StcU-2 Gene Mutation via CRISPR/Cas9 Leads to Misregulation of Spore-Cyst Formation in Ascosphaera apis

文献类型： 外文期刊

第一作者： Aynalem, Tessema

作者： Aynalem, Tessema;Meng, Lifeng;Getachew, Awraris;Wu, Jiangli;Yu, Huimin;Tan, Jing;Li, Nannan;Xu, Shufa;Aynalem, Tessema;Getachew, Awraris

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关键词： honey bee; Ascosphaera apis; CRISPR; Cas9; sporulation; versicolorin reductase gene (StcU-2)

期刊名称：MICROORGANISMS （ 影响因子：4.926； 五年影响因子：5.143 ）

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年卷期： 2022 年 10 卷 10 期

页码：

收录情况： SCI

摘要： Ascosphaera apis is the causative agent of honey bee chalkbrood disease, and spores are the only known source of infections. Interference with sporulation is therefore a promising way to manage A. apis. The versicolorin reductase gene (StcU-2) is a ketoreductase protein related to sporulation and melanin biosynthesis. To study the StcU-2 gene in ascospore production of A. apis, CRISPR/Cas9 was used, and eight hygromycin B antibiotic-resistant transformants incorporating enhanced green fluorescent protein (EGFP) were made and analyzed. PCR amplification, gel electrophoresis, and sequence analysis were used for target gene editing analysis and verification. The CRISPR/Cas9 editing successfully knocked out the StcU-2 gene in A. apis. StcU-2 mutants had shown albino and non-functional spore-cyst development and lost effective sporulation. In conclusion, editing of StcU-2 gene has shown direct relation with sporulation and melanin biosynthesis of A. apis; this effective sporulation reduction would reduce the spread and pathogenicity of A. apis to managed honey bee. To the best of our knowledge, this is the first time CRISPR/Cas9-mediated gene editing has been efficiently performed in A. apis, a fungal honey bee brood pathogen, which offers a comprehensive set of procedural references that contributes to A. apis gene function studies and consequent control of chalkbrood disease.

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