enable comparisons of densities between sampling sections. Captured fish were 

 anesthetized with either tricaine methanesulfonate (MS-222) or clove oil, weighed (g) and 

 measured (mm) for total length (TL). For this report, we converted all weights and 

 lengths to standard units. 



Whirling Disease Sentinel Cage Studies 



Whirling disease surveys involving sentinel fish exposures were undertaken in the 

 Blackfoot Watershed in 2002 and 2003. Sentinel cage studies are controlled experiments 

 used to detect levels of whirling disease. Cages consist of an 18 x 24" cylindrical 

 screened container placed into a stream site, which allows stream water to flow through 

 the cage. Each cage contained 50 uninfected rainbow trout or WSCT (35-60 mm) 

 supplied by a state fish hatchery. In specific studies, brook and brown trout were also 

 used to detect levels of whirling disease infection. Timing of field exposure was based on 

 anticipated mean daily temperatures in the 50's (F), which correlates with peak 

 triactinomyxon (TAM) production, and corresponds to peak infection rates in fish 

 (Vincent 2000), except in spring creeks (Kleinschmidt and Nevada Spring Creek) where 

 recent research indicated peak infection occurred in late winter and early Spring 

 (Anderson 2004). The exposure period for each live cage was standardized at 10 days. 

 At the end of the 1 0-day exposure period, the trout were transferred to Pony, MT, where 

 they were held for an additional 80 days at a constant 50 ° F temperature to insure the WD 

 infection if present would reach its maximum intensity (Vincent 2000). At the end of the 

 holding period, all surviving fish were sacrificed and sent to the Washington State 

 University Animal Disease Diagnostic Laboratory at Pullman, WA. At the lab, the heads 

 were histologically examined using the MacConnell-Baldwin histological grading scale, 

 which ranks infection intensity from (absent) to 5 (severe) (Baldwin et al. 2000). The 

 results of this histological rating were presented as mean grade infection. Mean grade 

 infections above 2.7 are likely to result in population level declines (Vincent 2001). 

 Each sentinel cage also had an accompanying thermograph to establish mean daily water 

 temperatures during the exposure period. 



WSCT Genetic Investigations 



In 2002 and 2003, seventeen WSCT-bearing streams were tested for genetic 

 composiUon. Samples consisted of non-lethal tissue samples (fin-clip) taken from a 

 minimum 25 individual fish when possible. Samples collected were immediately 

 preserved in 95% ethyl alcohol and either placed in storage due to lack of funding, or 

 taken to the University of Montana, Salmon and Wild Trout Genetics Lab for 

 electrophoretic analysis. 



The Paired Interspersed Nuclear DNA Element-PCR (PINE-PCR) method is used 

 to determine each fish's genetic characteristics at 21 regions of nuclear DNA. This 

 method produces DNA fragments (PINE markers hereafter) that distinguish WSCT, from 

 rainbow trout and Yellowstone cutthroat trout. These species specific PINE markers, 

 therefore, can be used to determine whether a sample came from a genetically pure 

 population of one of these fishes or one in which hybridization between two or all three 

 of them has occurred. With a sample size of 25 fish, this testing method has a 95% 

 chance of identifying as little as 1% inlrogression. Results of WSCT genetic tests are in 



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