Tidal freshwater wetland soils contain large amounts of organic carbon, some of which is mineralized to carbon dioxide (CO2) and methane (CH4) by a diverse consortium of anaerobic microorganisms that includes fermenters, syntrophs, and methanogens (MG). These microbial groups are tightly linked and often rely on cooperative interspecies metabolisms (i.e., syntrophy) to survive. Environmental perturbations can disrupt these interactions and thus alter the rates and pathways of carbon cycling. One environmental change of particular concern in coastal wetlands is sea level rise, which can result in increased episodic saltwater intrusion events into these ecosystems. These events cause an influx of sulfate (SO4-2) to the soils and may stimulate sulfate-reducing bacteria (SRB), which can directly compete with syntrophs for energy sources (e.g., fermentation products such as butyrate). Since syntroph metabolism generates byproducts that serve as the energy source for many MG, this competition can have indirect negative effects on methanogenesis. In addition, SRB can directly compete with MG for these byproducts, particularly formate, H2, and/or acetate. The goal of this study was to understand how both MG and syntroph-MG consortia respond to and recover from SRB competition during an episodic saltwater intrusion event. To achieve this, microcosms containing soil slurry from a freshwater wetland were subjected to simulated saltwater intrusion, and metabolic inhibitors were used to isolate the activity of the various functional groups. This study focused on the breakdown of butyrate, which is a key energy source in syntroph‑MG consortia metabolisms. The observed changes in butyrate breakdown rates and byproduct accumulation during butyrate degradation assays confirmed that butyrate breakdown was mediated through syntroph-MG consortia, and that formate, rather than H2, was likely used as an electron carrier during syntrophic activity. Additions of SO4‑2 (as Na2SO4) to the freshwater microcosms stimulated SRB activity and shifted the MG community to favor acetoclastic members. These changes were accompanied by a 24% increase in CO2 production and an 80% decrease in CH4 production. Interestingly, when NaCl was added to achieve similar ionic strength, CH4 production decreased by ~32%, suggesting SRB competition is not the only factor affecting methanogenesis. Butyrate degradation rates demonstrated that while SRB were strong competitors for butyrate, concurrent syntrophic metabolism was possible. Further, data show that SRB were poor competitors for acetate, which could explain the increase in acetoclastic MG. Following removal of SRB competition, CH4 production recovered but only by ~50% after 28 days, which suggests that some MG communities in tidal freshwater wetlands may not be resilient to saltwater intrusion events. Over this same time, rates of syntrophic butyrate breakdown largely recovered, but butyrate breakdown resulted in the production of less CH4 and acetate and more CO2 and formate, indicating saltwater intrusion events may lead to persistent changes in the byproducts and pathways of carbon breakdown in tidal freshwater wetlands.

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