The diamondback moth, Plutella xylostella represents a worldwide threat to Brassicaceae crops and has developed substantial resistance to conventional insecticides. Entomopathogenic fungi (EPF) have emerged as environmentally sustainable alternatives to chemical insecticides. Since insect immunity constitutes the primary defense against fungal pathogens, understanding these mechanisms could advance biocontrol strategies. Nevertheless, research on the immune functions of galectins in insects remains limited. This study identifies a Galectin-4 homolog in P. xylostella (PxGalectin-4) and systematically examines its immunological functions against an EPF Isaria cicadae infection. The open reading frame of PxGalectin-4 encoded 338 amino acids with a carbohydrate recognition domain (CRD). PxGalectin-4 expression exhibited peak levels in late-instar larval stages and fat body, and increased significantly following I. cicadae challenge. Functional characterization demonstrated that recombinant PxGalectin-4 (rPxGalectin-4) directly bound cells and cell wall components of microbes, and displayed Ca²⁺-dependent microbial agglutination. Additionally, rPxGalectin-4 enhanced hemocyte-mediated immune responses by promoting nodulation and encapsulation, and increased phenoloxidase activity of hemolymph. Knockdown of PxGalectin-4 significantly increased the susceptibility of P. xylostella larvae to I. cicadae infection. In conclusion, PxGalectin-4 serves a vital immune function in P. xylostella defense against I.cicadae, and presents a potential target for novel pest control strategies.