Staying productive under pressure: systems evaluations of β-carotene production in Yarrowia lipolytica under continuous fermentation
Alyssa M. Worland, Vincent Xu, Maria F. Duran, Philip Gitman, Kristen Hunter-Cevera, Cinzia Klemm, Yufei Sun, Diego Ruiz Sanchis, Rodrigo Ledesma‐Amaro, Kyle Pomraning, Deepti Tanjore, Mark Blenner, Yinjie Tang
Abstract
Scaling biomanufacturing from laboratory to industrial scale poses significant challenges, especially for continuous fermentation. This study investigates these challenges using a β-carotene-producing Yarrowia lipolytica strain. Through fermentation experiments and proteomics, we have assessed how fermentation modes, carbon sources, dissolved O 2 , and media composition influence long-term bioproduction. In shaking flask subcultures, the strain maintained β-carotene production for over ~30 generations. However, in continuous fermentations, subpopulation shifted toward faster-growing low-producers, leading to significant production losses within just ~18 growth generations. This process was accelerated by O 2 limitation and high bioreactor dilution rates. Using canola oil as a carbon source increases population heterogeneity but enhances β-carotene biosynthesis and prolongs production compared with glucose-based media. Kinetic modeling suggests that strains optimized for the highest production in laboratory settings may be less robust in industrial environments, where suboptimal yet faster-growing variants gain a competitive edge under prolonged stress and ultimately shape overall continuous fermentation performance.