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首页> 美国卫生研究院文献>Applied and Environmental Microbiology >Increased Insect Virulence in Beauveria bassiana Strains Overexpressing an Engineered Chitinase
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Increased Insect Virulence in Beauveria bassiana Strains Overexpressing an Engineered Chitinase

机译：球孢白僵菌过表达工程化几丁质酶的昆虫致病力增加

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摘要

Entomopathogenic fungi are currently being used for the control of several insect pests as alternatives or supplements to chemical insecticides. Improvements in virulence and speed of kill can be achieved by understanding the mechanisms of fungal pathogenesis and genetically modifying targeted genes, thus improving the commercial efficacy of these biocontrol agents. Entomopathogenic fungi, such as Beauveria bassiana, penetrate the insect cuticle utilizing a plethora of hydrolytic enzymes, including chitinases, which are important virulence factors. Two chitinases (Bbchit1 and Bbchit2) have previously been characterized in B. bassiana, neither of which possesses chitin-binding domains. Here we report the construction and characterization of several B. bassiana hybrid chitinases where the chitinase Bbchit1 was fused to chitin-binding domains derived from plant, bacterial, or insect sources. A hybrid chitinase containing the chitin-binding domain (BmChBD) from the silkworm Bombyx mori chitinase fused to Bbchit1 showed the greatest ability to bind to chitin compared to other hybrid chitinases. This hybrid chitinase gene (Bbchit1-BmChBD) was then placed under the control of a fungal constitutive promoter (gpd-Bbchit1-BmChBD) and transformed into B. bassiana. Insect bioassays showed a 23% reduction in time to death in the transformant compared to the wild-type fungus. This transformant also showed greater virulence than another construct (gpd-Bbchit1) with the same constitutive promoter but lacking the chitin-binding domain. We utilized a strategy where genetic components of the host insect can be incorporated into the fungal pathogen in order to increase host cuticle penetration ability.
机译：昆虫病原真菌目前被用于控制几种害虫，作为化学杀虫剂的替代品或补充品。通过了解真菌的发病机理和遗传修饰靶基因，可以提高毒力和杀灭速度，从而提高这些生物防治剂的商业功效。致病性真菌（例如球孢白僵菌）利用大量水解酶（包括几丁质酶）渗透昆虫表皮，而水解酶是重要的毒力因子。曾在B. bassiana中鉴定了两个几丁质酶（Bbchit1和Bbchit2），它们都不具有几丁质结合域。在这里，我们报告了几个B.bassiana杂交几丁质酶的构建和表征，其中几丁质酶Bbchit1与衍生自植物，细菌或昆虫的几丁质结合域融合在一起。与其他杂化几丁质酶相比，含有家蚕家蚕几丁质酶与Bbchit1融合的包含几丁质结合结构域（BmChBD）的杂化几丁质酶显示出最大的结合几丁质的能力。然后将该杂合几丁质酶基因（Bbchit1-BmChBD）置于真菌组成型启动子（gpd-Bbchit1-BmChBD）的控制下，并转化成球孢杆菌。昆虫生物测定显示，与野生型真菌相比，转化体的死亡时间缩短了23％。与具有相同组成型启动子但缺乏几丁质结合结构域的另一种构建体（gpd-Bbchit1）相比，该转化体还显示出更高的毒力。我们采用了一种策略，可以将宿主昆虫的遗传成分掺入真菌病原体中，以增加宿主表皮的渗透能力。

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