1. Utilizing Skein Eggs as an Alternative Egg Source For Bloater Reintroduction into Lake Ontario K. J. NASH, M. A. CHALUPNICKI, J. H. JOHNSON USGS Great Lakes Science Center, Tunison Laboratory of Aquatic Science Cortland, NY 13045 N. H. RINGLER SUNY College of Environmental Science and Forestry Syracuse, NY 13210 ABSTRACT: Restoration of native fish populations into Lake Ontario is a multi-agency effort within New York State. Populations of bloater (Coregonus hoyi), a deep water cisco, over the past century have become extirpated in Lake Ontario due to overharvesting and competition with invasive species. The reintroduction of bloater as an alternative forage fish to non-endemic species has given rise to new culture methods utilizing the availability of current stable populations in Lake Michigan. Specific efforts for new culture techniques have been made for potential utilization of skein-bound eggs to produce viable offspring. Retrieving skein eggs for the purpose of reintroducing bloater back into Lake Ontario would be beneficial due to the large quantity available. In 2015, we evaluated the growth and survival of spawned ripe and skein-bound bloater eggs. The egg diameter of skein eggs was significantly smaller than the ripe eggs but did not appear to effect survival or development. Overall, the survival of ripe eggs was slightly higher than skein eggs but was not significantly different. In the first two months post-hatch survival was greater for “skein” fish, with “ripe” fish survival becoming slightly higher by the fall. Implementation by management agencies of skein-bound eggs as an alternative egg source may benefit restoration efforts of bloater into Lake Ontario. METHODS: Adult bloater were captured near Two Rivers (Wisconsin ) in Lake Michigan by commercial fishing trawlers in January with the assistance of the USFWS Green Bay Office and Jordan River National Fish Hatchery. The adults were evaluated for readiness to spawn as “ripe” where eggs released freely or “skein-bound” where eggs were not fully detached from the ovary and were physically removed. Eggs were fertilized and kept in separate containers and shipped overnight to TLAS and WLFCS for rearing (Figure 2). Once received, eggs were immersed for 10 minutes in culture water containing 25mg/L polymeric iodine (Argentyne, Argent Chemical Labs, Redmond, Washington) while gently being stirred and then rinsed 3x. To prevent egg adhesiveness, eggs were then immersed for 15 minutes in water containing 66 units of bacterial protease (Protease, Bacillus lichenifarmes, Sigma Chemical Co., St. Louis, Missouri) with occasional gentle stirring to prevent clumping. The eggs were again rinsed 3x and transferred to McDonald jars (Figures 3-4). Egg motion was evaluated and flow rates were measured and recorded daily. Floating, dead, and fungus eggs were siphoned off daily. Eggs were treated with 0.17% formalin for 15 min 2-3 times a week to prevent and control fungus levels. All effluent underwent UV treatment in order to prevent any potential disease spread. Embryogenesis was evaluated and monitored weekly throughout the incubation period using a (10- 100x) Leica digital microscope camera with digital imaging software. Weekly sub-samples of eggs from both ripe and skein adults were evaluated for developmental milestones and overall survival. Evaluation consisted of viewing and enumerating 1 ml of eggs from each sample under a dissecting microscope equipped with the digital microscope camera. Live and dead eggs were counted and developmental progress was noted. Embryonic milestones were defined as 1) Fertilization with cell division, 2) Notochord formation, 3) Body and head formation, 4) Eye up, 5) Heart beat observed, 6) Myomeres/pigmentation completed, 7) Hatch (Figure 5). Once hatch occurred evaluation of ripe versus skein egg success was complete. Hatched fry were transferred to green circular takes where they were allowed to grow. Growth and survival were recorded until fish were stocked in November. Differences in egg diameter between ripe and skein eggs were evaluated at fertilization and eye up using paired t-tests (Statistix 8.0, Tidepool Scientific, Tallahassee, Florida). Paired t-tests were also used to determine if the percent of viable eggs between ripe and skein at each developmental milestone were significantly different. We considered p<0.05 to be significantly different. RESULTS: Bloater skein eggs were significantly smaller than ripe eggs at fertilization (p<0.01, T Stat=3.17, DF=69) and at eye up (p=0.02, T Stat=2.38, DF=42) (Table 1). Although smaller, skein eggs enlarged 3x more than ripe eggs over the course of development (0.043mm v 0.015mm). Through all developmental milestones, ripe eggs had a higher percent survival than skein eggs but were not significantly different (Table 2). Survival after hatch was greater for skein fry initially for the first two months, but leveled off with ripe fry survival becoming slightly higher in the fall. INTRODUCTION: Bloater (Coregonus hoyi) is a native species within the Great Lakes whose populations have been extirpated from Lake Ontario as a result of overharvesting by commercial fishing and competition by non-endemic species including Alewife (Alosa pseudohorengus). Currently, there is a multi-agency and binational initiative between the US Fish and Wildlife Service (USFWS), Ontario Ministry of Natural Resources and Forestry (OMNRF), N.Y. Department of Environmental Conservation (NYSDEC), Great Lakes Fisheries Commission, and U.S. Geological Survey (USGS) to reestablish bloater back into Lake Ontario. The Tunison Lab of Aquatic Science (TLAS) in Cortland, NY and White Lake Fish Culture Station (WLFCS) in Ontario, Canada are developing specific hatchery techniques to rear bloater for re- introduction into Lake Ontario. By utilizing a stable population captured from Lake Michigan, we have developed several techniques for yielding successful bloater within a hatchery setting. We experimented with “skein-bound” eggs; eggs enveloped within ovarian tissue, as a potential alternative egg source to produce viable offspring (Figure 1). Acquiring large quantities of viable eggs from Lake Michigan has been previously problematic due to poor environmental conditions, low catch rates, and low collection rates resulting from the expelling of free flowing (ripe) eggs from stress during capture. The utilization of skein eggs due to the large quantity available may boost bloater restoration efforts. The objective of this study was to highlight the early life ontogenetic milestones of bloater development from skein-bound and ripe eggs through hatch as means to compare their overall survival. CONCLUSIONS: The potential utilization of skein-bound eggs as a viable alternative egg source allows the reintroduction effort to substantially increase the overall quantity of eggs obtained. These results show that although skein-bound eggs are smaller in diameter, the overall survival was not significantly different through all egg developmental milestones. This information enables the reintroduction effort to successfully utilize skein-bound eggs as an additional source for yielding viable offspring. Implementation of these results will allow management agencies the capability of increasing overall bloater restoration efforts and have the potential to achieve targeted egg quantity goals allowing for greater stocking potential into Lake Ontario. Further observation of long term survival including behavioral and physiological deficiencies still needs to be assessed. Figure 2: Bloater collection sight and distributions Figure 1: Skein-bound eggs being removed from bloater females Figure 3: Skein-bound eggs in jar Figure 4: Ripe eggs in jar Figure 5: Developmental milestones of bloater egg development Table 1: Ripe versus skein egg diameter at fertilization and eye up stages of embryogenesis. Ripe eggs contained an overall larger diameter but a drastic increase in diameter between these two developmental milestones occurred with skein-bound eggs. Table 2: Overall survival of ripe versus skein-bound eggs from embryogenesis through stocking. Although ripe eggs had an overall higher survival, no significant difference occurred. Between May-July skein-bound fry experienced greater survival than ripe fry.