Anhui Provincial Key Laboratory of Microbial Pest Control, Anhui Agricultural University, Hefei, 230036, China.
School of Plant Protection, Anhui Agricultural University, Hefei, 230036, China.
Microb Biotechnol. 2021 Jul;14(4):1433-1444. doi: 10.1111/1751-7915.13748. Epub 2021 Jan 18.
The slow lethality of fungal biopesticides to insects restrains their widespread application as a strategy of pest control. In this study, unary, binary and ternary transgenic Metarhizium robertsii were created by integrating genes that encode the scorpion neurotoxin BjαIT, the cuticle-degrading protease Pr1A, and a double-stranded RNA (dsRNA) that targets host gnbp3, individually or collectively under a constitutive promoter to enhance virulence. Compared with the parental wild type, all unary transgenic strains had increased virulence against four insect species, Tenebrio molitor, Locusta migratoria, Plutella xylostella and Galleria mellonella, whereas the binary transgenic strain expressing both pr1A and BjαIT had increased virulence to T. molitor and L. migratoria, with no change in virulence against P. xylostella and G. mellonella. Importantly, all ternary transgenic strains simultaneously expressing pr1A, BjαIT, and the dsRNA specific to host gnbp3 exhibited the highest increase in insect-specific virulence. This finding highlights a novel strategy for genetic engineering of dsRNAs that target genes associated with the host immune response alongside virulence genes to maximize fungal virulence and lethality against insect pests.
真菌生物农药对昆虫的缓慢致死作用限制了它们作为害虫防治策略的广泛应用。在这项研究中，通过整合编码蝎神经毒素 BjαIT、角质层降解蛋白酶 Pr1A 和靶向宿主 gnbp3 的双链 RNA (dsRNA) 的基因，分别或共同在组成型启动子下，创建了一元、二元和三元转基因玫烟色棒束孢，以增强其毒力。与亲本野生型相比，所有一元转基因株系对 4 种昆虫（黄粉虫、东亚飞蝗、小菜蛾和家蚕）的毒力均有所提高，而同时表达 pr1A 和 BjαIT 的二元转基因株系对 T. molitor 和 L. migratoria 的毒力有所提高，但对 P. xylostella 和 G. mellonella 的毒力没有变化。重要的是，同时表达 pr1A、BjαIT 和靶向宿主 gnbp3 的 dsRNA 的所有三元转基因株系对昆虫的特异性毒力均有显著提高。这一发现为针对与宿主免疫反应相关的基因以及毒力基因的 dsRNA 的遗传工程提供了一种新策略，以最大限度地提高真菌对昆虫害虫的毒力和致死率。
