Amylostereum areolatum is the main symbiotic fungus of woodwasp (Sirex noctilio) and is an active participant in the large-scale death of Pinus sylvestris. Woodwasps and associated fungi resort to active detoxification mechanisms to overcome the toxicity of host defence chemicals, which consist of a multitude of monoterpenes and diterpenes. Since cytochrome P450 (CYP) is considered to have extensive detoxification capabilities in fungi, 35 CYP genes were identified from A. areolatum by Iso-Seq. Moreover, we analysed the transcriptional levels of 11 chosen CYPs in the mycelia of fungi grown on different carbon sources or sprayed with different terpene mixtures or extracts to explore the relationship between CYPs and utilization of terpenoids or the detoxification capabilities for terpenoids. Molecular modeling and docking were also employed to predict the interaction between the P450 protein structure and substrate. The results showed that the 35 CYPs of A. areolatum belong to 22 families and 12 clans, and all had the typical P450 conserved domains. RT-qPCR revealed that most CYPs were down-regulated with monoterpenes as the sole carbon source and up-regulated with diterpenes. CYPs were induced after spraying with xylem extract, and venom can increase the expression levels of CYPs. Molecular docking predicted that three P450 proteins (CYP5037BM3, CYP5144KC1, and CYP5152T1) bind tightly to diterpenes through hydrogen bonds and π-alkyl interactions, they have a higher binding affinity for diterpenes than for monoterpenes, suggesting that they may preferentially metabolize diterpenes. This indicates that A. areolatum regulates the expression levels of key P450 enzymes through a special response pattern to reduce the toxicity of the chemical defence of the host on itself and its symbiotic insect, Sirex noctilio.