Evidence that infectious stages of Tetracapsula bryosalmonae for rainbow trout Oncorhynchus mykiss are present throughout the year

Proliferative kidney disease (PKD) is a hyperplastic condition of the lymphoid tissue of salmonids infected with the spores of Tetracapsula bryosalmonae, a myxozoan parasite formerly designated PKX, which has recently been described as a parasite of several species of bryozoans. The occurrence of PKD is generally associated with seasonal increase in water temperature, with research indicating that transmission of the disease does not occur below 12 to 13 degrees C. This suggested that the infectious stages are absent from about November to March/April. Here we document the transmission of PKD at water temperatures and seasons previously considered to be non permissive for PKD infection. The exposure of naive rainbow trout Oncorhynchus mykiss (Walbaum) to PKD-infected water ranging from 8 to 13 degrees C during the Autumn, Winter and early Spring, resulted in the infection of kidney interstitium once the trout were transferred to 16 degrees C. In addition, cohabitation studies were conducted with the bryozoan host Fredericella sultana collected from a river at times of low seasonal temperatures because this bryozoan species overwinters as living colonies. Cohabitation of trout with colonies of F sultana in parasite-free city water at 16 degrees C, also led to renal lymphoid tissue infection with the parasite and even to nephromegaly. Our results provide evidence that the infectious stages of T bryosalmonae for rainbow trout were present in the water throughout the entire year and that the impact of temperature on the development of PKD is primarily a result of the kinetics of Tetracapsula multiplication in bryozoan and fish hosts.     

Kidney pathology and parasite intensity in rainbow trout Oncorhynchus mykiss surviving proliferative kidney disease: time course and influence of temperature

Proliferative kidney disease (PKD) is an endoparasitic disease of salmonids caused by the myxozoan parasite Tetracapsuloides bryosalmonae. We recently described the development of the disease from initial infection until manifestation of clinical disease signs in rainbow trout held at 2 water temperatures, 12 and 18°C. The aim of the present study is to investigate whether (1) infected fish surviving the clinical phase would recover from renal pathological changes, (2) whether they would be able to reduce the parasite load in the kidneys, and (3) whether water temperatures would influence renal recovery and parasite clearance. At 18°C, fish showed a gradual recovery of normal kidney morphology which was associated with a decline in parasite numbers and infection prevalence. Fish kept at 12°C initially showed an enhancement of kidney lesions before recovery of normal kidney morphology took place. The decrease in renal parasite load was retarded compared to 18°C. The results from the present study provide evidence that rainbow trout surviving the clinical phase of PKD are able to (1) fully restore renal structure, and (2) significantly reduce renal parasite loads, although 100% clearance was not achieved within the experimental period of this study. Water temperature influences the rate but not the outcome of the recovery process.     

Transmission of Tetracapsuloides bryosalmonae (Myxozoa: Malacosporea) to Fredericella sultana (Bryozoa: Phylactolaemata) by various fish species

Tetracapsuloides bryosalmonae is a myxozoan parasite of salmonids and freshwater bryozoans, which causes proliferative kidney disease (PKD) in the fish host. To test which fish species are able to transmit T. bryosalmonae to bryozoans, an infection experiment was conducted with 5 PKD-sensitive fish species from different genera. Rainbow trout Oncorhynchus mykiss, brown trout Salmo trutta, brook trout Salvelinus fontinalis, grayling Thymallus thymallus and northern pike Esox lucius were cohabitated with T. bryosalmonae-infected Fredericella sultana colonies and then subsequently cohabitated with statoblast-reared parasite free Bryozoa. Statoblasts from infected colonies were tested by PCR to detect cryptic stages of T. bryosalmonae, which may indicate vertical transmission of the parasite. In this study, brown trout and brook trout were able to infect Bryozoa, while there was no evidence that rainbow trout and grayling were able to do so. Few interstitial kidney stages of the parasite were detected by immunohistochemistry in brown trout and brook trout, while rainbow trout and grayling showed marked proliferation of renal interstitial tissue and macrophages with numerous parasite cells. Intraluminal stages in the kidney tubules were only detected in brown trout and rainbow trout. In contrast to previous observations, pike was not susceptible to PKD in these trials according to the results of T. bryosalmonae-specific PCR. No DNA of T. bryosalmonae was detected in any statoblast.     

The phylogeography of salmonid proliferative kidney disease in Europe and North America

Salmonid proliferative kidney disease (PKD) is caused by the myxozoan Tetracapsuloides bryosalmonae. Given the serious and apparently growing impact of PKD on farmed and wild salmonids, we undertook a phylogeographic study to gain insights into the history of genealogical lineages of T. bryosalmonae in Europe and North America, and to determine if the global expansion of rainbow trout farming has spread the disease. Phylogenetic analyses of internal transcribed spacer 1 sequences revealed a clade composed of all North American sequences plus a subset of Italian and French sequences. High genetic diversity in North America and the absence of genotypes diagnostic of the North American clade in the rest of Europe imply that southern Europe was colonized by immigration from North America; however, sequence divergence suggests that this colonization substantially pre-dated fisheries activities. Furthermore, the lack of southern European lineages in the rest of Europe, despite widespread rainbow trout farming, indicates that T. bryosalmonae is not transported through fisheries activities. This result strikingly contrasts with the commonness of fisheries-related introductions of other pathogens and parasites and indicates that fishes may be dead-end hosts. Our results also demonstrate that European strains of T. bryosalmonae infect and induce PKD in rainbow trout introduced to Europe.     

Temperature-driven proliferation of Tetracapsuloides bryosalmonae in bryozoan hosts portends salmonid declines

Proliferative kidney disease (PKD) is an emerging disease of salmonid fishes. It is provoked by temperature and caused by infective spores of the myxozoan parasite Tetracapsuloides bryosalmonae, which develops in freshwater bryozoans. We investigated the link between PKD and temperature by determining whether temperature influences the proliferation of T. bryosalmonae in the bryozoan host Fredericella sultana. Herein we show that increased temperatures drive the proliferation of T. bryosalmonae in bryozoans by provoking, accelerating and prolonging the production of infective spores from cryptic stages. Based on these results we predict that PKD outbreaks will increase further in magnitude and severity in wild and farmed salmonids as a result of climate-driven enhanced proliferation in invertebrate hosts, and urge for early implementation of management strategies to reduce future salmonid declines.     

Severe mortality in wild Atlantic salmon Salmo salar due to proliferative kidney disease (PKD) caused by Tetracapsuloides bryosalmonae (myxozoa)

Extensive mortality in Atlantic salmon fry was reported in the River Aelva from 2002 to 2004. Dead fish were collected in late summer 2006, and live fish were sampled by electrofishing in September the same year. At autopsy and in histological sections, the fish kidneys were found to be pale and considerably enlarged. Proliferative lesions with characteristic PKX cells were seen in a majority of the fish. DNA from kidney samples of diseased fish was subjected to PCR and sequencing, and the amplified sequences matched those of Tetracapsuloides bryosalmonae. We concluded that this myxozoan transmitted from bryozoans was the main cause of the observed mortality in salmon fry in 2006. Results from quantitative electrofishing in 2005 and 2006, combined with the observed fry mortality from 2002 to 2004, show that the smolt production in the river is severely reduced and that T. bryosalmonae is the most likely explanation for this decline. The present study is the first to report a considerable negative population effect in wild Atlantic salmon due to proliferative kidney disease (PKD). It also represents the northernmost PKD outbreak in wild fish. The river is regulated for hydroelectric power purposes, causing reduced water flow and elevated summer temperatures, and the present PKD outbreak may serve as an example of increased disease vulnerability of northern fish populations in a warmer climate.     

Seasonal occurrence of the infectious stage of proliferative kidney disease (PKD) and resistance of rainbow trout, Salmo gairdneri Richardson, to reinfection

The seasonality of proliferative kidney disease (PKD) in rainbow trout, Salmo gairdneri Richardson, at the American River Hatchery, California was found to be primarily dependent on the presence or absence of the infectious stage in the water supply. This was determined by introducing sentinel trout into the hatchery water supply on a monthly basis, followed by their transfer to the laboratory for subsequent holding in 18° C, pathogen-free water for one month prior to examination. These exposures demonstrated that infections were obtained from April through October at ambient temperatures 12–20° C. Trout which had recovered from clinical infections were found to be resistant to reinfection. Resistance was induced by active infection and not just previous exposure to the infectious stage. Trout surviving PKD were also found to harbour later sporogonic stages of the parasite for at least one year following initial infection, but fully formed spores, as judged by well-developed valves, were not observed.     

PKX, the causative agent of proliferative kidney disease (PKD) in Pacific salmonid fishes and its affinities with the Myxozoa

Proliferative kidney disease (PKD), caused by an unclassified protozoan (PKX), is reported from Pacific salmon, Oncorhynchus tshawytscha (Walbaum) and O. kisutch (Walbaum), and steelhead trout, Salmo gairdneri Richardson, held at the Mad River Hatchery in California, USA. The cumulative mortality attributed to the disease was 95, 13, and 18% respectively. The mortalities were greatest at mean water temperatures of 12-14 degrees C during July 1983. The ultrastructure of the PKX organism and its associated pathology during clinical disease in all three species were consistent with those of the parasite in rainbow trout (Salmo gairdneri) as described in European outbreaks. Significant mortalities did not occur after August, at which time the parasite could no longer be detected in the salmon species. The steelhead continued to exhibit parasites in the kidney interstitium and epithelium and lumens of the tubules. Myxosporidan trophozoites and developing spores were also observed in the lumens of the kidney tubules of these fish. Although a mixed infection with another parasite may have occurred, evidence suggests that the myxosporidans are later stages of PKX. They were only observed in fish exposed to water with the infective stage and were particularly prominent in recovering fish. The PKX organism is similar to UBO, an unclassified protozoan of carp suspected to be an early stage of Sphaerospora renicola Dykovà & Lom. Both parasites infect the blood and kidney, divide by endogeny, and are released by disintegration of the primary mother cell. The intraluminal myxosporean forms show similarities to Sphaerospora spp. in that they are monosporous and sporoblasts are formed within pseudoplasmodia.(ABSTRACT TRUNCATED AT 250 WORDS)     

Proliferative, presaccular stages of Tetracapsuloides bryosalmonae (Myxozoa: Malacosporea) within the invertebrate host Fredericella sultana (Bryozoa: Phylactolaemata)

Proliferative kidney disease (PKD), caused by the malacosporean parasite, Tetracapsuloides bryosalmonae, is a major disease of salmonid culture both in western Europe and North America. The fish are infected from spores that develop within freshwater bryozoans and are released into the water column. Although sporogenesis has been studied in the bryozoan host and occurs within sacs, the formation of these sacs from presaccular stages has only been hypothesized. Examination of infected bryozoans by using a range of techniques identified proliferating, presaccular amoeboid stages of T. bryosalmonae on the body wall of the bryozoan Fredericella sultana. These stages possessed unique electron-dense bodies and were observed as aggregating within the bryozoan metacoel, differentiating to form spore sacs. Spore sac growth was associated with the assimilation of the presaccular parasites rather than through cryptomitosis of sac mural cells. This sac formation through aggregation and assimilation suggests an intriguing mechanism by which T. bryosalmonae can cross-fertilize.     

Fish infected with Sphaerospora spp. Thélohan (Myxosporea) from waters enzootic for proliferative kidney disease of salmonids

Sphaerospores were found among three species of fish examined from waters known to be enzootic for proliferative kidney disease (PKD) of salmonids. They were detected in the renal tubules of both hatchery-reared rainbow trout (Salmo gairdneri) exposed to the infectious stage of PKD and in chubs (Gila bicolor) in the headwaters of a hatchery where PKD is enzootic. Sticklebacks (Gasterosteus aculeatus) collected near net pens where Pacific salmon had experienced a PKD epizootic were also found to harbor sphaerospores in the lumen of the kidney tubules. The latter two host species contained developmental stages of a myxosporidan in the blood and in the lumen of the kidney tubules which are similar to those of PKX, the causative agent of PKD in salmonid fish. The sphaerospores observed in the rainbow trout are the first to be observed in this species. The similarity to previously observed developmental stages, rarity, and presence of these sphaerospores in salmonid fish from a hatchery where PKD is enzootic suggest that they are the most mature stage of the PKX myxosporidan yet observed.     

Effects of fish age versus size on the development of whirling disease in rainbow trout

We examined the effects of both fish age and size on the development of resistance to whirling disease in Erwin strain rainbow trout. Previously, we demonstrated that juvenile rainbow trout became resistant to development of the disease when first exposed to triactinomyxons of the parasite Myxobolus cerebralis at about 9 wk post-hatch when raised at 12 degrees C, but ages and sizes of fish used in that experiment were confounded (Ryce EKN, Zale AV, MacConnell E [2004] Dis Aquat Org 59:225-233). In this study, rainbow trout of the same age and different sizes, and the same size and different ages, were exposed to the parasite to distinguish the influences of age and size. Fish were reared at 3 different water temperatures prior to exposure to produce groups with different growth rates and were exposed to the parasite at 7 or 9 wk post-hatch. Disease severity was affected by both age and size at first exposure, but the effects were not independent. An increase in fork length from 36 to 40 mm among fish exposed at 7 wk post-hatch did not confer increased resistance, but the same increase in size at 9 wk post-hatch did. Similarly, an increase in age from 7 to 9 wk post-hatch among fish exposed at 36 mm fork length did not confer increased resistance, but the same increase in age at 40 mm did. Rainbow trout must be both 9 wk post-hatch or older and at least 40 mm in fork length at time of exposure to exhibit enhanced resistance to whirling disease. Resistance to disease was not associated with the level of skeletal ossification.     

Fish Infected with Sphaerospora spp. Thélohan (Myxosporea) from Waters Enzootic for Proliferative Kidney Disease of Salmonids1

ABSTRACT. Sphaerospores were found among three species of fish examined from waters known to be enzootic for proliferative kidney disease (PKD) of salmonids. They were detected in the renal tubules of both hatchery-reared rainbow trout (Salmo gairdneri) exposed to the infectious stage of PKD and in chubs (Gila bicolor) in the headwaters of a hatchery where PKD is enzootic. Sticklebacks (Gasterosteus aculeatus) collected near net pens where Pacific salmon had experienced a PKD epizootic were also found to harbor sphaerospores in the lumen of the kidney tubules. The latter two host species contained developmental stages of a myxosporidan in the blood and in the lumen of the kidney tubules which are similar to those of PKX, the causative agent of PKD in salmonid fish. The sphaerospores observed in the rainbow trout are the first to be observed in this species. The similarity to previously observed developmental stages, rarity, and presence of these sphaerospores in salmonid fish from a hatchery where PKD is enzootic suggest that they are the most mature stage of the PKX myxosporidan yet observed.     

PKX,the Causative Agent of Proliferative Kidney Disease (PKD) in Pacific Salmonid Fishes and Its Affinities with the Myxozoa1

Proliferative kidney disease (PKD), caused by an unclassified protozoan (PKX), is reported from Pacific salmon, Oncorhynchus tshawytscha (Walbaum) and O. kisutch (Walbaum), and steelhead trout, Salmo gairdneri Richardson, held at the Mad River Hatchery in California, USA. The cumulative mortality attributed to the disease was 95, 13, and 18% respectively. The mortalities were greatest at mean water temperatures of 12-14°C during July 1983. The ultrastructure of the PKX organism and its associated pathology during clinical disease in all three species were consistent with those of the parasite in rainbow trout (Salmo gairdneri) as described in European outbreaks. Significant mortalities did not occur after August, at which time the parasite could no longer be detected in the salmon species. The steelhead continued to exhibit parasites in the kidney interstitium and epithelium and lumens of the tubules. Myxosporidan trophozoites and developing spores were also observed in the lumens of the kidney tubules of these fish. Although a mixed infection with another parasite may have occurred, evidence suggests that the myxosporidans are later stages of PKX. They were only observed in fish exposed to water with the infective stage and were particularly prominent in recovering fish. The PKX organism is similar to UBO, an unclassified protozoan of carp suspected to be an early stage of Sphaerospora renicola Dyková & Lom. Both parasites infect the blood and kidney, divide by endogeny, and are released by disintegration of the primary mother cell. The intraluminal myxosporean forms show similarities to Sphaerospora spp. in that they are monosporous and sporoblasts are formed within pseudoplasmodia. It is possible that PKX migrates to the lumen of the kidney tubule and subsequently sporulates. If the myxosporean forms are later stages of PKX, then it would belong to the phlyum Myxozoa.     

Sporogony of Tetracapsuloides bryosalmonae in the brown trout Salmo trutta and the role of the tertiary cell during the vertebrate phase of myxozoan life cycles

Tetracapsuloides bryosalmonae is the myxozoan that causes the commercially and ecologically important proliferative kidney disease of salmonid fish species. Immunohistochemistry and electron microscopy were used to examine the development of this parasite within the kidney of the brown trout Salmo trutta. The main replicative phase of T. bryosalmonae is a cell doublet composed of a primary cell and a single secondary cell. Engulfment of one secondary cell by another to form a secondary-tertiary doublet (S-T doublet) heralded the onset of sporogony whereupon the parasite migrated to the kidney tubule lumen. Within the tubule, the parasite transformed into a pseudoplasmodium and anchored to the tubule epithelial cells via pseudopodial extensions. Within each pseudoplasmodium developed a single spore, composed of 4 valve cells, 2 polar capsules and 1 sporoplasm. The pseudoplasmodia formed clusters suggesting that large numbers of spores develop within the fish. This examination of T. bryosalmonae suggests that the main replicative phase of freshwater myxozoans within vertebrates is via direct replication of cell doublets rather than through the rupturing of extrasporogonic stages, while tertiary cell formation relates only to sporogony. Taken in conjunction with existing phylogenetic data, 5 distinct sporogonial sequences are identified for the Myxozoa.     

Occurrence of Flavobacterium psychrophilum in fish-farming environments

Occurrence of Flavobacterium psychrophilum in fish farms and fish-farming environments was studied using agar plate cultivation, the immunoflourescence antibody technique (IFAT) and nested PCR. Characteristics of 64 F. psychrophilum isolates from rainbow trout Oncorhynchus mykiss, fish farm rearing water, ovarian fluid and wild fish were serotyped, ribotyped and compared biochemically. Virulence of F. psychrophilum isolates from different sources was compared by injection into rainbow trout. Additionally, the number of F. psychrophilum cells shed by naturally infected rainbow trout was determined. F. psychrophilum was detected and isolated from skin mucus, skin lesions and internal organs of diseased rainbow trout and from fish without clinical disease. The pathogen was also present in wild perch Perca fluviatilis, roach Rutilus rutilus, and ovarian fluids of farmed rainbow trout brood fish. Isolates were biochemically homogenous, excluding the capability to degrade elastin. Five different agglutination patterns with different antisera against F. psychrophilum were found among the isolates studied. Although several different ribopatterns were found (ClaI: 12 ribopatterns and HaeIII: 9 ribopatterns), ribotype A was the most dominant. Farmed rainbow trout brood fish carried a broad-spectrum of serologically and genetically different F. psychrophilum in ovarian fluids. Virulence of the tested isolates in rainbow trout varied and naturally infected rainbow trout shed 10(4) to 10(8) cells fish(-1) h(-1) of F. psychrophilum into the surrounding water.     

Distribution of the flagellate Hexamita salmonis Moore, 1922 and the microsporidian Loma salmonae Putz, Hoffman and Dunbar, 1965 in brown trout, Salmo trutta L., and rainbow trout, Salmo gairdneri Richardson, in the River Itchen (U.K.) and three of its fish farms

The occurrence of Hexamita salmonis Moore, 1922 and Loma salmonae Putz, Hoffman and Dunbar, 1965 was investigated at 10 sites on the R. Itchen (five for brown trout only, three for rainbow trout only, and two for both brown trout and rainbow trout) and at three of its nine fish farms (two for rainbow trout, one for brown trout). Hexamita salmonis was recorded in brown trout from three river sites and the farm, and in rainbow trout from both farms and four river sites. Prevalence of Hexamita salmonis in farmed rainbow trout was higher than in farmed brown trout and was consistent with the former species being more susceptible to infection. H. salmonis was at significantly higher prevalence in rainbow trout from farm no. 5 than farm no. 2 for three size classes of fish. In wild brown trout and feral rainbow trout, the highest prevalences of H. salmonis were recorded at sites in the vicinity of farm no. 2. This distribution was consistent with an area of naturally high infection levels, and with infected fish unintentionally released from farm no. 2 serving as a source of infection, the infection subsequently becoming established in the river fish. Loma salmonae was recorded in wild brown trout and in rainbow trout from both farms. This appears to be the first recording of this parasite from British salmonids and also the first recording of the parasite from brown trout. The distribution of the parasite (particularly the prevalence being higher at farm no. 2 than farm no. 5) was consistent with it being introduced into the R. Itchen via rainbow trout from farm no. 2 (and probably no. 3) much of whose stock derived from imported Californian 'Shasta' rainbow trout.     

Ultraviolet irradiation inactivates the waterborne infective stages of Myxobolus cerebralis: a treatment for hatchery water supplies

The effects of ultraviolet (UV) irradiation on the viability of the waterborne triactinomyxon stages of Myxobolus cerebralis were evaluated by vital staining and the infectivity for juvenile rainbow trout Oncorhynchus mykiss. A dose of 1300 mWs cm-2 was required to inactivate 100% of the triactinomyxons held under a static collimated beam of UV as determined by vital staining. Juvenile rainbow trout were protected from infections with M. cerebralis when exposed to 14,000 or 1400 triactinomyxon spores per fish that had been treated with the collimating beam apparatus (1300 mWs cm-2). Among all fish receiving UV-treated triactinomyxons, none had clinical signs of whirling disease, or evidence of microscopic lesions or spores of M. cerebralis after 5 mo at water temperatures of 15 degrees C. In contrast, 100% of the fish receiving the higher dose of untreated triactinomyxons developed clinical signs of whirling disease and both microscopic signs of infection and spores were detected in all of the high and low dose trout receiving untreated triactinomyxon exposures. Two additional trials evaluated the Cryptosporidium Inactivation Device (CID) for its ability to treat flow-through 15 degrees C well water to which triactinomyxons were added over a 2 wk period. CID treatments of a cumulative dose exceeding 64,000 triactinomyxons per fish protected juvenile rainbow from infections with M. cerebralis. Rainbow trout controls receiving the same number of untreated triactinomyxons developed both microscopic lesions and cranial spore concentrations up to 10(4.6) per 1/2 head, although no signs of clinical whirling disease were observed. UV (126 mWs cm-2, collimated beam apparatus) was also effective in killing Flavobacterium psychrophilum, the agent causing salmonid bacterial coldwater disease, as demonstrated by the inability of bacterial cells to grow on artificial media following UV treatment.     

Effects of fish age and parasite dose on the development of whirling disease in rainbow trout

We determined the ages at which juvenile rainbow trout Oncorhynchus mykiss became resistant to the effects of whirling disease following exposure to a range of parasite doses. Heretofore, the development and severity of whirling disease in salmonids was known to be generally dependent on the age or size of fish when first exposed to the triactinomyxon spores of Myxobolus cerebralis; larger, older individuals tended to be less diseased. However, no systematic determination had been made of the exact age at which fish become resistant to the development of the disease. We exposed rainbow trout at 9 ages (1 to 17 wk post-hatch) to 4 parasite dose levels (0, 100, 1000 and 10,000 triactinomyxons per fish). Disease severity was measured using mortality, clinical signs, microscopic pathology, and myxospore counts. Disease and mortality were substantially reduced when exposure to the parasite occurred for the first time at 9 wk post-hatch (756 degree-days at 12 degrees C) or older. High doses elicited more disease among the younger age groups, but the effect was dampened in groups exposed at about 9 to 11 wk post-hatch and absent thereafter. Rainbow trout reared in M. cerebralis-free waters for 9 wk post-hatch or longer, whether in the wild or in a hatchery situation, should experience greater survival and less disease than fish first exposed to the parasite at younger ages.     

Invasion and initial replication of ultraviolet irradiated waterborne infective stages of Myxobolus cerebralis results in immunity to whirling disease in rainbow trout

Myxobolus cerebralis is a microscopic metazoan parasite (Phylum Myxozoa: Myxosporea) associated with salmonid whirling disease. There are currently no vaccines to minimise the serious negative economical and ecological impacts of whirling disease among populations of salmonid fish worldwide. UV irradiation has been shown to effectively inactivate the waterborne infective stages or triactinomyxons of M. cerbralis in experimental and hatchery settings but the mechanisms by which the parasite is compromised are unknown. Treatments of triactinomyxons with UV irradiation at doses from 10 to 80 mJ/cm(2) either prevented (20-80 mJ/cm(2)) or significantly inhibited (10 mJ/cm(2)) completion of the parasite life cycle in experimentally exposed juvenile rainbow trout (Oncorhynchus mykiss). However, even the highest doses of UV irradiation examined (80 mJ/cm(2)) did not prevent key steps in the initiation of parasite infection, including attachment and penetration of the epidermis of juvenile rainbow trout as demonstrated by scanning electron and light microscopy. Furthermore, replication of UV-treated parasites within the first 24h following invasion of the caudal fin was suggested by the detection of concentrations of parasite DNA by quantitative PCR comparable to that among fish exposed to an equal concentration of untreated triactinomyxons. Subsequent development of parasites treated with an 80 mJ/cm(2) dose of UV irradiation however, was impaired as demonstrated by the decline and then lack of detection of parasite DNA; a trend beginning at 10 days and continuing thereafter until the end of the study at 46 days post parasite exposure. Treatments of triactinomyxons with a lower dose of UV irradiation (20 mJ/cm(2)) resulted in a more prolonged survival with parasite DNA detected, although at very low concentrations, in fish up to 49 days post parasite exposure. The successful invasion but only short-term survival of parasites treated with UV in rainbow trout resulted in a protective response to challenges with fully infective triactinomyxons. Prior treatments of juvenile rainbow trout with UV-treated triactinomyxons (10 and 20 mJ/cm(2)) resulted in a reduced prevalence of infection and significantly lower concentrations of cranial myxospores (two direct measures of the severity of whirling disease) compared with trout receiving no prior treatments when assessed 5 months post parasite exposure to fully infective triactinomyxons.