Huang Yong-Ju, Qi Aiming, King Graham J, Fitt Bruce D L
School of Life and Medical Sciences, University of Hertfordshire, Hatfield, Hertfordshire, United Kingdom ; Department of Plant Pathology and Microbiology, Rothamsted Research, Harpenden, Hertfordshire, United Kingdom.
Department of Plant Pathology and Microbiology, Rothamsted Research, Harpenden, Hertfordshire, United Kingdom ; Southern Cross Plant Science, Southern Cross University, Lismore, Australia.
PLoS One. 2014 Jan 15;9(1):e84924. doi: 10.1371/journal.pone.0084924. eCollection 2014.
Quantitative resistance against Leptosphaeria maculans in Brassica napus is difficult to assess in young plants due to the long period of symptomless growth of the pathogen from the appearance of leaf lesions to the appearance of canker symptoms on the stem. By using doubled haploid (DH) lines A30 (susceptible) and C119 (with quantitative resistance), quantitative resistance against L. maculans was assessed in young plants in controlled environments at two stages: stage 1, growth of the pathogen along leaf veins/petioles towards the stem by leaf lamina inoculation; stage 2, growth in stem tissues to produce stem canker symptoms by leaf petiole inoculation. Two types of inoculum (ascospores; conidia) and three assessment methods (extent of visible necrosis; symptomless pathogen growth visualised using the GFP reporter gene; amount of pathogen DNA quantified by PCR) were used. In stage 1 assessments, significant differences were observed between lines A30 and C119 in area of leaf lesions, distance grown along veins/petioles assessed by visible necrosis or by viewing GFP and amount of L. maculans DNA in leaf petioles. In stage 2 assessments, significant differences were observed between lines A30 and C119 in severity of stem canker and amount of L. maculans DNA in stem tissues. GFP-labelled L. maculans spread more quickly from the stem cortex to the stem pith in A30 than in C119. Stem canker symptoms were produced more rapidly by using ascospore inoculum than by using conidial inoculum. These results suggest that quantitative resistance against L. maculans in B. napus can be assessed in young plants in controlled conditions. Development of methods to phenotype quantitative resistance against plant pathogens in young plants in controlled environments will help identification of stable quantitative resistance for control of crop diseases.
由于病原菌从叶片病斑出现到茎上出现溃疡症状的无症状生长周期较长，因此很难在幼苗期评估甘蓝型油菜对甘蓝链格孢的定量抗性。通过使用双单倍体（DH）系A30（感病）和C119（具有定量抗性），在可控环境下的两个阶段对幼苗期甘蓝型油菜对甘蓝链格孢的定量抗性进行了评估：阶段1，通过叶片接种使病原菌沿叶脉/叶柄向茎部生长；阶段2，通过叶柄接种使病原菌在茎组织中生长以产生茎溃疡症状。使用了两种接种物（子囊孢子；分生孢子）和三种评估方法（可见坏死程度；使用绿色荧光蛋白（GFP）报告基因可视化无症状病原菌生长；通过聚合酶链反应（PCR）定量病原菌DNA量）。在阶段1评估中，观察到A30和C119品系在叶片病斑面积、通过可见坏死或观察GFP评估的沿叶脉/叶柄生长距离以及叶柄中甘蓝链格孢DNA量方面存在显著差异。在阶段2评估中，观察到A30和C119品系在茎溃疡严重程度和茎组织中甘蓝链格孢DNA量方面存在显著差异。与C119相比，GFP标记的甘蓝链格孢在A30中从茎皮层向茎髓扩散得更快。使用子囊孢子接种比使用分生孢子接种能更快产生茎溃疡症状。这些结果表明，在可控条件下可以在幼苗期评估甘蓝型油菜对甘蓝链格孢的定量抗性。开发在可控环境下对幼苗期植物病原菌定量抗性进行表型分析的方法，将有助于鉴定用于控制作物病害的稳定定量抗性。
