Newly isolated Sistotrema brinkmannii and Agaricus arvensis were co-cultured using rice straw (20 g/L) as a carbon source and yeast extract (10 g/L) as a nitrogen source to produce high levels of cellobiohydrolase (12.6 U/mL), beta-glucosidase (21 U/mL), and endoglucanase (16.4 U/mL). The filter paper activity (FPU) of the mixed fungal culture was enhanced (1.61 FPU/mL) compared to that of S. brinkmannii (0.3 FPU/mL) or A. arvensis (0.5 FPU/mL) monoculture. Enzyme loading, substrate concentration, pH, and temperature were optimized by response surface methodology (RSM) to improve the saccharification yield of alkali-pretreated plant biomasses. The highest enzymatic hydrolysis (76.7%) was obtained from Pinus densiflora under the following conditions: crude enzyme loading 22.5 FPU/g-substrate, substrate 3.75%, temperature 35 degrees C, and pH 5. The enzymatic hydrolyzate of pretreated P. densiflora was used for ethanol production using Saccharomyces cerevisiae, Pichia stipitis, and a co-culture of both the strains. The co-culture of S. cerevisiae and P. stipitis produced 23% more ethanol than that produced by S. cerevisiae alone and 38% more ethanol than that produced by P. stipitis alone. This study shows the potential of exploiting a microbial consortium for the cost-effective production of cellulases for bioethanol processes. (c) 2013 Elsevier Ltd. All rights reserved.