BACKGROUND Hemicellulose is considered one of the most abundant renewable bio-resources and has been utilized as substrate for the manufacture of numerous bio-products through microbiological approaches. However, it has rarely been reported that acetoin was produced directly from hemicellulose by microbial fermentation via consolidated bioprocessing (CBP). RESULTS To achieve efficient xylan utilization, four endo-1,4-beta-xylanase and three beta-xylosidase were co-overexpressed in Bacillus subtilis. Among the resulting engineered strains, BSL24, harboring xyn10B (from Clostridium stercorarium) and xsa (from Selenomonas ruminantium GA192) expression plasmid pHP13-E2X4 exhibited the best fermentation performance in xylan degradation with relatively high endo-1,4-beta-xylanase and beta-xylosidase activities, reaching 937.5 and 68 U L-1, respectively. The activities of the two enzymes were further improved by 83.0% and 124.6%, respectively, by using yeast extract as nitrogen source. Moreover, the addition of 0.25% v/w D-valine can further stimulate the total activity of endo-1,4-beta-xylanase in the cultures. Finally, pHP13-E2X4 was introduced in an acetoin-producing strain (168ARSRCP Delta acoA Delta bdhA), and the resulting strain BSL24AC produced 6.9 g L-1 acetoin from 1 g L-1 xylose and 19 g L-1 xylan in 96 h, with a yield of 0.345 g g(-1), corresponding to 72.6% of the theoretical yield. When the xylan concentration was increased to 49 g L-1, BSL24AC produced 15 g L-1 acetoin in 132 h, corresponding to an acetoin yield of 0.300 g g(-1) total sugar. CONCLUSIONS Bacillus subtilis was engineered to directly utilize hemicellulose by co-overexpression of different xylan hydrolases. To the best of our knowledge, this is the first study on efficient production of acetoin from hemicellulose via CBP in Bacillus subtilis. (c) 2018 Society of Chemical Industry