An evolved strain of Spathaspora passalidarum produces ethanol from sugarcane bagasse and switchgrass lignocellulosic hydrolysates

Barros, K. O.; Fisher, K. J.; Crandall, J. G.; Magni, G.; Serate, J.; Xie, D.; Zhang, Y.; Silva, S. S.; Sato, T. K.; Hittinger, C. T.; Rosa, C. A.

doi:10.1101/2025.09.26.678828

bioRxiv · 10.1101/2025.09.26.678828

An evolved strain of Spathaspora passalidarum produces ethanol from sugarcane bagasse and switchgrass lignocellulosic hydrolysates

Barros, K. O.·Fisher, K. J.·Crandall, J. G.·Magni, G.·Serate, J.·Xie, D.·Zhang, Y.·Silva, S. S.·Sato, T. K.·Hittinger, C. T.·Rosa, C. A.

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Abstract

Lignocellulosic hydrolysates, derived from plant biomass, contain various inhibitors that can hinder microbial growth. This study aimed to enable the growth and ethanol production by the xylose-fermenting yeast Spathaspora passalidarum in the presence of lignocellulosic hydrolysate inhibitors, particularly acetic acid. Ultraviolet (UV)-induced mutagenesis and adaptive laboratory evolution (ALE) were used to select for mutants with higher tolerance to these inhibitors. The initial mutant strain, MT01, was selected for increased growth in medium containing xylose and acetic acid. This strain underwent further evolution, resulting in the strain ME3.5.5, which showed significant improvements in both growth and ethanol production compared to the parental strain when tested in sugarcane bagasse hemicellulosic hydrolysate (SBHH). Genomic analysis identified non-synonymous and frameshift mutations in four genes, including CYR1 (encoding adenylate cyclase). These findings suggest that genetically optimized S. passalidarum strains could play a crucial role in advancing industrial bioethanol production from lignocellulosic biomass by overcoming the inhibitory effects of compounds found in lignocellulosic hydrolysates.

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Barros, K. O., Fisher, K. J., Crandall, J. G., Magni, G., Serate, J., Xie, D., Zhang, Y., Silva, S. S., Sato, T. K., Hittinger, C. T., Rosa, C. A.. 2025-09-28. An evolved strain of Spathaspora passalidarum produces ethanol from sugarcane bagasse and switchgrass lignocellulosic hydrolysates. https://doi.org/10.1101/2025.09.26.678828

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