Novel trends of genome evolution in highly complex tropical sponge microbiomes

Abstract

Background: Tropical members of the sponge genus Ircinia possess highly complex microbiomes that perform a broad spectrum of chemical processes that influence host fitness. Despite the pervasive role of microbiomes in Ircinia biology, it is still unknown how they remain in stable association across tropical species. To address this question, we performed a comparative analysis of the microbiomes of 11 Ircinia species using whole-metagenomic shotgun sequencing data to investigate three aspects of bacterial symbiont genomes-the redundancy in metabolic pathways across taxa, the evolution of genes involved in pathogenesis, and the nature of selection acting on genes relevant to secondary metabolism.

Results: A total of 424 new, high-quality bacterial metagenome-assembled genomes (MAGs) were produced for 10 Caribbean Ircinia species, which were evaluated alongside 113 publicly available MAGs sourced from the Pacific species Ircinia ramosa. Evidence of redundancy was discovered in that the core genes of several primary metabolic pathways could be found in the genomes of multiple bacterial taxa. Across hosts, the metagenomes were depleted in genes relevant to pathogenicity and enriched in eukaryotic-like proteins (ELPs) that likely mimic the hosts' molecular patterning. Finally, clusters of steroid biosynthesis genes (CSGs), which appear to be under purifying selection and undergo horizontal gene transfer, were found to be a defining feature of Ircinia metagenomes.

Conclusions: These results illustrate patterns of genome evolution within highly complex microbiomes that illuminate how associations with hosts are maintained. The metabolic redundancy within the microbiomes could help buffer the hosts from changes in the ambient chemical and physical regimes and from fluctuations in the population sizes of the individual microbial strains that make up the microbiome. Additionally, the enrichment of ELPs and depletion of LPS and cellular motility genes provide a model for how alternative strategies to virulence can evolve in microbiomes undergoing mixed-mode transmission that do not ultimately result in higher levels of damage (i.e., pathogenicity) to the host. Our last set of results provides evidence that sterol biosynthesis in Ircinia-associated bacteria is widespread and that these molecules are important for the survival of bacteria in highly complex Ircinia microbiomes. Video Abstract.

Keywords: Ircinia; Marine; Metagenomics; Microbiome; Sponge; Sterols; Tropical.

Fig. 1

Whole-genome phylogenetic tree of MAGs sourced from bacterial symbionts of Ircinia spp. and North Atlantic and South Pacific Tara Oceans MAGs, produced using the MAG dataset that was dereplicated within each source using GTDB-Tk. The inner ring indicates the source of the MAGs, the middle ring denotes the presence or absence of CSGs, and the outer ring denotes the GTDB-Tk phylum-level taxonomic annotations. Tree plotting and annotation were performed with the R package ggtree v2.0.4 [51]

Fig. 2

Plots depicting the genes and domains that are enriched or depleted in tropical Ircinia spp. Each circle corresponds to a gene that was determined as being differentially abundant in either MAGs sourced from the microbiomes of Ircinia or Tara Oceans pelagic seawater samples in terms of normalized average gene copy number. The size of each circle is scaled to the log₁₀-transformed ratio of the gene calculated as (avg. copy number in Ircinia MAGs)/(avg. copy number in Tara Oceans MAGs) for genes under the y = x diagonal and (avg. copy number in Tara Oceans MAGs)/(avg. copy number in Ircinia MAGs) for genes above the y = x diagonal. The ELP graph was constructed from the Interproscan annotations; all others were constructed using the KO annotations produced using EnrichM

Fig. 3

Representative CSGs are depicted on the alphaproteobacterial clade of the GTDB-Tk-inferred phylogeny. CSGs are depicted with other genes found in the eukaryotic steroid biosynthesis pathway (PATH:ko00100) that were commonly found flanking the three core genes (TM7SF2/ERG24, CYP51, and LSS/ERG7) and DFR (dihydroflavonol-4-reductase). “Others” refers to the genes not found in pathway ko00100. If no operon is shown for a given MAG, then a CSG was not detected. Arrows denote the direction of the coding sequence. Genes that overlap with the “steroid biosynthesis” panels of Fig. 2 are colored to match. Tree plotting and annotation were performed with the R packages ggtree [51]

Fig. 4

Histogram of omega values for genes with tree-wide average bootstrap scores > 70%. Gray bars correspond to non-steroid biosynthesis genes. The dark gray line marks the density distribution of omega values calculated for non-steroid genes

Fig. 5

Plots depicting the proportions of nucleotide variability in CSGs compared to MAG-wide values for bacterial classes that had at least three MAGs with CSGs. The lines link the values within each MAG. FDR-corrected p-values are reported (*p < 0.05, **p < 0.01, and ***p < 0.001) for pairwise permutational t-tests each run for 10,000 iterations comparing the proportion of variable sites within each gene to the genome-wide proportion of variable sites that fall within coding regions. N.s., not significant. The CSG genes are positioned in the order in which they appear in the CSGs