RNA-based spatial community analysis revealed intra-reactor variation and expanded collection of direct interspecies electron transfer microorganisms in anaerobic digestion

Granular activated carbon (GAC) has been shown to mediate direct interspecies electron transfer (DIET) in anaerobic digestion. Adding GAC to up-flow anaerobic sludge bed reactor increased the total biomass slightly from 20.0 to 26.6 gVSS/reactor, and maximum organic removal capacity remarkably from 285 to 1660 mgCOD/L/d. Since GAC occupied 7% of reactor volume (denser than suspended sludge, settled to the reactor bottom), we used a spatial sampling strategy (sludge bed top/mid/bottom layers, and tightly attached GAC-biofilm) and DNA- and RNA-based community analyses. RNA-based analysis demonstrated significant community differences between the non-GAC and GAC-amended reactors (p < 0.05) based on ANOSIM statistical analysis. In comparison, DNA-based analysis showed little community difference between these reactors (p > 0.05). RNA-based analysis revealed active enrichments in GAC-biofilm, including bacteria Geobacter, Syntrophus, Desulfovibrio and Blvii28, and archaea Methanosaeta and Methanospirillum. These are potential electro-active syntrophic microorganisms related with DIET, which expand the previously defined list of DIET microorganisms.