Objective The red imported fire ant (Solenopsis invicta) is recognized as one of the most dangerous invasive species worldwide, posing severe threats to agricultural and forestry production, ecosystem stability, and public safety in China. This study is aim to systematically analyze the critical environmental conditions and temporally regulated cellular developmental characteristics during extra-nest embryonic development in S. invicta, providing a essential developmental biology insights and key operational parameters to controlling their population expansion through gene intervention strategies such as CRISPR/Cas9.

Method Under controlled laboratory conditions (28±1℃, 80±1% relative humidity), the morphological progression and developmental phenotypes of S. invicta embryos, from fertilized egg to first-instar larva, were observed and documented using a deep-focus stereomicroscopy system. Embryos were cultivated in different media environments, specifically mineral oil and ddH₂O, to assess their effects on embryonic survival and development. Additionally, the dynamics of nuclear migration during early embryogenesis (0.5-4.0 h post-oviposition) were analyzed using DAPI fluorescence staining.

Result Mineral oil treatment significantly enhances the extra-nest embryonic hatching rate of S. invicta to 70%, markedly surpassing the control group (6.7%) and the ddH₂O group (3.3%). Morphological observations reveal that the mineral oil environment promotes normal embryogenesis, with embryos exhibiting regular developmental progression, including early blastula formation and establishment of larval primordia. In contrast, embryos in the ddH₂O and control groups displayed hydration imbalance phenotypes, such as larval tissue edema and condensation of water droplets on the embryonic surface. During early development, embryonic polarity is established within 0.5 h, followed by the formation of a syncytial blastoderm at 0.5-2.0 h, during which clear axial polarity becomes apparent. Gastrulation is initiated at 3.0-4.0 h, accompanied by the appearance of densely clustered nuclear signals.

Conclusion Maintaining water homeostasis is a critical limiting ecological factor for the extra-nest embryonic development of S. invicta. As a hydrophobic medium, mineral oil can effectively mimic the nest environment's function in preserving water homeostasis through surface coverage. This study delineates the nuclear migration pattern during early embryogenesis and identifies the period of 1.0-3.0 h after oviposition as the optimal operational window for gene-editing procedures. These results provide essential embryonic developmental data to support the development of targeted control technologies for the red imported fire ant.