Serratia marcescens is a microorganism that exhibits insecticidal activity against various insects, including the migratory fall armyworm (FAW), Spodoptera frugiperda. This article investigates the insecticidal mechanism of S. marcescens through gastric toxicity. The study involved midgut tissue sectioning, hemolymph observation, and microbiome and transcriptome analysis of both infected and uninfected FAW. The findings revealed that S. marcescens effectively disrupted the structure of the midgut, causing midgut shrinkage and rupture, as well as inducing pseudopodia formation in granulocytes. Moreover, it increased the diversity of gut microbiota. Transcriptome analysis indicated an upregulation of metabolic-related genes and tissue repair genes, while there was a downregulation of fat synthesis genes, some immune genes, hormone synthesis genes, etc. The disruption of the midgut structure negatively affects the metabolism and immune function of the FAW, potentially resulting in midgut rupture, systemic sepsis, and ultimately mortality. In conclusion, our study has elucidated the insecticidal mechanism of S. marcescens against the FAW and demonstrated its potential as a biological control agent for managing this pest.