马铃薯青枯病菌生化型研究及菌株接种方法的比较

试验测定了我国山东、广东和广西3省的马铃薯青枯病菌生化型,结果表明:被测菌株包含了生化型Ⅰ、Ⅱ、Ⅲ和Ⅳ4种生化型,其中生化型Ⅱ和Ⅲ为优势生化型,均占被测菌株的40%,被测菌株中未测定出生化亚型。同时,比较了通过伤根灌注法和刺茎法这两种方法在番茄幼苗上接种马铃薯青枯病菌的接种效果,结果表明:与刺茎法相比较,伤根灌注法操作简便,幼苗发病快,发病级数高,是理想的青枯病菌接种方法。     

青枯病菌研究进展

细菌性青枯病是由茄科雷尔氏菌(Ralstonia solanacearum)引起的一种世界范围的细菌性土传病害,广泛分布于热带、亚热带及部分温带地区。青枯病菌寄主广泛,可侵染50多个科的200余种植物,给作物生产带来巨大损失。充分了解青枯病菌是进行青枯病防治的重要前提。本文从青枯病菌的菌群分类、基因组结构、致病机制与致病途径、病菌检测等方面做了系统的阐述,并对基于青枯病菌及寄主植物基因组序列信息,研究和探讨青枯病菌致病机制与青枯病防治进行了展望。     

烟草青枯病菌巢式PCR检测方法的建立及应用

利用细菌16S-23S r DNA内源转录间隔区通用引物L1/L2扩增烟草青枯病菌基因组DNA,并对其扩增产物进行克隆测序,经与近缘种序列多重比对分析后,设计1对特异性引物Rs F/Rs R,用于包括烟草青枯病菌在内的15种不同细菌、5种真菌、3种卵菌基因组DNA的PCR扩增。结果表明:在优化的反应体系与程序条件下,该对引物只能从烟草青枯病菌中扩增出241 bp的特异片段,并通过序列测定验证了其准确性;将引物Rs F/Rs R与细菌通用引物L1/L2进行巢式PCR扩增后,其检测灵敏度在DNA水平上可达0.4 fg/μL,较常规PCR提高1 000倍,表明该对引物能有效地用于烟草组织及土壤中青枯病菌的检测。此结果对烟草青枯病的早期诊断、快速检测及病害流行学研究具有重要意义。     

花生种子带青枯病菌对传播青枯病的影响

从我国花生主要产区广东、湖北两省的花生青枯病病株的荚果种子中分离出了花生青枯病菌,病株种子带菌率为5％。采用人工接种花生种子的方法证明花生种子所带病菌的存活力与其含水量有着密切的关系,当种子含水量下降时,病菌的生存力也随之降低。当花生种子含水量降到8.9％时青枯病菌失去了在花生体内的生存能力。表明在播种时花生种子含水量在8.9％以上,则病菌在种子体内仍可存活,并有传播病害的能力。若在花生收获后使含水量控制在8.9％以下,有可能防止带菌种子对青枯菌的传播。     

马铃薯环腐病菌鉴定检测技术研究进展

传统的马铃薯环腐病菌鉴定方法主要是革兰氏染色法、茄苗接种鉴定法以及根据菌体的各种特征特性进行鉴定应用血清学鉴定检测马铃薯环腐病菌方法主要是乳胶致敏试验(AG)、凝胶双扩散试验(DD)、间接荧光抗体染色试验(IFAS)以及酶联免疫吸附测定试验;分子生物学方法检测马铃薯环腐病菌技术主要是应用RFLP分子标记、基因探针技术、核酸斑点杂交方法以及ITS-PCR方法.对于马铃薯环腐菌的鉴定检测不能单单依靠一种方法,因为任何一种方法都有其缺点和局限性,为得到真实、可靠的判定结果,应将多种方法取得的结果结合起来进行最终判定,因此针对马铃薯环腐菌鉴定检测建立一套完整的技术体系十分必要.     

广州地区番茄青枯病菌的系统发育分析及致病力评价

2012~2013年,于广州地区采集番茄青枯病病样并进行病原分离工作,经分子鉴定后获得9个菌株。通过PCR扩增获得了其中9个菌株的egl基因序列,采用国际新兴的青枯菌演化型分类框架进行系统发育分析,以NCBI数据库中分离自不同寄主的青枯菌菌株egl基因序列为参考序列进行系统发育树构建。结果表明:9个菌株属于青枯菌演化型I型即亚洲分支菌株的4个序列变种,分别为序列变种13、14、34、44。以高抗青枯病番茄材料‘兴农021’和敏感材料‘金冠3号’为试材,评价了其中5个菌株的致病力,结果表明3-1和18-6致病力较高。     

马铃薯环腐病菌快速检测方法(NCM-ELISA)的建立

建立检测马铃薯环腐病菌NCM-ELISA方法并组装成试剂盒,应用于检测、检疫和流行病学调查具有实践意义。本研究以马铃薯环腐病菌为抗原,制备兔抗血清,利用碱性磷酸酯酶标记的羊抗兔血清(GAR-AP)为酶标二抗,建立了马铃薯环腐病菌NCM-ELISA快速检测方法。结果表明:抗血清的最佳工作浓度为1:400,最低检出菌液浓度为1.0×106个.mL-1,特异性也比较强。因此,该方法具有敏感性高、特异性强、速度快、实用方便等特点,可以应用于马铃薯环腐病菌的快速检测和诊断。     

NCM-ELISA检测马铃薯Y病毒(PVY)技术的研究及应用

血清学方法是病毒检测的主要手段。本试验通过对马铃薯Y病毒(PVY)的提纯,免疫家兔制备PVY抗血清,并提取PYV免疫球蛋白IgG作为NCM-ELISA反应为一抗,以市售羊抗兔抗血清为二抗,在硝酸纤维素膜(NCM)上进行NCM-ELISA反应检测PVY,建立PVY NCM-ELISA检测反应体系。试验结果显示,NCM-ELISA具有反应特异性强,灵敏度高的优点,检测植物叶片样品的最高稀释度可达到1:250。通过对田间40份样品的NCM-ELISA和DAS-ELISA检测比较,其检测结果吻合率达到100%。由于NCM-ELISA方法可以将样品点在硝酸纤维素膜上,并且可贮存几个星期或将膜送到其他实验室进行检测,因此具有操作简单,使用方便,检测成本低等优点。     

Symptom expression and seed transmission of alfalfa mosaic virus and potato yellowing virus (SB-22) inSolanum brevidens andS. etuberosum

Summary Solanum brevidens showed calico mosaic symptoms in all the leaves, butS. etuberosum yellowing and yellow mosaic only in the top leaves, following graft-inoculation with alfalfa mosaic virus (AMV). The South American potato virus SB-22 infected latentlyS. brevidens following graft-inoculation, whereasS. etuberosum was not infected. AMV was not transmitted to the ovules of AMV-infectedS. brevidens andS. etuberosum. However, AMV was transmitted to the seeds ofS. etuberosum through pollen, resulting in 0.9% infected seeds.S. brevidens infected by SB-22 produced up to 20.2% of virus-infected germinable seed, and in some of the seed lots of infectedS. brevidens the germination rate was as low as 1.0%.     

青枯菌对花生发芽率及芽长的影响

花生青枯病严重影响花生的产量和品质,了解青枯菌对花生发芽和生长的影响是研究青枯病作用机制以及信号转导途径的前提.为检测强致病力青枯菌对“日花1号”种子发芽的影响,采用发芽试验对种子发芽(露白)率以及芽长进行检测,并用荧光定量PCR方法检测了植物抗病相关基因非病程相关基因表达子1(NPR1)在青枯菌胁迫条件下的表达变化规律.结果表明,经过青枯菌浸泡的种子,发芽受到明显抑制,芽长较对照组亦明显变短.在胁迫条件下NPR1的表达量在第12h达到最高,随后缓慢降低,第48h的表达量仍未恢复到初始水平,是初始水平的1.7倍.     

Optimisation of sample size for the detection of latent infection byRalstonia solanacearum in potato seed tubers in the highlands of Peru

Summary A sampling strategy was evaluated in the Andean highlands of Peru to optimise the detection ofRalstonia solanacearum in seed tubers harvested from symptomless crops. A sensitive and specific serological method developed at CIP was used to detect the pathogen in latently infected tubers. Optimum sample size was evaluated for symptomless crops after analysing various numbers of composite samples and using a binomial distribution model to calculate the detection probabilities.R. solanacearum was detected in all lots from fields with visible symptoms, so validating the detection technique. About half of the seed lots from apparently healthy fields at altitudes of up to 3,100 m were found positive for the pathogen.R. solanacearum was detected with 99% probability in samples of 350 tubers from seed lots from symptomless crops. This number of seed tubers could feasibly be processed in a seed-health test without incurring too high a cost for labour and materials.     

Response of Potato Genotypes to Bacterial Wilt Caused by Ralstonia Solanacearum (Smith)(Yabuuchi et al.) In the Tropical Highlands

Potato varietal resistance to bacterial wilt disease caused by Ralstonia solanacearum Yabuuchi et al., (Microbiology and Immunology 39:897–904, 1995) is the best management of the disease. Because the causal pathogen exhibits strong host-pathogen-environment interaction, screening the potential parents for resistance under the target growing environmental conditions is the first important step for effective resistance breeding. The objective of this study was to determine the response to bacterial wilt of selected potato genotypes currently grown by farmers in Kenya and candidate clones from the International Potato Center to identify parents that can be used in the local breeding program to develop resistant cultivars. A study was carried out at Kabete, Kenya for three consecutive seasons between November 2011 and February 2013. Thirty six potato genotypes were planted on an inoculated field at the Kenya Agricultural Research Institute (KARI), Kabete using alpha lattice experimental design with three replications. Data collected were days from planting to onset of wilting (DTOW), bacterial wilt incidence (BWI), total tuber weight (ton ha^?1) (TTW), total tuber numbers/hectare (TTN), proportion of ware sized tubers (PWTTW), proportion of symptomatic tubers based on weight (PSTTW), proportion of symptomatic tubers based on tuber numbers (PSTTN) and latent infection (LI) of the tubers. Almost, all the potato genotypes evaluated in this study were susceptible to bacerial wilt. Ranking of genotypes based on resistance differed among the three seasons. On average, the three most resistant genotypes were Kenya Karibu, Kenya Sifa and Ingabire. The study identified eight potato genotypes (Meru, Ingabire, Kenya Karibu, Sherekea, Kihoro, Tigoni, Bishop Gitonga and Cangi) to be used as promising parents for subsequent crosses. The chosen genotypes are prolific in pollen production and popularly grown by Kenyan farmers.     

Assessment of latent infection frequency in progeny tubers of advanced potatoclones resistant to bacterial wilt: A new selection criterion

Summary This paper reports results of a 3-year evaluation of CIP advanced potato clones in a bacterial wilt-infested field (race 3) in Peru. Clones resistant or moderately resistant to wilt were selected and all tubers harvested from each clone were tested for latent infection byRalstonia solanacearum using a sensitive serological technique developed at CIP. A sampling strategy to estimate accurately the frequency of infected tubers in the clones has been evaluated. This method will allow consideration of tuber latent infection as a new selection criterion in breeding for resistance to bacterial wilt. Thirteen clones were found resistant to wilt in all three evaluations (i.e.≤6% wilt), from which five had no wilt in all trials. However, all clones harboured latent infection in tubers averaging 30%. Analysing 30 tubers/clone provides an accurate estimation of the proportion of infected tubers with a high precision level.     

甜菜品种联合区域试验的试点效应分析

本文运用品种稳定性测定的原理分析了黑龙江省甜菜品种联合区域试验中各试险点对块根产量和含糖率的判别力及其稳定性。分析结果表明，具备一般的自然条件和生产条件、参试品种平均表现中等的试验点有较好的判别力和稳定性，是合适的试验点。     

茶多酚诱导铜绿假单胞菌交叉耐受性研究

检测了亚致死浓度的茶多酚（Tea Polyphenols,TP）处理对铜绿假单胞菌交叉耐受性的诱导作用。铜绿假单胞菌暴露于1βmg·mL^-1茶多酚1βh后能够显著增强细菌对多种环境条件的耐受性,包括氧化剂（1βmmol·L^-1 H₂O₂）、高温（47℃）,及酸性溶液[磷酸缓冲液（pH4.0）、含有有机酸（60βmmol·L^-1柠檬酸、60βmmol·L^-1乳酸、80βmmol·L^-1乙酸）的磷酸缓冲液（pH4.0）]。另外,通过荧光定量RT-PCR技术分析了茶多酚诱导下铜绿假单胞菌胁迫相关基因的表达情况。研究发现,茶多酚能够显著诱导铜绿假单胞菌氧化胁迫相关基因katB、sodM、ohr、lexA和recN的表达,以及热激蛋白基因dnaK、groEL、htpG、grpE和groES的表达。这些胁迫相关基因的表达很可能在细菌交叉耐受性形成过程中起到重要作用。以上研究结果表明,虽然茶多酚作为天然食品添加剂具有较高的安全性,但在实际应用过程中应充分考虑茶多酚诱导细菌交叉耐受性所导致的潜在风险,以优化食品保鲜策略。     

荞麦品种区域试验的精确度探讨

以第五轮(1997~1999年)、第六轮(2000~2002年)和第七轮(2003~2005年)全国荞麦良种区域试验(太原点)数据为资料,分组对9年试验的误差变异系数(CEV)和相对最小显著差数(RLSDα)进行计算,结果表明,苦荞组的CEV在13.7%~36.4%,平均为20.5%;甜荞组的CEV在12.1%~30.7%,平均为18.2%。相对而言,区域试验苦荞组的误差大于甜荞组。从9年的平均数来看,苦荞组试验中只能鉴别出的品种最小差异为34.1%,甜荞组中只能鉴别出品种最小差异为29.8%。