作 者 ：于洋,于涛,王洋,阎秀峰
期 刊 ：生态学报 2012年 32卷 5期 页码：1370~1377
Keywords:Camptotheca acuminata seedlings, arbuscular mycorrhiza fungi, camptothecin,
摘 要 ：在前期工作中利用蜜色无梗囊霉(Acaulospora mellea)和根内球囊霉(Glomus intraradices)从接种时期角度分析了喜树碱含量与菌根形成过程对应关系的基础上,通过温室盆栽接种试验,继续观察了这两种丛枝菌根真菌接种后与喜树(Camptotheca acuminata)幼苗的共培养时间对喜树幼苗喜树碱积累的影响。分别用两种菌根真菌每隔7d接种一批喜树幼苗,第5批接种7 d后采样,获得菌根真菌与喜树幼苗共培养时间分别为35、28、21、14、7 d的喜树幼苗样品,测定了菌根浸染状况和喜树碱含量。结果表明:(1)接种两种丛枝菌根真菌均促进了喜树幼苗喜树碱的积累,表现为喜树碱含量和产量(单株幼苗所含的喜树碱量,喜树碱含量与幼苗生物量的乘积)的显著提高。(2)从接种后共培养时间的效果看,两种菌根幼苗各器官(根、茎、叶)及全株喜树碱含量和产量均呈现随着丛枝菌根真菌与喜树幼苗共培养时间的增加而增加的趋势。两种菌根幼苗的根和茎、根内球囊霉菌根幼苗的叶片和全株的的喜树碱含量和产量,在共培养时间增加至21 d时趋于稳定,而蜜色无梗囊霉菌根幼苗的叶片和全株的喜树碱含量和产量在共培养时间增加至28 d时达到最高,其后略有降低。(3)两种丛枝菌根真菌的侵染率和侵染强度同样随共培养时间的增加而增加,至共培养28 d后无显著变化。在一定共培养时间范围内,喜树碱含量和产量的变化与丛枝菌根真菌的侵染及菌根形成之间具有对应性。
Abstract:Arbuscular mycorrhizal (AM) is the most widespread form of plant symbiosis and has been found to enhance photosynthesis rate, biomass accumulation, pathogen defense, and tolerance to heavy metals and cold stress. In addition, it affects the production of plant secondary metabolites, one of which is camptothecin, an anti-cancer compound used in clinical practice. Camptothecin is a water-insoluble cytotoxic monoterpene derived from indole alkaloid and was initially isolated from the Chinese tree Camptotheca acuminate (Nyssaceae). It has gained great attention for its remarkable inhibitory activity against tumor cells. Based on our previous study on the correlation between the formation of mycorrhiza and camptothecin content in mycorrhizal C. acuminata seedlings, the effect of co-cultivation time on camptothecin accumulation in C. acuminata seedlings after inoculation with Acaulospora mellea and Glomus intraradices was investigated in the present study. Seeds of C. acuminata were sterilized and sown in sterilized matrix (a mixture of soil and sand) in the greenhouse. Seedlings with similar height and crown size were selected and divided into three groups, inoculation with arbuscular mycorrhizal fungi A. mellea, inoculation with G. intraradices and mock inoculation control. Each group was divided into five sub-groups, with an inoculation of each sub-group every seven days. The seedlings were sampled at the seventh day of the last inoculation, i.e. the co-cultivation time of each subgroup was 35, 28, 21, 14 and 7 days, respectively. Mycorrhizal colonization frequency, colonization intensity of roots and camptothecin contents and yields in C. acuminata seedlings were determined. The results showed that: (1) Camptothecin contents and yields in mycorrhizal seedlings increased significantly. This implies that the accumulation of camptothecin in C. acuminata seedlings was enhanced after inoculation with the two mycorrhizal fungi. (2) Camptothecin contents and yields in roots, stems, leaves and the whole plants increased with the co-cultivation time of C. acuminata seedlings with the mycorrhizal fungi. Camptothecin contents and yields in roots and stems of the two arbuscular mycorrhizal seedlings, and in leaves and whole plants of G. intraradices seedlings reached the highest levels after 21 days of co-cultivation, and then remained constant. While in leaves and whole plants of A. mellea seedlings camptothecin contents and yields were the highest on the 28th day after co-cultivation and dropped slightly thereafter. (3) Mycorrhizal colonization frequency and colonization intensity in two arbuscular mycorrhizal seedlings were enhanced with the increase of co-cultivation time till the 28th day, after which no significant differences were observed. Therefore, a significant correlation was observed between camptothecin contents and yields in C. acuminata seedlings and co-cultivation with arbuscular mycorrhizal fungi for up to 28 days. These results demonstrate that there is a precise correlation of camptothecin accumulation and fungal development. Our future studies will be focused on understanding of the fungal-dependent regulation of the temporal metabolic activities during mycorrhiza development and the spatial distribution of camptothecin in different tissues and cell types. Ultimately, we aim to conduct molecular genetics and engineering of camptothecin -AM fungus interactions.