Ralstonia solanacearum preferential colonization in the shoot apical meristem explains its pathogenicity pattern in tomato seedlings

Ralstonia solanacearum causes a lethal bacterial wilt disease in many plants by colonizing the vascular tissues of the hosts. Upon inoculation of tomato seedlings through either leaf or root, the wilting symptoms occur first at the apical region and then proceed downward along the shoot. The systemic order of the disease initiation and progression in the host, independent of the site of pathogen inoculation, is yet to be investigated. To understand the disease progression more clearly, we have carried out a systematic study of the pathogen localization by GUS staining of inoculated tomato seedlings, at 24-hour intervals from 0 days post-inoculation (dpi) to 5 dpi. In both inoculation methods, pathogen colonization was observed at 1 dpi at the apical meristem as well as the cotyledon leaves, where the disease initiates. As the disease progressed, colonization by the pathogen towards the lower region of the shoot was observed. Disease consistency and pathogenicity magnitude were observed to be higher using the leaf inoculation method than the root inoculation method. Several R. solanacearum transposon-induced mutants that were reduced in virulence by root inoculation but virulent by leaf inoculation were obtained. Using GUS staining, it was observed that these mutants were unable to localize in the shoot region when inoculated in the root. Our study indicates that the apical meristem and the cotyledon leaves are the first regions to be colonized in inoculated tomato seedlings, which might explain the disease initiation from this region.     

青枯菌胞外酶对木麻黄的致病作用研究

对三个致病性不同的木麻黄青枯菌ＰｓｅｕｄｏｍｏｎａｓｓｏｌａｎａｃｅａｒｕｍＥ．Ｆ．Ｓｍｉｔｈ产生的胞外酶活性测定显示,聚半乳糖醛酸酶（果胶酶）活性在各菌株之间无明显差别,但纤维素酶活性差别明显,并随菌株的致病性上升而增加,菌株致病性与酶活性具有高度正相关（ｒ＝０．９６）。     

青枯假单胞菌的一段12.8kb DNA带有特异性控制病菌在花生上致病的基因

在本工作中,通过平板筛选分离得到两个来自马铃薯的青枯假单胞菌菌株Ｔ２００２和Ｔ２０５３（对花生不致病）的利福平抗性自发突变株Ｔ２０１６和Ｔ２０１７,通过三亲本接合,ｐＧＸ１２５２分别以９．０７×１０－７、２．５４×１０－６频率被导入到Ｔ２０１６和Ｔ２０１７中,所得接合子Ｔ２１２３（Ｔ２０１６／ｐＧＸ１２５２）和Ｔ２１２４（Ｔ２０１７／ｐＧＸ１２５２）均分别能使花生汕油５２３发生青枯病,但它们侵染的植株出现病症的时间比Ｔ２００５晚３～５ｄ,说明ｐＧＸ１２５２能扩大Ｔ２０１６和Ｔ２０１７的寄主范围使之包括花生。从侵染Ｔ２１２３、Ｔ２１２４的病株中重新分离病菌,四环素抗性检测和质粒检测表明所有细菌均含有ｐＧＸ１２５２,说明在植物体内无抗生素选择压力情况下ｐＧＸ１２５２能在Ｔ２０１６、Ｔ２０１７中稳定存在。本工作进一步证实ｐＧＸ１２５２带有特异性控制青枯假单胞菌在花生上致病的与寄主专一性有关的基因。     

几种杀菌剂对番茄青枯病菌的抑菌活性

采用番茄青枯病防治中3种已登记杀菌剂和7种未登记杀菌剂对番茄青枯病菌进行室内毒力测定,并通过EC50和病原菌的抗性水平进行抑菌作用对比。结果表明:在10种药剂中,以已登记的3%中生菌素WP抑菌效果最好,EC50值为1.308 mg/L,番茄青枯病菌对其表现敏感;在未登记的7种杀菌剂中,0.5%氨基寡糖素AS的抑菌效果最好,EC50值为2.180 mg/L,番茄青枯病菌对其表现敏感。     

茄科作物新病原Candidatus Liberibacter solanacearum入侵我国的风险分析

从传入和定殖可能性两方面分析了为害茄科作物的新病原Candidatus Liberibacter solanacearum入侵我国的风险性,并利用Max Ent模型预测了其在我国的潜在适生区。结果表明:该病原菌传入和定殖于中国的风险极高;重庆、陕西大部、贵州北部、四川东部、河南大部以及山东半岛等地区为该病原菌入侵的高风险区。因此我国应加强对该病原菌的检疫工作,实行严格的风险管理。     

非寄主植物可培养内生细菌防治桉树青枯病研究

为筛选对桉树青枯病有防治效果的内生细菌,采用抑菌圈法,测定从青枯菌的非寄主植物中分离的内生细菌对青枯菌的拮抗作用,并分别在水培和土壤系统中测定其防治作用以及内生特性。结果表明:从蒜、葱、洋葱和葡萄内分离的14种可培养内生细菌中,只有菌株SH、SB、PT、C1和C2对青枯病菌具有拮抗作用;SH对C1具有拮抗作用,C2对SH有拮抗作用;在水培系统中,菌株SH对桉树青枯病的防治效果最好;在土壤系统中菌株PT、SH具有防治作用;菌株SH具有内生性,将桉苗浸于SH菌液中24 h,内生菌能侵入到桉苗茎部。     

药剂防治烤烟根茎病害新方法研究初报

烟草青枯、病黑胫和根黑腐病是中国南方烟区多发性的主要土传病害。目前,生产上主要采取综合防治措施,包括栽培抗病品种、应用化学防治和生物防治等,但由于都存在着各自的局限性而无法从根本上进行防治。2004—2006年先后采用“单药灌茎”、“两插两灌”和“三药灌茎”等药剂防治方法对烤烟青枯病、黑胫病和根黑腐病进行防治。结果表明:“两插两灌”和“三药灌茎”对三种根茎性病害的防治效果明显。     

Changes in the nitrogen localized in the lamellae systems and stroma of rice chloroplast accompanying the stages of growth

The change in the nitrogen localized in the lamellae systems and stroma region of the rice chloroplast was pursued throughout the stages of growth and, further, the relationship between the composition of the chloroplast and leaf N content was examined. A common trend accompanying the stages of growth is recognized: that in the early stages of growth the amount of nitrogen existing as a form of stroma N is larger than that of lamellae N. Thereafter, stroma N tends to decrease in relation to lamellae N and finally, lamellae N becomes a dominant nitrogenous fraction in the chloroplast. The change in the composition of the chloroplast seems to be brought about by a change in the nitrogen metabolism of the chloroplast by which the distribution of absorbed nitrogen among the nitrogenous fractions of the chloroplast differs.     

Suppression of Ralstonia solanacearum in soil following colonization by other strains of R. solanacearum

The effect of prior colonization of a sterile loam soil and a sterile clay loam soil by individual soil bacteria on the subsequent growth of a bacterial wilt pathogen Ralstonia solanacearum YU1Rif43 (tRNA type III: Seal et al. 1992: Appl. Environ. Microbiol., 58, 3759–3761) was investigated. Various strains, belonging to the same type, the same species, the same genus, Gram-negative, Grampositive, or fungi, were used. The degree of suppression of the growth of R. solanacearum YU1Rif43 was markedly different depending on the species that had previously colonized the soil, hereafter referred to as priorcolonists. All the strains belonging to R. solanacearum type III suppressed the growth of R. solanacearum YU1Rif43 markedly, while strains of R. solanacearum type I and type II showed a moderate suppressive effect on R. solanacearum YU1Rif43. The suppressive effect of the strains belonging to species other than R. solanacearum, including fungal strains, was relatively limited, or some strains did not show any suppressive effect. The production of bacteriocin did not appear to be related to the strong suppressive effect of the R. solanacearum type III strains. Possible mechanisms for the suppressive effect of priorcolonists on R. solanacearum YU1Rif43 are discussed in relation to nutrients and physical sites in soil available for growth.     

青枯菌铜抗性基因copA 的功能

【目的】 植物细菌性青枯病（bacterial wilt of plants）是由茄科雷尔氏菌（Ralstonia solanacearum ）引起的一种世界性重大土传病害。作为防治青枯病等细菌性病害的重要杀菌剂,铜制剂的广泛使用造成多种植物病原细菌群体中出现了铜抗性菌株。青枯菌Po82菌株大质粒上携带了丁香假单胞菌中的铜抗性编码基因copA 的同源物,论文旨在探明青枯菌Po82菌株copA 在铜抗性、致病性等方面的生物学功能。【方法】 以青枯菌Po82菌株为研究对象,利用MEGA6.0软件包,基于邻接法构建copA 的系统发育树,探究铜抗性基因copA 在青枯菌和其他植物病原细菌中的系统进化关系。通过反向遗传学的研究手段,采用基因同源重组双交换和电击转化法,构建copA 基因缺失菌株及相应互补菌株。通过最小抑制浓度（minimal inhibition concentration,MIC）测定、RT-qPCR、Biolog代谢芯片以及致病力等基础生物学测定研究手段,解析copA 与青枯菌铜胁迫应答、代谢活性、致病性和运动性等表型生物学特征之间的关系。【结果】 同源性比对分析结果显示,copA 广泛存在于青枯菌群体中,青枯菌copA 在亲缘关系上与耐金属贪铜菌最为紧密, 与稻黄单胞菌、丁香假单胞菌和大肠杆菌的亲缘关系较远。RT-qPCR结果显示copA 的表达受到铜离子的诱导,copA 的表达量随着CuSO₄浓度的增加而增加。CuSO₄浓度为1.0 mmol·L ^-1时,基因表达量最高。铜MIC测定结果显示copA 基因缺失菌株对铜离子的敏感性增加,copA 基因缺失菌株的MIC值为0.8 mmol·L ^-1,较野生型菌株的1.2 mmol·L ^-1下降了33.3%,互补菌株恢复了铜抗性能力,表明copA 在青枯菌的铜胁迫应答过程中发挥着重要作用。与野生型菌株相比,copA 基因缺失菌株在普通NA培养基及含0.6 mmol·L ^-1 CuSO₄的NA培养基中的对数生长期的生长速率降低,表明copA 与青枯菌的生长速率相关。copA 基因缺失菌株于发病前期病情指数较野生型菌株下降,接种第10天,copA 基因缺失菌株的病情指数较Po82野生型菌株下降了11.7%。copA 的缺失导致青枯菌对α -D-葡萄糖和D-海藻糖等碳源,L-丙氨酸和葡萄糖醛酰胺等氮源的代谢利用速率降低,使青枯病发病病程延长。与野生型菌株Po82相比,copA 基因缺失菌株中III型分泌系统的转录激活因子编码基因hrpG 和hrpB ,III型效应子RipX编码基因ripX 的表达量显著下调。 【结论】 铜抗性基因copA 在青枯菌铜胁迫应答、致病性等方面发挥一定的作用,研究结果可为进一步解析青枯菌的铜抗性分子机制以及铜抗性菌株的防治提供理论依据。