The class Labyrinthulomycetes constitutes a multitude of species found ubiquitously in the environment, and includes pathogens of corals, hard clams, turfgrasses, and seagrasses. Labyrinthula zosterae, the causative agent of seagrass wasting disease, has been associated with declines in seagrass coverage since the 1930s. However, pathogenic and nonpathogenic Labyrinthula spp. have been isolated from seagrass tissue. These isolates are difficult to distinguish morphologically, and the diversity of isolates where seagrass wasting disease is present is often unknown. This study aimed to increase knowledge on the pathogenicity and phylogeny of Labyrinthula spp. in Washington and Oregon, USA where a high prevalence of seagrass wasting disease has been associated with eelgrass, Zostera marina, declines. We tested the pathogenicity of 14 Labyrinthula isolates and compared partial 18S rRNA gene sequences of 12 isolates to sequences from around the world through the NCBI database. We found that pathogenic isolates could be identified as Labyrinthula zosterae, while nonpathogenic isolates did not form a clade with any previously identified SSU ribotypes. These results add to the growing data on Labyrinthula and seagrass wasting disease and can improve our understanding of pathogen evolution and spread in the future.

In the abalone and Candidatus Xenohaliotis californiensis (Ca. Xc) system, the Ca. Xc bacterium infects abalone digestive tissues and leads to extreme starvation and a characteristic "withering" of the gastropod foot. First identified in black abalone in California after an El Niño event, withering syndrome (WS) has caused large declines in wild black and captive white abalone on the northeastern Pacific coast, but disease resistance levels are species-, and possibly population-specific. This study compared gene expression patterns in the digestive gland of Ca. Xc-exposed and unexposed (control) Pinto abalone (Haliotis kamtschatkana), a particularly susceptible species. Lab-induced Ca. Xc infections were followed over 7 months and RNAseq was used to identify differential gene expression. Exposed Pinto abalone showed distinct changes in expression of 68 genes at 3 and 7 months post-infection relative to those in control animals. Upregulation of an orexin-like receptor (which is involved in feeding signaling) and a zinc peptidase-like region (many amino peptidases are zinc peptidases) in animals infected for 7 months indicates that animals with Ca. Xc infection may be starving and upregulating processes associated with feeding and digestion. Other groups of differentially expressed genes (DEGs) were upregulated or downregulated across control and exposed individuals over the 7-month experiment, including DEG groups that likely correspond to early disease state and to general stress response of being held in captivity. No patterns emerged in genes known to be involved in molluscan immune response, despite this being an expectation during a 7-month infection; digestion-related genes and unannotated DEGs were identified as targets for future research on potential immune response to WS in abalone.

Oyster aquaculture and seagrasses often co-occur and are each vital to the ecological and economic value of coastal ecosystems. Global declines in seagrasses, including Zostera marina, have recently been observed in association with multiple factors, including infection with diseases such as seagrass wasting disease (SWD), caused by the protist Labyrinthula zosterae. Protection of seagrasses has led to restrictions on oyster aquaculture due to perceived negative impacts on seagrass beds; however, positive impacts may also occur. An important aquaculture species, the Pacific oyster Crassostrea gigas, can filter L. zosterae from the water, potentially reducing pathogen transmission, although oysters may vector infection if they accumulate and release live L. zosterae into the water. We investigated whether oyster presence decreases lesion severity and infection intensity in eelgrass, or acts as a vector of L. zosterae, via laboratory and field experiments. In the laboratory, oysters and eelgrass were exposed to L. zosterae for 24 h and kept at 11°C or 18°C for 13 d. In the field, eelgrass ramets were deployed with and without oysters for 28 d adjacent to eelgrass known to have SWD. In the laboratory experiment, the presence of oysters significantly decreased lesion severity and infection intensity, but oysters previously exposed to L. zosterae did transmit the pathogen to naïve eelgrass. Temperature did not affect oyster ability to mitigate SWD; however, increased temperature significantly increased lesion severity. Oysters had no effect on SWD in the field. Further research is needed regarding the potential for oysters to vector L. zosterae and to quantify when oysters reduce SWD in the field.

We examined the risk of withering syndrome (WS) rickettsia-like organism (WS-RLO) infection in sentinel red abalone (Haliotis rufescens) deployed in modules at two Southern California field sites, one adjacent to an abalone farm and one adjacent to wild abalones. WS-RLO DNA was detected in seawater near modules at the wild abalone site but not near the farm (WS-RLO DNA was detected in the farm effluent). More WS-RLO DNA was detected in tissue from abalone near the farm relative to those near wild abalones (p < 0.05). However, infection prevalence and intensity based on histology were low and similar between sites (p > 0.05) and were independent of WS-RLO DNA loads in abalone tissue and seawater. More stippled (ST)-RLO than WS-RLO were observed with more ST-RLO infections near wild abalone than near the abalone farm (p < 0.05). We demonstrate the utility of caged sentinel abalone to better understand pathogen transmission patterns in the field.

To keep pace with rising opportunities for disease emergence and spread, surveillance in aquaculture must enable the early detection of both known and new pathogens. Conventional surveillance systems (designed to provide proof of disease freedom) may not support detection outside of periodic sampling windows, leaving substantial blind spots to pathogens that emerge in other times and places. To address this problem, we organized an expert panel to envision optimal systems for early disease detection, focusing on Ostreid herpesvirus 1 (OsHV-1), a pathogen of panzootic consequence to oyster industries. The panel followed an integrative group process to identify and weight surveillance system traits perceived as critical to the early detection of OsHV-1. Results offer a road map with fourteen factors to consider when building surveillance systems geared to early detection; factor weights can be used by planners and analysts to compare the relative value of different designs or enhancements. The results were also used to build a simple, but replicable, model estimating the system sensitivity (SSe) of observational surveillance and, in turn, the confidence in disease freedom that negative reporting can provide. Findings suggest that optimally designed observational systems can contribute substantially to both early detection and disease freedom confidence. In contrast, active surveillance as a singular system is likely insufficient for early detection. The strongest systems combined active with observational surveillance and engaged joint industry and government involvement: results suggest that effective partnerships can generate highly sensitive systems, whereas ineffective partnerships may seriously erode early detection capability. Given the costs of routine testing, and the value (via averted losses) of early detection, we conclude that observational surveillance is an important and potentially very effective tool for health management and disease prevention on oyster farms, but one that demands careful planning and participation. This evaluation centered on OsHV-1 detection in farmed oyster populations. However, many of the features likely generalize to other pathogens and settings, with the important caveat that the pathogens need to manifest via morbidity or mortality events in the species, life stages and environments under observation.

The spread, emergence, and adaptation of pathogens causing marine disease has been problematic to fisheries and aquaculture industries for the last several decades creating the need for strategic management and biosecurity practices. The Pacific oyster (Crassostrea gigas), a highly productive species globally, has been a target of disease and mortality caused by a viral pathogen, the Ostreid herpesvirus 1 (OsHV-1) and its microvariants (OsHV-1 mu vars). During routine surveillance to establish health history at a shellfish aquaculture nursery system in San Diego, California, the presence of OsHV-1 in Pacific oyster juveniles was detected. Quantification of OsHV-1 in tissues of oysters revealed OsHV-1 viral loads > 10(6) copies/mg. We characterized and identified the OsHV-1 variant by sequencing of ORFs 4 (C2/C6) and 43 (IA1/IA2), which demonstrated that this variant is a novel OsHV-1 microvariant: OsHV-1 mu var SD. A pilot transmission study indicates that OsHV-1 mu var SD is infectious with high viral loads similar to 7.57 x 10(6) copies/mg detected in dead individuals. The detection of OsHV-1 mu var SD in a large port mirrors previous studies conducted in Australia where aquaculture farms and feral populations near port locations may be at a higher risk of OsHV-1 emergence. Further research is needed to understand the impacts of OsHV-1 mu var SD, such as transmission studies focusing on potential vectors and characterization of virulence as compared to other OsHV-1 mu vars. To increase biosecurity of the global aquaculture industry, active and passive surveillance may be necessary to reduce spread of pathogens and make appropriate management decisions.

Economically devastating mortality events of farmed and wild shellfish due to infectious disease have been reported globally. Currently, one of the most significant disease threats to Pacific oyster Crassostrea gigas culture is the ostreid herpesvirus 1 (OsHV-1), in particular the emerging OsHV-1 microvariant genotypes. OsHV-1 microvariants (OsHV-1 µvars) are spreading globally, and concern is high among growers in areas unaffected by OsHV-1. No study to date has compared the relative virulence among variants. We provide the first challenge study comparing survival of naïve juvenile Pacific oysters exposed to OsHV-1 µvars from Australia (AUS µvar) and France (FRA µvar). Oysters challenged with OsHV-1 µvars had low survival (2.5% exposed to AUS µvar and 10% to FRA µvar), and high viral copy number as compared to control oysters (100% survival and no virus detected). As our study was conducted in a quarantine facility located ~320 km from the ocean, we also compared the virulence of OsHV-1 µvars using artificial seawater made from either facility tap water (3782 µmol kg-1 seawater total alkalinity) or purchased distilled water (2003 µmol kg-1). Although no differences in survival or viral copy number were detected in oysters exposed to seawater made using tap or distilled water, more OsHV-1 was detected in tanks containing the lower-alkalinity seawater, indicating that water quality may be important for virus transmission, as it may influence the duration of viral viability outside of the host.

The Ostreid herpesvirus 1 (OsHV-1) and variants, particularly the microvariants (μVars), are virulent and economically devastating viruses impacting oysters. Since 2008 OsHV-1 μVars have emerged rapidly having particularly damaging effects on aquaculture industries in Europe, Australia and New Zealand. We conducted field trials in Tomales Bay (TB), California where a non-μVar strain of OsHV-1 is established and demonstrated differential mortality of naturally exposed seed of three stocks of Pacific oyster, Crassostrea gigas, and one stock of Kumamoto oyster, C. sikamea. Oysters exposed in the field experienced differential mortality that ranged from 64 to 99% in Pacific oysters (Tasmania>Midori = Willapa stocks), which was much higher than that of Kumamoto oysters (25%). Injection trials were done using French (FRA) and Australian (AUS) μVars with the same oyster stocks as planted in the field and, in addition, two stocks of the Eastern oyster, C. virginica. No mortality was observed in control oysters. One C. virginica stock suffered ~10% mortality when challenged with both μVars tested. Two Pacific oyster stocks suffered 75 to 90% mortality, while one C. gigas stock had relatively low mortality when challenged with the AUS μVar (~22%) and higher mortality when challenged with the French μVar (~72%). Conversely, C. sikamea suffered lower mortality when challenged with the French μVar (~22%) and higher mortality with the AUS μVar (~44%). All dead oysters had higher viral loads (~1000×) as measured by quantitative PCR relative to those that survived. However, some survivors had high levels of virus, including those from species with lower mortality. Field mortality in TB correlated with laboratory mortality of the FRA μVar (69% correlation) but not with that of the AUS μVar, which also lacked correlation with the FRA μVar. The variation in response to OsHV-1 variant challenges by oyster species and stocks demonstrates the need for empirical assessment of multiple OsHV-1 variants.

Pacific oysters, Crassostrea gigas, are one of the most productive aquaculture species in the world. However, they are threatened by the spread of Ostreid herpesvirus-1 (OsHV-1) and its microvariants (collectively “µvars”), which cause mass mortalities in all life stages of Pacific oysters globally. Breeding programs have been successful in reducing mortality due to OsHV-1 variants following viral outbreaks; however, an OsHV-1-resistant oyster line does not yet exist in the United States (US), and it is unknown how OsHV-1 µvars will affect US oyster populations compared to the current variant, which is similar to the OsHV-1 reference, found in Tomales Bay, CA. The goals of this study were to investigate the resistance of C. gigas juveniles produced by the Molluscan Broodstock Program (MBP) to three variants of OsHV-1: a California reference OsHV-1, an Australian µvar, and a French µvar. This is the first study to directly compare OsHV-1 µvars to a non-µvar. The survival probability of oysters exposed to the French (FRA) or Australian (AUS) µvar was significantly lower (43% and 71%, respectively) than to the reference variant and controls (96%). No oyster family demonstrated resistance to all three OsHV-1 variants, and many surviving oysters contained high copy numbers of viral DNA (mean ~3.53 × 108). These results indicate that the introduction of OsHV-1 µvars could have substantial effects on US Pacific oyster aquaculture if truly resistant lines are not achieved, and highlight the need to consider resistance to infection in addition to survival as traits in breeding programs to reduce the risk of the spread of OsHV-1 variants.

Withering syndrome (WS) is a chronic bacterial disease that affects numerous northeastern Pacific abalone Haliotis spp. The causative agent of WS is an obligate intracellular Rickettsiales-like bacterium (WS-RLO) that remains unculturable, thereby limiting our understanding of WS disease dynamics. The objectives of our study were to (1) determine the temporal stability of WS-RLO DNA outside of its abalone host in 14°C and 18°C seawater, (2) develop a standardized protocol for exposing abalones to known concentrations of WS-RLO DNA, and (3) calculate the dose of WS-RLO DNA required to generate 50% infection prevalence (ID50) in the highly cultured red abalone Haliotis rufescens. The WS-RLO stability trials were conducted in October 2016, February 2017, and June 2017. A quantitative PCR (qPCR) analysis was used to quantify bacterial DNA for 7 d in seawater collected at an abalone farm in southern California, where the pathogen is now endemic. For all trials and temperature treatments, WS-RLO DNA was unstable in seawater for longer than 2 d. To determine an ID50, groups of uninfected juvenile red abalone were subjected to 3-h bath exposures with four concentrations of WS-RLO at 0, 103 , 104 , and 105 DNA copies/mL. Abalone feces were tested biweekly for the presence of WS-RLO DNA, and abalone tissues were sampled 9 weeks postinfection for histological and qPCR analyses. The ID50 results indicated that our protocol was successful in generating WS-RLO infections; a pathogen dose of 2.3 × 103 DNA copies/mL was required to generate a 50% infection prevalence in red abalone tissue. These findings are critical components of disease dynamics that will help assess WS transmission risk within and among abalone populations and facilitate appropriate management and restoration strategies for both wild and cultured abalone species in WS-endemic areas.

Abstract Background Variants of the Ostreid herpesvirus 1 (OsHV-1) cause high losses of Pacific oysters globally, including in Tomales Bay, California, USA. A suite of new variants, the OsHV-1 microvariants (μvars), cause very high mortalities of Pacific oysters in major oyster-growing regions outside of the United States. There are currently no known Pacific oysters in the United States that are resistant to OsHV-1 as resistance has yet to be evaluated in these oysters. As part of an effort to begin genetic selection for resistance to OsHV-1, 71 families from the Molluscan Broodstock Program, a US West Coast Pacific oyster breeding program, were screened for survival after exposure to OsHV-1 in Tomales Bay. They were also tested in a quarantine laboratory in France where they were exposed to a French OsHV-1 microvariant using a plate assay, with survival recorded from three to seven days post-infection. Results Significant heritability for survival were found for all time points in the plate assay and in the survival phenotype from a single mortality count in Tomales Bay. Genetic correlations between survival against the French OsHV-1 μvar in the plate assay and the Tomales Bay variant in the field trait were weak or non-significant. Conclusions Future breeding efforts will seek to validate the potential of genetic improvement for survival to OsHV-1 through selection using the Molluscan Broodstock Program oysters. The lack of a strong correlation in survival between OsHV-1 variants under this study’s exposure conditions may require independent selection pressure for survival to each variant in order to make simultaneous genetic gains in resistance.

Pinto abalone (Haliotis kamtschatkana), the widest ranging abalone species in North America, occurs from Alaska, United States to Central Baja California, Mexico. The species has been observed in intertidal and subtidal habitats from 0 to 40 m depth. The best available data indicate that pinto abalone abundance has declined in many areas throughout the species' range due to fisheries harvest. Subsistence and personal use fisheries in Alaska and a commercial fishery in Mexico persist. Preliminary data from 2008 to 2016 indicate signs of recovery for some pinto abalone populations along the British Columbia coast due to multiple contributing factors including a reduction in illegal harvest, natural recovery following fishery closure, and low predation pressure. By contrast, pinto abalone populations at the San Juan Islands in Washington are experiencing recruitment failure and continuing to decline, despite closure of the fisheries and no evidence of poaching. Throughout the remainder of the species' range, trends are less clear, due to the lack of regular, long-term monitoring surveys for pinto abalone. The limited data from surveys and/or opportunistic sightings indicate that pinto abalone populations are small, patchily distributed, and/or fluctuate episodically in Alaska, California, and Mexico, with evidence of recent recruitment in a number of locations within these three areas. Baseline abundance and trend data for the species before the advent of commercial fisheries and, in some areas, the local extirpation of sea otters is lacking. Without a clear baseline with which to compare the current abundance levels and trend information, it is difficult to interpret what these levels mean for the status and viability of the species. Threats to pinto abalone were evaluated and characterized using a qualitative rating (i.e., low, moderate, high, very high) based on the threats' scope, severity, and persistence and the sufficiency of the data to support the rating. Several threats that posed a moderate level of risk to pinto abalone were identified including the following: low densities as a result of historical overfishing; the potential threat posed by ocean acidification; and illegal take because of poaching and inadequate law enforcement. The overall risk that pinto abalone face throughout their range was evaluated, and it was determined that they have a low to moderate level of extinction risk now and in the foreseeable future (over both the 30-y and 100-y time horizons). There is a high level of uncertainty regarding demographic factors, in particular regarding whether abundance and productivity levels are sufficient to support the persistence and recovery of the species in the face of continuing and potential future threats. Although recruitment failure may be occurring in some areas (e.g., San Juan Islands Archipelago), in other areas throughout the range recurring and/or recent recruitment events have been observed, despite low densities, and have even resulted in increased densities (across all size classes) at several index sites in British Columbia. Limitations in using demographic data to guide conservation actions and help ensure species persistence could be overcome by conducting consistent monitoring of pinto abalone populations throughout their range.

Parasites of the genus Ichthyophonus infect many fish species and have a non-uniform distribution within host tissues. Due in part to this uneven distribution, the comparative sensitivity and accuracy of using molecular-based detection methods versus culture to estimate parasite prevalence is under debate. We evaluated the analytical and diagnostic performance of an existing qPCR assay in comparison to the 'gold standard' culture method using Pacific herring Clupea pallasii with known exposure history. We determined that the assay is suitable for use in this host, and diagnostic specificity was consistently high (> 98%) in both heait and livei tissues. Diagnostic sensitivity could not be fully assessed due to low infection rates, but our results suggest that qPCR is not as sensitive as culture under all circumstances. Diagnostic sensitivity of qPCR relative to culture is likely affected by the amount of sample processed. The prevalence values estimated by the 2 methods were not significantly different when sample amounts were equal (heart tissue), but when the assayed sample amounts were unequal (liver tissue), the culture method detected a significantly higher prevalence of the parasite than qPCR. Further, cultui e of liver also detected significantly more Ichthyophonus infections than culture of heart, suggesting that the density and distribution of parasites in tissues also plays a role in assay sensitivity. This sensitivity issue would be most problematic foi fish with light infections. Although qPCR does not detect the presence of a live organism, DNA-based pathogen detection methods provide the opportunity for alternate testing strategies when culture is not possible.

Climate change is affecting the health and physiology of marine organisms and altering species interactions. Ocean acidification (OA) threatens calcifying organisms such as the Pacific oyster, Crassostrea gigas. In contrast, seagrasses, such as the eelgrass Zostera marina, can benefit from the increase in available carbon for photosynthesis found at a lower seawater pH. Seagrasses can remove dissolved inorganic carbon from OA environments, creating local daytime pH refugia. Pacific oysters may improve the health of eelgrass by filtering out pathogens such as Labyrinthula zosterae (LZ), which causes eelgrass wasting disease (EWD). We examined how co-culture of eelgrass ramets and juvenile oysters affected the health and growth of eelgrass and the mass of oysters under different pCO2 exposures. In Phase I, each species was cultured alone or in co-culture at 12°C across ambient, medium, and high pCO2 conditions, (656, 1,158 and 1,606 μatm pCO2 , respectively). Under high pCO2 , eelgrass grew faster and had less severe EWD (contracted in the field prior to the experiment). Co-culture with oysters also reduced the severity of EWD. While the presence of eelgrass decreased daytime pCO2 , this reduction was not substantial enough to ameliorate the negative impact of high pCO2 on oyster mass. In Phase II, eelgrass alone or oysters and eelgrass in co-culture were held at 15°C under ambient and high pCO2 conditions, (488 and 2,013 μatm pCO2 , respectively). Half of the replicates were challenged with cultured LZ. Concentrations of defensive compounds in eelgrass (total phenolics and tannins), were altered by LZ exposure and pCO2 treatments. Greater pathogen loads and increased EWD severity were detected in LZ exposed eelgrass ramets; EWD severity was reduced at high relative to low pCO2 . Oyster presence did not influence pathogen load or EWD severity; high LZ concentrations in experimental treatments may have masked the effect of this treatment. Collectively, these results indicate that, when exposed to natural concentrations of LZ under high pCO2 conditions, eelgrass can benefit from co-culture with oysters. Further experimentation is necessary to quantify how oysters may benefit from co-culture with eelgrass, examine these interactions in the field and quantify context-dependency.

Population declines in wild and cultured abalones (Haliotis spp.) due to a bacterial disease called withering syndrome (WS) have been documented along the northeastern Pacific Ocean. However, observed differences in species susceptibility to the disease are not well understood. Here, we examined the susceptibility of three temperate abalone species, the cool water (4-14 degrees C) pinto or northern abalone (Haliotis kamtschatkana), the intermediate water (8-18 degrees C) red abalone (H. rufescens), and the warm water (12-23 degrees C) pink abalone (H. corrugata), to experimental WS infection at temperatures facilitating disease proliferation. Mortality data paired with histological and molecular detection of the WS pathogen confirmed that these abalone species exhibit different levels of susceptibility to infection and resistance to WS development ranging from high susceptibility and low resistance in pinto abalone to moderate/low susceptibility and resistance in red and pink abalones. The temperature associated with WS induced mortalities also varied among species: pinto abalone died at the lowest experimental temperature (17.32 +/- 0.09 degrees C), while red abalone died at an intermediate temperature (17.96 +/- 0.16 degrees C), and pink abalone required the highest temperature (18.84 +/- 0.16 degrees C). When data from the current and previous studies were examined, susceptibility to WS was inversely related to phylogenetic distance from white abalone (H. sorenseni), which had the highest susceptibility and lowest resistance of all abalone species tested prior to the current study. These results provide further evidence that an abalone's thermal optima and phylogenetic relationship can determine its susceptibility to WS; species with cool water evolutionary histories are most susceptible to WS and the most susceptible species appear to be closely related. Differences among the thermal ranges of abalone species have broad implications for WS disease dynamics and highlight the importance of understanding the mechanisms governing the abalone-WS relationship in order to properly manage declining abalone populations.

The Dungeness crab, Cancer magister, is an important resource species, and in Puget Sound, USA, where the adults occur in inshore waters that have summer pH as low as 7.6, future levels are predicted as low as 7.1. Using eggs and larvae from females captured in Puget Sound in late 2012, this laboratory study examined hatching success, larval survival, and larval development rate at target pH of 8.0, 7.5, and 7.1, which represent present open ocean, present coastal upwelling, and projected upwelling conditions. Toward the end of their development, the eggs of one C. magister were exposed to the three treatments and they began to hatch after 22 days. Hatching probability was unaffected by lower pH, but hatching was delayed at pH 7.1. In a second experiment, significantly more C. magister larvae survived after 45 days at pH 8.0 than at the two lower pH: 58, 14, and 21 %. The sizes of the zoeae were unaffected by treatment, but larvae in the low-pH treatments progressed through larval stages more slowly. This study shows that low-pH seawater slows embryonic and early larval development and causes appreciable larval mortality. It suggests that ocean acidification could have a measurable impact on the population dynamics of C. magister.

Linking marine epizootics to a specific aetiology is notoriously difficult. Recent diagnostic successes show that marine disease diagnosis requires both modern, cutting-edge technology (e.g. metagenomics, quantitative real-time PCR) and more classic methods (e.g. transect surveys, histopathology and cell culture). Here, we discuss how this combination of traditional and modern approaches is necessary for rapid and accurate identification of marine diseases, and emphasize how sole reliance on any one technology or technique may lead disease investigations astray. We present diagnostic approaches at different scales, from the macro (environment, community, population and organismal scales) to the micro (tissue, organ, cell and genomic scales). We use disease case studies from a broad range of taxa to illustrate diagnostic successes from combining traditional and modern diagnostic methods. Finally, we recognize the need for increased capacity of centralized databases, networks, data repositories and contingency plans for diagnosis and management of marine disease.

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.