Determination of Microbial Maintenance in Acetogenesis and Methanogenesis by Experimental and Modeling Techniques

Abstract

For biogas-producing continuous stirred tank reactors, an increase in dilution rate increases the methane production rate as long as substrate input can be converted fully. However, higher dilution rates necessitate higher specific microbial growth rates, which are assumed to have a strong impact on the apparent microbial biomass yield due to cellular maintenance. To test this, we operated two reactors at 37°C in parallel at dilution rates of 0.18 and 0.07 days-1 (hydraulic retention times of 5.5 and 14 days, doubling times of 3.9 and 9.9 days in steady state) with identical inoculum and a mixture of volatile fatty acids as sole carbon sources. We evaluated the performance of the Anaerobic Digestion Model No. 1 (ADM1), a thermodynamic black box approach (TBA), and dynamic flux balance analysis (dFBA), to describe the experimental observations. All models overestimated the impact of dilution rate on the apparent microbial biomass yield when using default parameter values. Based on our analysis, a maintenance coefficient value below 0.2 kJ per carbon mole of microbial biomass per hour should be used for the TBA, corresponding to 0.12 mmol ATP per gram dry weight per hour for dFBA, which strongly deviates from the value of 9.8 kJ Cmol h-1 that has been suggested to apply to all anaerobic microorganisms at 37°C. We hypothesized that a decrease in dilution rate might select taxa with minimized maintenance expenditure. However, no major differences in the dominating taxa between the reactors were observed based on amplicon sequencing of 16S rRNA genes and terminal restriction fragment length polymorphism analysis of mcrA genes. Surprisingly, Methanosaeta dominated over Methanosarcina even at a dilution rate of 0.18 days-1, which contradicts previous model expectations. Furthermore, only 23–49% of the bacterial reads could be assigned to known syntrophic fatty acid oxidizers, indicating that unknown members of this functional group remain to be discovered. In conclusion, microbial maintenance was found to be much lower for acetogenesis and methanogenesis than previously assumed, likely due to the exceptionally low growth rates in anaerobic digestion. This finding might also be relevant for other microbial systems operating at similarly low growth rates.

Simultaneous Oxidation of Atmospheric Methane, Carbon Monoxide and Hydrogen for Bacterial Growth

“…Thus, the numbers have possibly been underestimated, as DNA extractions from soils may provide less than 100% yields, qPCR quantification can be inhibited by soil-derived impurities in the DNA extract, and primer mismatches can result in an underrepresentation of copy numbers. Methanogenic archaea and acetogenic bacteria were recently found to require much less maintenance energy (0.2 kJ Cmol h −1 ) than previously believed (9.8 kJ Cmol h −1 ) [ 29 ]. According to the authors, the low maintenance energy was based on the low growth rates of these organisms, a feature that had not previously been taken into account.…”

Microbial Resource Management for Ex Situ Biomethanation of Hydrogen at Alkaline pH

“…Modified mineral medium DSMZ1036 containing yeast extract (0.2 g L −1 ), as described by Porsch and colleagues [36], was used for the enrichment and is designated as medium A hereafter. For further experiments, the medium was used in two variants: medium B did not contain yeast extract, but was supplemented with a vitamin solution, as described by [37], and cysteine-HCl as reducing agent in the same concentration as in medium A. Medium C contained vitamins, like medium B, but sodium sulfide as a reducing agent, as described by [37].…”

“…For further experiments, the medium was used in two variants: medium B did not contain yeast extract, but was supplemented with a vitamin solution, as described by [37], and cysteine-HCl as reducing agent in the same concentration as in medium A. Medium C contained vitamins, like medium B, but sodium sulfide as a reducing agent, as described by [37]. After preparing the media as described in Text S1 (Supplementary Materials), the pH for all media variations was adjusted to nine with a sterile anoxic stock solution of 2 M KOH.…”

Augmenting Biogas Process Modeling by Resolving Intracellular Metabolic Activity

“…While higher values have been estimated before, including 0.9 mmol ATP/gDW/h for M. maripaludis , and 3.6 mmol ATP/gDW/h for acetate converting M. barkeri [collected in (Koch et al (2016)], maintenance energies have also been reported to be lower than theoretical predictions under methanogenic conditions (Scholten and Conrad, 2000). A combination of experimental and modeling approaches has recently suggested even smaller values of below 0.116 mmol ATP/gDW/h for short hydraulic retention times, likely being applicable to also longer hydraulic retention times (Bonk et al, 2019). Fitting with this observation, maintenance demands in a binary propionate utilizing syntrophic methanogenic culture were experimentally determined to be 0.14 mmol ATP/gDW/h for Syntrophobacter fumaroxidans and 0.025 mmol ATP/gDW/h for Methanospirillum hungatei (Hamilton et al, 2015).…”