FIRST REPORT IN CANADA OF CERATONOVA GASTEROSTEA (CNIDARIA: MYXOZOA) IN THREESPINE STICKLEBACK GASTEROSTEUS ACULEATUS FROM THE FRASER RIVER, BRITISH COLUMBIA.

A survey of threespine stickleback (Gasterosteus aculeatus L.) and juvenile Chinook salmon (Oncorhynchus tshawytscha (Walbaum)) from the lower Fraser River, British Columbia, Canada and adjacent marine waters was conducted for the presence of myxozoan parasites of the genus Ceratonova Atkinson, Foott, and Bartholomew, 2014 (Bivalvulida, Ceratomyxidae). Quantitative polymerase chain reaction (PCR; small subunit ribosomal RNA gene) combined with conventional PCR and sequencing (ribosomal internal transcribed spacer region 1) detected Ceratonova gasterostea Atkinson, Foott, and Bartholomew, 2014 in 89 of 269 (33.1%) sticklebacks from the Fraser River and in 1 of 51 (2%) from Howe Sound. In contrast, Ceratonova shasta (Atkinson, Foott, and Bartholomew, 2014) was detected in 2 of 26 (7.7%) Chinook salmon from the Fraser River. Reciprocal infections were not detected. Light microscopic examination of the stickleback intestine revealed the presence of an intense infection with large numbers of pseudoplasmodia which interacted with enterocytes through pseudopodia-like projections. Myxospores with characteristic elongate valve cells were visible within mature pseudoplasmodia and free in the intestinal lumen. This is the first report of C. gasterostea in Canada.

Parvicapsula pseudobranchicola in the northeast Pacific Ocean is rare in farmed Atlantic salmon Salmo salar despite widespread occurrence and pathology in wild Pacific salmon Oncorhynchus spp.

BACKGROUND: Infection with the myxozoan parasite Parvicapsula pseudobranchicola causes disease in wild and farmed salmonids in Norway. In the northeast Pacific Ocean, the parasite has been reported in Pacific salmon Oncorhynchus spp. without evidence of disease. The objectives of the present study were to confirm the identity of P. pseudobranchicola in the Pacific, document its host and geographic ranges, and describe associated pathological changes. METHODS: Ocean-entry year wild pink salmon Oncorhynchus gorbuscha, chum salmon O. keta, Chinook salmon O. tshawytscha, coho salmon O. kisutch and sockeye salmon O. nerka were collected in summer and autumn surveys near Vancouver Island (VI) and from a winter survey in the Gulf of Alaska. Samples were also obtained from farmed Atlantic salmon Salmo salar and Chinook salmon near VI. Samples were analysed by qPCR and histology using conventional staining or in situ hybridisation. Parasite sequence was obtained from small subunit ribosomal RNA gene (SSU rDNA). RESULTS: Identical 1525 base-pair SSU rDNA sequences from infected pink salmon, chum salmon and Chinook salmon shared 99.93% identity with a P. pseudobranchicola sequence from Norwegian Atlantic salmon. In autumn surveys, the prevalence was greatest in chum salmon (91.8%) and pink salmon (85.9%) and less so in Chinook salmon (68.8%) and sockeye salmon (8.3%). In farmed salmon, the prevalence was zero in Atlantic salmon (n = 967) and 41% in Chinook salmon (n = 118). Infections were preferentially sited in pseudobranch and visualised by in situ hybridisation. Heavy parasite burdens in all species of Pacific salmon were inconsistently associated with focal granulomatous pseudobranchitis. CONCLUSIONS: In the northeast Pacific, widespread occurrence of P. pseudobranchicola in Pacific salmon together with its absence or sporadic occurrence in farmed Atlantic salmon differs from its epidemiology in Norway, despite similar pathological development in the pseudobranch. Consequences of the infections to the health of wild Pacific salmon, identity of the invertebrate host and the distribution and abundance of infective actinospores are unknown and remain high priorities for research.

Increased prevalence and severity of Kudoa thyrsites (Cnidaria: Myxosporea) in Atlantic salmon Salmo salar exposed to deeper seawater.

Kudoa thyrsites is a myxozoan parasite of marine fish with a global distribution. In British Columbia (BC), Canada, severe infections are associated with an economically significant degradation of fillet quality in farmed Atlantic salmon. Exposures to naturally occurring actinospores at a coastal research laboratory were used to test the hypothesis that the prevalence and severity of K. thyrsites infections acquired by exposure of Atlantic salmon to seawater (SW) of various depths are not different. In Expt 1, fish were exposed to SW from 1, 7 or 13 m below the surface. Following exposure to deeper-sourced SW, the prevalence of K. thyrsites, determined from microscopic examination of muscle histology sections, was greater in all 4 trials and the severity of infection was greater in 2 trials. In Expt 2, infections were compared over time among salmon held in tanks supplied with deep-sourced SW (raw or UV-irradiated) or in a surface net-pen. The infection was observed in 35 of 40 fish sampled between 3 and 6 mo after tank exposure to raw SW. Coincidentally, the parasite was observed in 4 of 40 fish maintained in the net-pen. No consistent association of the parasite infection was observed with temperature; however, reduced salinity and solar radiation were not ruled out as factors which may reduce the risk of infection from surface SW. These findings require verification at commercial aquaculture sites in BC, as they will inform considerations related to farm siting and net-pen configuration.

Elevated Seawater Temperature and Infection with Neoparamoeba perurans Exacerbate Complex Gill Disease in Farmed Atlantic Salmon (Salmo salar) in British Columbia, Canada.

Gill disorders and diseases are emergent health concerns affecting marine-farmed salmon, for which the causal factors are poorly understood in British Columbia (BC), Canada. This study sought to describe and compare spatial and temporal patterns of infection with Neoparamoeba perurans, the causal agent of amoebic gill disease, and visually assessed gill health scores in farmed Atlantic salmon. Gill tissue obtained during the Fisheries and Oceans Canada's Fish Health Audit and Intelligence Program (DFO-FHAIP) between 2016 and 2020 were screened for N. perurans by qPCR. Semi-quantitative visual gill health assessments were conducted during the audits, and farms were assigned clinical AGD status based on microscopic visualization of N. perurans together with histopathological lesions. Seawater temperature and salinity data were collected from all active farms in the region during the study period. Trends in gill scores and associations with N. perurans infections were described and tested using an ordinal logistic mixed model. The amoeba was detected in 21% of 345 audited farms and in 12% of 1925 fish samples. Most (56%, n = 1898) samples had no visible gill damage (score = 0), and 23% had scores ≥ 2 (high). Distinct patterns of spatial and temporal variability in the rates of high gill scores and N. perurans infections are demonstrated. The model supported the statistically significant relationship observed between seawater temperature and the proportion of samples with elevated gill scores. The model also revealed a direct relationship between salinity and gill score but only in the presence of N. perurans. While the data suggest that histopathological lesions contributed to the gill scores, temperature and, to a lesser extent, salinity were significant risk factors of increased gill score. The results are discussed in the context of recently frequent thermal anomalies in the northeastern Pacific Ocean.

Piscirickettsia salmonis shedding and tissue burden, and hematological responses during cohabitation infections in chum Oncorhynchus keta, pink O. gorbuscha and Atlantic salmon Salmo salar.

BACKGROUND: Salmonid rickettsial septicemia is an emergent and geographically widespread disease of marine-farmed salmonids caused by infection with the water-borne bacterium Piscirickettsia salmonis. Very little is known about the route, timing, or magnitude of bacterial shedding from infected fish. METHODOLOGY/PRINCIPAL FINDINGS: A cohabitation challenge model was used to assess shedding from chum Oncorhynchus keta, pink O. gorbuscha and Atlantic salmon Salmo salar. Infections in donor fish were established by intraperitoneal injection of P. salmonis. Naïve recipients were cohabitated with donor fish after which cumulative percent morbidity and mortality (CMM) was monitored, and bacterial burdens in kidney and in tank water were measured by qPCR. All donor fish died with mean days-to-death (MDD) among species ranging from 17.5 to 23.9. Among recipients, CMM ranged from 42.7% to 77.8% and MDD ranged from 49.7 to 56.4. In each trial, two peaks of bacterial DNA concentrations in tank water closely aligned with the MDD values of donor and recipient fish. Bacterial tissue burden and shedding rate, and plasma physiological parameters were obtained from individual donors and recipients. Statistically significant positive correlations between the shedding rate and P. salmonis kidney burden were measured in donor pink and in donor and recipient chum salmon, but not in donor or recipient Atlantic salmon. In Atlantic salmon, there was a negative correlation between kidney bacterial burden and hematocrit, plasma Ca++ and Mg++ values, whereas in infected chum salmon the correlation was positive for Na+ and Cl- and negative for glucose. CONCLUSIONS: A dependency of bacterial shedding on species-specific patterns of pathogenesis was suggested. The coincidence of bacterial shedding with mortality will inform pathogen transmission models.

Development of a Piscirickettsia salmonis immersion challenge model to investigate the comparative susceptibility of three salmon species.

Piscirickettsia salmonis, the aetiological agent of salmonid rickettsial septicaemia (SRS), is a global pathogen of wild and cultured marine salmonids. Here, we describe the development and application of a reproducible, standardized immersion challenge model to induce clinical SRS in juvenile pink (Oncorhynchus gorbuscha), Atlantic (Salmo salar) and sockeye salmon (O. nerka). Following a 1-hr immersion in 105 colony-forming units/ml, cumulative mortality in Atlantic salmon was 63.2% while mortality in sockeye salmon was 10%. Prevalence and levels of the bacterium in kidney prior to onset of mortality were lower in sockeye compared with Atlantic or pink salmon. The timing and magnitude of bacterial shedding were estimated from water samples collected during the exposure trials. Shedding was estimated to be 82-fold higher in Atlantic salmon as compared to sockeye salmon and peaked in the Atlantic salmon trial at 36 d post-immersion. These data suggest sockeye salmon are less susceptible to P. salmonis than Atlantic or pink salmon. Finally, skin lesions were observed on infected fish during all trials, often in the absence of detectable infection in kidney. As a result, we hypothesize that skin is the primary point of entry for P. salmonis during the immersion challenge.

Myosporidium ladogensis n. comb. in burbot Lota lota from Finland: fine structure and microsporidian taxonomy.

Infections with microsporidian parasites are described in skeletal muscle of burbot Lota lota from Lake Haukivesi, Finland. Infected myocytes contained spores within sporophorous vesicles (SPVs) in contact with host cell cytoplasm, similar to Pleistophora ladogensis in L. lota and smelt Osmerus eperlanus in western Russia and northern Germany. Analysis of small subunit ribosomal RNA (SSU rRNA) gene sequences indicated identity with Myosporidium spraguei in burbot and pike-perch from this lake. The latter is considered a junior synonym of P. ladogensis. Phylogenetic analysis of SSU rRNA sequences resolved the burbot parasite apart from a clade containing the type species P. typicalis, but together with M. merluccius. The parasite is renamed Myosporidium ladogensis (Voronin, 1978) n. comb. Networks of tubular appendages arising from developing meronts and SPVs were associated with degradation of host cell cytoplasm.

Factors associated with severity of naturally occurring piscirickettsiosis in netpen- and tank-reared juvenile Atlantic salmon at a research aquarium in western Canada.

The opportunistic examination of factors associated with an outbreak of piscirickettsiosis (SRS) is described in Atlantic salmon Salmo salar post-smolts held in an open netpen or in tanks supplied with raw sea water at a research aquarium in western Canada. During the outbreak, seawater temperature was significantly higher and salinity significantly lower in the netpen compared with the tanks. Mortality in the netpen began approximately 3 weeks prior to that in the tanks, and cumulative mortality in the netpen (34%) was significantly higher than in the tanks (12%). Piscirickettsia salmonis was confirmed by qPCR in tissues from moribund and dead fish and from colonies grown on enriched blood agar medium. Neither P. salmonis nor SRS were observed in salmon held concurrently in UV-irradiated sea water. The elevated mortality was curtailed by treatment with oxytetracycline. These observations further indicate warmer, less saline and periodically hypoxic seawater are risk factors for SRS. UV irradiation of sea water is shown to be a tool for SRS management in fish-holding facilities.

Host size influences prevalence and severity of Kudoa thyrsites (Cnidaria: Myxosporea) infection in Atlantic salmon Salmo salar.

Kudoa thyrsites is a cosmopolitan myxozoan parasite of marine fish. The infection causes an economically important myoliquefaction in farmed Atlantic salmon in British Columbia, Canada. Laboratory exposure of Atlantic salmon smolts to infectious seawater was used to test the hypothesis that infection with K. thyrsites is more severe in age-matched, smaller salmon. In each of 2 trials approximately 4 mo apart, smolts were graded into small (80 and 68 g), medium (117 and 100 g) and large (142 and 157 g) initial weight groups (IWGs) and concurrently exposed to infectious seawater. The effects of IWG and time on fish size and infection severity were assessed by linear mixed-effects models. The fish were screened for infection by histological examination at intervals following exposure. Increases in mean length and weight were statistically significant in all IWG during both trials. The infection was detected in fish in both trials, and in Trial 2, the prevalence was significantly greater in larger fish 1000 degree-days (DD) after exposure. The severity of infection (plasmodia mm-2 muscle) was significantly higher in larger smolts: between medium and large IWGs at 2500 DD in Trial 1 and between small and medium IWGs at 1500 and 2000 DD in Trial 2. The hypothesis is rejected and possible explanations for the greater occurrence of K. thyrsites in larger smolts are discussed.

Synergistic osmoregulatory dysfunction during salmon lice (Lepeophtheirus salmonis) and infectious hematopoietic necrosis virus co-infection in sockeye salmon (Oncorhynchus nerka) smolts.

While co-infections are common in both wild and cultured fish, knowledge of the interactive effects of multiple pathogens on host physiology, gene expression and immune response is limited. To evaluate the impact of co-infection on host survival, physiology and gene expression, sockeye salmon Oncorhynchus nerka smolts were infected with the salmon louse Lepeophtheirus salmonis (V-/SL+), infectious hematopoietic necrosis virus (IHNV; V+/SL-), both (V+/SL+), or neither (V-/SL-). Survival in the V+/SL+ group was significantly lower than the V-/SL- and V-/SL+ groups (p = 0.024). Co-infected salmon had elevated osmoregulatory indicators and lowered haematocrit values as compared to the uninfected control. Expression of 12 genes associated with the host immune response was analysed in anterior kidney and skin. The only evidence of L. salmonis-induced modulation of the host antiviral response was down-regulation of mhc I although the possibility of modulation cannot be ruled out for mx-1 and rsad2. Co-infection did not influence the expression of genes associated with the host response to L. salmonis. Therefore, we conclude that the reduced survival in co-infected sockeye salmon resulted from the osmoregulatory consequences of the sea lice infections which were amplified due to infection with IHNV.

Differential Effects of Adult Salmon Lice Lepeophtheirus salmonis on Physiological Responses of Sockeye Salmon and Atlantic Salmon.

The salmon louse Lepeophtheirus salmonis, a type of sea lice (family Caligidae), is enzootic in marine waters of British Columbia and poses a health risk to both farmed Atlantic Salmon Salmo salar and wild Pacific salmon Oncorhynchus spp. At the adult stage, sea lice infections can often result in severe cutaneous lesions in their salmonid hosts. To evaluate and compare the physiological consequences of adult L. salmonis infections, smolts of Atlantic Salmon and Sockeye Salmon O. nerka were exposed to 2 (low), 6 (medium), or 10 (high) adult female lice/fish. Mean lice abundance decreased over time in all groups. Skin disruption due to parasite infection was observed in both species. Plasma samples were collected from infected fish and uninfected controls at 1, 3, 5, and 7 d postinfection and measured for indicators of osmoregulatory function and stress. Sockeye Salmon, regardless of L. salmonis exposure level, showed a rapid onset of elevated osmolality and sodium and chloride ion concentrations which were sustained until 7 d postinfection when values returned to levels comparable with the unexposed controls. Conversely, these effects were not measured in Atlantic Salmon. Additionally, differential host effects in blood glucose levels were observed, with Sockeye Salmon displaying immediate elevation in glucose. Relative to Atlantic Salmon, infection with L. salmonis caused a profound physiological impact to Sockeye Salmon characterized by loss of osmoregulatory integrity and a stress response. This work provides the first comprehensive report of the physiological consequences of infections with adult L. salmonis in Sockeye Salmon smolts and helps to further define the mechanisms of susceptibility in this species.

Acquired Protective Immunity in Atlantic Salmon Salmo salar against the Myxozoan Kudoa thyrsites Involves Induction of MHIIß+ CD83+ Antigen-Presenting Cells.

The histozoic myxozoan parasite Kudoa thyrsites causes postmortem myoliquefaction and is responsible for economic losses to salmon aquaculture in the Pacific Northwest. Despite its importance, little is known about the host-parasite relationship, including the host response to infection. The present work sought to characterize the immune response in Atlantic salmon during infection, recovery, and reexposure to K. thyrsites After exposure to infective seawater, infected and uninfected smolts were sampled three times over 4,275 degree-days. Histological analysis revealed infection severity decreased over time in exposed fish, while in controls there was no evidence of infection. Following a secondary exposure of all fish, severity of infection in the controls was similar to that measured in exposed fish at the first sampling time but was significantly reduced in reexposed fish, suggesting the acquisition of protective immunity. Using immunohistochemistry, we detected a population of MHIIß+ cells in infected muscle that followed a pattern of abundance concordant with parasite prevalence. Infiltration of these cells into infected myocytes preceded destruction of the plasmodium and dissemination of myxospores. Dual labeling indicated a majority of these cells were CD83+/MHIIß+ Using reverse transcription-quantitative PCR, we detected significant induction of cellular effectors, including macrophage/dendritic cells (mhii/cd83/mcsf), B cells (igm/igt), and cytotoxic T cells (cd8/nkl), in the musculature of infected fish. These data support a role for cellular effectors such as antigen-presenting cells (monocyte/macrophage and dendritic cells) along with B and T cells in the acquired protective immune response of Atlantic salmon against K. thyrsites.

Effects of the vertically transmitted microsporidian Facilispora margolisi and the parasiticide emamectin benzoate on salmon lice (Lepeophtheirus salmonis).

BACKGROUND: Microsporidia are highly specialized, parasitic fungi that infect a wide range of eukaryotic hosts from all major taxa. Infections cause a variety of damaging effects on host physiology from increased stress to death. The microsporidian Facilispora margolisi infects the Pacific salmon louse (Lepeophtheirus salmonis oncorhynchi), an economically and ecologically important ectoparasitic copepod that can impact wild and cultured salmonids. RESULTS: Vertical transmission of F. margolisi was demonstrated by using PCR and in situ hybridization to identify and localize microsporidia in female L. salmonis and their offspring. Spores and developmental structures of F. margolisi were identified in 77% of F1 generation copepods derived from infected females while offspring from uninfected females all tested negative for the microsporidia. The transcriptomic response of the salmon louse to F. margolisi was profiled at both the copepodid larval stage and the pre-adult stage using microarray technology. Infected copepodids differentially expressed 577 transcripts related to stress, ATP generation and structural components of muscle and cuticle. The infection also impacted the response of the copepodid to the parasiticide emamectin benzoate (EMB) at a low dose of 1.0 ppb for 24 h. A set of 48 transcripts putatively involved in feeding and host immunomodulation were up to 8-fold underexpressed in the F. margolisi infected copepodids treated with EMB compared with controls or either stressor alone. Additionally, these infected lice treated with EMB also overexpressed 101 transcripts involved in stress resistance and signalling compared to the other groups. In contrast, infected pre-adult lice did not display a stress response, suggesting a decrease in microsporidian virulence associated with lice maturity. Furthermore, copepodid infectivity and moulting was not affected by the microsporidian infection. CONCLUSIONS: This study demonstrated that F. margolisi is transmitted vertically between salmon louse generations and that biological impacts of infection differ depending on the stage of the copepod host. The infection caused significant perturbations of larval transcriptomes and therefore must be considered in future studies in which impacts to host development and environmental factors are assessed. Fitness impacts are probably minor, although the interaction between pesticide exposure and microsporidian infection merits further study.

Enhanced transcriptomic responses in the Pacific salmon louse Lepeophtheirus salmonis oncorhynchi to the non-native Atlantic Salmon Salmo salar suggests increased parasite fitness.

BACKGROUND: Outcomes of infections with the salmon louse Lepeophtheirus salmonis vary considerably among its natural hosts (Salmo, Oncorhynchus spp.). Host-parasite interactions range from weak to strong host responses accompanied by high to low parasite abundances, respectively. Parasite behavioral studies indicate that the louse prefers the host Atlantic Salmon (Salmo salar), which is characterized by a weak immune response, and that this results in enhanced parasite reproduction and growth rates. Furthermore, parasite-derived immunosuppressive molecules (e.g., proteases) have been detected at higher amounts in response to the mucus of Atlantic Salmon relative to Coho Salmon (Oncorhynchus kisutch). However, the host-specific responses of the salmon louse have not been well characterized in either of the genetically distinct sub-species that occur in the Atlantic and Pacific Oceans. RESULTS: We assessed and compared the transcriptomic feeding response of the Pacific salmon louse (L. salmonis oncorhynchi,) while parasitizing the highly susceptible Atlantic Salmon and Sockeye Salmon (Oncorhynchus nerka) or the more resistant Coho Salmon (Oncorhynchus kisutch) using a 38 K oligonucleotide microarray. The response of the louse was enhanced both in the number of overexpressed genes and in the magnitude of expression while feeding on the non-native Atlantic Salmon, compared to either Coho or Sockeye Salmon. For example, putative virulence factors (e.g., cathepsin L, trypsin, carboxypeptidase B), metabolic enzymes (e.g., cytochrome B, cytochrome C), protein synthesis enzymes (e.g., ribosomal protein P2, 60S ribosomal protein L7), and reproduction-related genes (e.g., estrogen sulfotransferase) were overexpressed in Atlantic-fed lice, indicating heightened parasite fitness with this host species. In contrast, responses in Coho- or Sockeye-fed lice were more similar to those of parasites deprived of a host. To test for host acclimation by the parasite, we performed a reciprocal host transfer experiment and determined that the exaggerated response to Atlantic Salmon was independent of the initial host species, confirming our conclusion that the Pacific salmon louse exhibits an enhanced response to Atlantic Salmon. CONCLUSIONS: This study characterized global transcriptomic responses of Pacific salmon lice during infection of susceptible and resistant hosts. Similar parasite responses during infection of Coho or Sockeye Salmon, despite differences in natural immunity to infection between these host species, indicate that host susceptibility status alone does not drive the parasite response. We identified an enhanced louse response after feeding on Atlantic Salmon, characterized by up-regulation of virulence factors, energy metabolism and reproductive-associated transcripts. In contrast, the responses of lice infecting Coho or Sockeye Salmon were weaker, with reduced expression of virulence factors. These observations indicate that the response of the louse is independent of host susceptibility and suggest that co-evolutionary host-parasite relationships may influence contemporary host-parasite interactions. This research improves our understanding of the susceptibility of Atlantic Salmon and may assist in the development of novel control measures against the salmon louse.

Sex-biased gene expression and sequence conservation in Atlantic and Pacific salmon lice (Lepeophtheirus salmonis).

BACKGROUND: Salmon lice, Lepeophtheirus salmonis (Copepoda: Caligidae), are highly important ectoparasites of farmed and wild salmonids, and cause multi-million dollar losses to the salmon aquaculture industry annually. Salmon lice display extensive sexual dimorphism in ontogeny, morphology, physiology, behavior, and more. Therefore, the identification of transcripts with differential expression between males and females (sex-biased transcripts) may help elucidate the relationship between sexual selection and sexually dimorphic characteristics. RESULTS: Sex-biased transcripts were identified from transcriptome analyses of three L. salmonis populations, including both Atlantic and Pacific subspecies. A total of 35-43 % of all quality-filtered transcripts were sex-biased in L. salmonis, with male-biased transcripts exhibiting higher fold change than female-biased transcripts. For Gene Ontology and functional analyses, a consensus-based approach was used to identify concordantly differentially expressed sex-biased transcripts across the three populations. A total of 127 male-specific transcripts (i.e. those without detectable expression in any female) were identified, and were enriched with reproductive functions (e.g. seminal fluid and male accessory gland proteins). Other sex-biased transcripts involved in morphogenesis, feeding, energy generation, and sensory and immune system development and function were also identified. Interestingly, as observed in model systems, male-biased L. salmonis transcripts were more frequently without annotation compared to female-biased or unbiased transcripts, suggesting higher rates of sequence divergence in male-biased transcripts. CONCLUSIONS: Transcriptome differences between male and female L. salmonis described here provide key insights into the molecular mechanisms controlling sexual dimorphism in L. salmonis. This analysis offers targets for parasite control and provides a foundation for further analyses exploring critical topics such as the interaction between sex and drug resistance, sex-specific factors in host-parasite relationships, and reproductive roles within L. salmonis.

Differential modulation of resistance biomarkers in skin of juvenile and mature pink salmon, Oncorhynchus gorbuscha by the salmon louse, Lepeophtheirus salmonis.

Juvenile pink salmon larger than 0.7 g reject the sea louse, Lepeophtheirus salmonis, and are considered resistant to the infection. Robust innate defense responses in the skin contribute to the observed resistance. In contrast adult pink salmon captured at sea or shortly before spawning carry large numbers of the parasite, suggesting inability to control the infection. The purpose of this research is to better understand these apparently contradictory conclusions by comparing a suite of genetic and cellular markers of resistance to L. salmonis in the skin of juvenile and mature pink salmon. The expression of major histocompatibility factor II, C-reactive protein, interleukin-1ß, interleukin-8 and cyclooxygenase-2 was down-regulated in mature but not juvenile pink salmon. Similarly, skin at the site of parasite attachment in juvenile salmon was highly populated with MHIIß(+) and IL-1ß(+) cells that were either absent, or at reduced levels at similar sites in mature salmon. In addition, mucocyte density was relatively low in the skin of mature salmon, irrespective of louse infection. In juveniles, the higher mucocyte density decreased following louse attachment. We show that in mature pink salmon, genetic and histological responses in skin are depressed and speculate that salmonid defense against L. salmonis is modulated by maturation.

Specific PCR for Myxobolus arcticus SSU rDNA in juvenile sockeye salmon Oncorhynchus nerka from British Columbia, Canada.

A PCR for the specific detection of the salmon brain parasite Myxobolus arcticus (Pugachev and Khokhlov, 1979) was developed using primers designed to amplify a 1363 base pair fragment of the small subunit rDNA. The assay did not amplify DNA from 5 other Myxobolus species or from 7 other myxozoan species belonging to 5 other genera. For juvenile sockeye salmon Oncorhynchus nerka (Walbaum) collected from Chilko Lake, British Columbia (BC), Canada, in 2011, the prevalence by PCR was 96%, in contrast to 71% by histological examination of brain tissue. In 2010, the histological prevalence was 52.5%. Sequence identity between M. arcticus from Chilko Lake and other sites in BC ranged from 99.7 to 99.8% and was 99.6% for a Japanese sequence. In contrast, an M. arcticus sequence from Norway shared 95.3% identity with the Chilko Lake sequence, suggesting misidentification of the parasite. Chilko Lake sockeye salmon were previously reported free of infection with M. arcticus, and more research is required to understand the processes involved in the local and global dispersion of this parasite.

Transcriptomic responses to emamectin benzoate in Pacific and Atlantic Canada salmon lice Lepeophtheirus salmonis with differing levels of drug resistance.

Salmon lice Lepeophtheirus salmonis are an ecologically and economically important parasite of wild and farmed salmon. In Scotland, Norway, and Eastern Canada, L. salmonis have developed resistance to emamectin benzoate (EMB), one of the few parasiticides available for salmon lice. Drug resistance mechanisms can be complex, potentially differing among populations and involving multiple genes with additive effects (i.e., polygenic resistance). Indicators of resistance development may enable early detection and countermeasures to avoid the spread of resistance. Here, we collect sensitive Pacific L. salmonis and sensitive and resistant Atlantic L. salmonis from salmon farms, propagate in laboratory (F1), expose to EMB in bioassays, and evaluate either baseline (Atlantic only) or induced transcriptomic differences between populations. In all populations, induced responses were minor and a cellular stress response was not identified. Pacific lice did not upregulate any genes in response to EMB, but downregulated degradative enzymes and transport proteins at 50 ppb EMB. Baseline differences between sensitive and now resistant Atlantic lice were much greater than responses to exposures. All resistant lice overexpressed degradative enzymes, and resistant males, the most resistant group, overexpressed collagenases to the greatest extent. These results indicate an accumulation of baseline expression differences related to resistance.

Signatures of resistance to Lepeophtheirus salmonis include a TH2-type response at the louse-salmon interface.

Disease outbreaks with the salmon louse Lepeophtheirus salmonis cause significant economic losses in mariculture operations worldwide. Variable innate immune responses at the louse-attachment site contribute to differences in susceptibility among species such that members of Salmo spp. are more susceptible to infection than those of some Oncorhynchus spp. Relatively little is known about the mechanisms that contribute to disease resistance or susceptibility to L. salmonis in salmon. Here, we utilize histochemistry and transcriptomics in a comparative infection model with susceptible (Atlantic, sockeye) and resistant (coho) salmon. At least three cell populations (MHIIß+, IL1ß+, TNFα+) were activated in coho salmon skin during L. salmonis infection. Locally elevated expression of several pro-inflammatory mediators (e.g. IL1ß, IL8, TNFα, COX2, C/EBPß), and tissue repair enzymes (MMP9, MMP13) were detected in susceptible and resistant species. However, responses specific to coho salmon (e.g. IL4, IL6, TGFß) or responses shared among susceptible salmon (e.g. SAP, TRF, Cath in Atlantic and sockeye salmon) provide evidence for species-specific pathways contributing to resistance or susceptibility, respectively. Our results confirm the importance of an early pro-inflammatory TH1-type pathway as an initial host response during infection with Pacific sea lice, and demonstrate subsequent regulatory TH2-type processes as candidate defense mechanisms in the skin of resistant coho salmon.

Detection of a parasitic amoeba (Order Dactylopodida) in the female gonads of oysters in Brazil.

The impacts of oocyte parasites on the reproductive success of molluscs are largely unknown. In this study, we evaluated the presence of gonad parasites in 6 species of marine bivalve molluscs native to southern Brazil. Cultured bivalves included the mangrove oyster Crassostrea gasar (sometimes called C. brasiliana), the brown mussel Perna perna, the lion's paw scallop Nodipecten nodosus and the wing pearl oyster Pteria hirundo. Another species of mangrove oyster, C. rhizophorae, and the carib pointed venus clam Anomalocardia brasiliana (syn. A. flexuosa) were collected from the wild. Molluscs were collected in winter 2009 and summer 2010 for histopathological and molecular evaluation. An unknown ovarian parasite (UOP) was observed in histopathological sections of female gonads of C. gasar and C. rhizophorae. The UOP possessed features suggestive of amoebae, including an irregular outer membrane, frothy cytoplasm, a nucleus with a prominent central nucleolus and a closely associated basophilic parasome. PCR analysis was negative for Marteilioides chungmuensis, Perkinsus spp. and Paramoeba perurans. However, real-time PCR successfully amplified DNA from oyster gonads when using universal Paramoeba spp. primers. Also, conventional PCR amplified DNA using primers specific for Perkinsela amoebae-like organisms (syn. Perkinsiella), which are considered as endosymbionts of Parameoba spp., previously thought to be the parasome. Our results suggest that this UOP is a species of amoeba belonging to 1 of the 2 families of the order Dactylopodida, possibly related to Paramoeba spp. This study represents the first report of this type of organism in oysters. We found that C. gasar and C. rhizophorae were the most susceptible molluscs to these UOPs.