Molpeceres-Garcia_2026_Microb.Biotechnol_19_e70340

Polyethylene terephthalate (PET) waste remains a major environmental challenge due to its recalcitrance and low economic value. Here, we present an integrated biochemical approach that couples glycolysis with a synthetic microbial consortium to upcycle PET into polyhydroxyalkanoates (PHAs). Glycolysis efficiently depolymerized post-consumer PET into bis(2-hydroxyethyl) terephthalate (BHET) in 2 h, circumventing the limitations of in vivo PET degradation. We engineered a two-species microbial consortium composed of Comamonas testosteroni RW31, able to metabolize terephthalic acid, and Pseudomonas putida JM37, able to consume ethylene glycol, each modified for the extracellular secretion of PET- and MHET-hydrolases, employing different plasmid architectures. This division of labour creates a metabolic co-dependency, enabling rapid BHET hydrolysis and the subsequent upcycling of the released monomers into PHAs. The combination of the different strains allowed us to select C. testosteroni pSEVA354-MHETase and P. putida pSEVA234-PETase as the best consortium combination, based on growth and PHAs content. Overall, this work proposes a strategy for PET waste depolymerisation and valorisation, highlighting the potential of mixed chemical and biological approaches and the use of non-conventional microbial chassis within engineered consortia.