Microbial communities of the Lemon Creek Glacier show subtle structural variation yet stable phylogenetic composition over space and time

Glaciers are geologically important yet transient ecosystems that support diverse, biogeochemically significant microbial communities. During the melt season glaciers undergo dramatic physical, geochemical and biological changes that exert great influence on downstream biogeochemical cycles. Thus, we sought to understand the temporal melt-season dynamics of microbial communities and associated geochemistry at the terminus of Lemon Creek Glacier (LCG) in coastal southern Alaska. Due to late season snowfall, sampling of LCG occurred in three interconnected areas: proglacial Lake Thomas, the lower glacial outflow stream and the glacier’s terminus. LCG associated microbial communities were phylogenetically diverse and varied by sampling location. However, Betaproteobacteria, Alphaproteobacteria and Bacteroidetes dominated communities at all sampling locations. Strict anaerobic groups such as methanogens, SR1, and OP11 were also recovered from glacier outflows, indicating anoxic conditions in at least some portions of the LCG subglacial environment. Microbial community structure was significantly correlated with sampling location and sodium concentrations. Microbial communities sampled from terminus outflow waters exhibited day-to-day fluctuation in taxonomy and phylogenetic similarity. However, these communities were not significantly different from randomly constructed communities from all three sites. These results indicate that glacial outflows share a large proportion of phylogenetic overlap with downstream environments and that the observed significant shifts in community structure are driven by changes in relative abundance of different taxa, and not complete restructuring of communities. We conclude that LCG glacial discharge hosts a diverse and relatively stable microbiome that shifts at fine taxonomic scales in response to geochemistry and likely water residence time

Nitrogen fixation may provide a significant yet under quantified source of Nitrogen in the Great Lakes

Nitrogen fixation (NFix) is an important yet understudied microbial process in aquatic ecosystems and especially in the Laurentian Great Lakes (LGL). Early work suggested the contribution of NFix in the LGL is minimal to the nitrogen budget. However, recent work has shown during bloom events, NFix can help alleviate nitrogen limitations. Thus, we sought to revisit NFix in the LGL and comprehensively sample from near and offshore stations and with depth to understand the spatial variability of NFix. Nitrogen fixation rates were quantified using an adapted acetylene reduction assay (Stewart et al. 1967; doi:doi:10.1073/pnas.58.5.2071). Water samples were collected from discrete depths (Table 1) using a CTD-rosette water sampler equipped with 12 - 8L Niskin bottles and outfitted with a Sea-Bird multi-parameter profiler (conductivity, temperature and depth). Two liters of water were collected in triplicate from each depth and concentrated onto a 0.22µm pore size MF- Millipore Membrane filter using a vacuum manifold at low pressures to minimize cell breakage. Given the vast difference in lake depths between lakes and even within lakes, at each sampling station we took three depths, a surface (5m) sample (constant at all stations), mid- water column (based on max station depth) and near-bottom water (5m from bottom) (SI Table 1). The mid-water collection did not necessarily coincide with the thermocline, if it was even present. In preliminary testing in Lake Superior, we determined that approximately 2L of water was necessary to capture enough biomass to quantify rates in oligotrophic waters. In areas of high biomass, such as Lake Erie, water was filtered until the filter was clogged, which were all below 1L. Filtrate volume was noted, and final nitrogen fixation values were corrected based on volume filtered. Following filtration, filters were transferred to 50mL serum vials, submerged in 25mL of filtrate (lake water) from the same depth it originated from, and capped with a Teflon stopper. Acetylene gas was generated shipboard prior to sampling by combing 1g calcium carbide (CaC2) and 100ml of deionized water in a side arm flask attached to a 1L Tedlar gas sampling bag (EnviroSupply & Service). Samples were spiked with 1mL of acetylene gas and incubated at near in situ conditions (temperature and light) in the lab for 24 hours. For deep samples, incubations were done in the dark. After 24 hours, the incubations were terminated using 5mL of trichloroacetic acid (TCA) and stored in the dark at 4°C until measurements could be made in the lab. Acetylene gas concentration (peak area integration) was measured using an Agilent 6890 Plus GC System equipped with a flame deionized detector (FID) and a GS-Carbon Plot column 100/120 mesh (Agilent 113-3122). The GC-FID parameters are as follows: the carrier gas He2 was set at a flow rate of 30cm/s, the oven temperature was held isothermally at 125°C for 3 minutes, with an a split injection of 1:20 at 250°C. Sample injection volumes were 100 µL taken from the headspace of the vial. The retention times for acetylene and ethylene were observed at 1.8 and 1.9 minutes, respectively. For each station and at each depth, the following series of blanks and controls were used: 1) kill standard, 2) acetylene + DI water 3) acetylene + filtrate. Peak area integrations were calculated at retention times 1.9, and a correction ratio of 1:4 of N2 fixed to ethylene formed were factored in to determine molar N2 fixation rates (nmol/L/day) (Peterson and Burris 1976; doi:10.1016/0003-2697(76)90187-1)

Proteomic analysis reveals metabolic and regulatory systems involved the syntrophic and axenic lifestyle of Syntrophomonas wolfei.

Microbial syntrophy is a vital metabolic interaction necessary for the complete oxidation of organic biomass to methane in all-anaerobic ecosystems. However, this process is thermodynamically constrained and represents an ecosystem-level metabolic bottleneck. To gain insight into the physiology of this process, a shotgun proteomic approach was used to quantify the protein landscape of the model syntrophic metabolizer, Syntrophomonas wolfei, grown axenically and syntrophically with Methanospirillum hungatei. Remarkably, the abundance of most proteins as represented by normalized spectral abundance factor (NSAF) value changed very little between the pure and coculture growth conditions. Among the most abundant proteins detected were GroEL and GroES chaperonins, a small heat shock protein, and proteins involved in electron transfer, beta-oxidation, and ATP synthesis. Several putative energy conservation enzyme systems that utilize NADH and ferredoxin were present. The abundance of an EtfAB2 and the membrane-bound iron-sulfur oxidoreductase (Swol_0698 gene product) delineated a potential conduit for electron transfer between acyl-CoA dehydrogenases and membrane redox carriers. Proteins detected only when S. wolfei was grown with M. hungatei included a zinc-dependent dehydrogenase with a GroES domain, whose gene is present in genomes in many organisms capable of syntrophy, and transcriptional regulators responsive to environmental stimuli or the physiological status of the cell. The proteomic analysis revealed an emphasis macromolecular stability and energy metabolism to S. wolfei and presence of regulatory mechanisms responsive to external stimuli and cellular physiological status