The effects of oral administration of Astragalus polysaccharides (APS) and florfenicol (FFC), singly or in combination, on the survival performance, disease resistance, and immunity of Litopenaeus vannamei were investigated. After challenge with an AHPND-causing strain of Vibrio parahaemolyticus (VPAHPND), shrimp were immediately fed a drug-free diet, diets containing only APS (200 mg·kg-1) or FFC (15 mg·kg-1), or diets containing low-dose (7.5 mg·kg-1 FFC + 100 mg·kg-1 APS), medium-dose (15 mg·kg-1 FFC + 200 mg·kg-1 APS), and high-dose (30 mg·kg-1 FFC+400 mg·kg-1 APS) drug combinations for 5 days. The cumulative shrimp mortality over 5 days after injection of VPAHPND in the APS + FFC combination groups was significantly lower than that in the APS or FFC alone groups (p < 0.05). Immune parameters, including the total hemocyte counts (THCs), hemocyanin (HEM) concentration, antibacterial activity, activity levels of lysozyme (LZM), and levels of acid phosphatase (ACP), alkaline phosphatase (AKP), and phenoloxidase (PO) in cell-free hemolymph, and the expression levels of the immune-related genes anti-lipopolysaccharide factor (ALF), cathepsin B (catB), crustin, lectin (Lec), lysozyme (LZM), and Toll-like receptor (TLR) in hemocytes and hepatopancreas were determined in the shrimp. The values for these immune parameters in the drug combination groups were higher than those in the APS or FFC group (p < 0.05). Finally, in the histological examinations, the histological structural alignment and integrity of the hepatopancreatic tubules in the drug combination groups was better than that in the APS and FFC groups. Under the experimental conditions, dietary APS and FFC had a synergistic effect on immunity and disease resistance among shrimp after VPAHPND infection.