Engineering of a probiotic yeast for the production and secretion of medium-chain fatty acids antagonistic to an opportunistic pathogen Candida albicans

Abstract

<i>Candida albicans</i> is an opportunistic pathogen, with its infection as one of the causes of morbidity or mortality. Notably, the probiotic yeast <i>Saccharomyces cerevisiae</i> var. boulardii has shown the potential to fight against <i>Candida</i> infections. In this study, we aimed to engineer a commercial boulardii strain to produce medium-chain fatty acids (MCFAs) with antagonistic effects against <i>C. albicans</i>. First, we identified and characterized a boulardii strain and created its auxotrophic strain <i>Δura3</i>. Next, we constructed and expressed a heterologous MCFA biosynthetic pathway under the control of inducible and constitutive promoters. Aside from examining MCFA production and secretion, we confirmed MCFAs' effects on <i>C. albicans'</i> anti-biofilm and anti-hyphal formations and the immunomodulatory effect of MCFA-containing supernatants on Caco-2 cells. We found that under constitutive promoters, the engineered boulardii strain constitutively produced and secreted a mixture of C6:0, C8:0, and C10:0. The secreted MCFAs then reduced biofilm and hyphal formations in <i>C. albicans</i> SC5314. We also confirmed that MCFAs upregulated the expression of virulence-related genes in SC5314. Furthermore, we found that the constitutively produced MCFAs in the supernatant induced the upregulation of immune response genes in Caco-2 cells co-cultured with SC5314, indicating MCFAs' roles in immunomodulation. Overall, the engineered boulardii strain produced and secreted MCFAs, as well as demonstrated antagonistic effects against <i>C. albicans</i> SC5314 and immune-modulatory effects in Caco-2. To our knowledge, this represents the first study tackling the metabolic engineering of a commercial probiotic yeast strain to constitutively produce and secrete MCFAs showing anti-<i>Candida</i> effects. Our study forms the basis of the potential development of a live biotherapeutics probiotic yeast against <i>Candida</i> infections through metabolic engineering strategies.