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ABSTRACT While methane is typically produced under anoxic conditions, methane supersaturation in the presence of oxygen has been observed in both marine and fresh waters. The biological cleavage of methylphosphonate (MPn), which releases both phosphate and methane, is one pathway that may contribute to this paradox. Here, we explore the genomic and functional potential for oxic methane production (OMP) via MPn in Flathead Lake, a large oligotrophic freshwater lake in northwest Montana. Time series and depth profile measurements show that epilimnetic methane was persistently supersaturated despite high oxygen levels, suggesting a possiblein situoxic source. Metagenomic sequencing indicated that 10% of microorganisms in the lake, many of which are related to the Burkholderiales (Betaproteobacteria) and Actinomycetota, have the genomic capacity to cleave MPn. We experimentally demonstrated that these organisms produce methane stoichiometrically with MPn consumption across multiple years. However, methane was only produced at appreciable rates in the presence of MPn when a labile organic carbon source was added, suggesting that this process may be limited by both MPn and labile carbon supply. Members of the generaAcidovorax,Rhodoferax, andAllorhizobium, organisms which make up less than 1% of Flathead Lake communities, consistently responded to MPn addition. We demonstrate that the genomic and physiological potential for MPn use exists among diverse, resident members of Flathead Lake and could contribute to OMP in freshwater lakes when substrates are available. IMPORTANCEMethane is an important greenhouse gas that is typically produced under anoxic conditions. We show that methane is supersaturated in a large oligotrophic lake despite the presence of oxygen. Metagenomic sequencing indicates that diverse, widespread microorganisms may contribute to the oxic production of methane through the cleavage of methylphosphonate. We experimentally demonstrate that these organisms, especially members of the genusAcidovorax, can produce methane through this process. However, appreciable rates of methane production only occurred when both methylphosphonate and labile sources of carbon were added, indicating that this process may be limited to specific niches and may not be completely responsible for methane concentrations in Flathead Lake. This work adds to our understanding of methane dynamics by describing the organisms and the rates at which they can produce methane through an oxic pathway in a representative oligotrophic lake. 

Considerable attention is given to absolute nutrient levels in lakes, rivers, and oceans, but less is paid to their relative concentrations, their nitrogen:phosphorus (N:P) stoichiometry, and the consequences of imbalanced stoichiometry. Here, we report 38 y of nutrient dynamics in Flathead Lake, a large oligotrophic lake in Montana, and its inflows. While nutrient levels were low, the lake had sustained high total N: total P ratios (TN:TP: 60 to 90:1 molar) throughout the observation period. N and P loading to the lake as well as loading N:P ratios varied considerably among years but showed no systematic long-term trend. Surprisingly, TN:TP ratios in river inflows were consistently lower than in the lake, suggesting that forms of P in riverine loading are removed preferentially to N. In-lake processes, such as differential sedimentation of P relative to N or accumulation of fixed N in excess of denitrification, likely also operate to maintain the lake’s high TN:TP ratios. Regardless of causes, the lake’s stoichiometric imbalance is manifested in P limitation of phytoplankton growth during early and midsummer, resulting in high C:P and N:P ratios in suspended particulate matter that propagate P limitation to zooplankton. Finally, the lake’s imbalanced N:P stoichiometry appears to raise the potential for aerobic methane production via metabolism of phosphonate compounds by P-limited microbes. These data highlight the importance of not only absolute N and P levels in aquatic ecosystems, but also their stoichiometric balance, and they call attention to potential management implications of high N:P ratios. 

ABSTRACT Radiocarbon ( 14 C) is an isotopic tracer used to address a wide range of scientific research questions. However, contamination by elevated levels of 14 C is deleterious to natural-level laboratory workspaces and accelerator mass spectrometer facilities designed to precisely measure small amounts of 14 C. The risk of contaminating materials and facilities intended for natural-level 14 C with elevated-level 14 C-labeled materials has dictated near complete separation of research groups practicing profoundly different measurements. Such separation can hinder transdisciplinary research initiatives, especially in remote and isolated field locations where both natural-level and elevated-level radiocarbon applications may be useful. This paper outlines the successful collaboration between researchers making natural-level 14 C measurements and researchers using 14 C-labeled materials during a subglacial drilling project in West Antarctica (SALSA 2018–2019). Our strict operating protocol allowed us to successfully carry out 14 C labeling experiments within close quarters at our remote field camp without contaminating samples of sediment and water intended for natural level 14 C measurements. Here we present our collaborative protocol for maintaining natural level 14 C cleanliness as a framework for future transdisciplinary radiocarbon collaborations. 

Abstract Inorganic carbon fixation, usually mediated by photosynthetic microorganisms, is considered to form the base of the food chain in aquatic ecosystems. In high-latitude lakes, lack of sunlight owing to seasonal solar radiation limits the activity of photosynthetic plankton during the polar winter, causing respiration-driven demand for carbon to exceed supply. Here, we show that inorganic carbon fixation in the dark, driven by organisms that gain energy from chemical reactions rather than sunlight (chemolithoautotrophs), provides a significant influx of fixed carbon to two permanently ice-covered lakes (Fryxell and East Bonney). Fryxell, which has higher biomass per unit volume of water, had higher rates of inorganic dark carbon fixation by chemolithoautotrophs than East Bonney (trophogenic zone average 1.0 µg C l −1 d −1 vs 0.08 µg C l −1 d −1 , respectively). This contribution from dark carbon fixation was partly due to the activity of ammonia oxidizers, which are present in both lakes. Despite the potential importance of new carbon input by chemolithoautotrophic activity, both lakes remain net heterotrophic, with respiratory demand for carbon exceeding supply. Dark carbon fixation increased the ratio of new carbon supply to respiratory demand from 0.16 to 0.47 in Fryxell, and from 0.14 to 0.22 in East Bonney. 

Abstract The segregation of bacteria, inorganic solutes, and total organic carbon between liquid water and ice during winter ice formation on lakes can significantly influence the concentration and survival of microorganisms in icy systems and their roles in biogeochemical processes. Our study quantifies the distributions of bacteria and solutes between liquid and solid water phases during progressive freezing. We simulated lake ice formation in mesocosm experiments using water from perennially (Antarctica) and seasonally (Alaska and Montana, United States) ice‐covered lakes. We then computed concentration factors and effective segregation coefficients, which are parameters describing the incorporation of bacteria and solutes into ice. Experimental results revealed that, contrary to major ions, bacteria were readily incorporated into ice and did not concentrate in the liquid phase. The organic matter incorporated into the ice was labile, amino acid‐like material, differing from the humic‐like compounds that remained in the liquid phase. Results from a control mesocosm experiment (dead bacterial cells) indicated that viability of bacterial cells did not influence the incorporation of free bacterial cells into ice, but did have a role in the formation and incorporation of bacterial aggregates. Together, these findings demonstrate that bacteria, unlike other solutes, were preferentially incorporated into lake ice during our freezing experiments, a process controlled mainly by the initial solute concentration of the liquid water source, regardless of cell viability.