Graduate Thesis Or Dissertation


Bacteriophages Significantly Reduce Mortality of Larval Pacific Oysters Crassostrea gigas from Infection by the Pathogenic Bacterium Vibrio coralliilyticus RE98 Public Deposited

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  • Vibrio coralliilyticus (Vcor) is a bacterial pathogen that is well adapted to shellfish hatcheries and is very pathogenic to the larvae of the Pacific oyster Crassostrea gigas. Vcor has been associated with several large scale larval mortality events in the Pacific Northwest that interrupt the supply of seed oysters available to farmers. Vcor can exist in hatcheries at ~102 CFU ml-1 without causing any mortality, but when a shift in the microbial community or environmental conditions favor Vcor growth, it can rapidly increase to 103 or 104 CFU ml-1 and cause widespread larval mortality before hatchery managers can respond. There is a clear need to be able to control Vcor populations to prevent them from ever reaching lethal concentrations. This thesis assesses the ability of bacteriophages (phages) to reduce larval oyster mortality by killing Vcor. Phages are viruses that selectively infect bacteria. There are two scenarios that can lead to Vcor reaching a lethal concentration in a hatchery. The scenario that is likely the most common is for a low concentration of Vcor to increase in the larval culture water until it reaches lethal levels. The second scenario is for the Vcor to increase in concentration in the bay or intake water and reach a lethal concentration before coming into contact with the larvae. The first scenario was simulated by adding 600 and 6 x 103 colony forming units (CFU) ml-1 of Vcor to the larval culture water and allowing it the opportunity to increase in concentration by not performing water changes for six days before assessing larval mortality. The second scenario was simulated by adding 6 x 104 CFU ml-1 Vcor to the larval culture water and assessing mortality after 48 hours. The addition of 2 x 103 plaque forming units (PFU) ml-1 of a cocktail of two purified phages prevented 600 CFU ml-1 of Vcor from causing any larval mortality (38.3 ± 7.3% mortality in the absence of phage) and reduced the mortality caused by 6 x 103 CFU ml-1 Vcor from 81.4 ± 3.5% in the absence of phage to 46.3 ± 4.1%. Vcor added at 6 x 104 CFU ml-1 caused > 99% larval mortality within 48 hours in the absence of phage, and the lowest concentration of a cocktail of three purified phages that eliminated mortality was 2 x 107 PFU ml-1. Purified phages had no negative effects on larval growth or survival and eliminated any detectable negative effects from Vcor exposure. Larvae exposed to phage early in life had a higher metamorphosis rate than the larvae only controls even if they had been exposed to a lethal concentration of Vcor. This study demonstrates that phages can be very effective at reducing larval mortality and any negative effects on growth or metamorphosis from exposure to Vcor. Phages have the potential to benefit the shellfish industry by reducing the frequency and severity of larval mortality events in hatcheries.
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  • Funding provided by USDA Small Business Innovative Research Grant
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