Genomic and Transcriptome Analysis Reveals the Biosynthesis Network of Cordycepin in Cordyceps militaris

Cordycepin is the primary active compound of Cordyceps militaris. However, the definitive genetic mechanism governing cordycepin synthesis in fruiting body growth and development remains elusive, necessitating further investigation. This study consists of 64 C. militaris strains collected from northeast China. The high-yielding cordycepin strain CMS19 was selected for the analysis of cordycepin production and the genetic basis of cordycepin anabolism. First, the whole-genome sequencing of CMS19 yielded a final size of 30.96 Mb with 8 contigs and 9781 protein-coding genes. The genome component revealed the presence of four additional secondary metabolite gene clusters compared with other published genomes, suggesting the potential for the production of new natural products. The analyses of evolutionary and genetic differentiation revealed a close relationship between C. militaris and Beauveria bassiana. The population of strains distributed in northeast China exhibited the significant genetic variation. Finally, functional genes associated with cordycepin synthesis were identified using a combination of genomic and transcriptomic analyses. A large number of functional genes associated with energy and purine metabolism were significantly enriched, facilitating the reconstruction of a hypothetical cordycepin metabolic pathway. Therefore, our speculation of the cordycepin metabolism pathway involved 24 genes initiating from the glycolysis and pentose phosphate pathways, progressing through purine metabolism, and culminating in the core region of cordycepin synthesis. These findings could offer fundamental support for scientific utilizations of C. militaris germplasm resources and standardized cultivation for cordycepin production.