The global utilization of single-use, non-biodegradable plastics, such as bottles made of polyethylene terephthalate (PET), has contributed to catastrophic levels of plastic pollution. Fortunately, microbial communities are adapting to assimilate plastic waste. Previously, our work showed a full consortium of five bacteria capable of synergistically degrading PET. Using omics approaches, we identified the key genes implicated in PET degradation within the consortium’s pangenome and transcriptome. This analysis led to the discovery of a novel PETase, EstB, which has been observed to hydrolyze the oligomer BHET and the polymer PET. Besides the genes implicated in PET degradation, many other biodegradation genes were discovered. Over 200 plastic and plasticizer degradation-related genes were discovered through the Plastic Microbial Biodegradation Database (PMBD). Diverse carbon source utilization was observed by a microbial community-based assay, which, paired with an abundant number of plastic- and plasticizer-degrading enzymes, indicates a promising possibility for mixed plastic degradation. Using RNAseq differential analysis, several genes were predicted to be involved in PET degradation, including aldehyde dehydrogenases and several classes of hydrolases. Active transcription of PET monomer metabolism was also observed, including the generation of polyhydroxyalkanoate (PHA)/polyhydroxybutyrate (PHB) biopolymers. These results present an exciting opportunity for the bio-recycling of mixed plastic waste with upcycling potential.

Polyethylene terephthalate (PET) pollution has significant environmental consequences; thus, new degradation methods must be explored to mitigate this problem. We previously demonstrated that a consortium of three Pseudomonas and two Bacillus species can synergistically degrade PET in culture. The consortium more readily consumes bis(2-hydroxyethyl) terephthalate (BHET), a byproduct created in PET depolymerization, compared to PET, and can fully convert BHET into metabolically usable monomers, namely terephthalic acid (TPA) and ethylene glycol (EG). Because of its crystalline structure, the main limitation of the biodegradation of post-consumer PET is the initial transesterification from PET to BHET, depicting the need for a transesterification step in the degradation process. Additionally, there have been numerous studies done on the depolymerization reaction of PET to BHET, yet few have tested the biocompatibility of this product with a bacterial consortium. In this work, a two-step process is implemented for sustainable PET biodegradation, where PET is first depolymerized to form BHET using an orange peel ash (OPA)-catalyzed glycolysis reaction, followed by the complete degradation of the BHET glycolysis product by the bacterial consortium. Results show that OPA-catalyzed glycolysis reactions can fully depolymerize PET, with an average BHET yield of 92% (w/w), and that the reaction product is biocompatible with the bacterial consortium. After inoculation with the consortium, 19% degradation of the glycolysis product was observed in 2 weeks, for a total degradation percentage of 17% when taking both steps into account. Furthermore, the 10-week total BHET degradation rate was 35%, demonstrating that the glycolysis products are biocompatible with the consortium for longer periods of time, for a total two-step degradation rate of 33% over 10 weeks. While we predict that complete degradation is achievable using this method, further experimentation with the consortium can allow for a circular recycling process, where TPA can be recovered from culture media and reused to create new materials.

Global use of single-use non-biodegradable plastics, like bottles made of polyethylene tereph-thalate (PET), have contributed to catastrophic levels of plastic pollution. Fortunately, microbi-al communities are adapting to assimilate plastic waste. Previously, our work showed a full consortium of five bacteria capable of synergistically degrading PET. Using omics approaches we identified key genes implicated in PET degradation within the consortium’s pangenome and transcriptome. This analysis led to the discovery of a novel PETase, EstB, discovered to hydrolyze oligomer BHET, and polymer PET. Besides genes implicated in PET degradation, many other biodegradation genes were discovered. Over 200 plastic and plasticizer degrada-tion related genes were discovered through the Plastic Microbial Biodegradation Database (PMBD). Diverse carbon source utilization was observed by a microbial community-based as-say, which paired with an abundant number of plastic and plasticizer degrading enzymes in-dicates a promising possibility for mixed plastic degradation. Using RNAseq differential analysis, several genes were predicted to be involved in PET degradation including aldehyde dehydrogenases and several classes of hydrolases. Active transcription of PET monomer me-tabolism was also observed, including the generation of polyhydroxyalkanoate (PHA) bi-opolymers. These results present an exciting opportunity for the bio-recycling of mixed plastic waste with upcycling potential.

Polyethylene terephthalate (PET) pollution poses significant environmental consequences, and thus new degradation methods must be explored to mitigate this problem. We previously demonstrated that a consortium of three Pseudomonas and two Bacillus species can synergistically degrade PET in culture. The consortium more readily consumes Bis(2-Hydroxyethyl) terephthalate (BHET), a byproduct made in PET depolymerization, compared to PET, and can fully convert BHET into metabolically usable monomers terephthalic acid (TPA) and ethylene glycol (EG). Because of its crystalline structure, the main limitation of the biodegradation of post-consumer PET is the initial transesterification from PET to BHET, depicting the need for the transesterification step in the degradation process. Additionally, there have been numerous studies done on the depolymerization reaction of PET to BHET, yet few have tested the biocompatibility of that product with a bacterial consortium. In this report, a two-step process was implemented for sustainable PET biodegradation, where PET is first depolymerized to form BHET using an Orange Peel Ash (OPA) catalyzed glycolysis reaction, followed by complete degradation of the BHET glycolysis product by the bacterial consortium. Results show that OPA-catalyzed glycolysis reactions can fully depolymerize PET, with an average BHET yield of 92% (w/w) and that the reaction product is biocompatible with the bacterial consortium. After inoculation with the consortium, 19% degradation of the glycolysis product was observed in 2 weeks, for a total degradation percentage of 17% when taking both steps into account. Furthermore, the 10-week total BHET degradation rate was 35%, demonstrating that the glycolysis products are biocompatible with the consortium for longer periods of time, for a total two-step degradation rate of 33% over 10 weeks. While we predict complete degradation is achievable using this method, further experimentation with the consortium can allow for a circular recycling process, where TPA can be recovered from culture media and reused to create new materials.