This study investigated the bioconversion of dilute acid (2% H2SO4) pretreated red algae (Gelidium amansii) hydrolysate into H-2 by anaerobic fermentation in a continuous stirred tank reactor under mesophilic conditions using hybrid immobilized cells as microbial catalyst. Two different hydraulic retention times (HRT) of 24 h and 16 h with a feed concentration of 15 g/L hexose equivalent have been investigated over 85 days of operation to evaluate H2 production performance and stability of the continuous system. The highest hydrogen production rate (HPR) and hydrogen yield (HY) of 2.7 L/L/d and 1.3 mol/mol substrate hexose(added) was achieved at 24 h HRT, while further operation at 16 h HRT led to a significant drop in the hydrogen production with a HPR and HY values of 1.8 L/L/d and 0.7 mol/mol substrate hexoseadded, respectively. The bacterial community analysis characterized by 454 pyrosequencing revealed that the changes in HRT significantly influence the composition of the dominant microflora. At longer HRT (24 h), the phyla Firmicutes was abundant over 98%, whereas at shorter HRT (16 h), Proteobacteria being the dominant populations with 84%. These outcomes suggested that controlling appropriate HRT is prerequisite for efficient hydrogen production. (C) 2017 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.