Chinese cordyceps, a parasitic complex formed by the larvae of Hepialidae (Lepidoptera) and the fungus Ophiocordyceps sinensis, exhibits various therapeutic properties, such as anti-fatigue, anti-aging effects, and the treatment of pulmonary and renal diseases. Although artificial cultivation techniques for Chinese cordyceps have been established, the dimorphic transition of O. sinensis, from blastospores to hyphae after infecting the host, and ultimately forming mummified insect, remains a major bottleneck that directly affects the cultivation efficiency and cost. To promote this dimorphic transition, transcriptome analysis was performed on blastospores treated with exogenous compounds—kanamycin sulfate and naftifine hydrochloride, resulting in the identification of 2 801 differentially expressed genes (DEGs). Enrichment analysis revealed that, after three days of kanamycin sulfate treatment, both up- and down-regulated genes were predominantly associated with carbohydrate metabolism. In contrast, in naftifine hydrochloride-treated spores over three days, up-regulated genes were mainly enriched in glutathione metabolism, fatty acid degradation, and peroxisome pathways, while down-regulated genes were significantly enriched in glycolysis, the tricarboxylic acid (TCA) cycle, fatty acid biosynthesis, ribosome, and transcription/translation processes. These findings provide novel insights into the molecular mechanisms of the dimorphic transition in O. sinensis and suggest potential strategies for improving the efficiency of its artificial cultivation.