Activation of cryptic biosynthetic pathways in Saccharopolyspora spinosa through deletion of the spinosyn gene cluster: induction of cryptic and bioactive natural products

Saccharopolyspora spinosa, a member of the Pseudonocardiaceae family, was originally isolated from soil in the Virgin Islands and is renowned for producing spinosad, a broad-spectrum insecticidal secondary metabolite. While research on S. spinosa has historically focused on spinosad production, little is known about the broader spectrum of secondary metabolites encoded by its genome. Like Streptomyces, S. spinosa harbors numerous biosynthetic gene clusters (BGCs), many of which remain cryptic under standard laboratory conditions. In this study, the spinosyn gene cluster was deleted using the heat-sensitive vector pKC1139, generating the mutant strain S. spinosaΔSPN. The fermentation products of both the wild-type S. spinosa (B1) and S. spinosaΔSPN (B2) were analyzed through HPLC coupled with high-resolution tandem mass spectrometry (HRMS/MS). Data analysis was conducted using GNPS-based molecular networking and MestReNova. A total of seven metabolites were putatively annotated in the wild-type strain (B1), with spinosyns being the predominant compounds. In contrast, the mutant strain (B2) produced putatively linear and cyclic lipopeptides, including gageostatins and gageopeptins as the major metabolites. Additionally, the crude extract from S. spinosaΔSPN (B2) exhibited antibacterial activity, likely due to the production of lipopeptides, which are known for their antimicrobial properties. These findings indicate that deletion of the spinosyn gene cluster can activate cryptic biosynthetic pathways, leading to the discovery of novel bioactive compounds with potential applications in medicine.