Research on the Expression of Immune-Related Genes at Different Stages in the Third-Instar Larvae of Spodoptera frugiperda Infected by Metarhizium rileyi

<i>Spodoptera frugiperda</i> is a major migratory agricultural pest that poses a significant threat to global crop safety. <i>Metarhizium rileyi</i> has emerged as an effective biocontrol agent against lepidopteran pests. In this study, we examined the immune responses of third-instar <i>S. frugiperda</i> larvae at various stages of an <i>M. rileyi</i> infection. Using RNA-seq and microscopic observation, we identified the immune-related pathways enriched at different infection stages, which were further validated by a qRT-PCR. Our findings revealed the following immune responses during infection: During the stage when <i>M. rileyi</i> penetrated the host cuticle (0-48 h), the genes related to energy metabolism, detoxification, and melanization were upregulated. Meanwhile, the TOLL and IMD signaling pathways were activated to counter the infection. During the stage of <i>M. rileyi</i>'s internal infection (48-96 h), which was the peak expression period of the immune-related genes, cellular immunity predominated. Hemocytes encapsulated and phagocytosed the hyphal bodies. Phagocytosis was enhanced through the upregulation of the genes related to ROS and the melanization-related genes, as well as the genes involved in insect hormone biosynthesis. During the stage when <i>M. rileyi</i> grew from the inside to the outside of the host (96-120 h), immune system paralysis resulted in host mortality. These findings deepen our understanding of the immune interactions between <i>M. rileyi</i> and <i>S. frugiperda</i>, support the potential of <i>M. rileyi</i> as an effective biocontrol agent, and provide a theoretical foundation for the development of targeted biopesticides for pests using biotechnological approaches.