Abstract: 【Aim】 Micro-computed tomography (Micro-CT) is a high-resolution 3D imaging technology tailored for small samples, showing amazing potential in biological research, particularly in analyzing the fine structures and functions of tiny insects. However, this technology has not yet been widely applied in entomological research. This study presents the application and significance of Micro-CT in the biological research of parasitoids from multiple perspectives, thereby filling a research gap in this field within our country.【Methods】 Parasitoid samples under six different conditions were scanned, observed and analyzed using Micro-CT applied under various settings. Trissolcus japonicus adults parasitizing Halyomorpha halys eggs were instantaneously frozen with liquid nitrogen. The position of the T. japonicus ovipositor insertion into the eggshell of H. halys was observed by scanning the instantaneously frozen samples. The 3-day-old female adults of Anastatus japonicus were dehydrated and fixed prior to scanning, aiming to analyze the structures of their reproductive and digestive systems. The embryos of Trissolcus cultratus, which had developed for 18, 37, 54, 80, and 168 h within H. halys eggs, were placed in a -80 ℃ ultra-low temperature freezer for 2 min before scanning, and their morphological characteristics at different developmental time within host were monitored. Antheraea pernyi eggs that had been parasitized by Trichogramma chilonis and developed at room temperature for 5 d were randomly selected and stored in a -20 ℃ freezer for 6 months prior to scanning. Ooencyrtus utetheisae specimens that had developed for 7 d inside H. halys eggs were scanned to assess the number and survival status of the gregarious parasitoids present. Eight fully developed adult T. japonicus emerged from H. halys eggs were randomly selected and scanned to determine their sex. 【Results】Cross-sectional observation of the host eggshell revealed that, the ovipositor of T. japonicus adults only penetrated the outermost layer of H. halys eggshell from the start of drilling until the 90th second. The external genitalia of A. japonicus are located at the lowermost part of the abdomen, composed of ovipositor, inner valvulae and outer valvulae, with the upper valvifers separated by a chitinous septum. Its digestive system mainly includes crop, proventriculus, midgut, Malpighian tubules, ileum, rectum and anus, with digestive tract appearing spherical or sac-like. After 18-h development inside H. halys eggs, T. cultratus remained at the egg stage; after 37 h, it developed into larvae, and at 54 h it was still in the larval stage, with the volume of parasitoid wasp occupying 9.32% of the total volume of the host egg. At 80 h within H. halys eggs, T. cultratus reached the prepupal stage, with the volume of parasitoid wasp occupying 73.76% of the total volume of the host egg. After 168-h development, T. cultratus entered the pupal stage, at which point its volume occupied 58.64% of the total volume of the host egg. O. utetheisae reached the prepupal stage, after 7-d development within H. halys eggs. Each host egg contained five viable wasps. Scanning of A. pernyi eggs parasitized by T. chilonis revealed that the host eggs contained 31 individuals of T. chilonis all of which had deceased development at the pupal stage. The antennae of T. japonicus adults were distinctly geniculate shaped by micro-CT. In female adults of T. japonicus, all five segments at the end of the antennal flagellum were enlarged, whereas in male adults, the antennal flagellum lacked such enlargement. 【Conclusion】 Micro-CT provides exceptionally high resolution and imaging contrast. In this study, we have demonstrated from multiple perspectives the feasibility and practicality of applying this technology to parasitoid research, offering new technical support for the investigation of insect morphology and biological characteristics.

Key words: Micro-computed tomography, multi-dimensional internal imaging, egg parasitoids, morphological characteristics, biological characteristics