Spodoptera frugiperda is a common and severely damaging agricultural pest. In-depth analysis of its population genomics and transcriptomics is crucial for providing references for pest control efforts. This study, focused on the extensive variation in the genome size of S. frugiperda, constructed its pan-genome and identified 1.37 Gb of non-reference sequences, highlighting significant genetic variation within the population. Analysis of Long Terminal Repeat (LTR) Presence/Absence Variation (PAV) suggests that LTR alterations may be one of the driving factors for genome size variation. Additionally, population gene PAV analysis revealed that variable genes are enriched in functions like acetyltransferase activity, which might be associated with detoxification, implying diverse selection pressures related to detoxification in different S. frugiperda populations. Moreover, 19 horizontal gene transfer (HGT) acquired genes were identified in the reference genome used in this study, which responded to 16 different treatments. Notably, three HGT-acquired genes (SFR02618, SFR05248, and SFR05249) co-expressed with heat shock protein family and responded under treatments with Avermectin and Cypermethrin. This may indicate their involvement in a detoxification mechanism coordinated with heat shock proteins. These results offering new insights into its genomic evolution and the potential functions of HGT-acquired genes.