Lignocellulosic biomass is an abundant, renewable feedstock for biorefineries, but pretreatment is usually required to overcome its high recalcitrance to biodegradation. Fungal pretreatment, an alternative process to traditional pretreatments, can enhance the enzymatic digestibility of the lignocellulosic biomass. Fungal pretreatment can be performed at low temperature, without added chemicals, and no wastewater generation. However, in comparison with traditional pretreatments, longer residence times, lower yields, and feedstock sterilization requirements make it challenging to implement. This work investigated the fungal pretreatment of the dedicated energy crop Miscanthus × giganteus with the white rot fungus Ceriporiopsis subvermispora. Fungal pretreatment of non-sterile miscanthus was performed in batch using miscanthus previously colonized with the fungus as inoculum. The process enhanced the enzymatic digestibility of miscanthus by 2-fold over that of untreated miscanthus, and was comparable to the pretreatment of sterilized miscanthus inoculated with a pure culture of the white rot fungus. The finished material from the unsterilized pretreatment was used as inoculum for two more generations in a sequential fungal pretreatment process, where no increase in enzymatic digestibility was observed. A propagation of indigenous fungi that out-colonized C. subvermispora was observed through the generations, showing that sterilization is a required step for the stability and reproducibility of fungal pretreatment. A techno-economic analysis of the production of fermentable sugars from miscanthus using fungal pretreatment showed that the process was not feasible at full biorefinery scale due to the high capital cost caused by long residence time, low feedstock bulk density, and low sugar yields
