【作者】 林贵兵；

【导师】 万德光；

【作者基本信息】 成都中医药大学 ， 中药学， 2010， 博士

【摘要】 丹参来源于唇形科Labiatae植物丹参Salvia miltiorrhiza Bunge的干燥根及根茎,是常用的大宗中药材。具有活血化瘀,通经止痛,清心除烦,凉血消痈的功效。但该药材在生产种植中产生的连作障碍严重危害到丹参的品质与产量,威胁到丹参种植产区的持续发展,对当地经济带来巨大损失。基于连作障碍理论,通过丹参产地土壤微生物研究,对丹参产地土壤适宜性微生物结构特征进行评价,结果如下。对中江丹参产地休地土壤微生物研究,首次表明随着休地年限的增加,土壤细菌数量增加,真菌数量减少,土壤缓慢地细菌化。聚类分析表明,休地2年前后,微生物主要菌群数目变化明显。中江丹参产地土壤功能微生物群落在不同休地年限的差异显著。因休地年限不同,功能微生物群落数目改变显著；且多样性指数增加。中江丹参产地在休地2、3年间,土壤微生物生物量磷含量有着显著的变化。中江丹参产地休地不同年限的两批土壤样品的细菌与真菌群落遗传多样性聚类分析表明,2005年、2006年栽培过丹参即休地4年、3年的土壤微生物群落遗传距离较小,群落遗传多样性较为相似,聚类为第一支；而2008年、2009年栽培过丹参即休地1年、0年的土壤微生物群落遗传距离较小,遗传多样性较为相似,聚类为第二支；2007年栽培过丹参即休地2年的土壤微生物群落遗传多样性则分散于这二类中,似乎是这二类之间的过渡。这与土壤可培养微生物分析的结果基本一致。中江休地年限越长的土壤细菌DGGE条带丰富度越大,真菌DGGE条带丰富度越小。丹参区域空白土壤真菌数量显著少于丹参根际土壤真菌数量。从功能微生物群落看,丹参根际土壤的厌氧纤维素分解菌、亚硝化细菌和反硫化细菌数量均显著多于区域空白土壤的,其它功能菌群与土壤功能微生物多样性指数差异未达到统计意义水平。对全国的丹参产地土壤样品分析表明丹参根际土壤与区域空白土壤的微生物生物量磷含量差异不显著。全国丹参产地土壤的细菌与真菌群落遗传多样性较为复杂。聚类分析显示,丹参土壤微生物群落遗传距离较小,丹参根际之间的群落遗传距离差异较大,规律不明显。测序后与GENBANK序列数据库对比结果表明,细菌主要分布在alphproteobacterium,Sphingomonas,Arthrobacter属。真菌主要分布在Termitaria,Agaricus Cercozoan,Cordyceps,Geomyces,Penicillium,Iodophanus属。将根际土壤的功能微生物群落与丹参有效成分含量作逐步回归分析,结果表明丹参中咖啡酸含量与C循环功能菌相关,丹参酮IIA含量与N循环功能菌相关,丹酚酸A含量与N/P,N/S相关功能菌的比值相关。首次将土壤微生物群落结构与丹参有效成分相联系。通过对根际土壤与区域空白土壤的功能微生物群落作逐步LOGIT回归分析.首次从功能微生物数量角度以方程的形式对丹参产区土壤适宜性评价以及判断是否受连作障碍影响。Y=e(-1.4560191nyx-0.63422181nfs-5.300482)-0.4(e为自然数,2.71828).综上所述,导致丹参连作障碍的微生物因素是土壤微生物群落结构被破坏；丹参休地过程就是土壤微生物群落结构重新构建、恢复的过程,休地2-3年是月参产地土壤微生物群落结构重构恢复的关键时期。更多还原

【Abstract】 Radix Salviae Miltiorrhizae, Chinese herbal medicinal frequently used, was dry radix and rhizoma of Salvia miltiorrhiza Bunge, promoting blood circulation, menstrual pain, pure heart and arrest restless, cooling elimination effectiveness of carbuncle. Continuous cropping of Salvia miltiorrhiza seriously harms to the quality of Radix and Rhizoma Salviae Miltiorrhizae in cultivation process, a threat to the development of its economic, causing huge economic loss. This research article, based on the theory of continuous cropping obstacles, throngh the soil microbial research in the producing Salvia miltiorrhiza regions, evaluated microbial structure characteristic of soil suitability. Results were as follow.The research on soil microbial at the uncropping Salvia miltiorrhiza areas of Zhongjiang county indicated that soil bacteria was increasing, but soil fungi was slow with the years off. Cluster analysis showed that that population of soil microbial mainly changed after continuous uncropping Salvia miltiorrhiza 2 years. The functional microbial population of Zhongjiang county was obviously different among uncropping Salvia miltiorrhiza years. With years off, cellulytic bacteria and aromatic compound decomposed bacteria were significantly increasing while other microbial population tended to decrease. And biodiversity index was increasing. MBP contents were significantly different among years uncropping Salvia miltiorrhiza at Zhongjiang county, and MBP contents obviously changedafter uncropping Salvia miltiorrhiza, which was consistent with the results of culturable microbial. With the years on of uncropping Salvia miltiorrhiza, DGGE bands rechness of bacteria genetic diversity was decreasing, but DGGE bands rechness of fungi genetic diversity was increasing.The number of culturable microbial in the suitably producing Salvia miltiorrhiza areas was significant difference with number of culturable microbial in the insuitably producing Salvia miltiorrhiza areas. The number of fungi at the areas of suitably producing Salvia miltiorrhiza was obviously greater than that of insuitably producing. The numbers of anaerobic cellulytic bacteria, nitrification bacteria and anti-surfur bacteria at the areas of suitably producing Salvia miltiorrhiza were significantly more than that of insuitably producing Salvia miltiorrhiza. Other functional microbial population difference was not to the level of statistical significance, and as to biodiversity index, which was relevant to environmental factors in the regions of producing Salvia miltiorrhiza all over the country. MBP contents at the areas of suitably producing Salvia miltiorrhiza were significantly higher than that at the areas of insuitably producing Salvia miltiorrhiza, which was inconsistent with the research results of culturable microbial population. PCR-DGGE molecular analysis technology can be a good tool to analyse culturable and unculturable microbial community genetic structure, and complement and support the traditional analysis method. Fungi and bacteria genetic diversity cluster analysis showed that soil microbial genetic distance of cropping Salvia miltiorrhiza in 2005 and 2006 was smaller than that of other years, and similar, gathered each other; soil microbial genetic distance of cropping Salvia miltiorrhiza in 2008 and 2009 was smaller than that of 2005 and 2006, and similar, gathered each other, while soil microbial genetic diversity of cropping Salvia miltiorrhiza in 2007 distributed in the two categories, seeming transition between the two categories. This was consistent on the results of culturable microbial research. Soil microbial genetic diversity was complex all over the country. But cluster analysis showed that the genetic distance of the suitably producing Salvia miltiorrhiza areas was relatively smaller than that of others. The genetic distance of the insuitably producing Salvia miltiorrhiza areas was quite different.Sequencing, comparison with the GENEBANK sequence database, the results showed that bacteria was in the genus of alpha proteobacterium, Sphingomonas, Arthrobacter, and fungi was in the genus of Termitaria, Agaricus, Cercozoan, Cordyceps, Geomyces, Penicillium, Uncultured Iodophanus.Functional microbial population of rhizosphere and the chemical components of Salvia miltiorrhiza Bunge for stepwise regression analysis showed that caffeic acid comtent was associated with the population of C circulation, tanshinone IIA content was associated with the population of N circulation, salvianolic acid A was associated with the population of N/P, N/S circulation.It was the first time that microbial community structure associate with active ingredients of Salvia miltirrhiza Bunge.Through LOGIT stepwise regression analysis on the rhizosphere soil microbial and the control soil microbial, results showed that equations were used to determine whether soil suitability for implanting Salvia miltiorrhiza Bunge the first time. Y=e (-1.4560191nyx-0.6342218lnfs-5.300482)-0.4The main factor causing continuous cropping obstacle of Salvia miltiorrhiza was that the soil microbial community was destroyed. And uncontinuous cropping Salvia miltiorrhiza was to rebuild and restore soil microbial population balance. Continuous 2-3 years uncropping Salvia miltiorrhiza was the key period of soil microbial communitiy reconstruction and restoration.更多还原