The core microbiome of cultured Pacific oyster spat is affected by age but not mortality

Abstract

The Pacific oyster is the most widely cultured shellfish worldwide, but production has been affected by mortality events, including in hatcheries that supply the seed for growers. Several pathogens cause disease in oysters, but in many cases, mortality events cannot be attributed to a single agent and appear to be multifactorial, involving environmental variables and microbial interactions. As an organism's microbiome can provide resilience against pathogens and environmental stressors, we investigated the microbiomes in cohorts of freshly settled oyster spat, some of which experienced notable mortality. Deep sequencing of 16S rRNA gene fragments did not show a significant difference among the microbiomes of cohorts experiencing different mortality levels, but revealed a characteristic core microbiome comprising 74 taxa. Irrespective of mortality, the relative abundance of taxa in the core microbiomes changed significantly as the spat aged, yet remained distinct from the microbial community in the surrounding water. The core microbiome was dominated by bacteria in the families Rhodobacteraceae, Nitrosomonadaceae, Flavobacteriaceae, Pirellulaeceae, and Saprospiraceae. Within these families, 14 taxa designated as the "Hard-Core Microbiome" were indicative of changes in the core microbiome as the spat aged. The variability in diversity and richness of the core taxa decreased with age, implying niche occupation. As well, there was exchange of microbes with surrounding water during development of the core microbiome. The shift in the core microbiome demonstrates the dynamic nature of the microbiome as oyster spat age.IMPORTANCEThe Pacific oyster (Magallana gigas, also known as Crassostrea gigas) is the most widely cultivated shellfish and is important to the economy of many coastal communities. However, high mortality of spat during the first few days following metamorphosis can affect the seed supply to oyster growers. Here, we show that the microbiome composition of recently settled oyster spat experiencing low or high mortality was not significantly different. Instead, development of the core microbiome was associated with spat aging and was partially driven by dispersal through the water. These findings imply the importance of early-stage rearing conditions for spat microbiome development in aquaculture facilities. Furthermore, shellfish growers could gain information about the developmental state of the oyster spat microbiome by assessing key taxa. Additionally, the study provides a baseline microbiome for future hypothesis testing and potential probiotic applications on developing spat.

Keywords: 16S rRNA; Crassostrea gigas; Magallana gigas; Pacific oyster; amplicon sequencing; aquaculture; core microbiome; metamorphosis; spat.

Fig 1

Overview of the relative abundances of the 10 most abundant families of bacteria in the combined sequencing data based on 16S rRNA gene sequences. Water samples are labeled “W” followed by tank number and sample date (ddmm). Spat samples are labeled “OY” followed by cohort number and the number of days post settlement.

Fig 2

Constrained dbRDA of the community composition of water and oyster samples against the sample source; results of the PERMANOVA are shown. Blue triangles are water samples; red circles are oyster samples.

Fig 3

Principal Coordinate Analysis (PCoA, unconstrained dbRDA) of the community composition of core OTUs. Shapes indicate whether cohorts experienced low <50% /d (circles) or high >50% /d (triangles) mortality. Color indicates the age in days post settlement. Results of a subsequent PERMANOVA of community distance against sample age are shown.

Fig 4

Core microbiome diversity indices for spat age classes; indices were calculated based on the community compositions of samples per age class. Top graph depicts OTU richness; bottom graph shows alpha diversity, age classes (days post settlement), A (0 to 4), B (5 to 11), C (12 to 17), D (18 to 23), E (24 to 39), and F (≥30), which had respective sample sizes (n) of 6, 9, 8, 6, 5, and 5. Box plots describe the minimum and maximum values, lower and upper quartiles and median, outliers.

Fig 5

Heat map of the frequency of core OTUs across samples per age class. Color tone describes frequency per age class; white is absence and dark blue is presence. OTUs are defined at the genus level, but family annotation is labeled.

Fig 6

Mean abundance in water communities (source abundance) and frequency across oyster spat samples (target frequency) for OTUs included in the neutral community model analysis. Colors indicate the dispersion mode: red for OTUs overrepresented in oyster samples; blue for OTUs overrepresented in water samples; gray for OTUs where the observed frequency matched the predicted frequency in the neutral community model.