【作者】 李宁；

【导师】 王志刚；

【作者基本信息】 河北农业大学 ， 森林保护学， 2011， 硕士

【摘要】 为寻求防治光肩星天牛有效的昆虫病原线虫品系,本研究从保定市内采集的120份土样中分离线虫,并将分离到的昆虫病原线虫进行初步的筛选,测定了11个品系昆虫病原线虫抗干燥能力及其对光肩星天牛的致病性,以期筛选出最佳线虫品系,为光肩星天牛防治及昆虫病原线虫制剂的开发和应用提供理论依据。结果表明:1.昆虫病原线虫的分离及干燥性测定:在保定市郊及周边市、县25个地点采集120份土样,诱集并成功繁殖5个昆虫病原线虫种群且均为异小杆线虫（Heterohabditidae ）。不同昆虫病原线虫品系抗干燥能力不同,Steinernema carpocapsae GA品系的存活率最高,达到75.1%;Heterorhabditis sp. ZH经干燥后存活率达50%以上,且复苏后线虫对大蜡螟致死率达100%;Heterorhabditis sp. CZ和XH-9品系存活率最低为0%。2.线虫品系的分类鉴定:利用形态学特征和ITS-rDNA分析对ZH种群进行了分类鉴定。通过对该线虫的形态特征进行观察,并对其侵染性幼虫、雄性成虫的重要参数进行测量,表明该线虫与嗜菌异小杆线虫的形态特征最为相近;同时ITS1-rDNA序列比对和系统发育学分析结果也显示该线虫与嗜菌异小杆线虫亲缘关系最近。确定该线虫为嗜菌异小杆线虫涿州品系。3.线虫侵染对血淋巴蛋白质含量的影响:采用考马斯亮蓝G-250法测定线虫侵染光肩星天牛幼虫体内血淋巴蛋白质含量的变化,结果显示处理过的天牛血淋巴内蛋白质含量在侵染初期变化不大,侵染后12 h～24 h,蛋白浓度呈明显上升趋势,至24 h达到最大值后开始不断下降最后显著低于对照组蛋白含量。对照组天牛幼虫在不同处理时间蛋白质含量并无显著变化,仅有小幅度波动。4.线虫侵染对保护酶活性的影响:光肩星天牛感染线虫初期超氧化物歧化酶（SOD）,过氧化物酶（POD）和过氧化氢酶（CAT）活力均迅速提高,在侵染后期SOD和POD活力有不同程度的下降,而CAT活力下降后又上升到较高的水平而后开始下降。SOD和POD活性变化趋势大体相似,均表现为先升高后降低,CAT活性变化趋势可归纳为升高—降低—升高—降低。5.线虫侵染对解毒酶活性的影响:光肩星天牛感染线虫后体内主要解毒酶活力均产生不同程度的影响。羧酸酯酶（CarE）和谷胱甘肽-S-转移酶（GSTs）活性变化大体表现为激活→抑制→激活→抑制;对乙酰胆碱酯酶（AchE）活力的影响大体表现为抑制→激活→抑制→激活→抑制。6.昆虫病原线虫对光肩星天牛致病力测定:11个品系昆虫病原线虫对光肩星天牛成虫和幼虫均有一定防治效果,解剖死亡的天牛成虫和幼虫,可见大量活跃的侵染期线虫在虫尸体内。Heterorhabditis sp. ZH品系对光肩星天牛成虫和幼虫均有较高致病力,为最佳线虫品系。侵染10d后成虫累计校正死亡率为94.59%,LT₅₀为4.10d;侵染幼虫4d后累计校正死亡率为100%,LT₅₀为1.61d。不同浓度ZH品系线虫对光肩星天牛幼虫的致死能力不同,最优浓度为3500条/mL。更多还原

【Abstract】 In order to seek efficient strains of entomopathogenic nematodes to control Anoplophora glabripennis, more than 120 soil samples were collected from Baoding region in Hebei province and used for isolating entomopathogenetic nematodes （EPNs）. Preliminary screening of isolated EPNs was taken. Effects of 11 strains of entomopathogenic nematodes against Anoplophora glabripennis were determined. The results showed that:1. Five entomopathogenic nematodes population were isolated and successfully breeded from 120 soil samples, all of which belongs to Heterohabditidae. The anti-desiccation ability were different in different nematode strains. S. carpocapsae GA had the highest survival rate of 75.1%. The survival rate of Heterorhabditis sp. ZH was more than 50%. And the mortality rate to mellonella of recovery nematodes was 100%. Heterorhabditis sp.CZ and XH-9 strain survival rate was 0%.2. ZH population was identified by the morphological characteristics and ITS-rDNA sequence analysis. Through morphological observation and respond measurements of infective juveniles and males, the population was found that it was closely related to Heterorhabditis bacteriophora. At the same time, the results of ITS1-rDNA sequences alignment and phylogenetic analysis showed the population also had the closest genetic relationship with H. bacteriophora. Therefore, the population was identified as H. bacteriophora zhuozhou strain.3. Coomassie brilliant blue G-250 was used to determine the changes in hemolymph protein content of Anoplophora glabripennis larvae infected by nematodes. The results showed that hemolymph protein content of the Anoplophora glabripennis showed little changes in the early infection. After infection at 12 h 24h, the protein concentration showed a rising trend and reached the maximum at 24h, and then begin to decline. Finally it is significantly lower than that of the control group. The protein content of A. glabripennis larvae in the control group had no significant change with only minor fluctuations in different treatment time.4. During the initial infection stage, the activities of SOD, POD and CAT increased quickly, while during the later stage, the activities of SOD and POD decreased at different degrees, but that of CAT rose to a high level and then began to decline. The activities of SOD and POD which had similar trends, rose first and then gradually decreased, CAT activity trends could be summarized as increase -decrease - increase - decrease.5. The activities of the main detoxifying enzymes changed at different degrees due to invasion of entomopathogenic nematodes. The trend of changes of CarE and GSTs activity could be summarized as activated→suppressed→activated→suppressed. The impact on AchE activity could be summarized as suppressed→activated→suppressed→activated→suppressed.6. The control effects of 11 strains of entomopathogenic nematodes on A. glabripennis adult and larvae were detected.Heterorhabditis sp.ZH which showed highest virulence to both A. glabripennis adult and larvae among 11 strains. The mortality of adult was 94.59% with 4.10d LT₅₀. The mortality of larvae was 100% with 1.61d LT₅₀. The pathogenity of the Heterorhabditis sp.ZH of different concentrations to the Anoplophora glabripennis larvae was also determined. The results indicated that the optimum treatments were 3500 nematode of ZH/mL.更多还原