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Abstract Cultivated meat production requires bioprocess optimization to achieve cell densities that are multiple orders of magnitude higher compared to conventional cell culture techniques. These processes must maximize resource efficiency and cost-effectiveness by attaining high cell growth productivity per unit of medium. Microcarriers, or carriers, are compatible with large-scale bioreactor use, and offer a large surface-area-to-volume ratio for the adhesion and proliferation of anchorage-dependent animal cells. An ongoing challenge persists in the efficient retrieval of cells from the carriers, with conflicting reports on the effectiveness of trypsinization and the need for additional optimization measures such as carrier sieving. To surmount this issue, edible carriers have been proposed, offering the advantage of integration into the final food product while providing opportunities for texture, flavor, and nutritional incorporation. Recently, a proof of concept (POC) utilizing inactivated mycelium biomass derived from edible filamentous fungus demonstrated its potential as a support structure for myoblasts. However, this POC relied on a model mammalian cell line combination with a single mycelium species, limiting realistic applicability to cultivated meat production. This study aims to advance the POC. We found that the species of fungi composing the carriers impacts C2C12 myoblast cell attachment—with carriers derived fromAspergillus oryzaepromoting the best proliferation. C2C12 myoblasts effectively differentiated on mycelium carriers when induced in myogenic differentiation media. Mycelium carriers also supported proliferation and differentiation of bovine satellite cells. These findings demonstrate the potential of edible mycelium carrier technology to be readily adapted in product development within the cultivated meat industry. 

The growth and activity of adherent cells can be enabled or enhanced through attachment to a solid surface. For food and beverage production processes, these solid supports should be food-grade, low-cost, and biocompatible with the cell of interest. Solid supports that are edible can be a part of the final product, thus simplifying downstream operations in the production of fermented beverages and lab grown meat. We provide proof of concept that edible filamentous fungal pellets can function as a solid support by assessing the attachment and growth of two model cell types: yeast, and myoblast cells. The filamentous fungus Aspergillus oryzae was cultured to produce pellets with 0.9 mm diameter. These fugal pellets were inactivated by heat or chemical methods and characterized physicochemically. Chemically inactivated pellets had the lowest dry mass and were the most hydrophobic. Scanning electron microscope images showed that both yeast and myoblast cells naturally adhered to the fungal pellets. Over 48 h of incubation, immobilized yeast increased five-fold on active pellets and six-fold on heat-inactivated pellets. Myoblast cells proliferated best on heat-treated pellets, where viable cell activity increased almost two-fold, whereas on chemically inactivated pellets myoblasts did not increase in the cell mass. These results support the use of filamentous fungi as a novel cell immobilization biomaterial for food technology applications.