Tenebrio molitor gut-derived microbial consortia as an ex situ platform for expanded polystyrene transformation

Expanded polystyrene (EPS) is highly persistent and remains limited under controlled experimental conditions. Here, we evaluated ex situ EPS biodegradation by gut-derived microbiota isolated from Tenebrio molitor larvae by integrating microbial succession, metabolomics, gel permeation chromatography (GPC), and partial carbon accounting. Enriched consortia were incubated for 42 days in carbon-free Bushnell–Haas medium with PS films as the sole carbon source, producing reproducible gravimetric loss across replicates (20.7–44.1%). Community profiling showed progressive loss of diversity and convergence toward Serratia marcescens dominance by weeks 5–6, coinciding with dense biofilm formation on PS surfaces. Ultra-high-performance liquid chromatography with high-resolution Orbitrap mass spectrometry (UHPLC–Orbitrap MS) captured a time-ordered aromatic catabolic sequence, with early transient styrene-series intermediates (styrene, styrene oxide, benzyl alcohol, benzaldehyde, phenylacetaldehyde) followed by increasing concentrations of phenylacetic acid, catechol, muconic acid, and other ring-cleavage products, consistent with engagement of the β-ketoadipate pathway and downstream entry into central metabolism. Polymer-level transformation was corroborated by GPC: pre-experiment and abiotic controls showed single narrow distributions, whereas microbiome-treated samples developed dual-peak profiles with a retained high-MW fraction and a newly formed low-MW fraction. Carbon accounting recovered 17–25% of polymer-derived carbon in the aqueous phase, dominated by dissolved organic carbon (DOC): 6.3–11.25%, and particulate organic carbon (POC): 4.76–7.70%, with smaller dissolved inorganic carbon (DIC): 3.40–6.25%. Collectively, these results demonstrate substantial EPS biodegradation by ex situ gut consortia and succession of S. marcescens during incubation of the mixed consortium.