Effects of imposed salinity gradients on dissimilatory arsenate reduction, sulfate reduction, and other microbial processes in sediments from two California soda lakes

Abstract

Salinity effects on microbial community structure and on potential rates of arsenate reduction, arsenite oxidation, sulfate reduction, denitrification, and methanogenesis were examined in sediment slurries from two California soda lakes. We conducted experiments with Mono Lake and Searles Lake sediments over a wide range of salt concentrations (25 to 346 g liter(-1)). With the exception of sulfate reduction, rates of all processes demonstrated an inverse relationship to total salinity. However, each of these processes persisted at low but detectable rates at salt saturation. Denaturing gradient gel electrophoresis analysis of partial 16S rRNA genes amplified from As(V) reduction slurries revealed that distinct microbial populations grew at low (25 to 50 g liter(-1)), intermediate (100 to 200 g liter(-1)), and high (>300 g liter(-1)) salinity. At intermediate and high salinities, a close relative of a cultivated As-respiring halophile was present. These results suggest that organisms adapted to more dilute conditions can remain viable at high salinity and rapidly repopulate the lake during periods of rising lake level. In contrast to As reduction, sulfate reduction in Mono Lake slurries was undetectable at salt saturation. Furthermore, sulfate reduction was excluded from Searles Lake sediments at any salinity despite the presence of abundant sulfate. Sulfate reduction occurred in Searles Lake sediment slurries only following inoculation with Mono Lake sediment, indicating the absence of sulfate-reducing flora. Experiments with borate-amended Mono Lake slurries suggest that the notably high (0.46 molal) concentration of borate in the Searles Lake brine was responsible for the exclusion of sulfate reducers from that ecosystem.

FIG. 1.

(A) Potential arsenate reduction in SL sediment slurries at various salinities (in grams/liter). (B) Arsenate reduction rates as a function of salinity in ML (open symbols) and SL (closed symbols) sediment slurries. The incubation times were 20 days (d) for ML sediments and 30 days for SL sediments.

FIG. 2.

DGGE of 16S rRNA obtained from SL and ML sediment slurries following incubation with added arsenate at various salinities and compared to natural sediment samples. DGGE profiles of natural sediments are profiles of samples collected from sediment hand cores at the labeled depth intervals (in centimeters). The salinities range from 25 to 346 g liter⁻¹ and are shown after the lake. The similarities between samples are shown in an unweighted-pair group method with arithmetic mean dendrogram of Dice similarity coefficients based on band position. The scale bar indicates levels of percent similarity. The numbered bands refer to the sequences in Table S1 in the supplemental material. Open arrowheads indicate bands that match 16S rRNA gene sequences from uncultivated organisms previously discovered in ML water (see text for details). The band indicated by the solid arrowhead matches strain SLAS-3, a clone that is closely related to the halophilic As(V)-respiring strain SLAS-1 isolated previously from SL sediments (16). (DGGE profiles of natural sediments are reprinted from reference .)

FIG. 3.

Phylogenetic relationship of partial arrA DNA sequences retrieved from Mono Lake sediments incubated at low (25 g/liter NaCl; MLSG-25) and high salinities (325 g/liter NaCl; MLSG-325). SL and ML refer to sequences originating from Searles Lake and Mono Lake sediments, respectively. Partial arrA sequences (n = 23) from the clone libraries obtained in this study (italicized, boldface type) were aligned with various known sequences using ClustalW in MacVector (v. 8). The tree was constructed using ARB and neighbor-joining analysis with Jukes-Cantor correction for transitions and transversions. Bootstrap values of at least 50% from 1,000 replications are labeled on the corresponding nodes. Clones marked with asterisks indicate unique arrA phylotypes. The scale bar represents 0.1 nucleotide substitution per site. Strains and GenBank accession numbers used in the tree are as follows: Bacillus arseniciselenatis strain E1H, AY660885; Shewanella sp. strain ANA-3, AAQ01672; and other SL and ML environmental sequences previously reported by Kulp et al. (10).

FIG. 4.

Potential rates of As(III) oxidation (A), denitrification (measured as N₂O production) (B), methanogenesis (C), and sulfate reduction (D) versus total salinity in ML (open symbols) and SL (closed symbols) sediment slurries. The inset in panel D shows N₂O production at salinities above 200 g liter⁻¹ on a finer scale. Arsenite oxidation slurries were incubated 20 days for ML and 14 days for SL. Denitrification slurries were incubated 14 days for ML and 27 days for SL. Methanogenesis was measured in As(V) reduction slurries incubated 20 days for ML and 30 days for SL. Sulfate reduction slurries were incubated 7 days for both lakes.

FIG. 5.

Sulfate reduction activity in SL sediment slurries inoculated with ML sediment. Sulfate reduction was stimulated at 25 and 90 g liter⁻¹ salinity in SL sediments that received 3% (by volume) active ML sediment as an inoculum (open symbols). No activity was observed in SL slurries where sterilized ML sediment was used as the inoculum (closed symbols). Incubation time was 10 days.

FIG. 6.

Sulfate reduction rates in ML sediment slurries showing the effect of borate additions. Slurries were incubated at a salinity of 50 g liter⁻¹ for 10 days.