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The multiple nucleopolyhedrovirus of Spodoptera frugiperda (SfMNPV) plays an important role in regulating its natural host and has high potential for use as a bioinsecticide. However, information about how agricultural practices such as fertilization and plant biotic interactions affect the biocontrol efficacy of SfMNPV is limited. In this study, we examined how multitrophic maize-mycorrhiza-insect herbivore interactions affect the biocontrol efficacy of SfMNPV against S. frugiperda under full and reduced mineral nitrogen fertilization.