PROBIOTICS FOR EASTERN OYSTER HATCHERIES: COMMERCIAL FORMULATIONS AND EFFECT ON MICROBIAL COMMUNITIES

In the United States of America, oyster production is an important component of the seafood industry in many coastal communities. Oysters provide ecological, economic, and cultural services. Several hatcheries providing eastern oyster, C. virginica, seed to oyster farms face significant losses owing to Vibrio spp. infections causing massive larval mortalities. Probiotics have been proposed as a potential preventive measure to limit the impact of bacterial diseases in shellfish hatcheries. The probiotic bacterium Phaeobacter inhibens S4 has been shown to protect C. virginica larvae from Vibrio coralliilyticus RE22 (RE22) infection. A liquid formulation of this probiotic has been developed for ease of use in commercial hatcheries. The overall goal of this dissertation is to evaluate the efficacy and safety of the formulated probiotic S4 as an alternative management tool in disease prevention in oyster hatcheries. Chapter 1 provides an overview of the importance of oysters in aquaculture, the use of probiotics for disease prevention in bivalve hatcheries, and the potential role that the relationships between the environment, microbial communities, and probiotics could play on the larval oyster host. Chapter 2 describes the impact of treatment with a formulation of probiont Phaeobacter inhibins S4 on the growth and survival of oysters in several hatchery trials. Daily application of an S4 formulation mixed with algal feed to culture tanks in the hatchery consistently increased the survival of oyster larvae to experimental challenge with the bacterial pathogen V. coralliilyticus RE22, but had no detectable impact on larval growth and survival in the hatchery in the absence of a bacterial challenge. Treatment with S4 had no significant effect on the levels of total culturable vibrios in the larvae. This result suggests that the novel S4 formulation is safe, easy to use, and an effective tool in preventing larval losses to vibriosis. Chapter 3 characterized the effect of treatment with the S4 formulation on the bacterial community of oyster larvae in the hatchery through several trials spanning different hatcheries, years and seasons using 16S rDNA sequencing. Proteobacteria was consistently the most abundant phylum in oyster larval samples. Larval bacterial communities significantly differed mainly by hatchery and trial, and, to a lesser extent, by probiotic treatment. The addition of the S4 formulation caused subtle but significant changes in the structure of oyster larval bacterial communities but did not affect bacterial diversity. Probiotic treatment had a targeted impact on the relative abundance of a few selected taxa in oyster larvae, amplifying Alteromonas and decreasing Pseudomonas. This shows that the effect of probiont S4 is subtle and targeted to a few selected taxa (i.e. does not cause dysbiosis). Chapter 4 analyzed data collected from larval performance (survival and growth), microbial community, and environmental parameters during the trials reported in previous chapters to determine the variables that may impact larval performance in the hatchery. As previously described, variations in environmental parameters were associated with changes in larval survival, with the best larval performance being observed at temperatures of 25 - 26 C, salinities of 14 – 17 psu, and pH of 8.2-8.3. A principal component analysis (PCA) showed correlations between bacterial community composition, environmental variables, and larval growth and survival. Several taxa whose relative abundance correlated with changes in larval performance at the hatchery were identified. Bacterial taxa showing correlations with temperature and salinity also overlapped with larval performance, suggesting a complex interplay between temperature, salinity, microbial community composition, and larval performance in the hatchery. Results from this dissertation provide essential foundational knowledge for better understanding of the use of the probiotic S4 formulation in the hatchery and its interaction with the larval host