Diverse Bacteria Utilize Alginate Within the Microbiome of the Giant Kelp Macrocystis pyrifera

Abstract

Bacteria are integral to marine carbon cycling. They transfer organic carbon to higher trophic levels and remineralise it into inorganic forms. Kelp forests are among the most productive ecosystems within the global oceans, yet the diversity and metabolic capacity of bacteria that transform kelp carbon is poorly understood. Here, we use 16S amplicon and metagenomic shotgun sequencing to survey bacterial communities associated with the surfaces of the giant kelp Macrocystis pyrifera and assess the capacity of these bacteria for carbohydrate metabolism. We find that Macrocystis-associated communities are distinct from the water column, and that they become more diverse and shift in composition with blade depth, which is a proxy for tissue age. These patterns are also observed in metagenomic functional profiles, though the broader functional groups-carbohydrate active enzyme families-are largely consistent across samples and depths. Additionally, we assayed more than 250 isolates cultured from Macrocystis blades and the surrounding water column for the ability to utilize alginate, the primary polysaccharide in Macrocystis tissue. The majority of cultured bacteria (66%) demonstrated this capacity; we find that alginate utilization is patchily distributed across diverse genera in the Bacteroidetes and Proteobacteria, yet can also vary between isolates with identical 16S rRNA sequences. The genes encoding enzymes involved in alginate metabolism were detected in metagenomic data across taxonomically diverse bacterial communities, further indicating this capacity is likely widespread amongst bacteria in kelp forests. Overall, the M. pyrifera epibiota shifts across a depth gradient, demonstrating a connection between bacterial assemblage and host tissue state.

Keywords: Macrocystis; alginate; epibiota; kelp; metagenomics; microbiome.

FIGURE 1

Bacterial diversity associated with Macrocystis blades and the nearby water column. (A) Heatmap of the relative abundance of operational taxonomic units (OTUs) detected on Macrocystis blades from different depths and in the water column from cultured and uncultured samples. OTUs are ordered by taxonomy and the major bacterial clades detected are indicated on the y-axis. (B–E) Uncultured samples only. (B) Taxa summaries of uncultured communities on Macrocystis and in the water column at the Phylum level (^∗Class for Proteobacteria). All samples within a sample type merged. (C) Genus level plot of (B). (D) Bacterial species richness (Chao1 index) is higher on the Macrocystis surface compared to the water column (N = 11), and greater on blades at bottom depth (N = 14) compared to middle (N = 15) and top (N = 15) depths. Statistical tests summarized in text; ^∗p < 0.01. (E) Principle coordinate plot of Weighted UniFrac distance for kelp and water bacterial communities.

FIGURE 2

Bacterial taxonomic and functional profiles within a single kelp bed. Bacterial communities from kelp blades at middle (n = 4) and bottom (n = 5) depths, and in the surrounding water column (n = 5), in one kelp bed (Site 3). (A,B) Taxonomic profiles from 16S rRNA gene amplicon sequencing are variable between water and Macrocystis, and across depth. See Supplementary Figure S6 for corresponding taxonomic profiles derived from metagenomic data. (A) Phylum (^∗Class for Proteobacteria) level. (B) Genus level (most abundant genera shown in legend). (C,D) Relative abundance of Carbohydrate Active Enzyme (CAZy) annotations from these communities is largely consistent across sample types. (C) CAZy annotations largely fall within Carbohydrate Binding Modules (CBM), Glycosyl Transferases (GT), Glycoside Hydrolases (GH), and Polysaccharide Lyase classes (PL). Enzyme families within these broader groups are represented by stacked bars of the same color. (D) Relative abundance of CAZy families within the Polysaccharide Lyase class. Families PL6, 7, 15, and 17 are the most abundant and almost exclusively contain alginate and oligoalginate lyases. (E) Principle coordinate plot of CAZy annotations constructed from Bray–Curtis dissimilarity matrix shows differences between Macrocystis bottom and middle blades, and the water column, similar to taxonomic composition (statistical tests summarized in Table 2). (F) Functional richness (number of CAZy families) is correlated with taxonomic richness of OTUs in Macrocystis communities (r_s = 0.87).

FIGURE 3

Distribution of alginate utilizing taxa within the Gammaproteobacteria. Phylogenetic tree of Gammaproteobacteria genera detected in the M. pyrifera epibiota and surrounding water column in this study, constructed from 16S rRNA gene sequences by placing cultured bacterial isolates into a tree of reference sequences with RAxML EPA. See Supplementary Figure S3 for labeled tree with isolate identities, reference taxa, and accession numbers. Green: cultured bacterial isolates in this study; sequences from isolated bacteria with 99.8–100% 16S similarity were collapsed into operational taxonomic units (OTUs). Blue: cultured isolates from this study whose growth was enriched by at least 50% in the presence of alginate, or that grew with alginate as the sole carbon source (marked with ^∗). Red: cultured isolates in this study that degrade alginate. Purple: cultured isolates whose growth was enhanced and that degrade alginate. Gray: cultured isolates demonstrating neither enhanced growth nor degradation. Detailed information on cultured isolates in Supplementary Table S1.