## 4 Discussion  
Until recently, there have been only a few studies on the impacts of IR in hypoxic conditions on the reproduction and development of _B. dorsalis_[8; 10; 28]. The hypoxic environment diminishes the efficacy of IR on certain insects, leading the IPPC to withhold support for IR for the quarantine treatment of MAP commodities [12]. This study, however, demonstrates that the current phytosanitary IR dose is effective in inhibiting _B. dorsalis_ larvae even under HY/Sup-HY conditions (Figure 2), lending support to the administrative adjustments by the USDA-APHIS revising the minimum O2 level for treatments from 18% to 10% [29]. Our research findings also provide a robust basis for the IPPC to reassess the restrictions on phytosanitary IR applications for _B. dorsalis_ under hypoxic conditions. Figure 9: (**A**) The integrated schematic diagram of the key metabolic pathways and metabolites in the _B. dorsalis_ (Diptera: Tephrtidae) larvae exposed to different modified atmospheres under a phytosanitary irradiation dose, and (**B**) the changes in peak areas of 5 key compounds ((**a**). tetradecanoic acid, (**b**). n-hexadecanoic acid, (**c**). octadecanoic acid, (**d**). oleic acid and (**e**). squalene) across different treatment conditions. IR-induced DNA damage, a significant factor causing male sterility and impacting insect growth [30; 31], might suggest the activation of DNA repair genes under hypoxic conditions. However, the RNA-seq analysis conducted by [PERSON] et al. on DNA repair-related genes in the cowpea weeevil (_Callosobruchus maculatus_), including XRCC1, XRCC3, RAD23, and RAD51, indicates otherwise [8]. Their findings challenge the hypothesis that hypoxia aids in DNA-damage repair, suggesting that the mechanisms behind radioprotective effects in hypoxic conditions warrant further investigation. Our study first employed the DI-SPME-GC-MS technique, a validated and effective tool for metabolomic analysis, to explore this area. Topological analysis showed that treatments like IR-alone, HY, Sup-HY, HY + IR, and Sup-HY + IR primarily exhibit changes in the lipid metabolism pathways compared to the control group, especially in unsaturated fatty acids and fatty acid biosynthesis. These pathways are crucial for maintaining biological tissue structure and function [32; 33]. Interestingly, there was an overlap in the differential metabolic pathways observed across these five treatment methods, suggesting a certain correlation in the mechanisms of action of IR-alone, HY, Sup-HY, HY + IR and Sup-HY + IR on _B. dorsalis_ larvae. This also implies that the combined impacts of IR and MAs extend beyond a mere additive \"1 + 1\" effect. In this study, significant changes were observed in the free fatty acids of _B. dorsalis_ larvae across different treatment groups. These fatty acids are not only essential for maintaining cell membrane fluidity and permeability but are also vital in anti-oxidative stress [34; 35; 36]. Notably, \(n\)-hexadecanoic acid was consistently downregulated across all treatment groups, indicating a marked inhibition of the fatty acid biosynthesis pathway. As depicted in Figure 9B, n-hexadecanoic acid, a key differential regulatory metabolite, holds a central position in the differential metabolic pathways of the _B. dorsalis_ larvae subjected to IR-alone, HY, Sup-HY, HY + IR, and Sup-HY + IR treatments. N-hexadecanoic acid, a precursor to many long-chain and unsaturated fatty acids, can be transformed into significant amounts of free fatty acids like octadecanoic acid through the fatty acid elongation pathway, and their accumulation in the endoplasmic reticulum may inhibit certain protein synthesis pathways, negatively affecting insect growth and development [37; 38]. Furthermore, squalene is synthesized in the endoplasmic reticulum through the steroid biosynthesis pathway and promotes biological oxidation and metabolism, thereby enhancing the defense and stress-response capabilities of insects [39].