Based on the assessment of the trophic needs of basidiomycete Flammulina velutipes and the optimization of the composition of the nutrient medium using design of experimental approach, the yield of the submerged biomass of the fungus was increased to 41 g/L, the duration of the cultivation process was reduced to 5 days. For this purpose, the 24 full factorial design, the Box–Wilson steep ascent method and the construction of the response surface were used. Using the method of scanning electron microscopy, it was shown that the studied F. velutipes strain grew in the form of small spherical pellets with a diameter of 1–2 mm on an optimized medium. The surface of the pellets was loose; the inner part was filled with hyphae tightly adjacent to each other. The center of the pellets had no cavity. F. velutipes pellets were formed by septate hyphae with clamp connections. The micromorphological characteristics of the submerged F. velutipes mycelium ensured a high diffusion of nutrients and oxygen into the pellets and their maximum filling of the volume of the culture medium.
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