To explore the semi-lethal concentration （LC₅₀） of Vibrio anguillarum infection in Ruditapes philippinarum and its effects on gill tissues and immune related genes. In this study， R. philippinarum was challenged with five different concentrations of V. anguillarum for 168 h. Meanwhile， the LZM activity， bacterial load， pathological analysis and the expression of immune-related genes in the gills were conducted. The results showed that the challenge group injected with V. anguillarum at 5×10⁷ CFU/mL for 168 h corresponded to the semi-lethal concentration （LC₅₀） group， and the semi-lethal concentration （LC₅₀） was 6.08×10⁶ CFU/（mL·g）， with a survival rate of 52.3%. The survival rate was the lowest at 33.6% in the 5×10⁸ CFU/mL group and the highest survival rate of 67.0% was observed in the 5×10⁶ CFU/mL group. The results indicated that the bacterial load peaked at 24 h （7.53 CFU/mg）， and then gradually decreased， reaching 4.26 CFU/mg at 120 h in semi-lethal concentration （LC₅₀） groups. The LZM activity significantly increased in the gills and peaked at 12 h （123.03 U/mg prot/mL）， and then gradually decreased， reaching 77.92 U/mg prot/mL at 120 h. The pathological analysis revealed that the gills filaments showed disordered arrangement， severe shrinkage， and increased gaps at 24 h. Additionally， quantitative real-time PCR （qPCR） analysis revealed that V. anguillarum infection significantly upregulated the expression of several immune related genes， including heat shock protein （Hsp70）， Cytochrome p450 （Cyp1a1）， serine/threonine protein kinase pim3 （Pim3）， Lysozyme， Leukocyte differentiation antigen （Cd109） and tumor necrosis factor receptor associated factor 6 （Traf6） （P<0.05）， revealing their important roles in the immune defense response of R. philippinarum against V. anguillarum. This study preliminarily explored the effects of V. anguillarum on R. philippinarum， providing new insights for the innate immune mechanism of the R. philippinarum against V. anguillarum.