WINTER ET AL,; RESISTANCE OF COHO SALMON AND STEELHEAD TROUT 



with cysteine (0.1%) and calf serum (20%). Before 

 each experiment, cells were passed once in the 

 species being tested to produce a fresh isolate, and 

 this isolate was further cultured until sufficient 

 cells were available for an inoculum. 



All test fish received an intraperitoneal 

 injection of 0.1 ml of a suspension of kidney disease 

 bacteria in phosphate-buffered saline (PBS), and 

 all control fish received a 0.1 ml intraperitoneal 

 injection of only PBS. The approximate inocula 

 were 9 x 10' cells for the coho salmon (mean 

 weight, 23 g), and 3 x 10* cells for the steelhead 

 trout (mean weight, 36 g). The coho salmon were 

 injected on 17 March 1977 and the steelhead trout 

 on 12 September 1977. We examined all fish that 

 died and identified BKD as the causative agent on 

 the basis of presumptive diagnosis, using gram 

 stains of kidney smears. In addition, kidney 

 smears from 10% of the fish that died were 

 cultured on Mueller-Hinton media. Experiments 

 were terminated at the, end of 4 mo or earlier, 

 depending on the progress of infection. 



One week after the coho salmon had been 

 injected, an accidental exposure of the fish, 

 including the controls, to chlorine resulted in 

 mortalities as high as 50% in some stocks. The 

 study was nevertheless continued, but a second, 

 abbreviated test was begun on 24 August 1977. 

 Only Alsea and Big Creek stocks (mean weight, 

 33.2 g) were used; the Big Creek fish were obtained 

 directly from the hatchery. The inoculum for this 

 second experiment was increased to 3 x 10* cells. 



Vibriosis 



The V. anguillarum strain (LS-174) used in 

 these experiments was isolated on brain heart 

 infusion agar from a coho salmon at Lint Slough, 

 Waldport, Oreg., by J. S. Rohovec. The inocula 

 were either prepared from lyophilized cells or 

 recent passage isolates. Experimental fish were 

 exposed to the pathogen in 93 1 stainless steel 

 tanks at Oregon State University's Fish Disease 

 Laboratory. 



Two experiments were undertaken with the 

 coho salmon. In the first (8 October 1976), 225 fish 

 (mean weights for Big Creek and Alsea stocks 

 were 10.4 g and 14.5 g, respectively) from each 

 stock were divided equally among two test 

 replicates and an untreated control. The three 

 tanks contained fish from each stock to insure 

 identical treatment. The fish in this experiment, 

 having not been bled and tagged for transferrin 



genotype identification, were freeze branded to 

 differentiate the stocks in each tank. In the second 

 experiment (10 June 1977) the number offish per 

 tank was reduced to about 25 (mean weight, 36.6 

 g) because larger numbers were not available, but 

 transferrin genotypes had been determined. 



In the steelhead trout phase of the study (21 

 October 1977), 75 fish from each stock (mean 

 weight, 36 g) were divided equally among three 

 test replicates and 15 from each stock were placed 

 in a fourth tank for controls. A hatchery-reared 

 Rogue stock was also used in this steelhead trout 

 experiment. In a second experiment (27 December 

 1977) in which we used steelhead trout from the 

 Cole Rivers (Rogue), Alsea, and Marion Forks 

 (North Santiam) hatcheries, 50 fish (mean weight, 

 42.2 g) were divided equally between two 

 replicates. Transferrin genotypes were 

 distributed randomly among the tanks. 



The initial temperature in all experimental 

 tanks was 12.2° C, to which all fish had been 

 acclimated. The temperature was then raised to 

 17.7° C over a period of 1.5 h, and at this 

 temperature water flow was discontinued in all 

 tanks for 15 min. The bacteria suspended in brain 

 heart infusion broth (Difco Laboratories) were 

 then introduced into the test tanks (other than 

 those of the controls). The inocula were 5 x 10'' 

 cells/ml for the first coho salmon exposure and 8.6 

 X 10^ cells/ml for the second; the steelhead trout 

 received concentrations of 8.8 x 10" cells/ml in the 

 first experiment and 7.2 x 10" cells/ml in the 

 second. All fish that died were necropsied and 

 kidney smears were cultured on brain heart 

 infusion agar. Positive diagnosis of V. 

 anguillarum was confirmed by slide agglutination 

 with specific antiserum. The experiments were 

 terminated at the end of 1 wk. 



Statistical comparison of three or more stocks 

 involved a one-way analysis of variance based on 

 arcsin transformations of percentages and least 

 significant difference, and comparisons of 

 transferrin genotypes of two stocks were based on 

 X^ test employing a 2 x /t contingency table 

 (Snedecor and Cochran 1967). 



RESULTS AND DISCUSSION 



Bacterial Kidney Disease 



In the first experiment in which coho salmon 

 were infected with BKD, the Alsea stock and B x 

 U cross were about twice as resistant to the disease 



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