Development of microbial populations in the anaerobic hydrolysis of grass silage for methane production.

Six batch leach bed (LB) reactors, installed in parallel and connected to a common upflow anaerobic sludge blanket reactor, were fed with grass silage and operated at 35 (+/-1) degrees C. The development and distribution of microorganisms, which firmly and loosely attached to solid materials, and presented in the leachate in the LB reactors, were investigated by 16S rRNA gene-based terminal restriction fragment length polymorphism and clone library analyses. The phylotypes and their relative abundance changed in the respective bacterial community throughout the 49-day run and showed differences between the communities. Large numbers of phylotypes were detected from day 10 onwards. On day 17, the majority of phylotypes in the bacterial community firmly attached to solid residues affiliated to the classes Clostridia and Bacteroidetes. There were high numbers of the phylotypes in the leachate bacterial community. They were closely related to members of classes Clostridia, Bacteroidetes, Betaproteobacteria, Alphaproteobacteria, Gammaproteobacteria, and OP10. The Clostridium-like species clearly dominated the bacterial community. Archaea were only found in the solid residues on day 17 and in the leachate on days 10 and 17. The majority of the Archaea fell within the hydrogenotrophic genus Methanobacterium. The organism assigned to the aceticlastic genus Methanosarcina was only present in the solid residues.