Myxobolus cerebralis Causes Presporogonic Mortality in Juvenile Mountain Whitefish.

Recent range expansions of whirling disease impelled us to understand the impacts of its causative agent, the myxozoan parasite Myxobolus cerebralis, on lesser-studied fish hosts. Mountain Whitefish Prosopium williamsoni overlap broadly with M. cerebralis across the western United States and Canada, and populations have experienced widespread declines since the 1990s. To evaluate effects of the parasite on Mountain Whitefish, we revisit formerly unpublished work of the Colorado Division of Wildlife (now Colorado Parks and Wildlife), comparing infection in age-matched Mountain Whitefish, Rainbow Trout Oncorhynchus mykiss, and Brown Trout Salmo trutta. To complement the original report, we reanalyze mortality data and include additional SEM imagery. Infection of M. cerebralis in juvenile Mountain Whitefish was characterized by a brief but heavy period of mortality in the first 2 weeks after exposure, with limited pathology. This clinical effect is unique among the known salmonid hosts of M. cerebralis.

Accelerated deactivation of Myxobolus cerebralis myxospores by susceptible and non-susceptible Tubifex tubifex.

In the 1990s, the Tubifex tubifex aquatic oligochaete species complex was parsed into 6 separate lineages differing in susceptibility to Myxobolus cerebralis, the myxozoan parasite that can cause whirling disease (WD). Lineage III T. tubifex oligochaetes are highly susceptible to M. cerebralis infection. Lineage I, IV, V and VI oligochaetes are highly resistant or refractory to infection and may function as biological filters by deactivating M. cerebralis myxospores. We designed a 2-phased laboratory experiment using triactinomyxon (TAM) production as the response variable to test that hypothesis. A separate study conducted concurrently demonstrated that M. cerebralis myxospores held in sand and water at temperatures ≤15°C degrade rapidly, becoming almost completely non-viable after 180 d. Those results provided the baseline to assess deactivation of M. cerebralis myxospores by replicates of mixed lineage (I, III, V and VI) and refractory lineage (V and VI) oligochaetes. TAM production was zero among 7 of 8 Lineage V and Lineage VI T. tubifex oligochaete groups exposed to 12500 M. cerebralis myxospores for 15, 45, 90 and 135 d. Among 4 mixed lineage exposure groups, TAM production averaged 14641 compared with 2202495 among 12 groups of Lineage III oligochaetes. Among the 6 unexposed Lineage III experimental groups seeded into original Phase 1 substrates for the 45, 90 and 135 d treatments during the Phase 2 portion of the study, TAM production was reduced by 98.9, 99.9 and 99.9%, respectively, compared with the average for the 15 d exposure groups. These results are congruent with the hypothesis that Lineage V and Lineage VI T. tubifex oligochaetes can deactivate and destroy M. cerebralis myxospores.

Assessment of the Long-Term Viability of the Myxospores of Myxobolus cerebralis as Determined by Production of the actinospores by Tubifex tubifex.

While whirling disease was first observed in Rainbow Trout Oncorhynchus mykiss in 1893, the complete life cycle of Myxobolus cerebralis (Mc), the causative agent of the disease, was not understood until 1984, when it was shown to involve two obligate hosts, a salmonid fish and the aquatic oligochaete Tubifex tubifex (Tt). The viability of the triactinomyxon (TAM) actinospores produced by Tt has been well studied, and is known to be temperature dependent and measured in days and weeks. Assertions that Mc myxospores produced by infected fish remain viable for years or even decades were made during the mid-20th century, decades before the Mc life cycle was described. Moreover, the duration of myxospore viability has not been well studied since the life cycle was elucidated. In a series of time-delay treatments, we assessed the long-term viability of Mc myxospores by exposure to Mc-susceptible Tt oligochaetes and quantified TAM production. As the time delay between inoculation and incubation of Mc myxospores in sand and water and exposure to Tt oligochaetes increased, TAM production decreased exponentially. Production among the 15-d time-delay replicates was reduced 74.7% compared with the 0-d treatment. Likewise, total TAM production was reduced 94.5, 99.4, and 99.9%, respectively, in the 90-, 120-, and 180-d time-delay treatments. Linear regression analysis of our data and the absence of TAM production among replicates of Mc myxospores held at 5°C for 365 d prior to exposure to Mc-susceptible Tt oligochaetes indicate that the long-term viability of Mc myxospores is less than 1 year under the conditions of this study.

Survival and reproduction of Myxobolus cerebralis-resistant rainbow trout introduced to the Colorado river and increased resistance of age-0 progeny.

Myxobolus cerebralis caused severe declines in rainbow trout populations across Colorado following its introduction in the 1980s. One promising approach for the recovery of Colorado's rainbow trout populations has been the production of rainbow trout that are genetically resistant to the parasite. We introduced one of these resistant crosses, known as the GR×CRR (cross between the German Rainbow [GR] and Colorado River Rainbow [CRR] trout strains), to the upper Colorado River. The abundance, survival, and growth of the stocked GR×CRR population was examined to determine if GR×CRRs had contributed offspring to the age-0 population, and determine whether these offspring displayed increased resistance and survival characteristics compared to their wild CRR counterparts. Apparent survival of the introduced GR×CRR over the entire study period was estimated to be 0.007 (±0.001). Despite low survival of the GR×CRRs, age-0 progeny of the GR×CRR were encountered in years 2008 through 2011. Genetic assignments revealed a shift in the genetic composition of the rainbow trout fry population over time, with CRR fish comprising the entirety of the fry population in 2007, and GR-cross fish comprising nearly 80% of the fry population in 2011. A decrease in average infection severity (myxospores fish-1) was observed concurrent with the shift in the genetic composition of the rainbow trout fry population, decreasing from an average of 47,708 (±8,950) myxospores fish-1 in 2009 to 2,672 (±4,379) myxospores fish-1 in 2011. Results from this experiment suggest that the GR×CRR can survive and reproduce in rivers with a high prevalence of M. cerebralis. In addition, reduced myxospore burdens in age-0 fish indicated that stocking this cross may ultimately lead to an overall reduction in infection prevalence and severity in the salmonid populations of the upper Colorado River.

Genetic basis of differences in myxospore count between whirling disease-resistant and susceptible strains of rainbow trout.

We used a quantitative genetics approach and estimated broad sense heritability (h2b) of myxospore count and the number of genes involved in myxospore formation to gain a better understanding of how resistance to Myxobolus cerebralis, the parasite responsible for whirling disease, is inherited in rainbow trout Oncorhynchus mykiss. An M. cerebralis-resistant strain of rainbow trout, the German Rainbow (GR), and a wild, susceptible strain of rainbow trout, the Colorado River Rainbow (CRR), were spawned to create 3 intermediate crossed populations (an F1 cross, F2 intercross, and a B2 backcross between the F1 and the CRR). Within each strain or cross, h2b was estimated from the between-family variance of myxospore counts using full-sibling families. Estimates of h2b and average myxospore counts were lowest in the GR strain, F1 cross, and F2 intercross (h2b = 0.34, 0.42, and 0.34; myxospores fish-1 = 275, 9566, and 45780, respectively), and highest in the B2 backcross and CRR strain (h2b = 0.93 and 0.89; myxospores fish-1 = 97865 and 187595, respectively). Comparison of means and a joint-scaling test suggest that resistance alleles arising from the GR strain are dominant to susceptible alleles from the CRR strain. Resistance was retained in the intermediate crosses but decreased as filial generation number increased (F2) or backcrossing occurred (B2). The estimated number of segregating loci responsible for differences in myxospore count in the parental strains was 9 ± 5. Our results indicate that resistance to M. cerebralis is a heritable trait within these populations and would respond to either artificial selection in hatcheries or natural selection in the wild.

The effects of Myxobolus cerebralis on the physiological performance of whirling disease resistant and susceptible strains of rainbow trout.

The development of rainbow trout Oncorhynchus mykiss strains that are resistant to whirling disease has shown promise as a management tool for populations in areas where Myxobolus cerebralis is present. However, the physiological effects of the disease on characteristics necessary for fish survival in natural river conditions have not been tested in many of these strains. Five rainbow trout strains were evaluated for their swimming ability and growth characteristics in relation to M. cerebralis exposure: the resistant German rainbow trout (GR) strain (Hofer strain), the susceptible Colorado River rainbow trout (CRR) strain, and three intermediate (hybrid) strains (F1 = GR x CRR; F2 = F1 x F1; B2 = backcross of F1 x CRR). Three broad response patterns among strain and exposure were evident in our study. First, exposure metrics, growth performance, and swimming ability differed among strains. Second, exposure to the parasite did not necessarily produce differences in growth or swimming ability. Exposure to M. cerebralis did not affect batch weight for any strain, and critical swimming velocity did not differ between exposed and unexposed families. Third, although exposure did not necessarily affect growth or swimming ability, individuals that exhibited clinical deformities did show reduced growth and swimming performance; fish with clinical deformities were significantly smaller and had lower critical swimming velocities than exposed fish without clinical deformities. Research and management have focused on GR x CRR hybrid strains; however, given the performance of the GR strain in our study, it should not be discounted as a potential broodstock. Additional field trials comparing the GR and F1 strains should be conducted before wholesale adoption of the GR strain to reestablish rainbow trout populations in Colorado.