Broad-spectrum antifungal activity and genome-guided characterization of Paenibacillus polymyxa CACC1094 isolated from the bovine rumen

While Paenibacillus polymyxa is widely recognized for its biocontrol capabilities, most characterized strains originate from soil or rhizosphere environments, leaving animal-associated populations largely unexplored. In this study, we report the isolation of P. polymyxa strain CACC1094 from the bovine rumen and its genome-guided characterization to investigate its biosynthetic potential and antifungal activity. Whole-genome sequencing yielded a complete circular chromosome of 5.55 Mb with a GC content of 45.36%, comprising 5,099 coding sequences, 39 rRNA genes, and 111 tRNA genes. Comparative phylogenomic analysis placed CACC1094 within the P. polymyxa species complex, clustering most closely with the rumen-associated strain ND24 (ANI: 98.31%) and strain 188 (ANI: 96.92%), while forming a distinct branch within the species. Genome mining identified 13 biosynthetic gene clusters, including those associated with fusaricidin and tridecaptin biosynthesis, a paenicidin-like lanthipeptide cluster, and a hybrid NRPS-PKS cluster. In dual-culture assays, CACC1094 showed broad in vitro antifungal activity against multiple plant-pathogenic fungi and oomycetes, as well as selected fungal and yeast pathogens of clinical and veterinary relevance. LC-QTOF/MS analysis of culture supernatants confirmed the production of fusaricidin A and fusaricidin B, providing direct experimental validation of genome-derived predictions. Additionally, genome annotation revealed a complete CRISPR-Cas adaptive immune system, suggesting an integrated ecological strategy that combines antimicrobial biosynthesis with defense against mobile genetic elements. Together, these findings identify CACC1094 as a rumen-associated P. polymyxa strain with broad-spectrum antifungal activity and experimentally validated fusaricidin production, highlighting its potential as a source of antifungal metabolites for agricultural and veterinary applications.