Arundo donax hydrolysates shape hydrogen-producing microbial community in dark fermentation processs

Dark fermentation (DF) has great potential for development as a sustainable biohydrogen production system. Mixed anaerobic microbial consortia from sewage or wastewater sludge digester can be used as seeding for fermentative hydrogen (H2) production. Biomass such as energy crops are good substrates for H2 fermentation. Arundo donax (L.), a perennial non-food crop with high biomass, was chosen as a source of lignocellulosic biomass in a pilot-scale DF process. A mixed microbial consortium from a primary sludge digester, adapted to a synthetic medium amended with glucose, was used as inoculum. Before starting the hydrolysates fermentation, the inoculum was acclimated either on glucose or on A. donax hydrolysates. A glucose fermentation with glucose-acclimated inoculum was used as control. The microbial communities were characterized by Pyrotag sequencing of 16S rRNA genes. Hydrogen producing populations were quantified by Real-Time PCR of specific 16S rRNA and hydrogenase genes. Other coexisting microorganisms were also quantified. Performances of A. donax fermentation in presence of differently acclimated consortia were compared by analyzing molecular data in conjunction to H2 production. In glucose fermentation, where H2 production was fast, the bacterial consortium was characterized by the co-dominance of Enterobacteriaceae and Lactobacillaceae being 80% and 15% of the total community respectively. Microbial communities of both A. donax fermentations strongly differed from those of glucose-fermentation, being characterized by the dominance of Lactobacillaceae (40% of the total) and low percentages of H2 producing populations (<0.5% of the total). Quantification of hydrogenase genes of Clostridium spp. confirmed that these populations were present in low amount (104 gene copies). Nevertheless, remarkable H2 yields were recorded. The study evidence that the inhibitory compounds derived from the hydrolysis of A. donax had a prompt effect on the microbial community of the inoculum. However, this noticeable shift in the microbial communities did not affect H2 production. Finally, adaptation on different carbon substrates exerted a comparable selective pressure, leading to similar H2 yields.