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Marrowstone Marine Field Station

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Specific Pathogen Free (SPF)

Why the need for Specific Pathogen-Free (SPF) Marine Fishes?

In the past, fish health research has been limited primarily to understanding diseases in aquaculture and enhancement facilities, where abnormal conditions could be relatively easily observed, treated, and prevented. Disease impacts to populations of wild, marine fishes are difficult to observe because sick fish are often consumed by predators before they die directly from disease. Alternatively, fish that die directly from disease are often difficult to locate if carcasses sink to the bottom or if mortalities occur offshore. The problem is compounded by the unavailability of marine fish test animals with a known disease history that can be used as experimental subjects. For example, in the past, if we wanted to test the virulence of a suspected pathogen to Pacific herring, wild herring were captured and transported to the laboratory where experimental challenges would occur. Unfortunately, if the fish did not respond to the pathogen, we were left with the possibility that the captured test animals were previously exposed to the pathogen, survived the epizootic, and were immune as a result of previous exposures. This experimental uncertainty was overcome by rearing colonies of specific pathogen-free test animals in our laboratory. In addition to helping us understand the basic pathogenesis and kinetics of diseases, these fish are enabling us to develop predictive tools that forecast and prevent disease epizootics in wild populations. Our research approach involves a combination of field-based disease observations, followed by hypothesis formation and testing using SPF marine fish. This approach is similar to that utilized by the Centers for Disease Control and Prevention (CDC), with the notable exception that we are able to perform experiments with the specific pathogens and hosts of concern, rather than utilizing mammalian surrogates for human hosts, as is done with human diseases.

SPF Pacific Herring

During the late winter and early spring, massive schools of Pacific herring migrate to nearshore regions along the Pacific coast to spawn. Females deposit their adhesive eggs on submerged macrophytes and males fertilize the eggs by broadcasting milt in the vicinity of the eggs. The spawning events are often so large that the water turns white from release of the spawning products. Time from fertilization to hatch is approximately 14 days (longer in cooler water), when yolk-sac larvae emerge from the eggs. Herring then undergo a 3 month larval period, after which metamorphisis to juveniles is complete. In the Puget Sound region, herring begin to mature to adults after 2 years when they return to spawn with other adult age cohorts. Unlike Pacific salmon, which spawn once and die, herring are iteroparous and adults typically spawn for multiple years before they die.

Because of the importance of herring as a forage fish in the eastern North Pacific and concerns that infectious and parasitic diseases currently impact their population sizes and zoographics in Puget Sound and Prince William Sound, we have developed techniques to rear Pacific herring under specific pathogen-free conditions. Naturally deposited herring eggs are collected by colleagues at Washington Department of Fish and Wildlife and transported to flow-through tanks, supplied with processed seawater that is double sand filtered, particle filtered to 100um, and exposed to ultraviolet irradiation twice at the Marrowstone Marine Field Station. Food for the larvae, including live rotifers and Artemia, is raised in the same processed seawater. Once past metamorphosis, the juveniles are weaned to a specially-formulated dry pellet diet that is produced for us by colleagues at the USFWS - Abernathy Fish Technology Center. To avoid exposure of the SPF herring to killed antigens that likely occur in fish meal, the experimental pellet diet contains a combination of vegetable-based protein and euphausid meal.

Development of colonies of SPF Pacific herring has provided new tools for disease ecology research and enables us to empirically address fundamental principles that were previously based on speculation. As a result, researchers from, around the world routinely visit the Marrowstone Marine Field Station to utilize these SPF colonies in experimental trials.

photo of puget sound

Small herring spawning event in Puget Sound, evidenced by the presence of ‘white water’, caused by mass release of spawning products into the water.

photo of herring eggs

Naturally-deposited herring eggs attached to submerged macrophytes. Coiled embryos are evident inside the eggs.

photo of herring larvae

Newly-hatched day 0 herring larvae. Yolk sac is apparent behind the eyes.

photo of herring larvae

Herring larvae 15 days post-hatch. Food in the gut appears orange.

photo of herring larvae

Herring larvae 53 d post-hatch. Black pigmentation is evident.

photo of herring juvenile

Herring juvenile 103 days post-hatch. Larval metamorphosis to juveniles is complete.

photo of herring

Currently four age classes (0, 1, 2, and 4 year olds) of SPF herring are maintained at the Marrowstone Marine Field Station.

SPF Rockfishes

Commercially and recreationally important rockfishes have undergone dramatic population declines throughout the Pacific Northwest. “Stock assessments prepared from 1999-2001 indicate that the biomass of at least seven of the major commercial rockfish species (bocaccio, canary rockfish, cowcod, darkblotch rockfish, Pacific ocean perch, widow and yelloweye rockfish) are at or below 25% of that estimated in the 1970’s… Populations of copper, brown, and quillback rockfish have declined to very low levels in Puget Sound.” The Department of Commerce in January 2000 declared the West Coast rockfish fishery a disaster and the Pacific Fishery Management Council and the State of California began to restrict fishing for these species in 2002 and 2003.

A leading hypothesis to account for rockfish population declines involves excessive harvest from over-fishing because juveniles are often encountered as bycatch in commercial and recreational fisheries. Unintentional catch results in high post-release mortality due to pressure differentials encountered during hook-and-line and trawl retrieval to the surface. Recovery of an over-exploited rockfish population may take decades because average age of maturity is 8-10 years. The protection of juveniles from fishing-induced mortalities is increasingly seen as a strong justification for protection through Marine Protected Areas, national marine sanctuaries, and national parks. Recently cooperative research funded through the U.S. Geological Survey, Minerals Management Service, and California Artificial Reef Ecosystem Program has indicated the importance of habitat surrounding offshore oil and gas platforms in the Santa Maria Basin as refugia to some rockfish species.

photo of rockfishes

School of specific pathogen-free copper rockfish that were reared at the Marrowstone Marine Station

A supplemental hypothesis accounting for rockfish declines involves a parasitic disease caused by the protozoan parasite Ichthyophonus hoferi. In a recent survey of rockfishes from the Oregon, Washington, and British Columbia coasts, the parasite was detected in 51% of yellowtail rockfish and 48% of Pacific Ocean perch. The authors stated that “The role of Ichthyophonus and the putative Mycobacterium infections in the decline of S. aluticus and the potential future impact on S. flavidis is unknown at this point. However, the possibility that the former has some deleterious impact on these stocks should be considered because of the extremely high prevalence of infection, and many of the fish exhibited severe infections and associated tissue damage.” Although Ichthyophonus has been associated with massive epizootics in wild herring where it is highly pathogenic, the impacts of Ichthyophonus infections to the health and survival of rockfishes remains uninvestigated, primarily because of the unavailability of a rockfish laboratory model that is suitable for disease challenge studies. Basic pathogenicity studies being performed to begin to determine the population-level effects of ichthyophoniasis to Pacific coast rockfish populations.

Currently, SPF copper and brown rockfish are reared and maintained at the Marrowstone Marine Field Station. Pathogenicity studies with Ichthyophonus are underway.

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