North American Journal of Aquaculture 71:252–255, 2009 Ó Copyright by the American Fisheries Society 2009 DOI: 10.1577/A08-064.1
[Technical Note]
Observations Concerning Reproductive Temperature Requirements of Captive Lahontan Cutthroat Trout JOHN P. BIGELOW* U.S. Fish and Wildlife Service, Lahontan National Fish Hatchery Complex, 1340 Financial Boulevard, Suite 234, Reno, Nevada 89502, USA
WENDY M. RAUW
AND
LUIS GOMEZ-RAYA
Department of Animal Biotechnology, University of Nevada–Reno, 1664 North Virginia Street, Reno, Nevada 89557, USA Abstract.—The effects of holding temperature during late oocyte development and spawning on the timing of ovulation, fecundity, and egg survival in female Lahontan cutthroat trout Oncorhynchus clarkii henshawi in three postovulatory regimes were investigated. The timing of ovulation was similar for females held at 7.28C and 12.48C from 2.5 to 6.0 months before the time of ovulation until spawning. Mean fecundity was higher for females held at 7.28C. Mean egg eyeup, hatch, and swim-up rates were also higher for females held at 7.28C. Eggs collected 1, 4, and 7 d postovulation exhibited similar survival rates within females regardless of holding temperature.
The Lahontan National Fish Hatchery Complex maintains a captive broodstock of Lahontan cutthroat trout Oncorhynchus clarkii henshawi, a federally listed threatened species (USOFR 1975), for reintroduction efforts in the Lake Tahoe, Truckee River, and Walker River subbasins in northern Nevada and eastern California from which they were extirpated during the early 20th century (La Rivers 1962; Sigler and Sigler 1987; Behnke 1992). Before their extirpation, lacustrine populations of Lahontan cutthroat trout migrated into rivers to spawn during the spring. Adults were renowned for reaching sizes exceeding 18 kg (Behnke 1992). Egg survival for captive Lahontan cutthroat trout broodstock held in ambient water (12.48C) at the Lahontan National Fish Hatchery has been too low to meet hatchery requirements; however, eggs collected from wild Lahontan cutthroat trout in 78C waters and incubated in ambient hatchery water have consistently high survival rates (authors’ unpublished data). In several cultured salmonids, excessive temperatures during late oocyte development and spawning can cause: (1) inhibited ovulation resulting in later spawning (Gillet 1991; Taranger and Hansen 1993; Pankhurst and Thomas 1998), (2) oocyte resorption * Corresponding author:
[email protected] Received November 13, 2008; accepted November 17, 2008 Published online May 14, 2009
(i.e., atresia; Smith et al. 1983) resulting in reduced fecundity, and (3) poor egg quality (Hokanson et al. 1973; Smith et al. 1983; Gillet 1991; Pankhurst et al. 1996; King and Pankhurst 2004) and accelerated postovulatory oocyte aging (Aegerter and Jalabert 2004) resulting in reduced egg survival. Determining holding temperatures for females that can prevent these detrimental effects is necessary for successful management of the captive Lahontan cutthroat trout broodstock. The objective of this study was to compare the reproductive performance, under the spawning protocols employed at the Lahontan National Fish Hatchery, of Lahontan cutthroat trout brood females held in ambient (12.48C) and chilled (7.28C) hatchery well water during the final stages of oocyte maturation and spawning. Characteristics of reproductive performance included (1) timing of ovulation, (2) fecundity, and (3) egg survival while postovulatory aging was controlled. Methods Fish.—Forty females between 2.4 and 2.7 years of age (mean 6 SE ¼ 346 6 6.25 mm fork length [FL]) were randomly selected from the captive Lahontan cutthroat trout broodstock at Lahontan National Fish Hatchery, Gardnerville, Nevada. All fish were thirdgeneration progeny of trout captured from wild populations in the Pilot Mountains of western Utah and confirmed by phylogenetic analysis as recent descendents from the extirpated Truckee River subbasin populations (Peacock and Kirchoff 2007). All fish were reared in ambient (128C) artesian well water under natural photoperiod. On December 7, 2006, 20 females were assigned with no known bias to each of two 640-L circular tanks supplied with nonrecirculated well water at the hatchery. Temperatures in all tanks were monitored to the nearest 0.018C at 30-min intervals with Stowaway Tidbit Temperature Loggers (Onset, Bourne, Massachusetts). One group remained at ambient temperature (mean 6 SE ¼ 12.44 6 0.528C;
252
253
TECHNICAL NOTE
T12.4). The other group was chilled beginning on December 18, 2006 (mean 6 SE ¼ 7.20 6 0.508C; T7.2). These temperatures were maintained until all females were spawned. Mechanical problems resulted in a temperature increase to 10.558C in T7.2 on February 17 and 18, 2007. The temperature ranged from 5.448C to 10.558C in T7.2 and from 11.378C to 13.398C in T12.4 after December 18, 2006. Fifty 3-yearold males were held in one 2,800-L circular tank supplied with ambient water. Flows were maintained at 23 L/min for all tanks. Fish were fed Rangen trout brood diet (6.35-mm pellets; Rangen, Inc., Buhl, Idaho). Females were fed 0.5% body weight/d for the duration of the study. Males were fed 0.5% body weight/d until February 24, 2007, after which they remained off feed. Dissolved oxygen concentrations were monitored daily with a Hach HQ10 oxygen meter (Hach Co., Loveland, Colorado) and maintained between 8 and 10 mg/L in all tanks. One female died before ovulation in each temperature group. Ovulation and egg collection.—Females were checked daily for ovulation beginning on January 12, 2007. To minimize handling stress, females were fully sedated with 50 lg/mL of Finquel MS-222 (tricaine methanesulfonate; Argent Laboratories, Redmond, Washington) before handling. Sedated females were gently transferred by net to a recovery tank and checked for ripeness by manual abdominal palpitation (Erdahl 1994). Ovulation date was recorded. Approximately one-third of the eggs from each ripe female were spawned. Females were then Floy-tagged (Floy Tag and Manufacturing, Inc., Seattle, Washington) for subsequent individual identification and returned to their original tanks. Females were treated daily in a 2% solution of salt (NaCl) for 30 min after handling to offset electrolyte loss. Daily handling took less than 20 min per group. Floy-tagged females were partially spawned (as in Aegerter and Jalabert 2004; Rime et al. 2004) again 4 and 7 d postovulation producing a total of three batches of approximately equal numbers of eggs per female. To mitigate possible effects of partial spawning on egg survival, females were handled as gently as possible. Egg batches were kept segregated for the duration of the study. Eggs were stored in covered stainless-steel bowls and maintained at the same temperature as their female parents. Fork length of each female was measured (mm) after her final spawning. Eggs were fertilized within 30 min of egg collection. Males were randomly netted and sedated. Each male was spawned three times to fertilize the three egg batches from one single female. Milt was collected and expressed onto the eggs with a pipette. Males were then
Floy-tagged and returned to their tank. The combined gametes were gently stirred with a pipette and allowed to sit for 1 min. Water isothermic to the eggs was added to activate the sperm. The eggs were rinsed 1 min later and placed in a Heath-type egg tray. Ambient water was trickled into the tray acclimating T7.2 eggs to approximately 128C over 6 h, but maintaining constant temperature for T12.4 eggs. Eggs were treated the following day with 100 lg/mL of Argentyne (Argent Laboratories, Redmond, Washington) solution for 15 min, and were then assigned (with no known bias) to one of four compartments in one of eight Heath-type egg trays supplied with a flow of 7.6 L/min at a temperature of 128C. To minimize any detrimental effects of egg temperature acclimation and retraying on egg survival, these activities were performed after water-hardening as recommended by Wagner and Arndt (2006) and before the period of increased sensitivity, which usually occurs from 48 h after fertilization until eye-up (Wedemeyer 2001). Initially, eggs were treated with a 2,000-lg/mL formalin solution for 15 min/d until 16 d after fertilization to prevent fungal (Saprolegnia spp.) infection; however, eggs from the first ovulating female became infected after eye-up and before hatching, and were eliminated from the study. Subsequently, the period that eggs were not treated with formalin (between eye-up and hatching) was reduced from 8 to 6 d. Fungal infections did not occur after this adjustment. In each batch, live and dead eggs were hand-counted at eye-up 16 d (198 degree-days [8C]) after fertilization and dead eggs were discarded. Fecundity was calculated by summing the total number of eggs in the three batches from each female. Live hatchlings were counted 24 d (298 degree-days) after fertilization and assigned with no known bias to one of twenty-one 28L circular tanks supplied with a flow of 3 L/min at a temperature of 128C. Fry reaching yolk sac resorption without obvious deformities were counted 41 d (508 degree-days) after fertilization. Percent survival to eyeup, hatch, and yolk sac resorption was calculated for each batch by dividing the number of survivors at 16, 24, and 41 d after fertilization, respectively, by the total number of eggs counted 16 d after fertilization. Results and Discussion We investigated the effect of holding temperature on timing of ovulation, fecundity, and egg survival in female Lahontan cutthroat trout. Two limitations of this study are: (1) treatments were not replicated, and (2) fecundity estimates did not include ovulated eggs remaining in the coelomic cavity after spawning. Treatments were not replicated because of anticipated low power for detecting tank effects due to scarcity of
254
BIGELOW ET AL.
TABLE 1.—Timing of ovulation, fork length at spawning, fecundity, and number of eggs surviving to yolk sac resorption (YSR) for 3-year-old female Lahontan cutthroat trout held at 7.28C (T7.2) and 12.48C (T12.4) at the Lahontan National Fish Hatchery during the winter and spring of 2007. Values are means 6 SEs; sample sizes are given in parentheses. Characteristic Timing of ovulation (days after Feb 24, 2007) Fork length at spawning (mm) Fecundity (eggs per female) Eggs surviving to YSR a b
T7.2
T12.4 a
32.4 6 5.35 (19) 43.8 6 7.83 (19)b 345 6 9.04 (19) 872 6 73.2 (19) 536 6 29.9 (18)
354 6 8.81 (19) 706 6 61.9 (19) 142 6 18.4 (19)
March 28. Apr 8.
test fish and high variation in egg survival (authors’ unpublished data). Females were not sacrificed after spawning to determine true fecundity due to their scarcity. To ensure variation in the number of unspawned eggs per female was not systematic to a given treatment, all fish were spawned by the same person. Timing of Ovulation Nineteen females ovulated in each temperature group. Results were inconclusive concerning the effect of holding temperature on timing of ovulation. The 1st, 10th, and 19th ovulation dates were February 24, March 25, and May 14, respectively, for T7.2, and February 28, March 29, and June 3, respectively, for T12.4. Mean ovulation date was 11 d earlier for T7.2 (Table 1). Twenty-one percent of the females in T12.4
FIGURE 2.—Percent survival (mean þ SE) of Lahontan cutthroat trout eggs to eye-up, hatch, and yolk sac resorption (YSR). Eggs were spawned from 3-year-old females held at 7.28C (T7.2; N ¼ 18) and 12.48C (T12.4; N ¼ 19) during the winter and spring of 2007 at the Lahontan National Fish Hatchery.
ovulated after the last female in T7.2, extending the spawning season by 20 d (to June 3) for that group. Delayed or inhibited ovulation at higher than optimal holding temperatures have been reported for rainbow trout O. mykiss (Pankhurst and Thomas 1998), Atlantic salmon Salmo salar (Taranger and Hansen 1993), Arctic char Salvelinus alpinus (Gillet 1991), and westslope cutthroat trout O. clarkii lewisi (Smith et al. 1983). Fecundity Although FL at spawning was similar for females from T7.2 and T12.4, mean fecundity was higher in T7.2 than in T12.4 (Table 1). Longer females had higher fecundities in both groups (Figure 1). The reduced fecundity exhibited by T12.4 females suggests inhibition of ovulation, or resorption of oocytes into the ovaries (i.e., atresia). In rainbow trout, a delay in the expression of an enzyme necessary for oocyte maturation may be at least partially responsible for inhibited ovulation at holding temperatures higher than optimal (Pankhurst and Thomas 1998). Smith et al. (1983) observed many atretic eggs on the ovaries of westslope cutthroat trout held at higher-than-optimal temperatures. Egg Survival
FIGURE 1.—Fecundity (total number of eggs spawned) in relation to the fork length (FL) of 3-year-old Lahontan cutthroat trout females held at 7.28C (T7.2; N ¼ 19) and 12.48C (T12.4; N ¼ 19) during the winter and spring of 2007 at the Lahontan National Fish Hatchery. The estimated relationships at these two temperatures were fecundity ¼1268.80 þ 6.21 3 FL and fecundity ¼ 684.94 þ 3.97 3 FL, respectively.
Mean survival rates to eye-up, hatch, and yolk sac resorption were higher for eggs from T7.2 than from T12.4 (Figure 2) as was the mean number of eggs per female surviving to yolk sac resorption (Table 1). Egg survival rates appeared not to be affected by postovulatory aging at any of the three stages of egg
TECHNICAL NOTE
TABLE 2.—Percent (mean 6 SE) egg survival to eye-up, hatch, and yolk sac resorption (YSR) for 3-year-old female Lahontan cutthroat trout held at 7.28C (T7.2; N ¼ 19) and 12.48C (T12.4; N ¼ 18) and partially spawned at 1, 4, and 7 d postovulation (DPO) at the Lahontan National Fish Hatchery during the winter and spring of 2007. Treatment
DPO
T7.2
1 4 7 1 4 7
T12.4
% Hatch 67.5 70.4 70.2 29.1 39.3 22.0
6 6 6 6 6 6
5.11 5.88 5.34 6.25 7.95 6.58
% Eye-up 63.9 67.6 68.1 25.9 34.0 18.7
6 6 6 6 6 6
5.26 5.83 5.20 6.22 7.67 6.13
% YSR 57.0 61.5 59.8 21.3 27.2 15.4
6 6 6 6 6 6
4.66 5.72 4.95 5.87 7.55 5.83
development for either temperature treatment (Table 2). These results indicate a lack of detrimental effects of partial spawning on egg survival. Results also suggest detrimental effects of higher female holding temperature on egg survival, and these effects are manifested before postovulatory oocyte aging in Lahontan cutthroat trout when postovulatory oocyte aging is limited to 1 week. In rainbow trout, Pankhurst et al. (1996) hypothesized that decreased egg quality at elevated temperatures is manifested during final maturation and ovulation. Our results suggest egg survival would be higher for Lahontan cutthroat trout brood females held at 7.28C rather than 12.48C from mid-December through spawning. Our results also indicate that artificially spawning captive Lahontan cutthroat trout more often than weekly may not increase egg survival for brood females held at either 7.28C or 12.48C. Acknowledgments We thank Alvin Duncan, Sarah Bigelow, Giovanni Reyes, and Edward Kelly for their assistance with the study. Mary Peacock made many improvements to the manuscript. The research was supported by the U.S. Fish and Wildlife Service. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the U.S. Fish and Wildlife Service. Reference to trade names does not imply endorsement by the U.S. Government. References Aegerter, S., and B. Jalabert. 2004. Effects of postovulatory oocyte ageing and temperature on egg quality and on the occurrence of triploid fry in rainbow trout, Oncorhynchus mykiss. Aquaculture 231:59–71. Behnke, R. J. 1992. Native trout of western North America. American Fisheries Society, Monograph 6, Bethesda, Maryland. Erdahl, D. A. 1994. Inland salmonid broodstock management
255
handbook. U.S. Fish and Wildlife Service, 712 FW1, Bozeman, Montana. Gillet, C. 1991. Egg production in an Arctic charr (Salvelinus alpinus L.) brood stock: effects of temperature on the timing of spawning and the quality of eggs. Aquatic Living Resources 4:109–116. Hokanson, K. E. F., J. H. McCormick, B. R. Jones, and J. H. Tucker. 1973. Thermal requirements for maturation, spawning, and embryo survival of brook trout (Salvelinus fontinalis). Journal of the Fisheries Research Board of Canada 30:975–984. King, H. R., and N. W. Pankhurst. 2004. Effect of short-term temperature reduction on ovulation and LHRHa responsiveness in female Atlantic salmon (Salmo salar) maintained at elevated water temperatures. Aquaculture 238:421–436. La Rivers, I. 1962. Fishes and fisheries of Nevada. Nevada State Fish and Game Commission, Reno. Pankhurst, N. W., G. J. Purser, G. Van Der Kraak, P. M. Thomas, and G. N. R. Forteath. 1996. Effect of holding temperature on ovulation, egg fertility, plasma levels of reproductive hormones and in vitro ovarian steroidogenesis in the rainbow trout Oncorhynchus mykiss. Aquaculture 146:277–290. Pankhurst, N. W., and P. M. Thomas. 1998. Maintenance at elevated temperature delays the steroidogenic and ovulatory responsiveness of rainbow trout Oncorynchus mykiss to lutenizing hormone releasing hormone analogue. Aquaculture 166:163–177. Peacock, M. M., and V. S. Kirchoff. 2007. Analysis of genetic variation and population genetic structure in Lahontan cutthroat trout (Oncorhynchus clarki henshawi) extant populations. Final Report to U.S. Fish and Wildlife Service, Reno, Nevada. Rime, H., N. Guitton, C. Pineau, E. Bonnet, J. Bobe, and B. Jalabert. 2004. Postovulatory aging and egg quality: a proteomic analysis of rainbow trout coelomic fluid. Reproductive Biology and Endocrinology 2:26–36. Sigler, W. F., and J. W. Sigler. 1987. Fishes of the Great Basin. University of Nevada Press, Reno. Smith, C. E., W. P. Dwyer, and R. G. Piper. 1983. Effect of water temperature on egg quality of cutthroat trout. Progressive Fish-Culturist 45:176–178. Taranger, G. L., and T. Hansen. 1993. Ovulation and egg survival following exposure of Atlantic salmon, Salmo salar L., broodstock to different water temperatures. Aquaculture and Fisheries Management 24:151–156. USOFR (U.S. Office of the Federal Register). 1975. ‘‘Threatened’’ status for three species of trout, final rule. Federal Register 40:137(16 July 1975):29863–29864. Wagner, E. J., and R. E. Arndt. 2006. The effect of temperature changes and transport on cutthroat trout eggs soon after fertilization. North American Journal of Aquaculture 68:235–239. Wedemeyer, G. A., editor. 2001. Fish hatchery management, 2nd edition. American Fisheries Society, Bethesda, Maryland.
