Microbiome-Associated Regulation of Isoquinoline Alkaloids in Sinomenium acutum Reveals Functional Endophytes for Sinomenine Accumulation

Sinomenium acutum produces isoquinoline alkaloids such as sinomenine, but the role of endophytes in alkaloid biosynthesis and accumulation remains unclear. This study integrated 16S/ITS amplicon sequencing, untargeted metabolomics, and culture-dependent strain isolation to compare endophytic community structures and metabolite profiles between S. acutum roots with high (HC) and low (LC) isoquinoline alkaloid contents. Compared with the LC group, the HC group showed significantly higher levels of amino acids and their derivatives (e.g., Tyr-Trp), as well as isoquinoline alkaloids. A total of 49 bacterial strains and 20 fungal strains were isolated. Spearman’s correlation analysis revealed that two Microbacterium amplicon sequence variants (ASVs) were positively correlated with the contents of total tyrosine derivatives and sinomenine, while Cladosporium ASV40 was positively correlated with the content of sinoacutine, a precursor of sinomenine. Targeted LC-MS/MS analysis tentatively annotated isoquinoline and 8-hydroxyquinoline in G4 cultures and sinomenine in P37 cultures (MSI level 2). Importantly, in a six-month field trial, foliar spraying of Microbacterium sp. G4 significantly increased sinomenine content but significantly decreased (S)-reticuline content in S. acutum roots—consistent with enhanced channeling of this branch-point precursor toward sinomenine–supporting a functional association between G4 inoculation and sinomenine accumulation. These findings reveal previously unknown endophyte–metabolite associations in S. acutum and provide a basis for microbe-assisted improvement of the medicinal quality of S. acutum.