Biodiversity and culture of prokaryotes inhabiting haloalkaline and meromictic Soap Lake, Washington, USA

Abstract

Despite their potential for harboring novel microorganisms exhibiting beneficial metabolisms or that produce useful products for biotechnology and industry, alkaline lakes and soils are among the least studied extreme environments. With its high productivity and meromictic water column, haloalkaline Soap Lake (Washington, USA) is among the most intriguing soda lakes in the world. We sampled the water column of Soap Lake and used both culture-based and culture-independent (16S rRNA amplicon-based) methods to analyze the microbial diversity of both its oxic and anoxic waters. Cultivable aerobic heterotrophs were specifically targeted in enrichment cultures, and over 100 isolates were obtained. Small-subunit rRNA gene sequences were obtained for isolates that exhibited diverse colony morphologies and grew well on alkaline media containing varying concentrations of NaCl, and two of these isolates were chosen for in-depth characterization: strain 12SL-E129, which aligned within the genus Roseinatronobacter; and strain SL14, of the genus Vibrio. Both strains grew optimally at or above pH 9 and were halophilic-no growth was evident in the absence of NaCl for either isolate. In addition, strain SL14 exhibited impressive cold adaptation, showing a faster growth rate at 0°C than at 37°C. Community (16S rRNA) analyses conducted on Soap Lake water samples from both the mixolimnion (3 m) and the monimolimnion (23 m) revealed an extensive diversity of Bacteria, with the shallower depth dominated by species of Pseudomonadota (especially Alphaproteobacteria), Actinomycetota, and Bacteroidota; Deep anoxic waters were dominated by Bacillota, including many taxa containing endospore formers, as well as a marked increase in sulfate-reducing Deltaproteobacteria. Only low numbers of Archaea were identified in both the upper and lower waters of Soap Lake. Our data suggest that despite its extreme conditions (high alkalinity, steep salinity gradient, and reportedly extraordinarily high sulfide concentrations in the monimolimnion), Soap Lake is a highly productive aquatic system supporting thriving and diverse bacterial communities.

Keywords: Soap Lake; alkaliphilic bacteria; haloalkaliphile; halophilic bacteria; hypersaline; meromictic lake; soda lake.

Figure 1

Geography and sampling of Soap Lake, east central Washington, USA. Soap Lake is the final and lowest elevation lake in a chain of eight lakes in the Lower Grand Coulee Basin. The lake has a closed basin and no outlet. The bathymetric map [adapted from Edmondson and Anderson (1965)] indicates lake depths (m), with the anoxic monimolimnion indicated with red lines. The blue star indicates the sampling site. The water column was sampled on a calm day using a fine-resolution, peristaltic pump-driven sampling device (Sattley et al., 2017).

Figure 2

Soap Lake community sampling analysis. (A) Total taxonomic diversity of recovered DNA sequences from a 3-m Soap Lake water sample. “Other Phyla” are those bacterial taxa that individually represented <1% of total sequences and includes Acidobacteriota, Spirochaetota, Cyanobacteriota, Deinococcota, and “Candidatus Kaiseribacteriota.” Identified classes of Bacteria are shown in uncolored rows under each corresponding phylum. (B) Graphical representation of data in (A) with inner ring showing phyla and outer ring showing classes (classes not shown for phyla representing <8% of total recovered sequences). (C) Total taxonomic diversity of recovered DNA sequences from a 23-m Soap Lake water sample. “Other Phyla” are those bacterial or archaeal taxa that individually represented <1% of total sequences and includes Acidobacteriota, Balneolota, Chloroflexota, Nitrospirota, Fusobacteriota, Cyanobacteriota, Deinococcota, and Thermotogota, as well as the DPANN archaeal superphylum. Identified classes of Bacteria are shown in uncolored rows under each corresponding phylum. (D) Graphical representation of data in (C) with inner ring showing phyla and outer ring showing classes (classes not shown for phyla representing <5% of total recovered sequences). (E) Alpha diversity (Shannon and Simpson indexes) and beta diversity (Bray–Curtis dissimilarity and mean Aitchison distance) metrics at species level, indicating a greater degree of prokaryotic diversity at 23 m than 3 m in the Soap Lake water column, as well as substantial differences in the diversity between the two depths.

Figure 3

16S rRNA phylogenetic analysis of strain 12SL-E129 (red font). The sequenced 16S rRNA gene was aligned with closely related type strains using Muscle. The tree was constructed from 1,402 nucleotide positions in MEGA11 using the neighbor-joining method and the Kimura 2-parameter model. Bootstrap values (1,000 replicates) of >50% are indicated at the branching points, and GenBank accession numbers of all included taxa are shown in parentheses. The tree scale bar indicates a distance equivalent of 2 nucleotide changes per 100 nucleotides. Inset photo: Phase-contrast photomicrograph (1,000 × total magnification) of cells of strain 12SL-E129.

Figure 4

16S rRNA phylogenetic analysis of strain SL14 (red font). The sequenced 16S rRNA gene was aligned with closely related type strains using Muscle. The tree was constructed from 472 nucleotide positions in MEGA11 using the neighbor-joining method and the Kimura 2-parameter model. Bootstrap values (1,000 replicates) of >50% are indicated at the branching points, and GenBank accession numbers of all included taxa are shown in parentheses. The tree scale bar indicates a distance equivalent of 2 nucleotide changes per 100 nucleotides. Inset photos: Phase-contrast photomicrographs (1,000 × total magnification) of cells of strain SL14. Top photo: vibrio- to spirilla-shaped cells. Bottom photo: cells of strain SL14 with visible polar flagella (arrows).

Figure 5

Growth of Soap Lake bacterial isolates as a function of (A) temperature, (B) salinity, and (C) pH. Growth is presented as a percent of the maximum absorbance (OD₅₄₀) averaged from triplicate cultures for each parameter. Under optimal conditions, final turbidity maxima (OD₅₄₀) reached 0.416 for strain 12SL-E129 and 0.722 for strain SL14, with mean generation times of 20 h and 5.5 h, respectively.