Rainbow trout, Oncorhynchus mykiss, were infected with Ichthyophonus sp. and held at 10 degrees C, 15 degrees C and 20 degrees C for 28 days to monitor mortality and disease progression. Infected fish demonstrated more rapid onset of disease, higher parasite load, more severe host tissue reaction and reduced mean-day-to-death at higher temperature. In a second experiment, Ichthyophonus-infected fish were reared at 15 degrees C for 16 weeks then subjected to forced swimming at 10 degrees C, 15 degrees C and 20 degrees C. Stamina improved significantly with increased temperature in uninfected fish; however, this was not observed for infected fish. The difference in performance between infected and uninfected fish became significant at 15 degrees C (P = 0.02) and highly significant at 20 degrees C (P = 0.005). These results have implications for changes in the ecology of fish diseases in the face of global warming and demonstrate the effects of higher temperature on the progression and severity of ichthyophoniasis as well as on swimming stamina, a critical fitness trait of salmonids. This study helps explain field observations showing the recent emergence of clinical ichthyophoniasis in Yukon River Chinook salmon later in their spawning migration when water temperatures were high, as well as the apparent failure of a substantial percentage of infected fish to successfully reach their natal spawning areas.

Journal of Fish DiseasesVolume 31, Issue 4 p. 317-320 Potential for cross-contamination of in vitro explant cultures initiated from Ichthyophonus-infected rainbow trout, Oncorhynchus mykiss (Walbaum) S LaPatra, S LaPatra Research Division, Clear Springs Foods, Inc., Buhl, ID, USASearch for more papers by this authorR Kocan, R Kocan School of Aquatic and Fishery Sciences, University of Washington, Seattle, WA, USASearch for more papers by this authorP Hershberger, P Hershberger Marrowstone Marine Field Station, US Geological Survey, Nordland, WA, USASearch for more papers by this author S LaPatra, S LaPatra Research Division, Clear Springs Foods, Inc., Buhl, ID, USASearch for more papers by this authorR Kocan, R Kocan School of Aquatic and Fishery Sciences, University of Washington, Seattle, WA, USASearch for more papers by this authorP Hershberger, P Hershberger Marrowstone Marine Field Station, US Geological Survey, Nordland, WA, USASearch for more papers by this author First published: 18 March 2008 https://doi.org/10.1111/j.1365-2761.2007.00892.xCitations: 2 S LaPatra, Clear Springs Foods, Inc., PO Box 712, Buhl, ID 83316, USA(e-mail: scottl@clearsprings.com) Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume31, Issue4April 2008Pages 317-320 RelatedInformation

Two genetically distinct populations of chinook salmon, Oncorhynchus tshawytscha (Walbaum), were simultaneously sampled at the confluence of the Yukon and Tanana rivers in 2003. Upper Yukon-Canadian fish had significantly higher infection prevalence as well as more severe infections (higher parasite density in heart tissue) than the lower Yukon-Tanana River fish. Both populations had migrated the same distance from the mouth of the Yukon River at the time of sampling but had significantly different distances remaining to swim before reaching their respective spawning grounds. Multiple working hypotheses are proposed to explain the differences between the two stocks: (1) the two genetically distinct populations have different inherent resistance to infection, (2) genetically influenced differences in feeding behaviour resulted in temporal and/or spatial differences in exposure, (3) physiological differences resulting from different degrees of sexual maturity influenced the course of disease, and (4) the most severely infected Tanana River fish either died en route or fatigued and were unable to complete their migration to the Tanana River, thus leaving a population of apparently healthier fish.

Swimming stamina, measured as time-to-fatigue, was reduced by approximately two-thirds in rainbow trout experimentally infected with Ichthyophonus. Intensity of Ichthyophonus infection was most severe in cardiac muscle but multiple organs were infected to a lesser extent. The mean heart weight of infected fish was 40% greater than that of uninfected fish, the result of parasite biomass, infiltration of immune cells and fibrotic (granuloma) tissue surrounding the parasite. Diminished swimming stamina is hypothesized to be due to cardiac failure resulting from the combination of parasite-damaged heart muscle and low myocardial oxygen supply during sustained aerobic exercise. Loss of stamina in Ichthyophonus-infected salmonids could explain the poor performance previously reported for wild Chinook and sockeye salmon stocks during their spawning migration.

Two morphologically distinct forms of an intraerythrocytic parasite(s) were detected by microscopic observation of Giemsa-stained blood films in 45.7% of 119 rockfish (Sebastes emphaeus) from the San Juan Archipelago (Washington State, U.S.A.). Infection prevalence for both forms was 53% in males, 44% in females, and 33% in fish of undetermined gender. A binucleate "ring-stage" was present at all 4 geographic sites, with a mean prevalence of 45.7%, while mean prevalence of a larger gamont-like form from the same sites was 5.1%. The relationship of the 2 forms to each other could not be determined. Neither schizogony nor binary fission was evident in any of the infected erythrocytes and the parasites contained no obvious pigment. The possibility of the 2 morphologic forms being 2 distinct species is supported by the observation that no difference in parasitemia was seen in the binucleate form among sites (1.6-1.9%), while parasitemia of the gamont-like form varied significantly among sites, ranging from a high of 4% to a low of 0.1%. Taxonomic status of either form could not be determined at this time based on limited existing morphologic data.

In vitro explant cultures identified Ichthyophonus in 10.9% of 302 Puget Sound rockfish Sebastes emphaeus sampled from five sites in the San Juan Islands archipelago and Puget Sound, Washington, in 2003. None of the infected fish exhibited visible lesions and only a single fish was histologically positive. Significantly more females were infected (12.4%) than males (6.8%), and while infected males were only detected at two of the five sites, infected females were identified at all sites, with no significant differences in infection prevalence. Genomic sequences of Ichthyophonus isolates obtained from Puget Sound rockfish, Pacific herring Clupea pallasii, and Yukon River Chinook salmon Oncorhynchus tshawytscha were identical in both the A and B regions of the small subunit 18S ribosomal DNA but were different from Ichthyophonus sequences previously isolated from four different species of rockfish from the northeastern Pacific Ocean Ichthyophonu.s in Puget Sound rockfish may not have been previously detected because the infection is subclinical in this species and earlier investigators did not utilize in vitro techniques for diagnosis of ichthyophoniasis. However, since clinical ichthyophoniasis has recently been identified in several other species of northeast Pacific rockfishes, it is hypothesized that this either is an emerging disease resulting from changing marine conditions or the result of introduction by infected southern species that appear during periodic El Nino events.

keywords, and citation............................................................. 4 Introduction.................................................................................... 5 Objectives...................................................................................... 6 Methods......................................................................................... 6 Results........................................................................................... 13 Discussion...................................................................................... 17 Summary....................................................................................... 24 Conclusions.................................................................................... 24 Acknowledgements........................................................................... 25 Literature cited................................................................................. 25 Footnotes....................................................................................... 29

Before 1985, Ichthyophonus was unreported among Pacific salmon Oncorhynchus spp. from the Yukon River; now it infects more than 40% of returning adult Chinook salmon O. tshawytscha. Overall infection prevalence reached about 45% in the Yukon River and about 30% in the Tanana River between 1999 and 2003. Mean infection prevalence was greater in females than males in the main-stem Yukon River during each of the 5 years of the study, but the infection prevalence in males increased each year until the difference was no longer significant. Clinical signs of ichthyophoniasis (presence of visible punctate white lesions in internal organs) were least at the mouth of the Yukon River (similar to10%) but increased to 29% when fish reached the middle Yukon River and was 22% at the upper Tanana River. However, clinical signs increased each year from 7% in 1999 to 27% in 2003 at the mouth of the river. As fish approached the upper reaches of the Yukon River (Canada) and the spawning areas of the Chena and Salcha rivers (Alaska), infection prevalence dropped significantly to less than 15% in females on the Yukon River and less than 10% for both sexes in the Chena and Salcha rivers, presumably because of mortality among infected prespawn fish. Age was not a factor in infection prevalence, nor was the position of fish within the run. The source of infection was not determined, but Ichthyophonus was not found in 400 Pacific herring Clupea pallasi from the Bering Sea or in 120 outmigrating juvenile Chinook salmon from two drainages in Alaska and Canada. Freshwater burbot Lota Iota from the middle Yukon River were subclinically infected with Ichthyophonus, but the origin and relationship of this agent to the Chinook salmon isolate is unknown.

The initial characterization of a rhabdovirus isolated from a single, asymptomatic starry flounder (Platichthys stellatus) collected during a viral survey of marine fishes from the northern portion of Puget Sound, Washington, USA, is reported. Virions were bullet-shaped and approximately 100 nm long and 50 nm wide, contained a lipid envelope, remained stable for at least 14 days at temperatures ranging from -80 to 5degreesC and grew optimally at 15degreesC in cultures of epithelioma papulosum cyprini (EPC) cells. The cytopathic effect on EPC cell monolayers was characterized by raised foci containing rounded masses of cells. Pyknotic and dark-staining nuclei that also showed signs of karyorrhexis were observed following haematoxylin and eosin, May-Grunwald Giemsa and acridine orange staining. PAGE of the structural proteins and PCR assays using primers specific for other known fish rhabdoviruses, including Infectious hematopoietic necrosis virus, Viral hemorrhagic septicemia virus, Spring viremia of carp virus, and Hirame rhabdovirus, indicated that the new virus, tentatively termed starry flounder rhabdovirus (SFRV), was previously undescribed in marine fishes from this region. In addition, sequence analysis of 2678 nt of the amino portion of the viral polymerase gene indicated that SFRV was genetically distinct from other members of the family Rhabdoviridae for which sequence data are available. Detection of this virus during a limited viral survey of wild fishes emphasizes the void of knowledge regarding the diversity of viruses that naturally infect marine fish species in the North Pacific Ocean.

A recent decrease in the mean age of adult Pacific herring Clupea pallasi in Puget Sound was associated with a high prevalence of Ichthyophonus hoferi, a protistan parasite that can be highly pathogenic to Pacific herring. In Puget Sound, high intensities of 1. hoferi infection may be maintained in older cohorts of Pacific herring because the prevalence of 1. hoferi increased with age from 12% among juveniles to 58% among the oldest, age-6 and older cohorts. Low intensities of L hoferi infection in the region may be maintained in alternative fish hosts, such as surf smelt Hypomesus pretiosus, Puget Sound rockfish Sebastes emphaeus, Pacific tomeod Microgadus proximus. and speckled sanddab Cithanichthys stigmaeus.

Specific-pathogen-free Pacific herring Clupea pallasi were reared in the laboratory from eggs and then challenged at 5, 9, and 13 months of age by waterborne exposure to low (10(1.5-2.5) plaque-forming units [PFU] per milliliter), medium (10(3.5-4.5) PFU/mL), or high (10(5.5-6.5) PFU/mL) levels of a North American isolate of viral hemorrhagic septicemia virus (VHSV). The fish were extremely susceptible to the virus, showing clinical disease, mortality approaching 100%,, and only a limited increase in resistance with age. Mortality began 4-6 d after exposure and peaked at approximately day 7 in fish exposed to high levels of virus. Whereas the mean time to death showed a significant dose response (P < 0.001), the percent mortality and virus titers in dead fish were generally high in all groups regardless of initial challenge dose. External signs of disease were usually limited to 1-2-mm hemorrhagic areas on the lower jaw and isthmus and around the eye, but 2 of 130 infected fish exhibited extensive cutaneous hemorrhaging. Histopathologic examination of tissues from moribund fish sampled at 2-8 d after exposure revealed multifocal coagulative necrosis of hepatocytes, diffuse necrosis of interstitial hematopoietic tissues in the kidney, diffuse necrosis of the spleen, epidermis, and subcutis, and occasional necrosis of pancreatic acinar cells. Virus titers in tissues of experimentally infected herring were first detected 48 h after exposure and peaked 6-8 d after exposure at 10(7.7) PFU/g. Fish began shedding virus at 48 h after exposure with titers in the Row-through aquaria reaching 10(2.5) PFU/mL at 4-5 d after exposure, just before peak mortality. When the water flow was turned off for 3 h, titers in the water rose to 10(3.5) PFU/mL, and the amount of virus shed by infected fish (on average, greater than 10(6.5) PFU/h per fish) appeared sufficient to sustain a natural epizootic among schooling herring. Taken together, these data suggest that VHSV could be a significant limiting factor for populations of Pacific herring.