Effects of magnetite on nitrate-dependent anaerobic oxidation of methane in an anaerobic membrane biofilm reactor: Metatranscriptomic analysis and mechanism prediction

Magnetite has been widely reported to accelerate methane production in anaerobic digestion. However, the effects of magnetite on the process of methane consumption, specifically nitrate-dependent anaerobic oxidation of methane (AOM), and its under-lying mechanisms remain poorly understood. In this study, the addition of magnetite significantly improved the removal of nitrate in an anaerobic membrane biofilm reactor (AnMBfR) (25.2 & PLUSMN; 2.5 mg NO-3-N/L/d) compared to that in an AnMBfR without magnetite (8.7 & PLUSMN; 4.2 mg NO-3-N/L/d). Metatranscriptomic analysis showed that genes encoding type IV pilus assembly proteins were highly expressed in nitrate/nitrite-reducing bac-teria in the biofilms of the two AnMBfRs. Specifically, the transcript abundance of these genes was observed to be higher in the presence of magnetite (gene expression levels, log2FPKM = 13.08) compared to the absence of magnetite (12.04). Biofilms with magnetite exhibited a linear increase in electron transfer resistance with increasing temperature, and the decrease in pH significantly increased their conductivity, indicating a metallic-like conductivity. Contrarily, an increase in temperature or decrease in pH did not influence the resistance or conductivity of biofilms without magnetite. Some of these cytochrome c (CYC)-like proteins were electrically active in the electrochemical Fourier transform infrared spectra and were closely associated with the outer-membrane multi-heme c-type cytochromes (OMCs). The intensities of the OMC-like bands in the surface-enhanced resonance Raman spectra with biofilms in the presence of magnetite were higher than those without magnetite. This study suggests that magnetite pro -motes the nitrate-dependent AOM process because of the formation of methanotrophic consortia based on direct interspecies electron transfer (DIET).& COPY; 2023 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Automated summary

Magnetite has been found to accelerate methane production in anaerobic digestion. In this study, the addition of magnetite significantly improved nitrate removal in an anaerobic membrane biofilm reactor. The presence of magnetite enhanced the expression of genes encoding type IV pilus assembly proteins, suggesting the formation of methanotrophic consortia based on direct interspecies electron transfer. The study highlights the role of magnetite in promoting nitrate-dependent anaerobic oxidation of methane.