Enhanced virulence of Autographa californica multiple nucleopolyhedrovirus in Spodoptera frugiperda is mediated by an Ac34 mutation that promotes nucleocapsid envelopment within occlusion bodies

Spodoptera frugiperda poses a significant threat as a polyphagous agricultural pest. Autographa californica multiple nucleopolyhedrovirus (AcMNPV), a broad-spectrum baculovirus with established activity against multiple lepidopteran pests, represents an environmentally benign biocontrol candidate. However, its efficacy against S. frugiperda is constrained by limited oral infectivity. Here, serial passaging in S. frugiperda yielded a mutant strain of AcMNPV (AcMNPV-Mut) with significantly enhanced oral infectivity relative to the wild strain AcMNPV (AcMNPV-Wt). Importantly, while both oral infectivity and overall virulence (shortened LT₅₀ and increased mortality) against S. frugiperda were enhanced, the virus maintained its ability to effectively infect and cause disease in other insect hosts, like Helicoverpa armigera . Whole-genome sequencing identified a missense mutation in ac34 . Functional characterization of a recombinant virus harboring this mutation revealed that the ac34 variant contributes to virulence augmentation through dual pathways: elevated budded viruses (BVs) production facilitating secondary infection, and upregulated late and very late gene expression, which improves occlusion-derived virus (ODV) embedding within occlusion bodies (OBs), and may lead to an increase in OB size, contributing to a higher viral load per particle, and thereby enhancing primary infectivity. The ac34 mutation enhances primary and secondary infection, promoting virulence evolution and providing a molecular target for biopesticide design. IMPORTANCE In this study, we found that Autographa californica multiple nucleopolyhedrovirus (AcMNPV) undergoes adaptive mutations after serial passages in the semi-permissive host Spodoptera frugiperda , driven by mutation in ac34 . These mutations not only significantly increase viral replication within this pest but also improve the packaging and infectivity of viral particles. This coordinated optimization allows the virus to achieve higher effective doses in a short time, synergistically enhancing its control efficiency against S. frugiperda . Importantly, the mutation does not compromise the virus’s pathogenicity toward its original permissive hosts, underscoring its potential utility in single-application strategies for integrated pest management. These findings provide new molecular insights for designing efficient and broadly applicable biocontrol agents, contributing to more sustainable management of agricultural pests and reduced reliance on chemical pesticides.