Electron acceptors modulate methane oxidation and active methanotrophic communities in anoxic urban wetland sediments

Urban wetlands, although often overlooked, are hotspots for CH₄ cycling. However, the understanding of anaerobic CH₄ oxidation and microbial responses to different electron acceptors in urban wetlands remains limited. Here, we employed DNA-stable isotope probing (SIP) and metagenomic techniques to identify the core CH₄-oxidizing microbial groups in the presence of NO₃^-, SO₄^2-, or ferrihydrite (Fe(III)) in anoxic urban-lake sediments. The addition of NO₃^- to sediment microcosms promoted CH₄ oxidation, whereas SO₄^2- and Fe(III) had minimal or inhibitory effects on CH₄ oxidation rates in anoxic microcosms. The SIP-identified methanotrophs in response to the addition of CH₄ comprised the Marine Benthic Group D (MBG-D), "Candidatus Methylomirabilis Sh765B-TzT-35 and Z114MB74," and the putatively aerobic methanotrophs Methylobacter and Methylocystis. The presence of NO₃^- further enriched Methylocaldum, Methylomonas, 'Ca. Methylomirabilis Sh765B-TzT-35 and Z114MB74', Methylococcaceae, and Methylospira. In contrast, SO₄^2- and Fe(III) amendments notably shifted the dominant methanotrophs through the significant labeling of Methylocystis. Furthermore, metagenomic analysis of ¹³C-labeled DNA showed enrichment of genes for oxidation of CH₄ and its intermediates, as well as the reduction of NO₃^-, SO₄^2-, Fe(III), and fermentation. These results underscore the activity of NO₃^--dependent CH₄-oxidizing microorganisms and highlight their potential metabolic versatility in highly anthropogenically influenced urban wetlands.

Urban wetlands are critical ecosystems for CH₄ cycling but are increasingly impacted by complex pollutants from urban development, such as nitrates, sulfates, and Fe(III) from industrial runoff, atmospheric deposition, and wastewater discharge. This study reveals how these pollutants act as electron acceptors, modulating microbial metabolic pathways and reshaping methanotrophic communities under anoxic conditions. By uncovering the microbial mechanisms driving CH₄ oxidation in urban wetland sediments, our findings provide a deeper understanding of how anthropogenic pollution alters biogeochemical cycles. These insights are crucial for developing targeted strategies to mitigate CH₄ emissions and improve greenhouse gas control in rapidly urbanizing regions.