Acetylshikonin regulates the gut microbiota and inhibits the horizontal transmission of colistin-resistant plasmids.

BACKGROUND

The gut microbiota serves as a major reservoir for antibiotic resistance genes (ARGs), driving the spread of antimicrobial resistance (AMR) via horizontal gene transfer (HGT). Acetylshikonin (ASK), a naphthoquinone derived from the medicinal plant Lithospermum erythrorhizon, was proved to inhibit plasmid conjugation in vitro and in vivo. However, its impact on gut microbiota composition and precise HGT inhibition process within complex gut microbiota community remains unexplored.

PURPOSE: This study aims to clarify the precise inhibition effect of ASK on the transfer process of colistin-resistant plasmid in gut microbiota and its mechanisms.

METHODS

High-throughput cell sorting and 16S rRNA gene amplicon sequencing were employed to assess the precise gut microbiota species that ASK inhibited the resistant plasmid transfer to. The plasmid stability and re-transferability of transconjugants was evaluated by passaging culture and in vitro conjugative assay. The biochemical impact of ASK on donor cell and gut microbiota were tested by fluorescence assay and ELISA.

RESULTS

ASK changed the gut microbiota composition by enriching probiotics and reducing Gram-positive bacteria. In addition, ASK effectively inhibited the conjugative transfer of colistin-resistant plasmids to Proteobacteria (Escherichia and Ligilactobacillus) within the gut community. Furthermore, ASK weakened the stability and re-transferability of transconjugants, thereby limiting ARG further dissemination in gut. Moreover, ASK inhibited the electronic transport chain (ETC) and suppressed the ATP supply for both donor cells and the gut microbiota. Thus the plasmid conjugation processing in gut microbiota was inhibited by ASK.

CONCLUSION

This study demonstrated that ASK restructured gut microbiota and disarmed plasmid-mediated resistance spreading, offering a dual-targeted strategy against antimicrobial resistance.