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• 1. [Article] Characterization of a new Lactobacillus species infectious for salmonid fish

  A new Lactobacillus species is proposed to include 17 strains of a bacterium from rainbow (Salmo gairdneri) and cutthroat trout (Salmo clarki), and chinook salmon (Oncorhynchus tshawytscha). The bacterium ...

  Citation × Citation Citation Abstract Copy Title: Characterization of a new Lactobacillus species infectious for salmonid fish Author: Hiu, Stephen Fooklen A new Lactobacillus species is proposed to include 17 strains of a bacterium from rainbow (Salmo gairdneri) and cutthroat trout (Salmo clarki), and chinook salmon (Oncorhynchus tshawytscha). The bacterium was isolated most frequently from fish stressed by handling or spawn ing. Phenotypically, the strains belong to the family Lactobacillaceae and can be distinguished from other species of Lactobacil1 us by their morphology and physiological characteristics. Growth occurred between 6 and 40°C. DL-lactic acid was produced homofermentatively from glucose. Diaminopimelic acid was present in the cell wall peptidoglycan. The 17 isolates were closely related gene tically, as demonstrated by similar percent guanine plus cytosine content (35 mol%) and high DNA reassociation values, both characteristics of a single species. Isolates exhibited less than 10% DNA reassoci ati on to other reference Lactobacillus strains with similar mol% G+C. The bacterium was pathogenic for rainbow trout maintained at 18 or 21°C. A 73 % mortality after 18 days was achieved at each water temperature after the fish were injected intraperitoneally with 3.3 x 10⁸ cells. The mean time from infection to death was 4 days for fish held at 18°C and 3 days for fish held at 21°C. The 50% lethal dose (LD50) for rainbow trout injected intraperitoneally and maintained in 18°C water was 5.4 x 10⁷. Histopathological examination of experimentally in fected fish showed that the bacterium was localized in the visceral adipose tissue, the peritoneal membrane, the loose connective tissue surrounding the liver, esophagus and spleen, and in the cardiac muscle and muscularis of the esophagus. Necrosis of the spleen, liver, or kidney was not observed. Gross pathology included ascites fluid accumulation and hemorrhaging of the visceral adipose tissue and lower intestine. The isolates from fish best match the description of the genus Lactobacillus but differ from other members of the genus genetically, in their fermentation patterns, and by their association with disease in fish and should be recognized as a new species for which the name Lactobacil! us piscicol us is proposed. Strain B270, isolated from diseased cutthroat trout at Bandon Hatchery, Coos County, Oregon, U.S.A., is designated the type strain. Title: Characterization of a new Lactobacillus species infectious for salmonid fish Author: Hiu, Stephen Fooklen A new Lactobacillus species is proposed to include 17 strains of a bacterium from rainbow (Salmo gairdneri) and cutthroat trout (Salmo clarki), and chinook salmon (Oncorhynchus tshawytscha). The bacterium was isolated most frequently from fish stressed by handling or spawn ing. Phenotypically, the strains belong to the family Lactobacillaceae and can be distinguished from other species of Lactobacil1 us by their morphology and physiological characteristics. Growth occurred between 6 and 40°C. DL-lactic acid was produced homofermentatively from glucose. Diaminopimelic acid was present in the cell wall peptidoglycan. The 17 isolates were closely related gene tically, as demonstrated by similar percent guanine plus cytosine content (35 mol%) and high DNA reassociation values, both characteristics of a single species. Isolates exhibited less than 10% DNA reassoci ati on to other reference Lactobacillus strains with similar mol% G+C. The bacterium was pathogenic for rainbow trout maintained at 18 or 21°C. A 73 % mortality after 18 days was achieved at each water temperature after the fish were injected intraperitoneally with 3.3 x 10⁸ cells. The mean time from infection to death was 4 days for fish held at 18°C and 3 days for fish held at 21°C. The 50% lethal dose (LD50) for rainbow trout injected intraperitoneally and maintained in 18°C water was 5.4 x 10⁷. Histopathological examination of experimentally in fected fish showed that the bacterium was localized in the visceral adipose tissue, the peritoneal membrane, the loose connective tissue surrounding the liver, esophagus and spleen, and in the cardiac muscle and muscularis of the esophagus. Necrosis of the spleen, liver, or kidney was not observed. Gross pathology included ascites fluid accumulation and hemorrhaging of the visceral adipose tissue and lower intestine. The isolates from fish best match the description of the genus Lactobacillus but differ from other members of the genus genetically, in their fermentation patterns, and by their association with disease in fish and should be recognized as a new species for which the name Lactobacil! us piscicol us is proposed. Strain B270, isolated from diseased cutthroat trout at Bandon Hatchery, Coos County, Oregon, U.S.A., is designated the type strain. Close

• 2. [Article] Production and behavioral interactions of salmonids in an experimental stream

  The behavior of two species of stream salmonids, juvenile coho salmon, Oncorhynchus kisutch (Walbaurn) and cutthroat trout, Salmo clarki clarki Richardson, was studied in terms of the time spent by individual ...

  Citation × Citation Citation Abstract Copy Title: Production and behavioral interactions of salmonids in an experimental stream Author: McIntyre, John D. The behavior of two species of stream salmonids, juvenile coho salmon, Oncorhynchus kisutch (Walbaurn) and cutthroat trout, Salmo clarki clarki Richardson, was studied in terms of the time spent by individual fish in carrying out various activities. Studies were conducted from June 1968, through May 1969, at Berry Creek, a small woodland stream located approximately ten miles north of Corvallis, Oregon. A 1500 foot section of the stream was brought under complete flow control for experimental purposes by a concrete diversion dam and a bypass channel. Water flow into the experimental section was controlled by a large gate valve at the dam. The lower part of the controlled section was divided into experimental sections that were separated by inclined-screen fish traps. The Berry Creek Experimental Stream Laboratory is maintained as a field research station by the Department of Fisheries and Wildlife, Oregon State University. Behavioral observations were made from an observation booth located in an experimental section of Berry Creek. Time-budget analyses derived from observational data were used to evaluate the effects on production of the interaction between individual salmon stocked at different densities in separate stream sections and the interaction among populations comprising both species stocked separately and together at similar densities in different sections of the stream. The growth rates of juvenile coho salmon stocked during June and July 1968, at different biomasses in five of the experimental sections generally declined with increases of salmon biomass. Downstream movement and mean percentages of time spent in agonistic behavior by individual salmon increased with increases in biomass. Higher values of production and mean biomass were generally recorded for an experimental group comprising salmon and trout than for single species groups. Several of the trout on occasion occupied the riffle area of the observation section, whereas, coho salmon remained in the pool throughout the experiment. The use of the riffle area by the trout may have resulted in more efficient utilization of space and food and may explain, in part, the differences in production between the trout-salmon group and the single species group. The growth and location in the stream section of an individual trout or salmon appeared to depend on the fish's position in the integrated social structure. Fish of high social position benefitted in terms of increased amounts of feeding time, decreased activity and decreased agonistic behavior as compared with fish of lower social position. The effect of intraspecific interaction among salmon appeared to be greatest at those stocking densities which resulted in a decline of growth rate from the growth rates that were associated with the highest values of production. This interaction appeared to lead to a compromise between energy expenditures for agonistic behavior and efficient metabolism that resulted in stock densities between two and three grams/meter2 of section area. Title: Production and behavioral interactions of salmonids in an experimental stream Author: McIntyre, John D. The behavior of two species of stream salmonids, juvenile coho salmon, Oncorhynchus kisutch (Walbaurn) and cutthroat trout, Salmo clarki clarki Richardson, was studied in terms of the time spent by individual fish in carrying out various activities. Studies were conducted from June 1968, through May 1969, at Berry Creek, a small woodland stream located approximately ten miles north of Corvallis, Oregon. A 1500 foot section of the stream was brought under complete flow control for experimental purposes by a concrete diversion dam and a bypass channel. Water flow into the experimental section was controlled by a large gate valve at the dam. The lower part of the controlled section was divided into experimental sections that were separated by inclined-screen fish traps. The Berry Creek Experimental Stream Laboratory is maintained as a field research station by the Department of Fisheries and Wildlife, Oregon State University. Behavioral observations were made from an observation booth located in an experimental section of Berry Creek. Time-budget analyses derived from observational data were used to evaluate the effects on production of the interaction between individual salmon stocked at different densities in separate stream sections and the interaction among populations comprising both species stocked separately and together at similar densities in different sections of the stream. The growth rates of juvenile coho salmon stocked during June and July 1968, at different biomasses in five of the experimental sections generally declined with increases of salmon biomass. Downstream movement and mean percentages of time spent in agonistic behavior by individual salmon increased with increases in biomass. Higher values of production and mean biomass were generally recorded for an experimental group comprising salmon and trout than for single species groups. Several of the trout on occasion occupied the riffle area of the observation section, whereas, coho salmon remained in the pool throughout the experiment. The use of the riffle area by the trout may have resulted in more efficient utilization of space and food and may explain, in part, the differences in production between the trout-salmon group and the single species group. The growth and location in the stream section of an individual trout or salmon appeared to depend on the fish's position in the integrated social structure. Fish of high social position benefitted in terms of increased amounts of feeding time, decreased activity and decreased agonistic behavior as compared with fish of lower social position. The effect of intraspecific interaction among salmon appeared to be greatest at those stocking densities which resulted in a decline of growth rate from the growth rates that were associated with the highest values of production. This interaction appeared to lead to a compromise between energy expenditures for agonistic behavior and efficient metabolism that resulted in stock densities between two and three grams/meter2 of section area. Close

• 3. [Article] A chromosome analysis of five cell lines from four species of salmonid fishes

  Four cell lines derived from embryonic tissues of four species of salmonid fishes, and one from a hepatoma in rainbow trout have been established in this laboratory. The species represented are chinook, ...

  Citation × Citation Citation Abstract Copy Title: A chromosome analysis of five cell lines from four species of salmonid fishes Author: McCain, Bruce B. Four cell lines derived from embryonic tissues of four species of salmonid fishes, and one from a hepatoma in rainbow trout have been established in this laboratory. The species represented are chinook, coho, and sockeye salmon; and steelhead and rainbow trout. All of these lines have been cultivated for approximately three years. A chromosome analysis was performed on these cell lines and two primary cell cultures from embryonic tissues of chinook and coho salmon. Each analysis consisted of two parts, chromosome preparation and chromosome characterization. Chromosome preparation involved exposing a growing culture to colchicine. The cells were harvested, flattened, expanded, fixed, and stained. Chromosomes were characterized by selecting 100 metaphase cells from stained preparations of each cell culture, and determining the number of chromosomes per cell and each chromosome's morphology. At the time of analysis, the cell lines were about two years old, and the primary cultures were three to six weeks old. The modal numbers and ranges of chromosomes per cell for the five cell lines were as follows: chinook salmon line (TC-114), mode 71, range 18 to 190; coho salmon line (TC-119), mode 71, range 57 to 173; sockeye salmon line (SeE), mode 56, range 51 to 101; steelhead trout line (TC-137), mode 62, range 58 to 126; and rainbow trout line (TC-149), modes 54 and 60, range 18 to 144. Similar distributions for the two primary cell cultures were the following: chinook salmon culture, mode 68, range 35 to 172; and coho primary culture, mode 60, range 58 to 119. These data indicate that the chinook and coho salmon, and the steelhead and rainbow trout hepatoma lines were heteroploid. Also, the sockeye salmon line, and coho and chinook salmon primary cultures had normal diploid chromosome constitutions. All seven cell cultures showed ranges and/or distributions of chromosome numbers per cell that were greater than found in normal in vivo cells. Chromosome morphology was determined by classifying each counted chromosome as either metacentric or telocentric. The morphology of the chromosomes in each sample of 100 cells was represented by the average ratio of telocentrics to metacentrics per cell. The sockeye salmon and rainbow trout cell lines and the chinook primary cell culture had ratios similar to those reported for these species. The other cultures had ratios which differed slightly from those characteristic of the species. Title: A chromosome analysis of five cell lines from four species of salmonid fishes Author: McCain, Bruce B. Four cell lines derived from embryonic tissues of four species of salmonid fishes, and one from a hepatoma in rainbow trout have been established in this laboratory. The species represented are chinook, coho, and sockeye salmon; and steelhead and rainbow trout. All of these lines have been cultivated for approximately three years. A chromosome analysis was performed on these cell lines and two primary cell cultures from embryonic tissues of chinook and coho salmon. Each analysis consisted of two parts, chromosome preparation and chromosome characterization. Chromosome preparation involved exposing a growing culture to colchicine. The cells were harvested, flattened, expanded, fixed, and stained. Chromosomes were characterized by selecting 100 metaphase cells from stained preparations of each cell culture, and determining the number of chromosomes per cell and each chromosome's morphology. At the time of analysis, the cell lines were about two years old, and the primary cultures were three to six weeks old. The modal numbers and ranges of chromosomes per cell for the five cell lines were as follows: chinook salmon line (TC-114), mode 71, range 18 to 190; coho salmon line (TC-119), mode 71, range 57 to 173; sockeye salmon line (SeE), mode 56, range 51 to 101; steelhead trout line (TC-137), mode 62, range 58 to 126; and rainbow trout line (TC-149), modes 54 and 60, range 18 to 144. Similar distributions for the two primary cell cultures were the following: chinook salmon culture, mode 68, range 35 to 172; and coho primary culture, mode 60, range 58 to 119. These data indicate that the chinook and coho salmon, and the steelhead and rainbow trout hepatoma lines were heteroploid. Also, the sockeye salmon line, and coho and chinook salmon primary cultures had normal diploid chromosome constitutions. All seven cell cultures showed ranges and/or distributions of chromosome numbers per cell that were greater than found in normal in vivo cells. Chromosome morphology was determined by classifying each counted chromosome as either metacentric or telocentric. The morphology of the chromosomes in each sample of 100 cells was represented by the average ratio of telocentrics to metacentrics per cell. The sockeye salmon and rainbow trout cell lines and the chinook primary cell culture had ratios similar to those reported for these species. The other cultures had ratios which differed slightly from those characteristic of the species. Close

• 4. [Article] Interrenal and metabolic responses to stress and their modifying factors in juvenile salmonid fishes

  Genetic, ontogenetic, and environmental factors modified characteristic interrenal and glycemic responses to stress in juvenile salmonid fishes. During continuous confinement stress, plasma cortisol rose ...

  Citation × Citation Citation Abstract Copy Title: Interrenal and metabolic responses to stress and their modifying factors in juvenile salmonid fishes Author: Barton, Bruce Alexander Genetic, ontogenetic, and environmental factors modified characteristic interrenal and glycemic responses to stress in juvenile salmonid fishes. During continuous confinement stress, plasma cortisol rose more quickly in chinook salmon, Oncorhynchus tshawytscha, acclimated to high rather than medium or low temperatures; hyperglycemia following either acute or chronic stress was also highest in this group. When chinook salmon were subjected to three handling stresses, separated by 3-h intervals, increases of plasma cortisol, glucose and lactate and decreases in plasma sodium were cumulative. Healthy chinook salmon appeared sore able than diseased fish to elevate plasma cortisol after stress, but post-stress plasma glucose was higher in the unhealthy fish. Glycemic responses to stress in chinook salmon were related to diet type and to fasting. When rainbow trout, Salmo gairdneri, were exposed to pH 4.7 and then handled, the post-stress plasma cortisol elevation was more than twice that in those at ambient pH. Rainbow trout subjected to an acute disturbance daily for 10 wk exhibited habituation, evident from plasma cortisol and glucose levels after handling that were about one half of those in previously unstressed fish. The interrenal. response to handling in coho salmon, Oncorhynchus kisutch, increased twofold during the normal period of smoltification. A comparison of chinook salmon, coho salmon, rainbow trout and brook trout, Salvelinus fontinalis, subjected to identical handling stresses, suggested that there were species and stock differences affecting the megnitude of plasma cortisol elevations. Rainbow trout fed with cortisol-treated feed exhibited reduced growth, condition factor, liver glycogen and circulating lymphocytes, and higher plasma glucose and hemetocrit. The plasma cortisol elevation following handling was completely abolished in the cortisol-fed trout, indicating that continuous negative feedback of cortisol eliminated the fish's ability to initiate an interrenal stress response. When forced to swim, seen oxygen consumption rate in stressed steelhead, Salmo gairdneri, was about twice that in unstressed fish and individual rates were positively correlated with please cortisol elevations. The metebolic cost of this stress was estimated to reduce the energy available for other performance components within the fish's scope for activity by about one quarter. Title: Interrenal and metabolic responses to stress and their modifying factors in juvenile salmonid fishes Author: Barton, Bruce Alexander Genetic, ontogenetic, and environmental factors modified characteristic interrenal and glycemic responses to stress in juvenile salmonid fishes. During continuous confinement stress, plasma cortisol rose more quickly in chinook salmon, Oncorhynchus tshawytscha, acclimated to high rather than medium or low temperatures; hyperglycemia following either acute or chronic stress was also highest in this group. When chinook salmon were subjected to three handling stresses, separated by 3-h intervals, increases of plasma cortisol, glucose and lactate and decreases in plasma sodium were cumulative. Healthy chinook salmon appeared sore able than diseased fish to elevate plasma cortisol after stress, but post-stress plasma glucose was higher in the unhealthy fish. Glycemic responses to stress in chinook salmon were related to diet type and to fasting. When rainbow trout, Salmo gairdneri, were exposed to pH 4.7 and then handled, the post-stress plasma cortisol elevation was more than twice that in those at ambient pH. Rainbow trout subjected to an acute disturbance daily for 10 wk exhibited habituation, evident from plasma cortisol and glucose levels after handling that were about one half of those in previously unstressed fish. The interrenal. response to handling in coho salmon, Oncorhynchus kisutch, increased twofold during the normal period of smoltification. A comparison of chinook salmon, coho salmon, rainbow trout and brook trout, Salvelinus fontinalis, subjected to identical handling stresses, suggested that there were species and stock differences affecting the megnitude of plasma cortisol elevations. Rainbow trout fed with cortisol-treated feed exhibited reduced growth, condition factor, liver glycogen and circulating lymphocytes, and higher plasma glucose and hemetocrit. The plasma cortisol elevation following handling was completely abolished in the cortisol-fed trout, indicating that continuous negative feedback of cortisol eliminated the fish's ability to initiate an interrenal stress response. When forced to swim, seen oxygen consumption rate in stressed steelhead, Salmo gairdneri, was about twice that in unstressed fish and individual rates were positively correlated with please cortisol elevations. The metebolic cost of this stress was estimated to reduce the energy available for other performance components within the fish's scope for activity by about one quarter. Close

• 5. [Article] Endocrine regulation of final oocyte maturation and sex differentiation in salmonids

  Sexual maturation and sex differentiation comprise facets of a common theme: reproduction. The endocrine system regulates many of the critical physiological processes necessary for reproduction and offers ...

  Citation × Citation Citation Abstract Copy Title: Endocrine regulation of final oocyte maturation and sex differentiation in salmonids Author: Fitzpatrick, Martin S. Sexual maturation and sex differentiation comprise facets of a common theme: reproduction. The endocrine system regulates many of the critical physiological processes necessary for reproduction and offers a framework within which technologies can be developed for controlling sexual maturation and sex differentiation. The studies described in this thesis were undertaken to improve the understanding of the endocrine control of these critical stages of development in salmonids. Final ovarian maturation in salmon is accompanied by dynamic changes in plasma hormone levels. Ovulation can be accelerated through the use of hormones such as gonadotropin releasing hormone or its analogs (GnRHa). The effectiveness of GnRHa often depends on the timing of treatment. To determine if plasma concentrations of steroids can be used to predict the sensitivity of adult female coho salmon (Oncorhynchus kisutch) to GnRHa, circulating levels of testosterone, 17α,20β-dihydroxyprogesterone (DHP), and estradiol were measured before and after injection with GnRHa to accelerate ovulation. We found that high levels of testosterone were predictive of early response of coho salmon to GnRHa treatment. The correlation between testosterone and ovulatory response to GnRHa suggested a possible functional relation. However. implantation or injection of testosterone. 17α-methyltestosterone (MT), or the antiandrogen, cyproterone acetate (CA), before or with GnRHa treatment did not affect the ovulatory response of coho or chinook salmon ( 0. tshawytscha) to GnRHa. Chinook salmon treated with MT alone had accelerated ovulation in comparison to controls. If steroids are involved in sex differentiation. steroids must be produced early in development. In vitro production of steroids in both coho salmon and rainbow trout (0. mykiss) was assessed from hatch through sex differentiation. Cortisol, androstenedione, testosterone, and estradiol were produced just after hatching by tissue explants that contained anterior kidneys and gonads of coho salmon. To circumvent the problem of not knowing the sex of individuals until after sex differentiation, single-sex populations of rainbow trout were produced by gynogenesis or androgenesis. Tissue explants produced more androstenedione than testosterone or estradiol. More androgens were produced by testes and more estradiol was produced by ovaries within 6 to 10 weeks of hatching. Dietary treatment with estradiol or MT inhibited gonadal steroid secretion. Electrophoresis of gonadal homogenates from salmonids revealed several sex-specific bands. In particular, a prominent band of about 50,000 daltons was apparent in ovaries but not testes. Production of sex-specific proteins may be affected by dietary steroid treatment. Title: Endocrine regulation of final oocyte maturation and sex differentiation in salmonids Author: Fitzpatrick, Martin S. Sexual maturation and sex differentiation comprise facets of a common theme: reproduction. The endocrine system regulates many of the critical physiological processes necessary for reproduction and offers a framework within which technologies can be developed for controlling sexual maturation and sex differentiation. The studies described in this thesis were undertaken to improve the understanding of the endocrine control of these critical stages of development in salmonids. Final ovarian maturation in salmon is accompanied by dynamic changes in plasma hormone levels. Ovulation can be accelerated through the use of hormones such as gonadotropin releasing hormone or its analogs (GnRHa). The effectiveness of GnRHa often depends on the timing of treatment. To determine if plasma concentrations of steroids can be used to predict the sensitivity of adult female coho salmon (Oncorhynchus kisutch) to GnRHa, circulating levels of testosterone, 17α,20β-dihydroxyprogesterone (DHP), and estradiol were measured before and after injection with GnRHa to accelerate ovulation. We found that high levels of testosterone were predictive of early response of coho salmon to GnRHa treatment. The correlation between testosterone and ovulatory response to GnRHa suggested a possible functional relation. However. implantation or injection of testosterone. 17α-methyltestosterone (MT), or the antiandrogen, cyproterone acetate (CA), before or with GnRHa treatment did not affect the ovulatory response of coho or chinook salmon ( 0. tshawytscha) to GnRHa. Chinook salmon treated with MT alone had accelerated ovulation in comparison to controls. If steroids are involved in sex differentiation. steroids must be produced early in development. In vitro production of steroids in both coho salmon and rainbow trout (0. mykiss) was assessed from hatch through sex differentiation. Cortisol, androstenedione, testosterone, and estradiol were produced just after hatching by tissue explants that contained anterior kidneys and gonads of coho salmon. To circumvent the problem of not knowing the sex of individuals until after sex differentiation, single-sex populations of rainbow trout were produced by gynogenesis or androgenesis. Tissue explants produced more androstenedione than testosterone or estradiol. More androgens were produced by testes and more estradiol was produced by ovaries within 6 to 10 weeks of hatching. Dietary treatment with estradiol or MT inhibited gonadal steroid secretion. Electrophoresis of gonadal homogenates from salmonids revealed several sex-specific bands. In particular, a prominent band of about 50,000 daltons was apparent in ovaries but not testes. Production of sex-specific proteins may be affected by dietary steroid treatment. Close