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Background   Caldicellulosiruptor  species have gained a reputation as being among the best microorganisms to produce hydrogen (H 2 ) due to possession of a combination of appropriate features. However, due to their low volumetric H 2  productivities ( Q  H2 ),  Caldicellulosiruptor  species cannot be considered for any viable biohydrogen production process yet. In this study, we evaluate biofilm forming potential of pure and co-cultures of  Caldicellulosiruptor saccharolyticus  and  Caldicellulosiruptor owensensis  in continuously stirred tank reactors (CSTR) and up-flow anaerobic (UA) reactors. We also evaluate biofilms as a means to retain biomass in the reactor and its influence on  Q  H2 . Moreover, we explore the factors influencing the formation of biofilm.    Results  Co-cultures of  C. saccharolyticus  and  C. owensensis  form substantially more biofilm than formed by  C. owensensis  alone. Biofilms improved substrate conversion in both of the reactor systems, but improved the  Q  H2  only in the UA reactor. When grown in the presence of each other’s culture supernatant, both  C. saccharolyticus  and  C. owensensis  were positively influenced on their individual growth and H 2  production. Unlike the CSTR, UA reactors allowed retention of  C. saccharolyticus  and  C. owensensis  when subjected to very high substrate loading rates. In the UA reactor, maximum  Q  H2  (approximately 20 mmol  ·  L −1    ·  h −1 ) was obtained only with granular sludge as the carrier material. In the CSTR, stirring negatively affected biofilm formation. Whereas, a clear correlation was observed between elevated (>40 μM) intracellular levels of the secondary messenger bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) and biofilm formation.    Conclusions  In co-cultures  C. saccharolyticus  fortified the trade of biofilm formation by  C. owensensis , which was mediated by elevated levels of c-di-GMP in  C. owensensis . These biofilms were effective in retaining biomass of both species in the reactor and improving  Q  H2  in a UA reactor using granular sludge as the carrier material. This concept forms a basis for further optimizing the  Q  H2  at laboratory scale and beyond.    Electronic supplementary material  The online version of this article (doi:10.1186/s13068-015-0201-7) contains supplementary material, which is available to authorized users.

Biofilm formation by designed co-cultures of Caldicellulosiruptor species as a means to improve hydrogen productivity