Preserving Pichia: A Study on Its Longevity on Freeze-Dried Rodent Pellets

Our group has the goal of developing the yeast Pichia pastoris into a probiotic that can colonize human digestive tracts and deliver therapeutic proteins. To introduce Pichia pastoris into the mouse gut, a model for human beings, we freeze dried cultured P. pastoris into the food pellets of mice. This study examined the survival of P. pastoris on two different food pellets, grain-based and purified. We soaked both food pellets in concentrated liquid cultures of the yeast before freeze drying. To determine the viability of the yeast in these pellets, the freeze-dried pellets were ground, resuspended in liquid and then plated on selective medium. P. pastoris growth was tracked weekly by counting colonies on selective medium. While yeast viability was sufficiently high on the grain-based pellets during the first week, it decreased between weeks two and four. Nevertheless, four weeks after freeze drying, the concentration of surviving yeast remained sufficiently high to feed mice more than 1 × 108 colony forming units per day using the grain-based food pellet. The same method was used to freeze dry P. pastoris onto purified food pellets, which consequently showed lower survival of P. pastoris. The viability of P. pastoris in the purified food pellet suggests that significantly lower than 1 × 108 colony forming units per day would be achieved. Our results demonstrate that freeze drying P. pastoris onto a grain-based diet would enable us to treat mice with the desired concentration of living yeast, a key step toward developing P. pastoris into a probiotic that could potentially ameliorate several diseases.