Among the greatest risks of failure threatening shellfish hatcheries is the danger posed by outbreaks of pathogens, especially Vibrio bacteria and Oyster Herpesviruses, either of which can cause devastating losses during culture of larvae and spat. The most obvious source of pathogens invading seaside hatcheries is seawater obtained from nearby water sources. The conventional counter-measure against pathogens is aggressive treatment of the water by fine-filtration (typically 1 micron), exposure to powerful UV illumination, and sometimes dosing with antibiotics. These treatment processes require significant investments in infrastructure and operating costs, but their greatest cost is when they fail to prevent pathogen outbreaks.
Fortunately, there is a low-tech approach to water treatment that is proving to be more effective.
Pathogenic Vibrios
In recent years there have been anecdotal accounts of reduced Vibrio pathogen problems in both shellfish and finfish culture when a diverse bacterial population is allowed to develop in the hatchery. An illustrative example comes from the Auburn University Shellfish Laboratory (Alabama, USA), which experienced frequent oyster larviculture failures attributable to Vibrio outbreaks following installation of a new seawater system with powerful UV treatment. These failures continued for two years until the hatchery adopted the practice of first storing the water for 24 hours before use and stopping UV treatment, whereupon the larviculture failures ceased (see https://www.youtube.com/watch?v=nqerVTlch_w ). This contradicts the conventional wisdom that the best defense against Vibrio pathogens is stringent sanitary precautions to reduce the numbers of bacteria present, with the hope of preventing invasion by pathogens. But it can be difficult or impossible to prevent invasion of a hatchery by all pathogenic Vibrios, because all seawater entering the hatchery must be sanitized, broodstock and algae cultures must be kept free of pathogens, airborne transport of pathogens must be prevented (very difficult in a shoreline location), and all these measures must work without breaches, all the time.
Now there are published experimental studies that demonstrate how water storage prevents Vibrio pathogens from causing problems in hatcheries. The protective mechanism is the activity of Vibrio Predatory Bacteria (VPB), which appear to be ubiquitous in raw seawater. When raw seawater is stored for several days, during the first 24 hours pathogenic Vibrios bloom to very high numbers, but by 48 hours VPB bloom and progressively reduce the numbers of pathogenic Vibrios, until at 72 hours they are virtually undetectable. This succession of Vibrios and then VPB was found to occur consistently in seawater samples from multiple sites in Delaware, Alabama, Oregon, and Hawaii (see the very comprehensive study of Richards et al. 2012, Appl. Environ. Microbiol. 78:7455-7466, free download from http://aem.asm.org/content/78/20/7455.full; see also more recent studies by this laboratory).
Only simple and low-cost measures are required for hatcheries to take advantage of this phenomenon.
Oyster Herpesvirus
Storage of seawater for 48 h before use has been shown to also provide reliable protection against infection of Pacific Oyster spat by Oyster Herpesvirus (Whittington et al. 2015, Aquaculture 437:10-20). Studies have indicated that 48 h may not be sufficient time to eliminate the infectivity of viruses in the ocean, so virus inactivation in storage is presumably aided by enhanced bacterial degradation, adsorption to inactivating suspended matter, or sedimentation on larger particles. Furthermore, Whittington et al. found that 5 μm filtration of seawater also prevented Oyster Herpesvirus infection of spat, despite the fact that such relatively coarse filtration cannot block passage of the far smaller virus particles. This and other studies indicate that the primary route of infection by these viruses is via adherence to larger particles in the water that are filtered and ingested by oysters, rather than by contact of free virus particles with oyster surface tissues that are exposed to seawater.
Vibrios + Oyster Herpesvirus = Double Trouble
A recent study found that simultaneous infection of spat with Oyster Herpesvirus and a pathogenic Vibrio was much more deadly than with only one pathogen (Patrick et al. 2016, J. of Invertebrate Pathology 139:92-101, free download from https://archimer.ifremer.fr/doc/00347/45835/45918.pdf). Oyster Herpesvirus pathology may therefore often be a “polymicrobial infection” of the virus and pathogenic Vibrios (Petton et al. 2019, Diseases of Aquatic Organisms 135: 97–106). Considering these findings, it is most fortunate that the simple practice of storing seawater for 2-3 days is an effective countermeasure against both pathogenic Vibrios and Oyster Herpesviruses.
Pathogen genetic material can now be detected by molecular testing even when there are no viable bacteria or infective virus particles present, or they are present only at levels too low to cause infections. It should be understood that although these water treatment processes cannot ensure complete elimination of any pathogen, they have nevertheless been shown to be capable of reducing pathogen effects to insignificance in hatchery settings.
After Storage of Seawater
After storage has “neutralized” Vibrios and infective Herpesviruses, the seawater can be filtered (commonly to 5 or 1 micron) to remove larger organisms (zooplankton) and used for culture of larvae, spat, or broodstock. Broodstock oysters should be pathogen-free and maintained in stored seawater. Seawater storage for 2-3 days and any subsequent filtration will remove most of the phytoplankton, so algae must be added to feed broodstock, larval and spat cultures. All food algae should be free of pathogens, and so must be cultured using only stored or sanitized seawater, or commercial algae concentrates can be used.
Reed Mariculture Inc. produces pathogen-free algae concentrate feeds designed for bivalves. Our Shellfish Diet® 1800 provides a nutritionally balanced combination of five specially-selected strains of Pavlova, Tisochrysis (“T-Iso”), Tetraselmis, and Thalassiosira, providing a range of cell sizes from 4 – 12 microns. These marine microalgae strains have demonstrated success as feeds for a variety of bivalves including oysters, clams, mussels, and scallops. This mixed diet provides excellent nutrition for all life stages, from first-feeding larvae all the way through broodstock, increasing both growth rates and survival. Reed Mariculture also offers the algae species used in Shellfish Diet as single-species Instant Algae® products, so hatchery operators have the option to create their own custom mix of species for particular applications. Our new LPB Frozen Shellfish Diet® offers a more economical mix of Tetraselmis sp., Thalassiosira weissflogii and Thalassiosira pseudonana, with a 6 – 15 micron cell size range for late larvae, spat and broodstock.
By Eric Henry PhD, Research Scientist, Reed Mariculture Inc.