Multifaceted roles of NADPH oxidases in the growth and pathogenicity of .

Reactive oxygen species (ROS), synthesized by the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) complex, are vital molecules in biological cells, influencing various physiological processes such as fungal growth, development, and virulence. , an entomopathogenic fungus, is a promising biopesticide for agricultural, forestry, and urban pest control. This study focuses on the characterization of NADPH oxidases (Noxs) in . Gene expression profiles of Noxs in (BbNoxs) were analysed using RT-qPCR. Knockout strains of single BbNoxA, BbNoxB, BbNoxR, and double BbNoxA and BbNoxB were constructed via homologous recombination, and their phenotypic characteristics were examined. Fungal virulence was evaluated using Galleria mellonella larvae, and infection structures formation and penetration ability were assessed on cicada wings. ROS production and actin assembly during fungal growth and infection were detected using staining and marker methods. Expression analysis revealed significant upregulation of BbNoxs during fungal growth and infection. Compared to the wild-type strain, single knockouts (ΔBbNoxA/B/R) and double knockout (ΔBbNoxAB) of BbNoxs exhibited reduced conidial yields, accelerated conidial germination rates. Deletion of BbNoxB or BbNoxR decreased fungal virulence compared to the WT strain in topical inoculation experiments. Additionally, loss of BbNoxB or BbNoxR impaired infection structures formation, penetration ability, ROS production, and actin aggregation during fungal infection. BbNoxs are crucial for fungal growth, development, and virulence in , playing essential roles in infection structures formation, penetration, ROS production, and actin assembly. Understanding their functions provides insights into 's pathogenic mechanisms.