Genome-centric metagenomics reveals microbial adaptation under high propionate stress in anaerobic digestion

Abstract

Propionate is a key intermediate in anaerobic digestion (AD), and its accumulation may lead to over-acidification and process failure. This study assessed methanogenic tolerance to increased propionate concentrations. The specific methane yield ranged from 238.0 to 277.3 mL/g COD at the initial propionate concentration of 2 to 10 g COD/L, and slightly decreased to 260.6 and 233.3 mL/g COD at 13 and 16 g COD/L, respectively. Kinetic analysis indicates lower inoculum-to-substrate ratio primarily negatively affected digestion time rather than accumulated methane yield. No clear inhibition occurred, even at 16 g COD/L, indicating a substantial propionate tolerance in AD. 16S rRNA gene analysis revealed that Desulfobacterota, particularly the family Syntrophobacteraceae was positively associated with propionate degradation, showing a linear correlation (p < 0.01) with both initial propionate concentration and digestion duration. Genome-centric metagenomic analysis further identified Bin.002 (Syntrophobacteraceae JABUEY01) and Bin.012 (Syntrophobacteraceae sp.) as dominant syntrophic populations, exhibiting high RPKM abundance and encoding the complete methylmalonyl-CoA (mmc) pathway. A metabolically diverse methanogenic community was also enriched, including aceticlastic Methanosarcina mazei (Bin.006), Methanothrix spp. (Bin.009 and Bin.014), and hydrogenotrophic Methanobacterium spp. (Bin.011 and Bin.026), which rapidly consumed metabolic intermediates, thereby avoiding inhibition at high propionate concentrations. Overall, the batch experiments demonstrated strong propionate tolerance under stable anaerobic conditions, supporting the conclusion that propionate accumulation is more likely a consequence rather than a primary cause of anaerobic reactor imbalance.

Graphical abstract

Keywords

Anaerobic digestion / Methane / Hydrogenotrophic methanogens / Kinetic model / Syntrophic propionate oxidation

Highlight

	● Propionate tolerance and microbial adaptation in anaerobic digestion were evaluated.
	● Propionate degradation was not evidently inhibited even at 16 g COD/L and ISR = 1.25.
	● 16S rRNA reveal Syntrophobacteraceae enriched significantly during incubation.
	● Metagenomic analysis suggest the putative SPOB–methanogen interactions.

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