The current communication reports the development of a single-stage biosystem for biohythane production from wastewater treatment. A semi-pilot scale bioreactor with 34 L capacity was used for this study. Maximum biohythane production of 147.5 +/- 2.4 L was observed after five cycles of operation with production rate of 4.7 +/- 0.1 L/h. The biohythane composition (H-2/(H-2 + CH4)) varied from 0.60 to 0.23 during stabilized fifth cycle of operation. During each cycle of operation, higher H-2 fraction was noticed within 12 h of cycle period followed by CH4 production for rest of operation (36 h). During biohythane production, COD removal efficiency of 60 +/- 5% (SDR, 29.0 +/- 1.9 kg CODr/m(3)-day) was also achieved. The synergistic function of volatile fatty acids (VFA) production and consumption during process in hybrid biosystem played vital role on the composition of biohythane. The single-stage biosystem facilitates production of high valued and cost efficient biofuel (biohythane) with fewer controls than individual acidogenic and methanogenic processes. (C) 2015 Elsevier Ltd. All rights reserved.