Gut microbial communities of plastic-munching worms provide novel insights for the development of plastic-processing biotechnologies. Considering the complexity of worm maintenance and the gut microbial communities, it is challenging to apply the worms directly in plastic processing. Harnessing the power of microbial communities derived from the worm gut microbiomes in vitro may enable a promising bioprocess for plastic degradation. We performed two-stage enrichment of gut microbiomes from superworms (Zophobas atratus), establishing plastic-associated biofilm communities in vitro. We evaluated their plastic-degrading capabilities through multiple physicochemical characterizations and metagenomic analyses. We then integrated conventional sequence homology alignment and machine learning-based prediction to identify potential PDEs. Subsequent analysis of metagenome-assembled genomes (MAGs) was further employed to reveal the distinct microbial interaction patterns. Metagenomic profiling revealed more than a 15-fold increase in the abundance of PDE genes following in vitro incubation. Through machine learning approaches, we identified 3,454 putative PDE genes, along with 7 previously unreported
genera, 14 novel species, and 10 newly discovered strains harboring PDE genes. Notably, different types of plastic selectively enriched distinct microbial taxa and specific PDE genes, underscoring the substrate specificity and metabolic versatility of gut-derived microbiomes. Network analyses further revealed that plastic-degrading strains formed complementary metabolic interactions with fermentative microbes, collectively enhancing the efficiency of plastic degradation. Our findings reveal the largely untapped potential of superworm gut
microbiomes as a rich microbial reservoir for plastic degradation, highlighting the effectiveness of in vitro biofilm enrichment strategies. This work provides a foundation for future exploration of novel enzymes and the development of synthetic microbial consortia aimed at efficient plastic waste valorization.
