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Hills evaporation pond facilities contained between 0.005 and 5 mg of Se per liter. The objective of this study was to isolate Se-methylating organisms in evaporation pond water and to assess, through enrichment and manipulation of their optimal growth parameters, the environmental factors which govern microbial Se methylation. Alternaria alternata was isolated as an active Se-methylating organism. The volatile product was identified as dimethylselenide. The effects of pH, temperature, Se substrates, and methyl donors on the ability of A. alternata to methylate Se were investigated in liquid medium containing 100 mg of Se per liter. The optimum pH and temperature for methylation were 6.5 and 30°C, respectively. Selenate and selenite were methylated more rapidly than selenium sulfide and various organic Se compounds (6-selenoguanosine, 6-selenoinosine, seleno-DL-methionine, and 6-selenopurine). L-Methionine and methyl cobalamine (0.1 ,uM) stimulated dimethylselenide production. This study demonstrates that Se-methylating organisms are present in evaporation pond water and are capable of liberating substantial quantities of Se in the volatile dimethylselenide form. By determining the optimum environmental conditions which stimulate volatilization, it may be possible to design a way to remove Se from seleniferous water in situ. * Corresponding author. first time that A. alternata has been reported to be Se resistant as well as an active methylator of Se. MATERIALS AND METHODS Reagents. Sodium selenite and sodium selenate (Na2SeO4) were obtained from Alfa Products (Danvers, Mass.), and DMSe was from Strem Chemical Co. (Newburyport, Mass.). The following chemicals were obtained from Sigma Chemical Co. (St. Louis, Mo.): selenium sulfide, 6-selenoguanosine, 6-selenoinosine, seleno-DL-methionine, 6-selenopurine, methyl cobalamine, and L-methionine. Sodium tetrahydroborate was obtained from Aldrich Chemical Co. (Milwaukee, Wis.), and ammonium persulfate was from Mallinckrodt (Paris, Ky.). Water sampling. Water samples were collected from evaporation ponds 1, 2, 5, 6, and 7 at Kesterson Reservoir (Merced County, Calif.); from Lost Hills (Kern County, Calif.) A, B, and C evaporation ponds; and from cells 1 to 6 at the Peck Ranch (Fresno County, Calif.) (Fig. 1). The water was collected in Nalgene bottles, transported on blue ice, and stored at 5°C. Atomic absorption spectrometry. Water samples were analyzed for Se atomic absorption spectrometry with hydride generation. The instrument used was a Varian (Mulgrave, Victoria, Australia) Spectra AA/10 with a VGA-76 vapor generation assembly. The operational conditions were as follows: acetylene, 2.4 ml/min; air, 6.3 ml/min; nitrogen, 90 ml/min; sample flow, 6.5 ml/min; acid flow, concentrated

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A major concern in the San Joaquin Valley, CA, is that high levels of Se in agricultural drainage water are concentrating to hazardous levels in evaporation ponds. The primary objective of this study was to determine factors that affect Se biomethylation and thus permanently remove Se directly from evaporation pond water. Pond water samples (14–2000 µg Se L−1) were set up as laboratory mesocosms. The natural formation of dimethylselenide (DMSe) in unamended water was less than 1% of the total Se inventory after 40 d of incubation. No Se methylation took place in autoclaved, unamended pond water. L‐Methionine (10 µM) stimulated DMSe production in nonsterile pond water, and in autoclaved water (1 µM). Increasing the temperature to 35°C and the addition of 1% (wt/wt) glucose with a fungal inoculum, Alternaria alternata, doubled DMSe production over the controls after 25 d of incubation. Carbon sources such as glucose, maltose, sucrose, and galacturonic acid at 2 g C L−1 under ambient conditions slightly enhanced indigenous Se methylation (1.5‐fold). Of the amino acids tested, L‐methionine (0.02 g C L−1) stimulated DMSe evolution from pond water more so than L‐cysteine, L‐cystine, and L‐serine. The proteins, egg albumen, casein, and gluten (2 g C L−1) dramatically increased Se biomethylation causing a 23, 41, and 10% Se loss from the inventory, respectively, after 43 d of incubation. The stimulation of Se volatilization from evaporation pond water through specific amendments could prove to be promising as a field detoxification technique.

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