Cordyceps cicadae (Chanhua) is a parasitic fungus that grows on Cicada flammata larvae and is used to relieve exhaustion and treat numerous diseases, in part through its active constituent, cordycepin. We used de novo Illumina HiSeq 4000 sequencing to obtain transcriptomes of C. cicadae mycelium, fruiting body, and sclerotium, and identify differentially expressed genes. In the mycelium versus sclerotium libraries, 1576 upregulated and 2300 downregulated genes were identified. In the mycelium versus fruiting body and fruiting body versus sclerotium body libraries, 1604 and 1474 upregulated and 1365 and 1320 downregulated genes, respectively, were identified. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses identified 19 genes differentially expressed in mycelium versus fruiting body as related to the purine pathway, along with 28 and 16 genes differentially expressed in the mycelium versus sclerotium and fruiting body versus sclerotium groups, respectively. Gene expression of six key enzymes was validated by quantitative polymerase chain reaction. Specifically, 5′-nucleotidase (c62060g1) and adenosine deaminase (c35629g1) in purine nucleotide metabolism, which are involved in cordycepin biosynthesis, were significantly upregulated in the sclerotium group. These findings improved our understanding of genes involved in the biosynthesis of cordycepin and other characteristic secondary metabolites in C. cicadae.

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Diseases caused by soil-borne fungal pathogens result in significant crop yield losses and quality reduction. Streptomyces albidoflavus strain W68 is effective in controlling several soil-borne fungal diseases. To identify antifungal substances critical for biocontrol activity of W68, the genome of W68 was sequenced and a linear chromosome of 6.80 Mb was assembled. A total of 21 secondary metabolite biosynthesis gene clusters (BGCs), accounting for 12.27% of the genome, were identified. Core gene deletion mutants for each of all 8 BGCs for non-ribosomal peptide synthetases and polyketide synthases, were created. Among them, only the mutant lacking gene ctg1-5755 (the gene was renamed as fscDW68) in BGC 19, which shares 100% sequence similarity to the BGC for candicidin synthesis, showed obvious reduction in antifungal activity. A pot experiment revealed that biocontrol effects of the ΔfscDW68 mutant in Rhizoctonia rot of cucumber were also significantly compromised relative to W68. LC-MS analysis revealed that W68 but not ΔfscDW68 can produce candicidin isomers, indicating that the production of candicidin isomers is key for antifungal activity and biocontrol activity of S. albidoflavus W68. Importance This study provides the first report that candicidin-like secondary metabolites produced by microbial cells in natural soil environment can effectively control soil-borne fungal diseases, revealing a novel mechanism of microbial biocontrol agents. We demonstrated that the main antifungal activity and biocontrol activity of Streptomyces albidoflavus strain W68 is attributed to the production of candicidin isomers, suggesting gene clusters for candicidin-like compound biosynthesis might be used as molecular markers to screen and breed microbial strains for biocontrol agent development.