Genomic Inference Unveils Population Bottlenecks and a North-to-South Migration Pattern of Wild Cordyceps militaris Across China

The Ascomycete genus Cordyceps affects plant crops significantly, filling an important ecological niche. Cordyceps militaris (L.) Fr. presents many health benefits for humans, but its population history has not been reported. The objective of this research was to report the collection, population structure, demographic history, diversity, and cytosine deaminases of 43 wild strains of C. militaris in China through resequencing using an Illumina HiseqTM platform. All strains were assigned to the warm, subtropical, and middle temperate zone populations, confirmed by ADMIXTURE-1.3.0, PCA, and phylogenic analysis. Their population sizes declined historically, suggesting that this species suffered from bottlenecks in the wild. LD decays (r 2 ) revealed a north-to-south migration pattern of wild C. militaris , consistent with the MSMC2-v2.1.4 analysis. The regions of high Pi were aggregating at the chromosomes CP023325.1 (51) and CP023323.1 (9), playing a key role in adaptation, especially for the sites on cytosine deaminase. Within the species, genetic differentiation was relatively high among the three populations ( F st = 0.083, 0.092, and even 0.109). According to the artificial intelligence-assisted (RoseTTAFold) predicted structures of the cytosine deaminases, they were classified into eight clades with unique, distinct, and structurally conserved domains, offering a potential suite of single- and double-stranded deaminases of great promise as tunable base editors for therapeutic and agricultural breeding applications. These provided new insights for mining novel proteins from macrofungi, structurally and functionally.