Succession-informed enrichment of plastisphere communities from superworm larvae enables isolation and co-culture evaluation of polyethylene-associated bacteria

Polyethylene (PE) is a persistent environmental pollutant, yet how prolonged PE exposure restructures plastisphere communities and relates to surface-associated microbial transformation remains insufficiently understood. Here, gut-derived microbial communities from superworm larvae (Zophobas morio) were subjected to six months of enrichment on PE films under PE-only, carbon-limited conditions to examine plastisphere succession, predicted functional shifts, culture-based recovery of PE-associated bacteria, and co-culture effects on PE surface modification. 16S rRNA gene amplicon sequencing revealed stage-associated community restructuring, including an increase in Shannon diversity from 3.89 to 4.88, a shift from early Bacteroidota dominance (88.0% at month 1) to late Actinomycetota enrichment (0.7 to 42.0% by month 6), and persistent representation of Pseudomonadota. PICRUSt2 with MetaCyc profiling further indicated stage-associated functional shifts during prolonged PE exposure. After 180 days, enriched PE films showed dense biofilm formation, visible surface damage, oxidation-associated FT-IR bands, and increased carbonyl and hydroxyl indices relative to the blank and abiotic control, indicating measurable oxidation-associated surface transformation during enrichment. Stepwise DCPIP and gravimetric screening enabled recovery of five PE-associated strains. In co-culture evaluation, selected combinations, particularly Ko + As and treatments involving three or more strains, showed more sustained biomass persistence together with denser surface colonization, stronger oxidation-associated FT-IR signals, and higher CI and HI values than single-strain treatments. Overall, these results indicate that prolonged PE exposure drives stage-associated plastisphere restructuring and measurable PE surface oxidation, while succession-informed enrichment improves the recovery and evaluation of PE-associated culturable consortia. Key points Prolonged PE enrichment was associated with plastisphere succession and predicted functional shiftsStepwise screening enabled isolation of five culturable PE-degrading strainsCo-cultures enhanced PE colonization and oxidation-associated modification