We have investigated the pretreatment of softwood lignocellulosic feedstock using a controlled partial biological degradation in combination with enzymatic saccharification. It is known from studies of wood decay that brown rot fungi utilise a mechanism that causes a rapid and extensive depolymerisation of the carbohydrate polymers of the wood cell wall. The hyphae of these fungi act as delivery vectors to the wood cell wall for what is thought to be a combination of a localised acid pretreatment and a hydroxyl radical based depolymerisation of the cell wall carbohydrate polymers. After a controlled exposure of pine sapwood to brown rot fungi under mild conditions we have demonstrated glucose yields in the order of 70% of theoretical maximum and significant compromises to the mechanical strength of the biomass (reducing the energy requirements for size reduction). The results are specific to brown rot fungi and, in addition to significantly enhanced glucose release, offer further benefits including energy savings and flexible process configurations that are currently being developed by an early-stage spin-out company, Mycologix Ltd. Our results are discussed with regard to the potential of using brown rot fungi as a biological pretreatment for lignocellulosic biofuel production.