A coculture system of Epicoccum nigrum with Mucor racemosus induces red pigment biosynthesis in vinegar mash

Abstract Fungal pigments hold significant food industrial value due to their safe and sustainable properties. Here we identified a naturally occurring coculture pair Epicoccum nigrum SPDX618 and Mucor racemosus SPDX522 in vinegar mash that produced red pigments, which was not observed in either monoculture. Our results showed that E. nigrum acted as the pigment producer while M. racemosus served as the elicitor. Response surface methodology and pH stabilization strategy improved the yield of red pigment into 328.7 U/g. Through bioactivity-guided fractionation coupled with UHPLC-MS/MS and in silico structure reconstitution, four red pigments were preliminarily assigned as anthracycline-like compounds. RNA-seq analysis revealed a putative dual-PKS gene cluster (SPDX618009169–SPDX618009178) with exclusively upregulation in cocultured E. nigrum that is potentially involved in red pigment biosynthesis. Besides, the production of the yellow pigment epipyrone was reduced in cocultured E. nigrum , consistent with the downregulated transcription level of its biosynthesis genes. Comparative metabolomics analysis further revealed a broad downregulation in both of primary metabolism and secondary metabolism in coculture system, indicating that a competition interaction relationship also existed between E. nigrum and M. racemosus . Altogether, this study reports a fungal coculture system that exhibited red pigments production. These findings contribute to the understanding of microbial interaction-driven metabolite biosynthesis and may provide a foundation for future exploration of their potential application as a colorant. Key points • An Epicoccum nigrum–Mucor racemosus coculture system was identified to produce red pigment for the first time. • E. nigrum acts as the red pigment producer and M. racemosus serves as the elicitor. • RNA-seq analysis tentatively identified a putative dual-PKS gene cluster upregulated in coculture that is putatively involved in red pigment biosynthesis.