Population dynamics of methanogens during acidification of biogas fermenters fed with maize silage

Abstract

Limitations in the supply of essential trace elements for methanogenic Archaea can arise in biogas production from renewable resources in flow‐through systems particularly when no manure is added. Without compensating supplementation, primarily Co and Na+ can become limiting in long‐term mono‐digestion of maize silage at threshold values of ca. 0.03 and 10 mg/L, respectively. These deficiencies apparently triggered process acidification. Using molecular biological methods (PCR‐SSCP and quantitative real‐time PCR), microbial population dynamics were monitored qualitatively and quantitatively at distinct stages during the experiments, investigating mcrA/mrtA, coding for a subunit of the key enzyme of methanogenesis. An exponential correlation between mcrA/mrtA copies and methane productivity was obtained. Members of obligatley acetoclastic Methanosaetaceae were found only at low acetate concentrations (below 1 g/L). At organic loading rates>1 g volatile solids/(L×d) and without acidification symptoms, obligately hydrogenotrophic (oh) Methanobacteriales and versatile Methanosarcinaceae were dominating, and an abundance of up to 1010 methanogens per mL fermenter content was determined. In the acidified process, however, ca. 4 orders of magnitude less methanogens were detected, and Methanomicrobiales (oh), more specifically Methanospirillum hungatei or Methanoculleus spp., were dominating. Species diversity at the DNA level was highest at efficient process performance without stress symptoms and at a relatively low organic loading rate (1–2 g volatile solids/(L×d)). According to the quantitative real‐time PCR data, the process was not sustained below an availability of 10−8 to 10−9 μg Co per methanogenic cell.

Agricultural biogas production in Germany - from practice to microbiology basics

“…After feeding stop, adjustment of TE concentrations, process recovery and re-continued operation, secondary acidosis and process breakdown due to sodium deficiency occurred. To our knowledge, this was the first report [31] that the AD process collapses due to Na deficiency, with a threshold at ca. 10 mg · kg FM −1…”

“…In support of the conclusions, metal/micronutrient determination of the fed maize silage batches (Table 1) confirmed correspondingly low concentrations of Co and of Na in the batches fed before and at the secondary acidification. Ni, Se, Mo, and Fe concentrations were sufficiently provided by the maize silage in average, and threshold concentrations [31] were not reached in the fermenter sludges. However, synergistic detrimental effects on the biocenosis cannot be excluded if several TEs are concomitantly approaching limiting levels.…”

“…Grass silage typically has much higher contents of almost all TEs than maize silage (Table 1) and may thus be used to compensate TE deficits. It should be considered, however, that grass silage has also higher protein content (Table 2), ; values in italics are alarming with respect to levels reported in [21,31]; data are compiled from [30,31,34] and unpublished results.…”

“…By plotting process performance data against total concentrations of ICP-MS determined metal concentrations it turned out that Cobalt was the primarily limiting TE. At about 60 μg Co per kg fresh matter (FM) first and at about 30 μg Co · kg FM −1 , severe acidosis symptoms with process breakdown were seen [30,31]. Cart analysis (http://www.statmethods.…”

“…In such cases, necessary supplementation of specific TEs should be calculated by mass balances considering silage TE contents and fermenter sludge volume changes in order to avoid process acidification. However, adding more than required for optimum process performance [21,31] typically does not lead to further improvement and should be avoided because toxicity problems can arise. Improvement by higher dosage points to additional limitations or TE precipitation, and efforts should be made in this case to increase the bioavailability by withdrawal of precipitating anions or agents.…”