Microbiome succession during ammonification in eelgrass bed sediments

Abstract

Background: Eelgrass (Zostera marina) is a marine angiosperm and foundation species that plays an important ecological role in primary production, food web support, and elemental cycling in coastal ecosystems. As with other plants, the microbial communities living in, on, and near eelgrass are thought to be intimately connected to the ecology and biology of eelgrass. Here we characterized the microbial communities in eelgrass sediments throughout an experiment to quantify the rate of ammonification, the first step in early remineralization of organic matter, also known as diagenesis, from plots at a field site in Bodega Bay, CA.

Methods: Sediment was collected from 72 plots from a 15 month long field experiment in which eelgrass genotypic richness and relatedness were manipulated. In the laboratory, we placed sediment samples (n = 4 per plot) under a N₂ atmosphere, incubated them at in situ temperatures (15 °C) and sampled them initially and after 4, 7, 13, and 19 days to determine the ammonification rate. Comparative microbiome analysis using high throughput sequencing of 16S rRNA genes was performed on sediment samples taken initially and at seven, 13 and 19 days to characterize changes in the relative abundances of microbial taxa throughout ammonification.

Results: Within-sample diversity of the sediment microbial communities across all plots decreased after the initial timepoint using both richness based (observed number of OTUs, Chao1) and richness and evenness based diversity metrics (Shannon, Inverse Simpson). Additionally, microbial community composition changed across the different timepoints. Many of the observed changes in relative abundance of taxonomic groups between timepoints appeared driven by sulfur cycling with observed decreases in predicted sulfur reducers (Desulfobacterales) and corresponding increases in predicted sulfide oxidizers (Thiotrichales). None of these changes in composition or richness were associated with variation in ammonification rates.

Discussion: Our results showed that the microbiome of sediment from different plots followed similar successional patterns, which we infer to be due to changes related to sulfur metabolism. These large changes likely overwhelmed any potential changes in sediment microbiome related to ammonification rate. We found no relationship between eelgrass presence or genetic composition and the microbiome. This was likely due to our sampling of bulk sediments to measure ammonification rates rather than sampling microbes in sediment directly in contact with the plants and suggests that eelgrass influence on the sediment microbiome may be limited in spatial extent. More in-depth functional studies associated with eelgrass microbiome will be required in order to fully understand the implications of these microbial communities in broader host-plant and ecosystem functions (e.g., elemental cycling and eelgrass-microbe interactions).

Keywords: Ammonification; Decomposition; Eelgrass; Microbiome; Seagrass; Succession; Sulfur cycling; Zostera marina.

Figure 1. Alpha diversity decreases over time.

Four alpha diversity metrics are depicted as boxplots colored by timepoint: (A) observed number of OTUs, (B) Chao1, (C) Shannon and (D) Inverse Simpson indices. Timepoints shown are: 1 (initial samples), 2 (seven days), 3 (13 days), and 4 (19 days).

Figure 2. Microbial community composition changes over time.

Principal Coordinates Analysis (PCoA) of Weighted Unifrac distances of microbial communities are shown here with shapes and colors representative of respective timepoint. Timepoints shown are: 1 (initial samples), 2 (seven days), 3 (13 days), and 4 (19 days).

Figure 3. Initial microbial community composition is not correlated with eelgrass presence/absence.

Principal Coordinates Analysis (PCoA) of Weighted Unifrac distances of microbial communities at the initial timepoint (timepoint 1) are shown here. Points in the ordination are colored by eelgrass status in each plot (one genotype, multiple genotypes, absent).

Figure 4. Taxonomic composition varies over time.

The average relative abundance of taxonomic orders with a mean greater than two percent are shown across timepoints with the standard error of the mean represented by error bars and lines colored by taxonomic order. Panels group taxonomic orders by phylum (A) Bacteroidetes, (B) Chloroflexi, (C) Planctomycetes and (D) Proteobacteria. Timepoints shown are: 1 (initial samples), 2 (seven days), 3 (13 days), and 4 (19 days).