Cheryl A. Kelley

Methane transport mechanisms and isotopic fractionation in emergent macrophytes of an Alaskan tundra lake

The carbon isotopic composition of methane emitted by the Alaskan emergent aquatic plants Arctophila fulva, a tundra mid-lake macrophyte, and Carex rostrata, a tundra lake margin macrophyte, was −58.6 ± 0.5 (n=2) and −66.6±2.5 (n= 6) ‰ respectively. The methane emitted by these species was found to be depleted in 13C by 12‰ and 18‰, relative to methane withdrawn from plant stems 1 to 2 cm below the waterline. As the macrophyte-mediated methane flux represented approximately 97% of the flux from these sites, these results suggest the more rapid transport of 12CH4 relative to 13CH4 through plants to the atmosphere. This preferential release of the light isotope of methane, possibly combined with CH4 oxidation, caused the buildup of the heavy isotope within plant stems. Plant stem methane concentrations ranged from 0.2 to 4.0% ( x¯, 1.4; standard deviation (sd), 0.9; n=28) in Arctophila, with an isotopic composition of −46.1±4.3 ‰ (n = 8). Carex stem methane concentrations were lower, ranging from 150 to 1200 ppm ( x¯, 500; standard deviation, 360; n = 8), with an isotopic composition of −48.3±1.4‰ (n=3). Comparisons of the observed isotopic fractionations with those predicted from gas phase effusion and diffusion coefficients suggest a combination of one or both of these gas transport mechanisms with bulk (non-fractionationating) flow.

Carbon and hydrogen isotopic characterization of methane from wetlands and lakes of the Yukon-Kuskokwim Delta, western Alaska

The total methane flux to the troposphere from tundra environments of the Yukon-Kuskokwim Delta is dominated by emissions from wet meadow tundra (∼75%) and small, organic-rich lakes (∼20%). The mean δ13C value of methane diffusing into collar-mounted flux chambers from wet meadow environments near Bethel, Alaska, was −65.82 ± 2.21‰ (±1 sigma, n = 18) for the period July 10 to August 10, 1988. Detritus-rich sediments of Delta lakes, including margins of large lakes and entire submerged areas of smaller ones, are laden with gas bubbles whose methane concentration ranges from 11% to 79%. Lowest methane concentrations are found along heavily vegetated lake edge environments and highest through-out organic-rich, fibrous sediments of small lakes. A minimum ebullition flux estimated for the 5% of total Delta area comprised of small lakes ranges from 0.34 to 9.7 × 1010 g CH4 yr−1, which represents 0.6% to 17% of the total Delta methane emission. The δ13C and δD values of this ebullitive flux are −61.41 ± 2.46‰ (n = 38) and −341.8 ± 18.2‰ (n = 21), respectively. The methane in gas bubbles from two lakes is of modern, bomb carbon enriched, radiocarbon age. Gas bubble δ13C values varied from 2 to 5‰ seasonally, reaching heaviest values in midsummer, no such variations in δD values were observed. Combined isotope data reveal that higher δ13C values in heavily vegetated areas correlate with lower δD values, suggesting enhanced methane production via acetate fermentation. Spatial isotopic variations in lakes appear to be controlled by variations in production rather than oxidation processes.

Shifts in methanogen community structure and function associated with long-term manipulation of sulfate and salinity in a hypersaline microbial mat.

Methanogenesis was characterized in hypersaline microbial mats from Guerrero Negro, Baja California Sur, Mexico both in situ and after long-term manipulation in a greenhouse environment. Substrate addition experiments indicate methanogenesis to occur primarily through the catabolic demethylation of non-competitive substrates, under field conditions. However, evidence for the coexistence of other metabolic guilds of methanogens was obtained during a previous manipulation of sulfate concentrations. To fully characterize methanogenesis in these mats, in the absence of competition for reducing equivalents with sulfate-reducing microorganisms, we maintained microbial mats for longer than 1 year under conditions of lowered sulfate and salinity levels. The goal of this study was to assess whether observed differences in methane production during sulfate and salinity manipulation were accompanied by shifts in the composition of methanogen communities. Culture-independent techniques targeting methyl coenzyme M reductase genes (mcrA) were used to assess the dynamics of methanogen assemblages. Clone libraries from mats sampled in situ or maintained at field-like conditions in the greenhouse were exclusively composed of sequences related to methylotrophic members of the Methanosarcinales. Increases in pore water methane concentrations under conditions of low sulfate correlated with an observed increase in the abundance of putatively hydrogenotrophic mcrA, related to Methanomicrobiales. Geochemical and molecular data provide evidence of a significant shift in the metabolic pathway of methanogenesis from a methylotroph-dominated system in high-sulfate environments to a mixed community of methylotrophic and hydrogenotrophic methanogens under low sulfate conditions.

Substrate Limitation for Methanogenesis in Hypersaline Environments

Motivated by the increasingly abundant evidence for hypersaline environments on Mars and reports of methane in its atmosphere, we examined methanogenesis in hypersaline ponds in Baja California Sur, Mexico, and in northern California, USA. Methane-rich bubbles trapped within or below gypsum/halite crusts have δ¹³C values near -40‰. Methane with these relatively high isotopic values would typically be considered thermogenic; however, incubations of crust samples resulted in the biological production of methane with similar isotopic composition. A series of measurements aimed at understanding the isotopic composition of methane in hypersaline systems was therefore undertaken. Methane production rates, as well as the concentrations and isotopic composition of the particulate organic carbon (POC), were measured. Methane production was highest from microbial communities living within gypsum crusts, whereas POC content at gypsum/halite sites was low, generally less than 1% of the total mass. The isotopic composition of the POC ranged from -26‰ to -10‰. To determine the substrates used by the methanogens, ¹³C-labeled methylamines, methanol, acetate, and bicarbonate were added to individual incubation vials, and the methane produced was monitored for ¹³C content. The main substrates used by the methanogens were the noncompetitive substrates, the methylamines, and methanol. When unlabeled trimethylamine (TMA) was added to incubating gypsum/halite crusts in increasing concentrations, the isotopic composition of the methane produced became progressively lower; the lowest methane δ¹³C values occurred when the most TMA was added (1000 μM final concentration). This decrease in the isotopic composition of the methane produced with increasing TMA concentrations, along with the high in situ methane δ¹³C values, suggests that the methanogens within the crusts are operating at low substrate concentrations. It appears that substrate limitation is decreasing isotopic fractionation during methanogenesis, which results in these abnormally high biogenic methane δ¹³C values.

Methane oxidation potential in the water column of two diverse coastal marine sites

Methane oxidation in the water column was investigated at two nearshore marine environments with relatively high concentrations of dissolved methane. In the northern Gulf of Mexico, high methane oxidation rates were observed at the pycnocline, with the highest oxidation rate corresponding to the most negative bacterial δ13C values. These low isotopic values occurred during the winter when overall bacterial productivity was low, suggesting that at this time of the year, methanotrophs in the Gulf could make up a significant portion of the overall bacterial assemblage. Although methane oxidation also occurred during more productive times (i.e., summer), the isotopic signal of methane oxidation was not observed in the bacterial biomass because of the higher overall bacterial productivity. The other site, Cape Lookout Bight, NC, is a small marine embayment where methane is produced in the organic-rich sediments. No measurable rates of methane oxidation in the water column occurred, and no anomalously low δ13C values of the bacterioplankton were measured. In both environments, methane production and oxidation appear to be spatially coupled, occurring at/near the pycnocline in the northern Gulf of Mexico and at the sediment-water interface at Cape Lookout Bight, NC.

Isotopic composition of methane and inferred methanogenic substrates along a salinity gradient in a hypersaline microbial mat system.

The importance of hypersaline environments over geological time, the discovery of similar habitats on Mars, and the importance of methane as a biosignature gas combine to compel an understanding of the factors important in controlling methane released from hypersaline microbial mat environments. To further this understanding, changes in stable carbon isotopes of methane and possible methanogenic substrates in microbial mat communities were investigated as a function of salinity here on Earth. Microbial mats were sampled from four different field sites located within salterns in Baja California Sur, Mexico. Salinities ranged from 50 to 106 parts per thousand (ppt). Pore water and microbial mat samples were analyzed for the carbon isotopic composition of dissolved methane, dissolved inorganic carbon (DIC), and mat material (particulate organic carbon or POC). The POC delta(13)C values ranged from -6.7 to -13.5 per thousand, and DIC delta(13)C values ranged from -1.4 to -9.6 per thousand. These values were similar to previously reported values. The delta(13)C values of methane ranged from -49.6 to -74.1 per thousand; the methane most enriched in (13)C was obtained from the highest salinity area. The apparent fractionation factors between methane and DIC, and between methane and POC, within the mats were also determined and were found to change with salinity. The apparent fractionation factors ranged from 1.042 to 1.077 when calculated using DIC and from 1.038 to 1.068 when calculated using POC. The highest-salinity area showed the least fractionation, the moderate-salinity area showed the highest fractionation, and the lower-salinity sites showed fractionations that were intermediate. These differences in fractionation are most likely due to changes in the dominant methanogenic pathways and substrates used at the different sites because of salinity differences.

Trimethylamine and Organic Matter Additions Reverse Substrate Limitation Effects on the δ13C Values of Methane Produced in Hypersaline Microbial Mats

ABSTRACT Methane production has been observed in a number of hypersaline environments, and it is generally thought that this methane is produced through the use of noncompetitive substrates, such as the methylamines, dimethylsulfide and methanol. Stable isotope measurements of the produced methane have also suggested that the methanogens are operating under conditions of substrate limitation. Here, substrate limitation in gypsum-hosted endoevaporite and soft-mat hypersaline environments was investigated by the addition of trimethylamine, a noncompetitive substrate for methanogenesis, and dried microbial mat, a source of natural organic matter. The δ13C values of the methane produced after amendments were compared to those in unamended control vials. At all hypersaline sites investigated, the δ13C values of the methane produced in the amended vials were statistically lower (by 10 to 71‰) than the unamended controls, supporting the hypothesis of substrate limitation at these sites. When substrates were added to the incubation vials, the methanogens within the vials fractionated carbon isotopes to a greater degree, resulting in the production of more 13C-depleted methane. Trimethylamine-amended samples produced lower methane δ13C values than the mat-amended samples. This difference in the δ13C values between the two types of amendments could be due to differences in isotope fractionation associated with the dominant methane production pathway (or substrate used) within the vials, with trimethylamine being the main substrate used in the trimethylamine-amended vials. It is hypothesized that increased natural organic matter in the mat-amended vials would increase fermentation rates, leading to higher H2 concentrations and increased CO2/H2 methanogenesis.

Persistence of 17 β-Estradiol in Water and Sediment-Pore Water from Cave Streams in Central Missouri

Concentrations less than 10 ng/L of 17 β-estradiol (E2), a natural estrogen, have been linked to adverse health effects in fish, including skewed sex distributions, reproductive failure, and organ impairment. The persistence of E2 in carbonate aquifer systems is not well documented. Water and sediment from cave streams within the Ozark Plateau of Missouri, USA, were collected and analyzed for E2. The persistence of E2 in the water was examined in two separate experiments, in which the holding temperatures (20°C vs. 4°C), bottle type, exposure to light, and filtration were varied. During two trials, no statistical difference was observed in the concentration of E2, suggesting that E2 is stable within the water. The fate of E2 was also examined in sediment-pore water collected from the cave streams in two independent trials. In trial 1, a significant decrease in E2 was noted over the 29 days of the experiment. However, in trial 2, no change in E2 concentration was observed. The results indicate that E2 is relatively stable in cave stream water and may persist in the sediment.

Phytoplanktonic and Bacterial Carbon Pools and Productivities in the Gerlache Strait, Antarctica, during Early Austral Spring

A bstractPhytoplankton and bacterial biomass and productivities were investigated at four depths in the upper 500 m of the water column in the Gerlache Strait, Antarctica, during the prebloom period of early austral spring, from October 13 to November 4, 1995. The concentrations of all carbon pools were low, with the total particulate organic carbon (POC) concentration averaging 1.9 ± 0.9 μM. Bacterial, protozoan, and phytoplankton carbon accounted for 21% of the total POC, indicating that detritus or unenumerated organisms comprised the bulk of the POC during this period. Larger zooplankton or protozoa, such as ciliates, may account for this difference, since microzooplankton can represent a significant fraction of the total microbial biomass. Primary and bacterial secondary production rates were also low, less than 300 and 30 ng C L−1 h−1, respectively. However, when production was normalized to either chlorophyll or bacterial cell number, rates were similar to those recorded during the spring bloom periods. This indicates that the cells were metabolically active during the prebloom period. Chlorophyll specific primary production averaged over the upper 80 m of the water column was 1.28 ± 0.84 μg C μg chl−1 h−1, whereas the mean bacterial specific growth rate over the same depth interval was 0.34 ± 0.24 d−1. The overall production rates were low only because of the low abundance of cells during the prebloom period. When the site was reoccupied the following year, all measures of biomass and productivity were higher [36], emphasizing the large interannual variability in the Gerlache Strait.

Characterization of methane flux from photosynthetic oxidation ponds in a wastewater treatment plant

Photosynthetic oxidation ponds are a low-cost method for secondary treatment of wastewater using natural and more energy-efficient aeration strategies. Methane (CH(4)) is produced during the anaerobic digestion of organic matter, but only some of it is oxidized in the water column, with the remaining CH(4) escaping into the atmosphere. In order to characterize the CH(4) flux in two photosynthetic oxidation ponds in a wastewater treatment plant in northern California, the isotopic compositions and concentrations of CH(4) were measured in the water column, in bubbles and in flux chambers, over a period of 12 to 21 months to account for seasonal trends in CH(4) emissions. Methane flux varied seasonally throughout the year, with an annual average flux of 5.5 g CH(4) m⁻² d⁻¹ Over half of the CH(4) flux, 56.1-74.4% v/v, was attributed to ebullition. The oxidation efficiency of this system was estimated at 69.1%, based on stable carbon isotopes and a calculated fractionation factor of 1.028. This is the first time, to our knowledge, that a fractionation factor for CH(4) oxidation has been empirically determined for oxidation ponds. Quantifying CH(4) emissions from these systems is essential to properly identify their contribution and to mitigate their impact on global warming.