ABSTRACT Thermal oxo-degradation (TOD) of plastic can transform plastic waste into fermentable feedstocks. Bioconversion of the TOD products could be used as feedstocks in a biorefinery concept and lead to new avenues for plastic waste upcycling. Previous work demonstrated this concept with high-density polyethylene (HDPE), the most abundant type of plastic, and identified the nonconventional yeast Candida maltosa as a promising candidate for this application. Here, we describe the evolution of an improved strain of C. maltosa and characterize the uptake mechanisms of TOD products from HDPE (TODHDPE). Batch cultures in series passaged at the mid-exponential growth phase applied a selective pressure for faster growth and resulted in a >100% increase in specific growth rate when using TODHDPE as a carbon source compared to the wild-type strain used in previous work. Adaptive improvement in specific growth rate is an important step toward the development of an industrial strain, and it provides a basis for a mechanistic understanding of the TODHDPE uptake. The evolved strain was compared to the parent strain to identify the cellular and biochemical changes associated with the improved phenotype and the uptake mechanisms involved in the bioconversion of TODHDPE. This comparison found that C. maltosa secretes biosurfactants capable of solubilizing hydrocarbons. The adaptive evolution resulted in changes in biosurfactant production that translated to improved emulsification of alkanes and increased solubilization of fatty alcohols and alkanes. In addition to the changes in metabolites, the study identified increases in membrane permeability associated with a reduction in ergosterol that may also play a role in the improved phenotype. These findings support the development of C. maltosa and other potential microbial cell factories for plastic biorefineries and may inform future design strategies. IMPORTANCE Plastics are considered non-biodegradable because they take decades to break down into molecules that can enter the various carbon cycles. Thermal oxo-degradation can accelerate this rate-limiting step by turning plastic waste into a fermentable carbon source. In this work, we use Adaptive Laboratory Evolution to optimize the bioconversion of depolymerized HDPE by Candida maltosa. We also investigate the changes that resulted from the evolution process to learn about the molecular and cellular mechanisms involved in the bioconversion of hydrophobic substrates. Our findings show unique mechanisms in C. maltosa to overcome mass transfer limitations and metabolize long-chain hydrocarbons and fatty substrates that can possibly be extrapolated to industrial applications beyond plastic upcycling.
Rodriguez-Ocasio et al. (Wed,) studied this question.