Characterization of eukaryotic microbial diversity in hypersaline Lake Tyrrell, Australia

Abstract

This study describes the community structure of the microbial eukaryotic community from hypersaline Lake Tyrrell, Australia, using near full length 18S rRNA sequences. Water samples were taken in both summer and winter over a 4-year period. The extent of eukaryotic diversity detected was low, with only 35 unique phylotypes using a 97% sequence similarity threshold. The water samples were dominated (91%) by a novel cluster of the Alveolate, Apicomplexa Colpodella spp., most closely related to C. edax. The Chlorophyte, Dunaliella spp. accounted for less than 35% of water column samples. However, the eukaryotic community entrained in a salt crust sample was vastly different and was dominated (83%) by the Dunaliella spp. The patterns described here represent the first observation of microbial eukaryotic dynamics in this system and provide a multiyear comparison of community composition by season. The lack of expected seasonal distribution in eukaryotic communities paired with abundant nanoflagellates suggests that grazing may significantly structure microbial eukaryotic communities in this system.

Keywords: 18S rRNA; Colpodella; Dunaliella; diversity; hypersaline; microbial eukaryotes; saltern.

Figure 1

(A) Lake Tyrrell, Victoria Australia. (B) Map showing of the lake within Australia and the sampling site for this project (^*). (C) 20–3.0 μm water fraction showing filter biomass. (D) Layered salt crust sampled for comparative purposes. Scale shows 1 cm increments (photo: J. Banfield).

Figure 2

Microbial eukaryote sample rarefaction analysis at 97% similarity. (A) individual sample curves and (B) grouped sample curves for summer (n = 6), winter (n = 3), and crust (n = 1).

Figure 3

Relative abundance of 18S rRNA sequences from summer samples (n = 6 samples; 1392 sequences) and winter samples (n = 3 samples; 450 sequences) and a benthic halite crust sample (crust) (n = 1; 322 sequences). Relative taxonomic abundance data by sample is provided in Table 4.

Figure 4

S17 Bray-Curtis similarity multidimensional scaling (MDS) plots showing sample similarity. (A) Clustering of all samples. (B) Clustering of only water samples.

Figure 5

18S rRNA ML tree of Colpodella (Apicomplexa) tree with 100 iterated bootstraps. The tree was inferred from an alignment of 89 nucleotide sequences over 1678 bp based using the Hasegawa-Kishino-Yano model in MEGA5. All representative Lake Tyrrell sequences (n = 69) are found within the collapsed node. The scale bar represents the number of substitutions per site. The tree is rooted with Thraustochytrium multiudimentale.

Figure 6

Expansion of 69 Lake Tyrrell Colpodella sequences from Figure 5.

Figure 7

18S rRNA ML of Dunaliella sequences based on a 1496 bp alignment, including 11 representative LT sequences (^*) with 13 Dunaliella public reference sequences. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site using the Hasegawa-Kishino-Yano model and MEGA5 as described in the methods. Bootstrap values are shown as the percentages of 100 trees inferred in the analysis. The tree was rooted with Chlamydomonas reinha.

Figure 8

18S rRNA ML tree of Ciliophora sequences generated from 23 sequences and a 1390 bp alignment. There were 14 representative Lake Tyrell sequences (^*) and 8 reference sequences. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site using the Hasegawa-Kishino-Yano model and MEGA5 as described in the methods. Bootstrap values are shown as the percentages of 100 trees inferred in the analysis. The tree was rooted with Dictyostelium discoideum.

Figure 9

18S rRNA ML tree of stramenopiles from 31 sequences and a 1519 bp alignment. The tree was generated using a Hasegawa-Kishino-Yano model and MEGA5 as described in the methods. There were 4 Lake Tyrrell sequences (^*). The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. Bootstrap values are shown as the percentages of 100 trees inferred in the analysis, and the tree was rooted with the dinoflagellate, Prorocentrum micans.

Figure 10

Select light micrographs of lake protists collected in January 2010 when cells were exposed to high salinities and temperatures. (A) Microscopic images showing Dunaliella cell aplanospores (cycts) and (B) an empty cell coat (C,D) Cells observed in cultures 12 months after collection (E) Heterotrophic flagellate, Colpodella sp. with ingested Dunaliella cell (F) Colpodella spp. (G) unidentified diatom observed in the halite crust sample. (H) unidentified ciliate from water sample. (I,J) unidentified diatoms from water samples.