Adaptive laboratory evolution of Rhodosporidium toruloides to inhibitors derived from lignocellulosic biomass and genetic variations behind evolution

Using lignocellulosic biomass hydrolysate for the production of microbial lipids and carotenoids is still a challenge due to the poor tolerance of oleaginous yeasts to the inhibitors generated during biomass pretreatment. In this study, a strategy of adaptive laboratory evolution in hydrolysate-based medium was developed to improve the tolerance of Rhodosporidium toruloides to inhibitors present in biomass hydrolysate. The evolved strains presented better performance to grow in hydrolysate medium, with a significant reduction in their lag phases, and improved ability to accumulate lipids and produce carotenoids when compared to the wild-type starting strain. In the best cases, the lag phase was reduced by 72 h and resulted in lipid accumulation of 27.89 ± 0.80% (dry cell weight) and carotenoid production of 14.09 ± 0.12 mg/g (dry cell weight). Whole genome sequencing analysis indicated that the wild-type strain naturally contained tolerance-related genes, which provided a background that allowed the strain to evolve in biomass-derived inhibitors.