Diversity and Function of the Eastern Oyster (Crassostrea virginica) Microbiome

Abstract

Marine invertebrate microbiomes play important roles in various host and ecological processes. However, a mechanistic understanding of host-microbe interactions is so far only available for a handful of model organisms. Here, an integrated taxonomic and functional analysis of the microbiome of the eastern oyster, Crassostrea virginica, was performed using 16S rRNA gene amplicon profiling, shotgun metagenomics, and genome-scale metabolic reconstruction. A relatively low number of amplicon sequence variants (ASVs) were observed in oyster tissues compared to water samples, while high variability was observed across individual oysters and among different tissue types. Targeted metagenomic sequencing of the gut microbiota led to further characterization of a dominant bacterial taxon, the class Mollicutes, which was captured by the reconstruction of a metagenome-assembled genome (MAG). Genome-scale metabolic reconstruction of the oyster Mollicutes MAG revealed a reduced set of metabolic functions and a high reliance on the uptake of host-derived nutrients. A chitin degradation and an arginine deiminase pathway were unique to the MAG as compared to other closely related Mycoplasma genomes, indicating a distinct mechanism of carbon and energy acquisition by the oyster- associated Mollicutes. A systematic reanalysis of public eastern oyster-derived microbiome data revealed the Mollicutes as a ubiquitous taxon among adult oysters despite their general absence in larvae and biodeposit samples, suggesting potential horizontal transmission via an unknown mechanism.IMPORTANCEDespite well-documented biological significance of invertebrate microbiomes, a detailed taxonomic and functional characterization is frequently missing from many non-model marine invertebrates. By using 16S rRNA gene-based community profiling, shotgun metagenomics, and genome-scale metabolic reconstruction, this study provides an integrated taxonomic and functional analysis of the microbiome of the eastern oyster, Crassostrea virginica. Community profiling revealed a surprisingly low richness, as compared to surrounding seawater, and high variability among different tissue types and individuals. Reconstruction of a Mollicutes MAG enabled the phylogenomic positioning and functional characterization of the oyster-associated Mollicutes. Comparative analysis of the adult oyster gut, biodeposits, and oyster larvae samples indicated the potentially ubiquitous associations of the Mollicutes taxon with adult oysters. To the best of our knowledge, this study represented the first metagenomics derived functional inference of the eastern oyster microbiome. An integrated analytical procedure was developed for the functional characterization of microbiomes in other non-model host species.

Comparative Genomic Analysis of Three Pseudomonas Species Isolated from the Eastern Oyster (Crassostrea virginica) Tissues, Mantle Fluid, and the Overlying Estuarine Water Column

“…Conversely, strain MF28 was found to sustain only three incomplete prophage regions of 9.7 kb, 9.4 kb, and 8.1 kb in size ( Figure 5B). Similarly, this prophage analysis revealed the presence of five prophage regions in strain WC55 with one intact, three incomplete and one was questionable ( Figure 5C); these regions were of 42.7 kb, 42.4 kb, 22.8 kb, 19.9 kb, and 17.7 kb in size. These findings were also confirmed by RAST based annotation in which strain OT69 was found to harbor 29 phage and prophage-related genes; similar to strain WC55, which contained 24 phage-associated gene fragments.…”

“…Recently, it was also demonstrated that the oyster extrapallial fluid (EF), which is the site for shell formation, also harbors diverse and abundant microbial communities; moreover, the oyster EF microbiota were found to be enriched for dissimilatory nitrate reduction, nitrogen fixation, nitrification, and sulfite reductase genes and it was proposed that sulfate and nitrate reduction may have a synergistic effect on calcium carbonate precipitation, thus indirectly facilitating in shell formation [19]. Microbial differences within the different oyster tissues have also been demonstrated, such that the stomach and guts of the eastern oysters were shown to be distinct relative to the other organs such as gills, and even the water and sediments that feeds the oyster reefs [9,11,[20][21][22]. Specifically, substantial differences between stomach and gut microbiomes of oysters from Lake Caillou in Louisiana were shown such that bacteria belonging to Chloroflexi, Mollicutes, Planctomycetes, and Spartobacteria comprised the major core of the oyster stomach microbiome, whereas Chloroflexi, Firmicutes, α-proteobacteria, and Verrucomicrobia were more abundant in the gut [9].…”

Cross-talk and mutual shaping between the immune system and the microbiota during an oyster's life

“…temperature, pH, salinity, pCO 2 ) [34][35][36][37][38]. In addition to important fluctuations in response to environmental changes, physiological status and developmental stages, the oyster microbiota shows important inter-individual variability, both at the level of the whole animal and at the level of individual tissues throughout all life stages [8, [11][12][13]39]. This heterogeneity has made it difficult to identify the core bacterial microbiota in oysters, if one exists.…”

“…A fine-tuned dialogue between oyster immunity and its microbiota was evidenced from early life to adult stages [23][24][25]. Recently a first insight into microbial community gene functions has been made accessible, either by shotgun metagenomics [11] or metatranscriptomics [26]. The present article summarizes the knowledge accumulated on oystermicrobiota interactions over the last few years and reviews the dysregulations leading to dysbiosis and oyster death.…”

“…Benthic oysters not only host microbial communities on their body surfaces [8] but also in their circulating body fluids, i.e. the haemolymph (equivalent of the blood) [9], in the pallial/extrapallial cavity fluid [10,11] as well as in all the tissues and developmental stages studied [12,13]. As they are filter feeders able to concentrate microorganisms present in their environment, it has been argued that bivalves host transient microbial communities that vary with environmental conditions.…”