Process diagnosis using methanogenic Archaea in maize-fed, trace element depleted fermenters

A mesophilic maize-fed pilot-scale fermenter was severely acidified due to trace element (TE) deficiency. Mainly cobalt (0.07 mg * kg⁻¹ fresh mass (FM)), selenium (0.007 mg * kg⁻¹ FM) and sodium (13 mg * kg⁻¹ FM) were depleted. From this inoculum, three lab-scale flow-through fermenters were operated to analyse micronutrient deficiencies and population dynamics in more detail. One fermenter was supplemented with selenium, one with cobalt, and one served as control. After starvation and recovery of the fermenters, the organic loading rate (OLR) was increased. In parallel, the concentration (Real-Time PCR) of methanogens and their population composition (amplicon sequencing) was determined at the DNA and mRNA level. The parameters Metabolic Quotient (MQ) and cDNA/DNA were calculated to assess the activity of the methanogens.The control without TE supplementation acidified first at an OLR of 4.0 kg volatile solids (VS) * m⁻³ * d⁻¹ while the singular addition of selenium and of cobalt positively influenced the fermenter stability up to an OLR of 4.5 or 5.0 kg VS * m⁻³ * d⁻¹, respectively. In the stable process, the methanogenic populations were dominated by probably residual hydrogenotrophic Methanoculleus sp. (DNA-level), but representatives of versatile Methanosarcina sp. were most active (cDNA-level). When the TE supplemented fermenters began to acidify, Methanosarcina spp. were dominant in the whole (DNA-level) and the active (cDNA-level) community. The acidified control fermenter was dominated by Methanobacteriaceae genus IV. Until acidification, the concentration of methanogens increased with higher OLRs. The MQ indicated stress metabolism approximately one month before the TVA/TIC ratio reached a critical level of 0.7, demonstrating its suitability as early warning parameter of process acidification. The development of the cDNA/DNA ratio also reflected the increasing methanogenic activity with higher OLRs. Highest cDNA/DNA values (ca. 2) were obtained at metabolic strain of the methanogens, at the onset of acidification.