Continuous anaerobic oxidation of methane: Impact of semi-continuous liquid operation and nitrate load on N2O production and microbial community

Abstract

This work proves the feasibility of employing regular secondary activated sludge for the enrichment of a microbial community able to perform the anaerobic oxidation of methane coupled to nitrate reduction (N-AOM). After 96 days of activated sludge enrichment, a clear N-AOM activity was observed in the resulting microbial community. The methane removal potential of the enriched N-AOM culture was then studied in a stirred tank reactor (STR) operated in continuous mode for methane supply and semi-continuous mode for the liquid phase. The effect of applying nitrate loads of ∼22, 44, 66, and 88 g NO₃^- m^-3 h^-1 on (i) STR methane and nitrate removal performance, (ii) N₂O emission, and (iii) microbial composition was investigated. Methane elimination capacities from 21 ± 13.3 to 55 ± 12 g CH₄ m^-3 h^-1 were recorded, coupled to nitrate removal rates ranging from 6 ± 3.2 to 43 ± 14.9 g NO₃^- m^-3 h^-1. N₂O production was not detected under the three nitrate loading rates applied for the assessment of potential N₂O emission in the continuous N-AOM process (i.e. ∼22-66 g NO^-3 m^-3 h^-1). The lack of N₂O emissions during the process was attributed to the N₂O reducing capacity of the bacterial taxa identified and the rigorous control of dissolved O₂ and pH implemented (dissolved O₂ values ≤ 0.07 g m^-3 and pH of 7.6 ± 0.4). Microbial characterization showed that the N-AOM process was performed in absence of putative N-AOM archaea and bacteria (ANME-2d, M. oxyfera). Instead, microbial activity was driven by methane-oxidizing bacteria and denitrifying bacteria (Bacteroidetes, α-, and γ-proteobacteria).

Keywords: Anaerobic oxidation of methane; Greenhouse gas; Microbial community; Nitrate removal; Nitrous oxide production.