Methanogens in biogas production from renewable resources – a novel molecular population analysis approach

Abstract

The population structure of thermo- and mesophilic biogas reactors digesting maize silage as the sole substrate was investigated employing a novel, highly degenerated PCR-primer pair targeting mcrA/mrtA coding for the key enzyme of methanogens. No sequence affiliating with Methanococcales, Methanopyrales, ANME-, rice or fen soil clusters was detected. Direct MeA PCR-cloning results indicated that Methanobacteriales were the most important methanogens in the thermophilic reactors. 57% and 80% of the analysed sequences affiliated with this order, 14% and 20% with Methanosarcinaceae and 0% and 29% with Methanomicrobiales. Methanomicrobiales dominated in the mesophilic reactors at the given conditions, 69% and 84% of the sequences recovered from direct MeA primed cloning affiliated with this order, 31% and 0% with Methanosarcinaceae and 0% and 16% with Methanobacteriales. No sequence affiliating with Methanosaetaceae was found. MeA primed PCR-single-strand conformation polymorphism indicated that population fluctuations occurred. According to sequence analysis of excised bands, Methanosarcinaceae dominated and Methanobacteriales were significantly represented in the thermophilic fermenter. Only 1 Methanosaetaceae sequence was found. Hydrogenotrophs appear to have a much higher and obligate acetoclastic methanogens a much lower importance than previously thought in biogas production from renewable resources.

Agricultural biogas production in Germany - from practice to microbiology basics

“…H 2 and formate are of outstanding importance in electron transfer between syntrophic donor bacteria and the accepting methanogens [32,33,44,45]. Along with syntrophic oxidation of other process intermediates, syntrophic acetate oxidation (SAO) was shown by results of different approaches to prevail in high throughput biogas processes, high-performance biogas plants particularly at high temperature and at high ammonia concentration [46][47][48][49][50][51][52]. The predominance of SAO in these processes is in contradiction to various textbooks assigning 70% of the methane to be produced by acetate splitting and only 30% via the hydrogenotrophic pathway.…”

“…In the mesophilic grass silage fermenters, a hitherto undescribed clade of sequences affiliating with Methanosarcinaceae (operationally named genus II) was identified ( Figure 1). In acidified mesophilic maize-silage-fed fermenters, sequences affiliating with the hydrogenotrophic order Methanomicrobiales were abundant at the DNA level [31,46], most probably due to their stability in acidic environments, but they are not necessarily the most active at such conditions (see below). Another novel clade falling into the mrtA branch of Methanobacteriaceae, operationally named Methanobacteriaceae II genus IV, was also frequently present at process acidosis of mesophilic maize digestion [63].…”

“…According to current understanding, the most susceptible steps are the final methanogenesis and the upstream syntrophically linked intermediary metabolism [73,74]. The primary focus was thus on developing monitoring tools for methanogenic Archaea and syntrophic bacteria and their activity [31,46,[75][76][77]. Further developments will include refinements, alternatives [33], and systems for the primary fermenting microorganisms (see below).…”

Mesophilic Fermentation of Renewable Biomass: Does Hydraulic Retention Time Regulate Methanogen Diversity?

“…This underrepresentation of acetoclastic Euryarchaeota only indicates a minor role for these methanogens given their very low specific growth rate and the low rate of substrate conversion relative to those of hydrogenotrophs. Some recent studies indicate a similar scenario with dominating hydrogenotrophic methanogens in mesophilic anaerobic digesters fed with maize or rye (4,22,30,35). Both for sheep (46,47) and in the rumen of cows, a similar situation concerning the methanogen population, i.e., dominating hydrogenotrophs, was reported (17,31).…”

“…Wavering microbial populations at one trophic level might cause a metabolic imbalance, leading to an accumulation of intermediates, pH changes, or reduced methane production efficiency (37), which in turn might even cause a process failure. As information on the influence of process parameters upon population dynamics is scarce (21), this study aimed to verify whether either a substrate change or a change in hydraulic retention time (HRT) could impact the diversity of methanogenic Euryarchaeota generating CH 4 . Beets were chosen among other potential crops as a model energy crop because they almost completely lack nondigestible lignin and have the highest energy output (45).…”