Cyanobacterial ecotypes in the microbial mat community of Mushroom Spring (Yellowstone National Park, Wyoming) as species-like units linking microbial community composition, structure and function

Abstract

We have investigated microbial mats of alkaline siliceous hot springs in Yellowstone National Park as natural model communities to learn how microbial populations group into species-like fundamental units. Here, we bring together empirical patterns of the distribution of molecular variation in predominant mat cyanobacterial populations, theory-based modelling of how to demarcate phylogenetic clusters that correspond to ecological species and the dynamic patterns of the physical and chemical microenvironments these populations inhabit and towards which they have evolved adaptations. We show that putative ecotypes predicted by the theory-based model correspond well with distribution patterns, suggesting populations with distinct ecologies, as expected of ecological species. Further, we show that increased molecular resolution enhances our ability to detect ecotypes in this way, though yet higher molecular resolution is probably needed to detect all ecotypes in this microbial community.

Figure 1

Mushroom Spring showing cyanobacterial mat in foreground and spring in background. Inset shows DGGE analysis of PCR-amplified 16S rRNA segments for unsectioned samples collected at the four temperature sites. Band labels are discussed in text; letters and primes (e.g. A, B′ and NPE) indicate 16S rRNA genotypes and, except for genotype OP10, a number following these designations indicates the number of nucleotide differences from that genotype (e.g. A′1 and C″2 indicate sequences one and two nucleotides different from genotypes A′ and C″).

Figure 2

Schematics showing the evolution of an ecotype (blue) through a series of periodic selection events (dashed fans exhibiting extinct variants) and the divergence of a new distinct ecotype (green) through a niche-invasion mutation and subsequent private periodic selection events, to form two distinct clades of extant variants (solid fans at ends of lineages). a–d depict progressively ‘younger’ ecotypes with progressively smaller degrees of divergence, with d depicting a nascent ecotype in a population whose diversity has yet to be purged by periodic selection.

Figure 3

Detailed views of vertical profiles within samples of each temperature site. (Column 1, left-most) cross-section views of mat layers (bar is 1 mm); (Column 2) photomicrographs showing autofluorescence of native Synechococcus populations sampled from the upper yellow-green and deep dark-green layers (bar is 10 μm); (Column 3) DGGE profiles of samples collected at successive 100 μm intervals progressing from top (left-most lane) downwards (to right). Band labels are discussed in text and described in the legend of figure 1.

Figure 4

Neighbour-joining phylogenetic tree based on ITS sequence variation. Clades and subclades corresponding A, A′ and B′ Synechococcus lineages defined by 16S rRNA variation are indicated. These clades were mostly demarcated based on bootstrap values greater than or equal to 70%. The tree was rooted with the ITS sequence of Synechococcus strain C9. Scale bar indicates substitutions per site.

Figure 5

Community phylogeny analysis of ITS variants from 2001 to 2004 Mushroom Spring and Octopus Spring samples showing comparison of observed and modelled distributions of the number of sequence clusters (bins) versus sequence identity criterion.

Figure 6

Parsimony phylogenetic trees showing clades demarcated as ecotypes by community phylogeny analysis of ITS variants of the Synechococcus A, A′ and B′ clades. n is the most probable number of ecotypes and values in parentheses indicate the lower and upper 95% confidence interval bounds of ecotype number, respectively. Clones originating from the upper and lower parts of the photic zone are indicated by upward- and downward-pointing triangles, respectively. Shading indicates temperature: white, 60°C; grey, 65°C; and black, 68°C.

Figure 7

Microprofiles of oxygen, oxygenic photosynthesis and light at photosynthetically useful wavelengths (numbers on curves indicate nanometres) measured with microsensors through the vertical aspect of mats at each temperature site.

Figure 8

Microsensor measurements of light intensity versus wavelength through the vertical aspect of mats (numbers on curves denote depth in millimetres below the mat surface) at each temperature site. Troughs in the transmitted light spectra indicate absorption maxima of characteristic photopigments in the mats: Chl a, chlorophyll a; PBP, phycobiliproteins; Bchl c, bacteriochlorophyll c; Bchl a, bacteriochlorophyll a.