播经历了从种植一区到种植二区,再从种植二区到种植三区的传播过程。

从油菜植株体内分离出的内生细菌BY-2,经过生物学鉴定为枯草芽孢杆菌(Bacillus subtilis)。用BY-2回接油菜,重新分离到具有BY-2相同形态特征和抑制病原真菌能力的内生菌株。油菜接种后的第10 d,体内的BY-2菌数达(2.24~9.02)×10³ cfu/g鲜植株,25 d仍然保持在(3.13~8.59)×10³ cfu/g鲜植株。BY-2与油菜核盘菌[Sclerotinia sclerotiorum(Lib.) de Bary]对峙培养可以形成直径为3.1 cm的抑菌圈;可使油菜核盘菌菌丝细胞浓缩变短,细胞壁破裂,原生质外溢,从而抑制真菌生长发育;同时还能抑制菌核的萌发,抑制率达60%~70%;在油菜离体叶片试验中,BY-2对菌核病的防治效果达100%。

利用高效液相色谱筛选到能够同时产生IAA和GA₃的细菌菌株1株,产GA₃的细菌菌株2株。生测结果显示,菌株CX-5-2对小麦的促生长作用最明显,10倍稀释发酵液处理比清水对照处理可使麦苗高度增加10.54%。利用反相高效液相色谱法明确了菌株CX-5-2在不同培养条件下产生GA₃和IAA的积累情况;通过细菌16S rDNA的保守序列鉴定,菌株CX-5-2为假单胞菌属。本研究表明利用高效液相色谱可快速高效地筛选产激素类植物促生菌。

The 3' terminal genomic 1 611 nucleotides of Beet mosaic virus Xinjiang isolate (BtMV-XJ) from China was determined (GenBank accession number DQ345522). The sequence started within a single open reading frame which was expected to encode part C-terminus of NIb protein of 201 amino acids, complete cap-sid protein (CP) of 276 amino acids and followed by a 3' untranslated region (UTR) of 168 nucleotides. The comparison of sequence diversity among BtMV-XJ and other three previously reported isolates revealed that it shared 98.3%, 98.1%, 91.4% nucleotide identity, and 99.79%, 99.58%, 97.69% deduced aa sequence identity with the corresponding 3' terminal regions of BtMV-SL, BtMV-UK and BtMV-US, respectively, and that most nucleotide mutations were silent with no effect on aa sequence. In addition, a virus-specific and rapid RT-PCR detection method of BtMV was also developed based on the nucleotide sequence obtained.

The pathogen associated with root-rot disease of aloe, a severe disease of aloe in Yunnan pro-vince, was identified as Fusarium solani (Mart.) Sacc., according to morphology, cultural characters and pathogenicity.

Rhabditis sp. Bn strain was the first time found suppressive on plant parasitic nematodes (PPNs) in the rhizosphere of greenhouse cucumber. The nematode Bn was applied into soil at the rate of 1.0×10¹⁰ Juveniles/hm² 10 days before cucumber plants being planted in October 2004 and then once a month as treat-ment. The same volume of tap water was used as control. The density of PPNs increased sharply in control and reached 1 561 Js/100 mL soil in April, whereas it was less than 400 Js/100 mL soil in treatment, signifi-cantly lower than that of control (P<0.01). The densities of PPNs, Meloidogyne spp. and Tylenchorhynchus spp. were reduced by 70% and 87% respectively by applying Rhabditis sp. Bn strain. The root-knot index of cucumber in treatment was also reduced significantly (P<0.05) in April 2005.

The genetic diversity of 110 strains of Ustilaginoidea virens from Liaoning province and Beijing where the types of rice variety constitution were different, was assessed by AFLP. The results showed that the coefficient of strains from Liaoning and Beijing was 0.92 and 0.55 respectively, and the strains from Beijing were divided into two distinct groups. There was no specific DNA pattern for the isolates from the same rice varieties, and there was no correlation between the clusters based on genetic similarity coefficient and variety origin of isolates.

An isolate of PVS, HZ00P1, was obtained from nature infected Solanum tuberosum from Hang-zhou, Zhejiang province. It was primarily identified by DAS-ELISA and host reaction tests. 3' end partial sequence of the virus isolate was cloned. Nucleic acid sequence of the cp gene and deduced cp amino acid se-quence alignments were compared with 16 other isolates registered in GenBank. The results showed that the 17 PVS isolates were divided into two groups. Among them, two Andean isolates were classified to group Ⅱ, HZ00P1 and the other 14 isolates were assigned to group Ⅰ. Compared with other PVS isolates in group Ⅰ, PVS HZ00P1 had 93.1%-98.1% and 95.9%-99.3% homologies of nucleotide sequence and deduced amino acid sequence respectively, whereas its similarities to the group Ⅱwere 81.4%-81.7% and 93.5%-93.9%. A survey of PVS occurrence in Zhejiang province was achieved by RNA spot hybridization (RSH). Among the 30 samples collected from different areas of the province, 26.7% were found to have the infection of PVS. It is concluded that PVS has become a common virus in Zhejiang province.

The effects of inoculum concentration, leaf stages of host plant, and environmental factors on the infection of Exserohilum monoceras isolate X-27 to barnyardgrass were investigated. The results showed that the lesion was visibly appeared on the leaf when inoculum concentration above 1.00×10⁵ conidia mL-1, 1-2 leaf stage of barnyardgrass was most sensitive for infection of E. monoceras. The data also indicated that dew period was the key factor for E. monoceras infection. Almost 90% of barnyardgrass plants were killed when dew period prolonged to 48 h. Temperature and illumination period did not significantly affect the infec-tion of E. monoceras, 25-28℃ and 12 h dark period after inoculation were enough for infection.

利用细菌16S rDNA基因的通用引物对16个供试菌株进行PCR扩增,把扩增产物进行核苷酸序列测定。将获得的序列与GenBank中相关菌株的16S rDNA序列进行同源性分析。以此设计出检测A.a.c的特异性引物,并利用最大简约法构建了16S rDNA系统演化树。系统演化关系分析表明,6~9号供试菌株的16S rDNA序列与A.a.c标准菌株仅有3个位点的差异,其同源性均在99.8%以上,在构建的系统演化树上,它们聚为同一个族群。利用设计的一对特异性引物(BFB64/65),对各供试菌株进行PCR检测,结果只有A.a.c相关菌株产生扩增条带,产物大小与预期一致。

2004~2005年在云南保山香料烟上发现一种由细菌侵染引起的新病害,称为香料烟细菌斑点病。该病主要危害叶片,初为黑褐色水浸状圆形或多角形小斑点,以后病斑扩大、连片,形成不规则的较大坏死斑。从病叶上分离到了28株细菌菌株,菌株接种于香料烟上,发病症状与田间自然症状一致,并从回接病株上重新分离得到此病病原细菌。经过革兰氏染色反应、菌体形态、培养性状、LOPAT试验、生理生化特性分析和16S-23S rDNA序列分析,以及病原菌寄主范围测定,确定该病原菌为丁香假单胞菌烟草致病变种[Pseudomonas syringae pv. tabaci(Wolfet al.)Young et al.]。