【作者】 王燕；

【导师】 薛明； 祝国栋；

【作者基本信息】 山东农业大学 ， 资源利用与植物保护（专业学位）， 2021， 硕士

【摘要】 韭菜迟眼蕈蚊Bradysia odoriphaga Yang et Zhang幼虫俗称韭蛆,是我国重要的蔬菜地下害虫,严重危害韭菜生长。目前防治韭菜迟眼蕈蚊的主要方法是施用化学杀虫剂,但过度依赖化学药剂与生态农业的大发展相悖。与化学防治相比,生物防治具有对环境友好、对人畜安全的优点,是有害生物综合防治的重要发展方向。与其它昆虫相比韭菜迟眼蕈蚊的生物防治资源较为匮乏,实验室团队前期从罹病韭蛆幼虫中分离到一株对韭蛆具有高致病活性的病原真菌,以期作为韭菜迟眼蕈蚊新的生防资源加以开发利用。本文通过对该菌株进行形态特征及ITS同源性比对,对该菌株进行分类鉴定,进而探究该菌株对韭菜迟眼蕈蚊不同虫态的毒杀作用,并进行盆栽试验初步探究该菌株的田间防治作用,同时探究了不同温度对该菌株生长和产孢能力的影响,为韭菜迟眼蕈蚊的绿色防控技术提供可利用的资源。主要研究结果如下:菌株接种到PDA培养基上,在23℃下培养4 d后,菌落直径为7.16-7.32 cm,菌落为圆形,菌丝呈白色,细长毛状,显微观察菌丝为无隔菌丝,顶端有黑色的球状孢子囊,里面含有大量的孢囊孢子。经18S r DNA和ITS测序分析,Blast比对结果与已报道的冻土毛霉菌Mucor hiemalis诸多菌株序列相似度达99%以上,可定义为冻土毛霉菌的一株新菌株。根据试验发现,冻土毛霉菌生长适宜的温度范围在11℃-28℃之间,在此温度范围内,冻土毛霉菌菌丝生长最适宜的温度为23℃。冻土毛霉菌最适宜产孢的温度为18℃,28℃时冻土毛霉菌的产孢量最低,冻土毛霉菌的产孢量为:18℃>23℃>13℃>28℃。不同温度条件下冻土毛霉菌对韭菜迟眼蕈蚊二龄幼虫和四龄幼虫的死亡率试验证明,在28℃时冻土毛霉菌对韭菜迟眼蕈蚊二龄幼虫和四龄幼虫的致病力受到抑制,28℃时用10⁶孢子/ml冻土毛霉菌处理韭菜迟眼蕈蚊二龄幼虫第4 d死亡率为29.67%,经10⁷孢子/ml处理后的第4 d韭蛆的死亡率为72.50%;23℃时10⁶、10⁷孢子/ml冻土毛霉菌处理韭菜迟眼蕈蚊二龄幼虫的死亡率分别为93.34%、94.17%。23℃是冻土毛霉菌菌丝生长、产孢、侵染韭菜迟眼蕈蚊最适宜的温度条件。冻土毛霉菌对韭菜迟眼蕈蚊不同虫态的致病力存在显著差异,对韭菜迟眼蕈蚊幼虫的致病力明显,对于卵和蛹的致病力相对较弱,处理后第4 d,对1-4龄幼虫的LC₅₀分别为:2.9×10⁵、1.21×10⁵、0.7×10⁵、1.89×10⁵,对卵和蛹的LC₅₀分别为5.95×10⁸、1.25×10⁵当冻土毛霉菌孢子悬液的浓度为10⁷孢子/ml时,对韭菜迟眼蕈蚊1-4龄幼虫的LT₅₀分别为1.49、2.29、2.52、2.69,对卵和蛹的LT₅₀为1.37、2.10。在试验过程中发现韭菜迟眼蕈蚊幼虫受冻土毛霉菌侵染后,发病初期幼虫行动迟缓、取食量减少,消化道存有少量食物;发病中期基本不移动,不取食,虫体略透明,脂肪体和消化道溶解;后期虫体呈淡黄色或者黄褐色,虫体软化,用毛笔轻触即破,幼虫已死亡;末期从死亡幼虫体内长出白色菌丝,顶端有黑色球状孢子囊。在盆栽试验中,10⁷孢子/ml冻土毛霉菌处理韭蛆二龄幼虫第3 d的校正死亡率为48.64%,第5 d的校正死亡率为69.44%,10⁹孢子/ml冻土毛霉菌处理韭蛆二龄幼虫第3 d的校正死亡率为59.45%,第5 d的校正死亡率为86.11%。10⁷孢子/ml冻土毛霉菌处理韭菜迟眼蕈蚊四龄幼虫第3 d的校正死亡率为34.21%,第5 d的校正死亡率为64.86%,10⁹孢子/ml冻土毛霉菌处理韭菜迟眼蕈蚊四龄幼虫第3 d的校正死亡率为47.36%,第5 d的校正死亡率为78.37%。更多还原

【Abstract】 Brachysia odoriphaga Yang et Zhang commonly known as B.odoriphaga,is an important underground vegetable pest in my country,which seriously endangers the growth of leeks.At present,the main method to prevent and control the B.odoriphaga is the application of chemical insecticides,but excessive dependence on chemical agents is contrary to the great development of ecological agriculture.Compared with chemical control,biological control has the right.Compared with other insects,the biological control resources of B.odorata are less.In the early stage,the laboratory team isolated a pathogenic fungus with high pathogenic activity to B.odoriphaga,in order to develop and utilize it as a biocontrol resource.In this paper,the morphological characteristics and ITS homology of the strain were compared,classify and identify that strain,to explore the toxic effect of this strain on different insect States of B.odoriphaga,pot experiment was carried out to determine the field control effect of the strain,the effects of different temperatures on the growth and sporulation ability of the strain were studied,to provide available resources for the green prevention and control technology of B.odoriphaga.The main research results are as follows:The strain was inoculated on PDA medium and cultured at 23℃for 4 days,the colony diameter was 7.16-7.32 cm,the colony was round,and the hyphae were white and slender hairy,microscopic observation of hyphae as non-septal hyphae.According to the experiment,the temperature range for the growth of frozen soil Mucor is 11℃-32℃,when the temperature is higher than 32℃,the growth of frozen soil Mucor stops,and the most suitable temperature for the mycelium growth of frozen soil Mucor is23℃.According to the test,the most suitable temperature for permafrost Mucor to produce spores is 18℃.At 28℃,the sporulation of permafrost Mucor is the lowest.The sporulation of permafrost Mucor is:18℃>23℃>13℃>28℃.The mortality test of Mucor in frozen soil to the second and fourth instars of B.odoriphaga under different temperature conditions proved that,at 28℃,the pathogenicity of Mucor permafrost to the second and fourth instars of B.odoriphaga was inhibited.At 28℃,the mortality of the second instar larvae of B.odoriphaga was 29.67%,on the 4th day after treatment with Mucor permafrost with 10⁶spores/ml,and the mortality of B.odoriphaga after treatment with 10⁷spores/ml was 72.50%.At 23℃,the mortality rates of the second instar larvae of B.odoriphaga treated with 10⁶and 10⁷spores/ml Mucor in frozen soil were93.34%and 94.17%.23℃is the most suitable temperature condition for mycelia growth,sporulation and infection of Mucor.There are significant differences in pathogenicity of Mucor in frozen soil to different insect States of B.odoriphaga,the pathogenicity to the larvae of B.odoriphaga is obviously stronger than that of other insect States.On the 4th day after treatment,the LC₅₀of 1st-4th instar larvae were 2.9×10⁵,1.21×10⁵,0.7×10⁵and 1.89×10⁵,the LC₅₀of eggs and pupae are5.95×108 and 1.25×105,When the concentration of Mucor spore suspension in frozen soil was 10⁷spores/ml,the LT₅₀of the 1st-4th instar larvae of B.odoriphaga are 1.49,2.29,2.52and 2.69,and the LT50 of eggs and pupae are 1.37 and 2.10.After the larvae of B.odoriphaga are infected by the permafrost Mucor,the larvae are slow to move and eat less at the beginning of the disease,and there is a small amount of food in the digestive tract;In the middle stage,there is basically no movement,no feeding,the worm body is slightly transparent,and the fat body and digestive tract dissolve;In the later stage,the insect body is light yellow or yellowish brown,which softens and breaks with a brush,and the larvae have died;At the end,white hyphae grew from dead larvae with black spherical sporangia at the top.In the pot experiment,the corrected mortality of the 2nd instar larvae of B.odoriphaga with 10⁷spores/ml Mucor permafrost spore suspension was 48.64%on the 3rd day,69.44%on the 5th day,the corrected mortality of the second instar larvae of B.odoriphaga with 10⁹spores/ml Mucor permafrost spore suspension was 59.45%on the third day and 86.11%on the fifth day.The corrected mortality of the 4nd instar larvae of B.odoriphaga with 10⁷spores/ml Mucor permafrost spore suspension was 34.21%on the 3rd day,64.86%on the 5th day,the corrected mortality of the second instar larvae of B.odoriphaga with 10⁹spores/ml Mucor permafrost spore suspension was 47.36%on the third day and 78.37%on the fifth day.更多还原