糜子丝黑穗病菌遗传多样性及综合防控技术研究

发布时间：2019-05-25 04:01

【摘要】：糜子生育期短、抗旱、耐瘠、营养丰富,是干旱半干旱地区重要的粮食作物和经济作物。丝黑穗病是糜子生产上的主要病害,是制约糜子产量和品质的重要因素。因此,分析病原菌遗传多样性水平和毒力分化程度,研究糜子丝黑穗病抗性生理机制,筛选高效防治药剂,明确适宜栽培措施,对抗病品种选育及病害绿色防控具有十分重要的意义。由于糜子属区域性作物,目前国内外关于糜子及糜子丝黑穗病的相关研究鲜有报道,本试验利用RAPD和SSR分子标记技术对来自我国糜子主产区的病原菌进行遗传多样性分析,鉴定糜子种质资源和育成品种丝黑穗病抗性水平,筛选鉴别寄主并对病原菌进行毒力分析,比较分析糜子不同生育时期感染丝黑穗病后叶片抗氧化酶系、抗病相关酶活性,蛋白质、糖含量,及同工酶谱带的变化和差异,通过室内毒力测定和田间药效试验,筛选防治糜子丝黑穗病高效杀菌剂,并且研究不同播期对抗、感品种丝黑穗病发病及产量的影响。主要研究结论如下:(1)田间调查结果显示,糜子丝黑穗病为害的穗部症状主要表现为黑粉包状、簇叶状、刺猬头和黑粉粒4种类型。糜子丝黑穗病菌冬孢子在光学显微镜下,呈褐色,球形,扫描电子显微镜下可见壁表具微小突起。冬孢子形态特征和ITS序列比对结果表明,本试验所用菌株为Sporisorium destruens。冬孢子萌发的最适温度为25℃,最适pH为5,甲醛处理促进萌发。丝黑穗病菌最适碳源和氮源为葡萄糖和KNO3,25℃比较适合此病菌生长和产孢,pH为7时生长最快、产孢量最大,在Czapek和PDA培养基上生长较好。(2)来自我国黑龙江、吉林、辽宁、内蒙古、河北、陕西和甘肃的糜子丝黑穗病菌(S.destruens)遗传变异程度不高,遗传背景比较单一。选出的28条RAPD引物和40对SSR引物分别扩增出381和119条谱带,其中多态性谱带373和109条,多态率分别为97.90%和91.60%。RAPD的多态性水平及检测效率高于SSR。两种标记的聚类结果有差异。将两种标记的数据结合进行聚类,结果显示,在遗传相似系数为0.7425时,51个菌株可以分为3个系谱群。菌株的遗传多态性和地理分布及寄主品种之间没有明显的相关性。(3)连续3年丝黑穗病抗性鉴定结果显示,261份种质资源的抗性差异明显,抗病种质所占比重较大,其中12MD-2和12MD-250连续3年均表现免疫;66份育成品种中,来自俄罗斯的品种blest jachee和orlovski karlik表现高抗,而我国的品种均未表现出稳定抗性。参试糜子种质资源中,3份具有高抗、矮秆、早熟和高产等优良性状,可直接用于实际生产。我国S.destruens存在毒力分化现象,根据各鉴别寄主对各菌株的抗性等级,可将16个菌株分为3个致病类型。(4)糜子接种S.destruens后,SOD活性升高,三叶期抗病品种的升高幅度显著大于感病品种,且抗病品种的SOD活性显著高于感病品种;抗病品种POD活性持续升高,而感病品种POD活性在后期下降;抗病品种MDA含量低于感病品种。糜子受S.destruens侵染后,抽穗期和灌浆期APX活性与抗性呈负相关;GR活性与抗性成反比;抗病品种AsA含量迅速升高并维持在较高水平,而感病品种升高较慢且后期下降幅度较大;抗、感品种GSH含量在三叶期和拔节期升高,在抽穗期和灌浆期下降。在S.destruens诱导下,抗病品种PAL活性升高幅度在三叶期、拔节期和灌浆期均高于感病品种,三叶期和拔节期抗病品种几丁质酶活性显著高于感病品种,β-1,3-葡聚糖酶活性在抽穗期与抗性呈正相关。接种后,可溶性蛋白含量与发病率无显著相关性;在三叶期和灌浆期可溶性总糖含量与抗性呈负相关;而还原糖含量在三叶期和拔节期接种后与抗性呈负相关。丝黑穗病菌侵染能引起不同抗病类型糜子品种POD、EST、SOD和PPO同工酶谱的变化,抗病品种谱带增加数大于感病品种。综上,在糜子抗病育种和生产实践中,SOD、POD、APX、GR、PAL、几丁质酶和β-1,3-葡聚糖酶活性,MDA、AsA、GSH、可溶性总糖及还原糖含量,POD、EST和SOD同工酶均可作为鉴别糜子丝黑穗病抗性的辅助指标。(5)通过对戊唑醇、甲基硫菌灵、烯唑醇、多菌灵、苯醚甲环唑和福美双6种药剂的室内毒力测定和两年田间药效试验,结果发现,戊唑醇对冬孢子萌发的抑制作用最强,福美双最弱;烯唑醇对种子萌发抑制作用最大;戊唑醇田间防效最好,其次为烯唑醇、甲基硫菌灵、多菌灵,福美双防治效果最差。多菌灵显著降低主茎倒二叶面积、主茎倒三叶面积、叶干重和分蘖数,各药剂处理下的单株粒重均有不同程度的减少,但产量均增加,其中戊唑醇处理下的产量两年均最高。戊唑醇能以较低浓度获得较好的防治效果,建议在实际生产中推广利用。(6)不同糜子品种对播期的响应不同,但除05在5个播期均不发病外,其他品种均在播期Ⅰ发病率最高,其他播期无显著差异。抗病品种巴盟小黑糜和05在播期Ⅲ产量最高,因此6月15日左右可作为这两个品种的适宜播期;感病品种杂黍和黄硬糜在播期Ⅱ产量最高,因此可将6月1日作为这两个品种的适宜播期。发病率和5 cm土层温度呈显著负相关,低温有利于丝黑穗病的发生;发病率与20 cm土层水分含量无显著相关性。
[Abstract]:The millet has the advantages of short growth period, drought resistance, drought tolerance and rich nutrition, and is an important food crop and economic crop in the arid and semi-arid region. The main disease in the production of mylis is the main disease which restricts the yield and quality of the mylin. Therefore, it is of great significance to study the genetic diversity of pathogenic bacteria and the degree of virulence differentiation, to study the physiological mechanism of the resistance to the black-ear disease of mylin, to select the effective control agent, to identify the appropriate cultivation measures, and to have a very important significance to the breeding of the disease-resistant variety and the prevention and control of the disease. In the present study, the genetic diversity of the pathogenic bacteria from the main production areas of the Myli in China was analyzed by using the RAPD and SSR molecular markers. in that invention, the resistance level of the millet germplasm resource and the black spike of the bred variety is identify, the host is screened and the virulence analysis is carried out on the pathogenic bacteria, the anti-oxidation enzyme system, the disease-related enzyme activity, the protein and the sugar content of the leaf after the different growth period of the millet are compared and analyzed, And the effect of different sowing time on the incidence and the yield of the black-ear diseases of the variety silk is studied through the indoor toxicity measurement and the field effect test. The main results of the study are as follows: (1) The results of the field investigation show that the symptoms of the ear part of Mylin black are mainly in the form of black powder, cluster-leaf shape, hedgehog head and black powder. The microprojections of the wall surface were observed under the optical microscope, in the form of brown, spherical and scanning electron microscope. The results of the comparison of the morphological characteristics and the ITS sequence of the winter spore show that the strain used in this test is Sporisoria dessenens. The optimum temperature of the winter spore germination was 25 鈩，

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