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
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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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