Geology topics
Paul Hershberger
Publications and source records attributed to Paul Hershberger.
Influence of temperature on the efficacy of homologous and heterologous DNA vaccines against viral hemorrhagic septicemia in Pacific Herring
Homologous and heterologous (genogroup Ia) DNA vaccines against viral hemorrhagic septicemia virus (genogroup IVa) conferred partial protection in Pacific Herring Clupea pallasii . Early protection at 2 weeks postvaccination (PV) was low and occurred only at an elevated temperature (12.6°C, 189 degree days), where the relative percent survival following viral exposure was similar for the two vaccines (IVa and Ia) and higher than that of negative controls at the same temperature. Late protection at 10 weeks PV was induced by both vaccines but was higher with the homologous vaccine at both 9.0°C and 12.6°C. Virus neutralization titers were detected among 55% of all vaccinated fish at 10 weeks PV. The results suggest that the immune response profile triggered by DNA vaccination of herring was similar to that reported for Rainbow Trout Oncorhynchus mykiss by Lorenzen and LaPatra in 2005, who found interferon responses in the early days PV and the transition to adaptive response later. However, the protective effect was far less prominent in herring, possibly reflecting different physiologies or adaptations of the two fish species.
Optimization of a Plaque Neutralization Test (PNT) to identify the exposure history of Pacific Herring to viral hemorrhagic septicemia virus (VHSV)
Methods for a plaque neutralization test (PNT) were optimized for the detection and quantification of viral hemorrhagic septicemia virus (VHSV) neutralizing activity in the plasma of Pacific Herring Clupea pallasii. The PNT was complement dependent, as neutralizing activity was attenuated by heat inactivation; further, neutralizing activity was mostly restored by the addition of exogenous complement from specific-pathogen-free Pacific Herring. Optimal methods included the overnight incubation of VHSV aliquots in serial dilutions (starting at 1:16) of whole test plasma containing endogenous complement. The resulting viral titers were then enumerated using a viral plaque assay in 96-well microplates. Serum neutralizing activity was virus-specific as plasma from viral hemorrhagic septicemia (VHS) survivors demonstrated only negligible reactivity to infectious hematopoietic necrosis virus, a closely related rhabdovirus. Among Pacific Herring that survived VHSV exposure, neutralizing activity was detected in the plasma as early as 37 d postexposure and peaked at approximately 64 d postexposure. The onset of neutralizing activity was slightly delayed in fish reared at 7.4°C relative to those in warmer temperatures (9.9°C and 13.1°C); however, neutralizing activity persisted for at least 345 d postexposure in all temperature treatments. It is anticipated that this novel ability to assess VHSV neutralizing activity in Pacific Herring will enable retrospective comparisons between prior VHS infections and year-class recruitment failures. Additionally, the optimized PNT could be employed as a forecasting tool capable of identifying the potential for future VHS epizootics in wild Pacific Herring populations.
Disease surveillance of Atlantic herring: molecular characterization of hepatic coccidiosis and a morphological report of a novel intestinal coccidian
Surveillance for pathogens of Atlantic herring, including viral hemorrhagic septicemia virus (VHSV), Ichthyophonus hoferi , and hepatic and intestinal coccidians, was conducted from 2012 to 2016 in the NW Atlantic Ocean, New Jersey, USA. Neither VHSV nor I. hoferi was detected in any sample. Goussia clupearum was found in the livers of 40 to 78% of adult herring in varying parasite loads; however, associated pathological changes were negligible. Phylogenetic analysis based on small subunit 18S rRNA gene sequences placed G. clupearum most closely with other extraintestinal liver coccidia from the genus Calyptospora , though the G. clupearum isolates had a unique nucleotide insertion between 604 and 729 bp that did not occur in any other coccidian species. G. clupearum oocysts from Atlantic and Pacific herring were morphologically similar, though differences occurred in oocyst dimensions. Comparison of G. clupearum genetic sequences from Atlantic and Pacific herring revealed 4 nucleotide substitutions and 2 gaps in a 1749 bp region, indicating some divergence in the geographically separate populations. Pacific G. clupearum oocysts were not directly infective, suggesting that a heteroxenous life cycle is likely. Intestinal coccidiosis was described for the first time from juvenile and adult Atlantic herring. A novel intestinal coccidian species was detected based on morphological characteristics of exogenously sporulated oocysts. A unique feature in these oocysts was the presence of 3 long (15.1 ± 5.1 µm, mean ±SD) spiny projections on both ends of the oocyst. The novel morphology of this coccidian led us to tentatively name this parasite G. echinata n. sp.
Ichthyophonus parasite phylogeny based on ITS rDNA structure prediction and alignment identifies six clades, with a single dominant marine type
Despite their widespread, global impact in both wild and cultured fishes, little is known of the diversity, transmission patterns, and phylogeography of parasites generally identified as Ichthyophonus. This study constructed a phylogeny based on the structural alignment of internal transcribed spacer (ITS) rDNA sequences to compare Ichthyophonus isolates from fish hosts in the Atlantic and Pacific oceans, and several rivers and aquaculture sites in North America, Europe, and Japan. Structure of the Ichthyophonus ITS1–5.8S–ITS2 transcript exhibited several homologies with other eukaryotes, and 6 distinct clades were identified within Ichthyophonus. A single clade contained a majority (71 of 98) of parasite isolations. This ubiquitous Ichthyophonus type occurred in 13 marine and anadromous hosts and was associated with epizootics in Atlantic herring, Chinook salmon, and American shad. A second clade contained all isolates from aquaculture, despite great geographic separation of the freshwater hosts. Each of the 4 remaining clades contained isolates from single host species. This study is the first to evaluate the genetic relationships among Ichthyophonus species across a significant portion of their host and geographic range. Additionally, parasite infection prevalence is reported in 16 fish species.
Infecting Pacific Herring with Ichthyophonus sp. in the laboratory
The protistan parasite Ichthyophonus sp. occurs in coastal populations of Pacific Herring Clupea pallasii throughout the northeast Pacific region, but the route(s) by which these planktivorous fish become infected is unknown. Several methods for establishing Ichthyophonus infections in laboratory challenges were examined. Infections were most effectively established after intraperitoneal (IP) injections with suspended parasite isolates from culture or after repeated feedings with infected fish tissues. Among groups that were offered the infected tissues, infection prevalence was greater after multiple feedings (65%) than after a single feeding (5%). Additionally, among groups that were exposed to parasite suspensions prepared from culture isolates, infection prevalence was greater after exposure by IP injection (74%) than after exposure via gastric intubation (12%); the flushing of parasite suspensions over the gills did not lead to infections in any of the experimental fish. Although the consumption of infected fish tissues is unlikely to be the primary route of Ichthyophonus sp. transmission in wild populations of Pacific Herring, this route may contribute to abnormally high infection prevalence in areas where juveniles have access to infected offal.
Study 8: Prevalence and load of Nanophyetus salmincola infection in outmigrating steelhead trout from five Puget Sound rivers
Nanophyetus salmincola is a parasitic trematode, or flatworm, that infects salmonid fishes in the Pacific Northwest, including Washington, Oregon, and portions of California. The adult worm lives in the intestine of fish-eating birds and mammals. Eggs shed into the water hatch into miracidia which penetrate the first intermediate host, one of two species of snail Juga plicifera or J. silicula . Asexual reproduction occurs within the snail. Free-swimming cercaria are released from the snail and penetrate the secondary intermediate host, often a salmonid fish, in fresh and brackish water. The cercaria encyst as metacercaria in various organs of the fish, including gills, muscle and heart, but favor the posterior kidney. Penetration and migration by the cercaria through the fish causes damage to nearly every organ system. Once encysted, metacercaria survive the ocean phase of salmonid life cycle. N. salmincola is a likely contributor to mortality of juvenile coho salmon ( Oncorhynchus kisutch ) during the early ocean rearing phase, and it is the most prevalent pathogen of outmigrating steelhead in the estuaries of the Pacific Northwest. A field survey was implemented from March-June 2014 to compare the prevalence and parasite load of N. salmincola infections in outmigrating steelhead from five Puget Sound watersheds and to assess changes in infection levels that occurred during the smolt out-migration through each watershed. N. salmincola infection prevalence and parasite loads were determined by counting metacercaria in posterior kidney samples. Tissue samples were collected and examined by standard histological methods.
Development and evaluation of a blocking enzyme-linked immunosorbent assay and virus neutralization assay to detect antibodies to viral hemorrhagic septicemia virus
Viral hemorrhagic septicemia virus (VHSV) is a target of surveillance by many state and federal agencies in the United States. Currently, the detection of VHSV relies on virus isolation, which is lethal to fish and indicates only the current infection status. A serological method is required to ascertain prior exposure. Here, we report two serologic tests for VHSV that are nonlethal, rapid, and species independent, a virus neutralization (VN) assay and a blocking enzyme-linked immunosorbent assay (ELISA). The results show that the VN assay had a specificity of 100% and sensitivity of 42.9%; the anti-nucleocapsid-blocking ELISA detected nonneutralizing VHSV antibodies at a specificity of 88.2% and a sensitivity of 96.4%. The VN assay and ELISA are valuable tools for assessing exposure to VHSV.
Viability and infectivity of Ichthyophonus sp. in post-mortem Pacific herring, Clupea pallasii
Ichthyophonus -infected Pacific herring, Clupea pallasii , were allowed to decompose in ambient seawater then serially sampled for 29 days to evaluate parasite viability and infectivity for Pacific staghorn sculpin, Leptocottus armatus . Ichthyophonus sp. was viable in decomposing herring tissues for at least 29 days post-mortem and could be transmitted via ingestion to sculpin for up to 5 days. The parasite underwent morphologic changes during the first 48 hr following death of the host that were similar to those previously reported, but as host tissue decomposition progressed, several previously un-described forms of the parasite were observed. The significance of long-term survival and continued morphologic transformation in the post-mortem host is unknown, but it could represent a saprozoic phase of the parasite life cycle that has survival value for Ichthyophonus sp.
Infectious diseases of fishes in the Salish Sea
As in marine regions throughout other areas of the world, fishes in the Salish Sea serve as hosts for many pathogens, including nematodes, trematodes, protozoans, protists, bacteria, viruses, and crustaceans. Here, we review some of the better-documented infectious diseases that likely contribute to significant losses among free-ranging fishes in the Salish Sea and discuss the environmental and ecological factors that may affect the population-level impacts of disease. Demonstration of these diseases and their impacts to critical and endangered resources provides justification to expand pathogen surveillance efforts and to incorporate disease forecasting and mitigation tools into ecosystem restoration efforts.
Evidence for an amoeba-like infectious stage of ichthyophonus sp. and description of a circulating blood stage: a probable mechanism for dispersal within the fish host
Small amoeboid cells, believed to be the infectious stage of Ichthyophonus sp., were observed in the bolus (stomach contents) and tunica propria (stomach wall) of Pacific staghorn sculpins and rainbow trout shortly after they ingested Ichthyophonus sp.–infected tissues. By 24–48 hr post-exposure (PE) the parasite morphed from the classically reported multinucleate thick walled schizonts to 2 distinct cell types, i.e., a larger multinucleate amoeboid cell surrounded by a narrow translucent zone and a smaller spherical cell surrounded by a “halo” and resembling a small schizont. Both cell types also appeared in the tunica propria, indicating that they had recently penetrated the columnar epithelium of the stomach. No Ichthyophonus sp. pseudo-hyphae (“germination tubes”) were observed in the bolus or penetrating the stomach wall. Simultaneously, Ichthyophonus sp. was isolated in vitro from aortic blood, which was consistently positive from 6 to 144 hr PE, then only intermittently for the next 4 wk. Small PAS-positive cells observed in blood cultures grew into colonies consisting of non-septate tubules (pseudo-hyphae) terminating in multinucleated knob-like apices similar to those seen in organ explant cultures. Organ explants were culture positive every day; however, typical Ichthyophonus sp. schizonts were not observed histologically until 20–25 days PE. From 20 to 60 days PE, schizont diameter increased from ≤25 μm to ≥82 μm. Based on the data presented herein, we are confident that we have resolved the life cycle of Ichthyophonus sp. within the piscivorous host.
Synchronous cycling of Ichthyophoniasis with Chinook salmon density revealed during the annual Yukon River spawning migration
Populations of Chinook salmon Oncorhynchus tshawytscha in the Yukon River declined by more than 57% between 2003 and 2010, probably the result of a combination of anthropogenic and environmental factors. One possible contributor to this decline is Ichthyophonus , a mesomycetozoan parasite that has previously been implicated in significant losses of fish, including Chinook salmon. A multiyear epidemiological study of ichthyophoniasis in the Yukon River revealed that disease prevalence and Chinook salmon population abundance increased and decreased simultaneously (i.e., were concordant) from 1999 to 2010. The two values rose and fell synchronously 91% of the time for female Chinook salmon and 82% of the time for males; however, there was no significant correlation between Ichthyophonus prevalence and population abundance. This synchronicity might be explained by a single factor, such as a prey item that is critical to Chinook salmon survival as well as a source of Ichthyophonus infection. The host–parasite relationship between Ichthyophonus and migrating Chinook salmon from 2004 to 2010 was similar to that reported for the previous 5 years. During 2004–2010, overall disease prevalence was significantly higher among females (21%) than among males (8%), increased linearly with fish length for both males and females, and increased in both sexes as the fish progressed upriver. These regularly occurring features of host–parasite dynamics confirm a stable base of transmission for Ichthyophonus . However, from 2003 to 2010, disease prevalence decreased from 30% to just 8% in males and from 45% to 9% in females, paralleling a similar decline in Chinook salmon abundance during the same period. These findings may help clarify questions regarding the complex host–parasite dynamics that occur in marine species such as herrings Clupea spp., which have less well-defined population structures.
Exxon Valdez oil spill restoration project final report: Prince William Sound Herring disease program (HDP), restoration project 070819
Surveys of pathogens in Pacific herring from 2007 – 2010 indicated that Ichthyophonus , viral hemorrhagic septicemia virus, and erythrocytic necrosis virus are endemic in Prince William Sound and throughout the NE Pacific. Laboratory studies with VHSV indicated that multiple herring stocks are equally susceptible to the resulting disease, Pacific herring shed copious levels of VHSV (as high at 5x10 8 plaque-forming units/day) shortly after exposure, chronic and persistent infections can occur in Pacific herring, susceptibility of Pacific herring to VHS extends to the larval life stages but not the embryonic stages, and the prior exposure history of Pacific herring to VHSV can be determined post hoc. Laboratory studies involving Ichthyophonus indicated that schizonts can be inactivated with chlorine and iodine solutions, the parasite can survive for extended periods in saltwater but not freshwater, a low potential exists for cross contamination between in vitro explant cultures, infectious schizonts are released from the skin surface of infected herring, schizonts are not uniformly distributed throughout the skeletal muscle of infected Pacific herring, multiple types of Ichthyophonus exist with different genotypes and phenotypic traits, and temperature is an important factor influencing the infectivity of Ichthyophonus . Additional field and laboratory studies indicated that Ichthyophonus negatively influences the swimming performance of infected hosts and the negative impacts effects are exacerbated by increasing temperatures, American shad are an important reservoir of Ichthyophonus in the NE Pacific, Pacific herring are not susceptible to infectious hematopoietic necrosis (IHN), and Pacific herring will accept surgically implanted acoustic tags with negligible impacts on survival.
Amplification and transport of an endemic fish disease by an introduced species
The introduction of American shad from the Atlantic to the Pacific coast of North America in the late 1800’s and the subsequent population expansion in the 1980’s resulted in the amplification of Ichthyophonus sp., a Mesomycetozoean parasite of wild marine fishes. Sequence analysis of the ribosomal DNA gene complex (small subunit and internal transcribed spacer regions) and Ichthyophonus epidemiological characteristics indicate a low probability that Ichthyophonus was co-introduced with American shad from the Atlantic; rather, Ichthyophonus was likely endemic to marine areas of the Pacific region and amplified by the expanding population of a highly susceptible host species. The migratory life history of shad resulted in the transport of amplified Ichthyophonus from its endemic region in the NE Pacific to the Columbia River watershed. An Ichthyophonus epizootic occurred among American shad in the Columbia River during 2007, when infection prevalence was 72%, and 57% of the infections were scored as moderate or heavy intensities. The epizootic occurred near the record peak of shad biomass in the Columbia River, and corresponded to an influx of 1,595 mt of infected shad tissues into the Columbia River. A high potential for parasite spillback and the establishment of a freshwater Ichthyophonus life cycle in the Columbia River results from currently elevated infection pressures, broad host range, plasticity in Ichthyophonus life history stages, and precedents for establishment of the parasite in other freshwater systems. The results raise questions regarding the risk for sympatric salmonids and the role of Ichthyophonus as a population-limiting factor affecting American shad in the Columbia River.
Is disease an important mortality factor for Pacific herring?
Marine pelagic forage fishes, including sardines, anchovies, and herring, undergo large oscillations in population abundance. Although over-fishing can have a dramatic impact in reducing population size, this anthropogenic perturbation cannot fully account for all population declines because natural oscillations in marine pelagic fish biomasses occurred prior to the onset of commercial fishing. Among the herring metapopulation in Puget Sound / Straight of Georgia, the mean estimated annual mortality, exclusive of commercial fishing, increased from 20% in the late 1970’s and early 1980’s to 64-87% during 1996-1999. This natural mortality affects primarily the older age cohorts and resulted in recent decreased median ages of adult herring from age 4-6 cohorts to age 2-3 cohorts that die prior to iteroparous spawning. Surveys of wild herring from the region indicate that the protozoan parasite Ichthyophonus sp., is currently ubiquitous among Pacific herring populations in Washington and British Columbia, and prevalence of infection increase directly with herring age, from 12% among juveniles to 58% among the underrepresented age 6+ cohorts. Ichthyophonus can be highly pathogenic to immunologically naïve Pacific herring, causing 80% mortality 2 mo. after exposure in the laboratory. Current laboratory-based studies are underway to determine whether natural Ichthyophonus infections are terminal for the host, and whether these infections account for the decreasing median age of wild Pacific herring populations in recent years.