Environ. Sci. Techno/. 1990, 24, 1157-1 164
Measurement of in Situ Rates of Selenate Removal by Dissimilatory Bacterial Reduction in Sediments Ronald S. Oremland,**t Nisan A. Steinberg,t Ann S. Maest,t Laurence G. Miller,t and James T. Holllbaugh* U.S. Geological Survey, 345 Middlefield Road, Menlo Park, California 94025, and Tiburon Center, San Francisco State University, Tiburon, California 94920
A radioisotope method for measurement of bacterial respiratory reduction of selenate to elemental selenium in aquatic sediments was devised. Sediments were labeled with [75Se]selenate,incubated, and washed, and 75Seo(s) was determined as counts remaining in the sediment. Core profiles of selenate reduction, sulfate reduction, and denitrification were made simultaneously in the sediments of an agricultural wastewater evaporation pond. Most of the in situ selenate reduction (85%)and all the denitrification activities were confined to the upper 4-8 cm of the profile, whereas sulfate reduction was greatest below 8 cm (89% of total). The integrated areal rate of selenate reduction was 301 pmol m-2 dag1, which results in a turnover of water column selenate in 82.4 days.
Introduction Oxyanions of selenium have been identified as toxic constituents in drainage waters from irrigated, seleniferous agricultural soils (1). This environmental problem is apparently quite common in many regions of the western United States (2). Therefore, methods that address the question of removal of selenium oxyanions from soils and/or drainage waters by various means have received considerable attention because they offer the prospect of restoring water or soil quality while allowing for the continuation of irrigated farming. With regard to possible microbiological methods, volatilization of selenium by formation of alkylated gases (e.g., dimethyl selenide) has been suggested for treatment of soils (3) or impacted marsh waters, such as those of the Kesterson Wildlife Refuge (4, 5).
Recently, we reported on a novel process by which anaerobic bacteria respire selenate, which in turn biochemically reduces this oxyanion to selenite and ultimately to elemental selenium (6). This dissimilatory reduction was demonstrated to occur in sediments, was independent of sulfate, and was inhibited by nitrate and chromate. Bacterial cultures capable of selenate respiratory growth have been isolated (6, 7). Although we were able to show that this process holds the potential to rapidly remove and sequester considerable quantities (millimolar) of added selenate from sediment slurries, the questions remained as to how rapidly it proceeds in nature, and what was its 'U.S. Geological Survey.
* San Francisco State University.
vertical distribution in sediments relative to other forms of anaerobic bacterial respiration. An understanding of how the process of dissimilatory selenate reduction operates in these respects should ultimately aid the design of treatment schemes and the rehabilitation of seleniumcontaminated regions. In order to answer these questions and to achieve these long-term goals, it was first necessary to devise a method for measuring in situ selenate reduction. In situ measures of bacterial activities have been a focus of microbial ecology for the past 25 years. A variety of techniques employing radioisotopes, gas chromatography, and/or enzyme inhibitors or analogues have been exploited to assess environmental rates of methanogenesis (8),denitrification (9, IO), nitrogen fixation ( I I ) , and sulfate reduction (12). This latter process is analogous to the methods we report herein to measure selenate reduction. However, in the case of selenate reduction, our task was simplified because 75Seis a y-emitting radioisotope ( t I l 2 = 120 days), which circumvents the quenching/efficiency problem associated with liquid scintillation counting of weak /3-emitters like 35S. Thus, there was no need to volatilize and trap the product 75Seo(s)as there is for [35S]sulfide,but merely to wash away the added [75Se]selenate. Our results with core profiles from a seleniumrich agricultural wastewater evaporation pond represent "quasi" in situ assays because they were incubated in the laboratory under conditions approximating the field. However, our results indicate that selenate reduction occurs in the surficial layers of these sediments, in proximity to the region of active denitrification, yet spatially separated from sulfate reduction occurring deeper in the core. Areal rates of selenate removal obtained from this core indicate its rapid removal by reduction to Seo(s), and a quick turnover of water column selenate. When combined with previous observations with regard to inhibitors and stimulators of selenium reduction, the conceptual design of an efficient anaerobic selenate "digestor" can be discerned.
Experimental Section Sites and Sampling. Surficial (upper -20 cm) sediments were collected from Hunter Drain, an agricultural drain located in Stillwater, NV, where selenium contamination is documented (2),as well as from the littoral zone of Big Soda Lake, an alkaline/saline environment (13) having a diversity of microbial activities (14,15).Grabs of anoxic sediments were placed in Mason jars (completely
Not subject to U.S. Copyright. Published 1990 by the American Chemical Society
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filled) and were used in the preliminary experiments to determine the practicality of the method. They were stored at 6 "C for up to 8 weeks prior to use. Subsequently, intact cores for determination of in situ activities were taken from a selenium-impacted agricultural waste evaporation pond in the San Joaquin Valley (6). Cores (1m X 8.25 cm) were collected by hand, stored at 6 "C, and processed within 24 h of collection. These sediments were generally characterized by having a brown surface layer (
