Development of a microbial remediation system for the enhanced biodegradation of polyethylene bags

Abstract

The global escalating ecological burden of persistent polyethylene waste necessitates sustainable mitigation strategies. This study presents an innovative biodegradation approach utilising indigenous bacterial strains isolated from long-term plastic landfills, namely Alathur, Laloor, and Bhramapuram in Kerala, India. Soil samples in triplicate were collected from each dump site after recording physicochemical parameters such as pH and temperature and enriched in carbon-free liquid mineral salt medium (CFLMSM) for one month. These were exposed to non-pre-treated low-density polyethylene (LDPE) films under static incubation for 120 days. Preliminary screening with Coomassie Brilliant Blue staining followed by repeated quadrant streaking identified potent degraders, whose biodegradation potential was subsequently validated through weight reduction assays, BATH tests, FDA assays, and CO; evolution analysis. The most efficient strains, OBAS (42.73 + 1.105%, Alathur), OLA2 (39.7 + 1.27%, Laloor), and OBM3 (40.22 £ 2.04%, Bhramapuram) exhibited weight loss of LDPE films were identified by biochemical tests, 16S rDNA sequencing, phylogenetic tree construction, and MALDI-TOF analysis, and identified as Bacillus cereus NID1, Pseudomonas aeruginosa NID2, and Micrococcus luteus NID3. Strong biofilm formation on LDPE films was demonstrated by total cell count, EPS production, protein estimation, pH shifts of media, and FDA assays. Enzymatic quantifications of biofilms formed on the LDPE films further revealed the catalytic activities of laccases, lipases, proteases, lignin peroxidases, manganese peroxidases, and esterases. Moreover, biosurfactant production, contributing to the reduction of LDPE hydrophobicity, was demonstrated through oil displacement and emulsification index assays, and substantiated by FTIR analysis. Advanced validation of bacterial adherence and polymer surface alterations was conducted using FESEM, SEM-EDAX, AFM, roughness parameter calculation, and water contact angle analysis, all of which confirmed enhanced hydrophilicity. Mechanical degradation of LDPE films was verified by tensile testing, while chemical modifications were assessed through FTIR, ATR-FTIR-FPA, and GCMS analyses, with carbonyl index calculations highlighting strain-specific degradation efficiencies. To enhance biodegradation, a laboratory-formulated artificial consortium of B. cereus NIJD1, P. aeruginosa NJD2, and M. luteus NJD3 was developed. Its performance was evaluated through biofilm assays, weight loss experiments, and enzymatic profiling, revealing superior synergistic activity compared to individual strains. Further, the potency of the strains on the polymer films was authenticated through sophisticated analytical techniques such as FE-SEM, SEM-EDAX, AFM, roughness parameter calculations, WCA analysis, tensile testing, FTIR, ATR-FTIR-FPA and GC-MS analyses. Finally, four soil microcosm setups amended with LDPE films were established to evaluate bioremediation potential under biocaugmentation and biostimulation approaches. Physicochemical properties of the garden soil and heterotrophic counts in it were measured, while LDPE degradation was assessed via biomass quantification, FDA assays, CO, evolution, and weight loss analysis. SEM, AFM, FTIR, and GC-MS further confirmed the degradation of LDPE films buried in soil for four months. This multidisciplinary investigation demonstrates that indigenous bacterial strains and their formulated consortium can significantly accelerate LDPE degradation under both laboratory and soil conditions. These findings lay the foundation for eco-sustainable strategies to mitigate plastic waste accumulation and combat white pollution globally.