Columbia River Project Water Use Plan Columbia White Sturgeon Management Plan Implementation Year 1 Reference: CLBWORKS-24 and CLBWORKS-34 Sturgeon Conservation Aquaculture Study Period: May 2007 – May 2008

Author(s) and Project Manager: FFSBC

May 1, 2008

COLUMBIA WHITE STURGEON CONSERVATION FISH CULTURE PROGRAM

KOOTENAY STURGEON HATCHERY

2007 Annual Report

Columbia White Sturgeon 2007 Annual Report

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TABLE OF CONTENTS 1.0 Executive Summary.…………………………………………………....…………3 2.0 Brood Capture…………………………………………………………….………..6 3.0 Transport …………………………………………………………….…………….11 4.0 Adult Holding..………………………………………….………………………….11 5.0 Spawning………………………………………………………………………… 12 5.1 Spawning Summary …………………………………………...18 6.0 Broodstock Release……………………………………………………………….19 7.0 Vemco Tagging Adult Broodstock…….……………………….………………...20 8.0 Incubation & Larval Development……………………………….…...………….20 8.1 Record Keeping …………………………………………………………..22 8.2 Juvenile Rearing ………………………………………………………….22 8.3 Monthly Reports ………………………………………………………….23 9.0 Juvenile Releases…………………………………………………………………24 10.0 Fish Health Testing Summary …………………………………………………28 10.1 Viral Screening for Broodstock …………………….…………………28 10.2 Juvenile Screening History ……………………………………………..28 10.3 Deformities ……………………………………………………………...29 11.0 Permits …………………………………………………………………….……..29 13.0 Presentations and Meetings……………………………………………………30 14.0 Research Projects………………………………………………………….……30 14.1 Cryopreservation………………………………………………..……….30 14.2 Washington Hatchery …………………………………………..………30 14.3 DNA Sampling ………………………………………………….……….30 14.4 Sonic Tagging……………………………………………………………30 14.5 Vemco Tagging Adults…………………………………………..……...30 14.6 Juvenile Substrate Preferences Study ..……………………………..30 14.7 Toxicology Study- University of Sask & Teck Cominco……………...31 14.8 Multiple Spawning Experiment………………………………...……….37 Appendix 1: Monthly Hatchery Reports Appendix 2: Multiple Spawning Experiment Report

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1.0 Executive Summary Conservation fish culture and release of juvenile white sturgeon into the Upper Columbia River as part of the Upper Columbia White Sturgeon Recovery Initiative met and exceeded targets of 16,000 juvenile sturgeon in Brood Year 2007. Brood capture was conducted June 4th to 27th 2007 in the Lower Columbia River including: Waneta Eddy, Kootenay Eddy, and areas downstream of Hugh L. Keenleyside dam. Five female and nine male sturgeon were captured and transported in a trailer-mounted transport tank to the Kootenay Sturgeon Hatchery. Four of the five female sturgeon were spawned with five males creating nine families. One female sturgeon captured did not mature further in captivity and was released back into the Lower Columbia River at the location of capture. In addition, four surplus immature males did not mature in captivity and were returned to the river at the location of their capture. The TWG selected six of the families generated by the spawning events to be raised as juveniles and the remaining three families were culled. The spawning events of June 12th, June 27th and July 4th produced, 572,000 green eggs. The TWG decided to cull three families from June 27 spawning fish which eliminated 316,000 eggs; the FFS conducted a fertilization study using 136,000 eggs and researchers (UBC and USask) obtained 30,000 neurulated eggs. The remaining 90,000 eggs were used to produce juvenile fish for release. In past years first-feeding fry sturgeon were introduced to solid diet using the commercial diet Bio-Oregon. This year, Bio-Oregon feed was unavailable and after consulting other sturgeon hatchery managers a custom-made diet was formulated from a base diet of Skretting salmon starter feed to which krill hydrolysate, anchovy oil, freeze dried blood worms and the commercial product Cyclopeeze (zooplankton; Argent Chemicals Inc.) were added. The sturgeon appeared to find this mixture palatable and took well to first feeding. Columbia White Sturgeon 2007 Annual Report

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In the 2006 brood year, deformities were noted in many of the offspring. This phenomenon was not repeated in this brood year. From a culture perspective, only rearing density and diet were the apparent differences in early life rearing.

After culling three of the nine half-sib families, 17,011 juvenile sturgeon were reared further, tagged and marked and released. Release occurred in both the fall of 2007 (Lower Columbia) and the spring of 2008 (Mid and Lower Columbia). Average size of released fish was 20g and 50g (no SEM provided) for fall and spring release, respectively. A subset of the supplemental progeny were reared for a longer duration and grown to 150-200g to allow for the insertion of acoustic transmitters (Vemco model V7) for monitoring their movements following release. A total of 4,029 juvenile sturgeon were released into the Canadian Lower Columbia River in two release events on November 14th at Beaver Creek and November and 15th below Keenlyside Dam near Castlegar. As well, for the second consecutive year, juvenile sturgeon (total of 7,048) were released during daylight in the spring of 2008 into the Canadian Mid Columbia River in the Revelstoke area, Beaver Creek and below the Hugh Keenlyside Dam. A further 5,934 sturgeon juveniles were released at night in the Mid Columbia into what is locally referred to as Big Eddy, immediately downstream of Revelstoke dam. The night release also included 50 larger juvenile sturgeon that were implanted with Vemco transmitter tags. In addition, a small number of juvenile fish were released during the day by four local elementary school classes to foster public involvement and awareness.

Two large public events release events took place in 2008. The first event was held at Beaver Creek on May 1st with 350 school children and many volunteers helping to release the sturgeon. The second event took place in Castlegar on May 2nd with at least as many children involved.

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In terms of fish health work, the FFSBC Health Lab again tested sturgeon samples for several viruses (IPNV, WSHV1, WSHV2 and WSIV). This testing was done on adults and juveniles from each family as defined by Federal and Provincial fish transplant permits. Results from these tests were negative.

In the past, milt samples had been collected and sent to Dr. Joe Cloud at the University of Southern Idaho for cryopreservation work. Because of funding limitations no milt was collected for this purpose. As numbers of potential broodstock decline in the Lower Columbia River, cryopreservation is likely to become an important tool and it will be imperative in the near future to find resources to support this work. DNA samples were collected by taking tissue samples from the dorsal fin of all adults and from a portion of each half-sib family. These tissues are stored on site at the Kootenay Sturgeon Hatchery for future use and reference.

The challenges of this year’s white sturgeon recovery work included reduced capture success that was met by increased effort in known spawner staging areas and altering set line equipment. Transport stress for adults was ameliorated by oxygen supplementation (to excess) and the addition of salt (NaCl; 5 ‰) to the transport water. Fertility, neurulation and hatch rates were as high as or higher than previous years. A decrease in deformed larvae was noted in comparison to the previous year with almost no deformed animals culled from the population. The use of natural feed supplements in the first feed and reduced rearing densities was thought to contribute to the decrease. The inability to obtain the type of first feed previously used was met by using a commercial salmon started feed augmented with several natural additives that were top-dressed onto feed. This diet resulted in high first-feeding rates.

Surplus fish were again produced and all quotas for released fish were achieved. There were two public daytime release events and two daytime, one nighttime stocking events. It was determined that nighttime releases are preferred Columbia White Sturgeon 2007 Annual Report

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because releasing fish during nighttime low flow events decreases the risk of stranding released fish during dewatering.

Fish health was excellent and is attributable to fish husbandry practices and support from the FFS Fish Health Unit.

Experimental work included toxicological studies conducted by the University of Saskatoon and investigations into fertilization delay post-initial ovulation conducted by FFS staff.

2.0 BROOD CAPTURE FFSBC staff assisted B.C. Hydro and the UCWSRI TWG personnel in the capture and identification of mature Columbia River sturgeon. Fishing efforts were concentrated downstream of Hugh L. Keenleyside dam (HLK). Two methods were used to capture mature adult white sturgeon. Sturgeon capture was by the use of baited hooks either connected to a set line and left over night, or presented to fish through active angling. Once captured, FFSBC staff ensured that sturgeon were handled in accordance with the Upper Columbia White Sturgeon Recovery Initiative (UCWSRI) Upper Columbia Adult White Sturgeon Capture, Transport and Handling Manual (UCWSRI 2006), and that data collection from prospective spawners met the protocol provided by the UCWSRI and regulators. Mature adults were examined for reproductive condition, and where acceptable, transported to the Fort Steele, FFSBC Kootenay Sturgeon Hatchery (KSH) facility for holding and spawning. The established annual spawner target is for 10 females and 10 males. Suitable holding and transport facilities were provided to move spawners from the river to the KSH in a safe and timely fashion. The details of the 2007 sturgeon capture and transport efforts follow.

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Capture techniques for adult sturgeon were as detailed in the Upper Columbia Adult White Sturgeon Capture, Transport and Handling Manual, Section 2.0 Capture Techniques using the longline method described. The observation of stripped hooks has been attributed to the presence of smaller sturgeon which remove the bait from the hooks which for some younger fish are too large for the sturgeon to take into their mouths. To address the issue of sub-adult sturgeon taking bait, the programme was altered to: a) using a larger hook to exclude sub adult fish, b) using larger bait sizes, and c) tying the bait onto the hook to make it harder for the small sturgeon to remove it. These practices have been shown to be effective in other brood capture areas such as the Nechako River. To increase adult sturgeon catch rates, the smaller 12/0 hooks were all replaced with larger 16/0 hooks. Bait used was 150 – 250g fillets of Eastern brook charr (pickled in salt and garlic; increases ‘scent’) or thawed Kokanee filletts. Lines were set in the afternoon, left overnight and checked the following morning.

Brood captures were conducted from June 4th to June 27th 2007 and resulted in nine males and five females being taken back to the KSH. Brood capture was not as successful as in the past few years as high numbers of KSH hatchery released juveniles are collected as by-catch. These hatchery released fish are readily identified by scute marks in addition to PIT tags. Lower capture rate of maturing sturgeon resulting in additional effort (i.e. more set lines) at Waneta and expansion of the sampling distribution to include sites upstream of Kootenay Eddy and below HLK. These late season efforts were successful in capturing two additional mature females at the Waneta and Kootenay Eddy areas.

Most of the males captured and chosen as suitable hatchery candidates, were flowing with milt at the time of capture, but three males checked by incision and examination by using an otoscope were suspect for later maturation. These fish were transported back to the hatchery as an experiment to see if they could be brought to full maturity by hormone injection. Subsequent hormonal treatment with IM 50µg/kg LHRHa in saline for these fish proved unsuccessful in bringing Columbia White Sturgeon 2007 Annual Report

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these three males to spermiation. Based on experiences with brood collection in this year and in previous years, we recommended that only flowing males or males with extremely large, white, lobed testes be taken back to the hatchery.

Female fish identified by either PIT tag (catch/use history) or external examination (larger size, rounded abdomen) were given a biopsy using an otoscope through an incision and if eggs were present, a sample removed. Immature fish were sutured if examined using the otoscope and returned to the river. Gravid fish were retained for transport. Samples of eggs collected from each gravid female were boiled in Ringers’ solution for 5 min and fixed in 10% formalin on the boat. These samples were transported back to the KSH so that initial polarization index (PI) calculations could be determined. Further procedural details and information on brood capture practices that were followed this year can be found in the Upper Columbia River Adult White Sturgeon Capture, Transport and Handling Manual 2003 produced by Golder Associates. Capture information for adult white sturgeon hatchery candidates including dates, size, location, PIT tag number and release dates are presented in Table 1. Table 2 displays capture history of the fish caught and held for spawning in 2007.

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Table 1: Adult sturgeon capture information Spring 2007. Male fish under column ‘Spawning Date’ with entries ‘Not spawned’ indicates that the fish were prospective candidates, but did not mature fully and were returned to the river after morphometric data were collected. One female (PIT tag 42657C7B47) captured June 5th was transferred to the hatchery, but failed to complete maturation and was returned to the river July 7th.

2007 Adult Information Sex

PIT

Capture Date

Weight kg

Capture Location

Release Date

Comments

Waneta

Spawning Date not spawned

F

42657C7B47

Jun-05

68

Jul-07

Spawned 2003

F

7F7C77126C

Jun-06

80

Waneta

Jun-12

Jun-25

F

985161000057769

Jun-19

73

Kootenay Eddy

Jul-04

Jul-06

F

415869408

Jun-22

120

Waneta

Jun-27

Jul-05

F

7F7D186E66

Jun-22

62

Kootenay Eddy

Jun-27

Jul-28

M

985120025166158

Jun-07

35

Waneta

Jul-06

M

426465351D

Jun-07

52

Ft Shepard

Jul-04 Not spawned

Jul-05

Not Spawning 07

M

7F7D490B02

Jun-08

37

Ft Shepard

Jul-04

Jul-06

Flowing

M

7F7D4F2F69

Jun-08

40

Waneta

June 12+27

Jul-07

Flowing

M

985161000065272

Jun-15

51

Waneta

Jul-08

M

985120019237290

Jun-15

33

Kootenay Eddy

Jun-28

M

985120019232551

Jun-15

37

HKL

Jul-04 not spawned not spawned

Jun-28

Flowing Not Spawning 07 Not Spawning 07

M

7F7D21605B

Jun-16

45

Ameircan Eddy

Jun-27

Jun-29

Flowing

M

4265646344

Jun-26

39

Waneta

Jun-27

Jun-29

Flowing

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Table 2: Broodstock capture history of fish caught during the 2007 brood acquisition and successfully spawned at the Kootenay Sturgeon Hatchery.

Capture Date

Fork Weight Length kg 142 28.2 142 168.5 52.3 181.5 62

PIT Tag Number

Sex

7F7D186E66

F

07-Jul-92 07-Nov-92 09-May-03 22-Jun-07

4158696408

F

26-Jun-92 18-Dec-97 16-Apr-98 16-Apr-02 22-Jun-07

185 199 194 226.5 225.5

59.1 65.5 64.1 86.2 120

1F6C77126C

F

16-May-97 06-Jun-07

173 207

43.2 80

985161000057769

F

19-Jun-07

205

73

7F7D4F2F69

M

11-Aug-93 08-Jun-07

123 170

18.2 40

7F7D21605B

M

04-Oct-96 19-Jun-07

170 181

45 45

4265646344

M

15-Mar-01

111

11.1

26-Jun-07

166.5

39

985120025166158

M

07-Jun-07

166

35

7F7D490B02

M

19-Jul-92 01-Jul-93 08-Jun-07

156.5 155.5 168

N/A 35 37

Personnel involved on the boats this year included: Ron Ek, Owen Schoenberger, Chad Fritz (FFSBC) Bob Chapman (Golder and Associates), Columbia White Sturgeon 2007 Annual Report

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Dean den Beisen and Sean Ord (BC Hydro), Matt Neufeld (Min of Environment),. It was made a priority this year to have Chad Fritz (under the supervision of Ron Ek) conduct the surgeries on as many of the adults as possible, so that he became accustomed with the adult maturity assessments and suturing surgical incisions.

3.0 TRANSPORT When an adult was captured and determined to be mature or maturing, it was loaded into the sturgeon transport tank using a stretcher or a tube net at the nearest access point to the river. The transport tank was filled with ambient river water using a fire pump. To minimize stress during transport and to facilitate healing abrasions that may have occurred during capture and handling, salt (heavy metal free sodium chloride) was added to the tank water to make a solution of 5 ‰. Oxygen was supplied to saturation to the tank through aeration stones that were recessed into the tank floor and was not monitored during transport. Twice during each transport, staff checked the fish for duress and none was noted. All fish faired well during transport from the site of capture to the KSH with no discernable negative effects of transport apparent on arrival and placement into tanks at the KSH. Transport times were approximately 4h. On arrival, hatchery tank temperatures were matched with transport tank temperatures. The fish were transferred from the transport tank to the culture tank using a tube net and physical labour. Fish were monitored hourly following arrival at the hatchery until staff were off duty and again the following morning. No adverse effects of transport were noted.

4.0 ADULT HOLDING Upon arrival at the hatchery, the adult sturgeon were placed in fiberglass tanks, and monitored for health and reproductive status as evidenced by the presence of milt or eggs. The tanks used are located in the KSH and in the C 1-4 area. Generally, tank densities are limited to up to four males or up to two females, Columbia White Sturgeon 2007 Annual Report

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depending on size and maturity of the fish. If a female is thought to be close to spawning (visual inspection) she may be placed in a separate tank in preparation for an early spawn.

All fish are initially held on flow through warmed ambient ground water at 10°C. When the brood capture is completed water temperature for female fish in holding is increased to by 1-2°C per day to 16°C. Water temperatures for female fish may remain elevated until maturation and spawning are complete. This period may encompass weeks to months depending on the maturation of the fish.

Female fish are not fed during their residency at KSH. Male fish are offered live Eastern brook trout fingerlings (5-10g) ad libitum starting with 40 fish on arrival and when depleted, an additional 40 fish may be added depending on how long the fish have been in captivity and the rate of consumption. Fish used as feed come from stocks designated as disease free.

Water flows through the holding tanks were held at levels to ensure near oxygen saturation levels. Oxygen levels and flow rates were monitored ad hoc on an ongoing basis as part of the daily routine and adjusted by supplementation from canisters where needed. Although tanks were covered with fiberglass lids, fish were not kept in total darkness and were exposed to a simulated natural photoperiod.

5.0 SPAWNING As stated, females undergo a maturation check when captured to determine the approximate state of maturation. The timing of breeding potential is tracked using egg stage polarization index (PI) of the germinal vesicle (GV) and germinal vesicle breakdown (GVBD). Based on a subjective evaluation of the PI conducted immediately after capture, the condition of the fish and a visual appraisal of the biopsy eggs, another biopsy will be done within 3-4 weeks after exposure to elevated water temperatures (16°C). Columbia White Sturgeon 2007 Annual Report

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For PI checks, a small sample of eggs (~20) is removed through a small surgical incision in the abdominal wall. These eggs are placed into a 150ml beaker of Ringers’ solution, placed on a hot plate and boiled for five to eight minutes. This hardens the yolk and fixes the position of the nucleus (GV). The sample is cooled by placing it on ice and when cool, drained and placed in a 10% formalin solution overnight. To determine GV position relative to the center of the egg and the periphery, the egg is bisected and examined under a dissecting microscope. This procedure is accomplished by grasping the egg with a pair of tissue forceps and making a cut along the animal-vegetal axis using a sharp single-edged razor blade. The position of the GV is then observed, evaluated and represented as a proportion of 1.0. For example, a PI of 0.15 would mean a GV migration of 85% of the distance between the center of the egg and the periphery. As observed in common practice, PI levels less than 0.10 have been shown to be optimal to begin a spawning induction treatment with the peptide human luteinizing hormone releasing hormone analogue (LHRHa). When a female has been determined to be ready for spawning induction she may be placed into a separate tank and injected with LHRHa.

In the 2007 spawning season, all four female fish captured had initial PIs that were near or below the benchmark of 0.10 needed for induction of spawning. Female 1 (PI = 0.073) was induced to spawn (see below for procedural details) four days after arrival at KSH (June 10/11th) and was spawned June 12th. The three successive females were captured on June 19th (Female 2), June 21st (Female 3) and June 22nd (Female 4). The PI values for these fish on capture were, respectively: 0.110, 0.101 and 0.029. Females 3 & 4 were induced on June 25/26th and spawned on June 27th. Female 2 was induced to spawn July 2nd/3rd and was spawned July 4th. Table 3 details adult spawner information for male and female fish.

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Table 3: Female and male white sturgeon spawning information in 2007. Female No. Capture Date Location Weight (kg) Pit Tag No. Vemco No. Tag Code PI Start PI Finish LHRHa Date Dose Temperature Spawn Date Male Crosses Male No. Capture Date Weight (kg) Pit Tag No. Vemco No. Tag Code LHRHa Date Dose: 40/10µg/kg Temperature Spawn Date

1 June 6

June 19

3 June 21

4 June 22

Waneta 80

Kootenay Eddy 73

Waneta 120

Kootenay Eddy 62

7F7C77126C

985161000057769

4158696408

7F7D186E66

1041279 5417 0.073

1041281 5419 0.110

1041278 5416 0.101

1041280 5418 0.029

Not Recorded

Not Recorded

Not Recorded

Not Recorded

June 10/11 40/10µg/kg 10 – 15°C June 12 4

July 2/3 40/10µg/kg 10 – 15°C July 4 1&2

June 25/26 40/10µg/kg 10 – 15°C June 27 6

June 25/26 40/10µg/kg 10 – 15°C June 27 4&5

1

2

2

3

4

5

6

June 2

June 8

June 15

June 8

June 26

June 16

35 25166158 1041272 5411

37 490B02 1041273 5412

51 65272 1041272 5410

40 F2F69 1041269 5407

39 646344 1041267 5405

45 7F71605B 1041268 5406

July 2

July 2

July 2

June 10 June 24

June 24

June 24

15°C July 4

15°C July 4

15°C Not used

15°C June 12 June 27

15°C June 27

15°C June 27

The LHRHa treatment regimen for female sturgeon consists of two doses of LHRHa given 8 hours apart: a loading (90%) and resolving (10%) dose. Total dose is 40/10µg/kg or on occasion higher depending on the size of the female and a subjective evaluation of overall state of maturation. Female fish will begin to ovulate and release eggs 24 hours after the resolving injection of LHRHa. Once a female has been observed to be releasing eggs, as evidenced by the presence of eggs on the tank floor, the water level is dropped in the tank. Staff enter the tank and place the fish ventral side up onto a hooded stretcher with a Columbia White Sturgeon 2007 Annual Report

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water hose providing fresh water flowing over the gills. Sufficient egg volumes to provide for the targeted juvenile numbers are collected from the female using manual expression through the urogenital opening or by extraction through an incision using a modified cesarean section method. To meet mid Columbia larval targets, cesarean sections may be more frequently used as this method can sometimes more fully permit the expression of ovulated eggs.

Male fish are held at 10°C until they are needed to supply milt. When a female has been assessed as mature and ready for induced spawning, 1-3 days prior to intended use male fish are intramuscularly injected with a single bolus dose (50µg/kg) of LHRHa in saline. At the time of the injection, the water temperature in the male tanks is increased to 16°C for the time remaining until after spawning. Once spawning is over, the water temperature in the male sturgeon tanks is decreased back down to 10°C.

Milt is collected several hours before egg fertilization is conducted to minimize workload and milt remains viable for several hours when kept on ice. To collect milt the male is held ventral side up on a hooded stretcher with the fishes’ vent exposed. The urogenital area is dried using paper towels, and care is taken to prevent contamination of the sperm with water as it is collected. Milt is collected using a 5.0 cm length of Tygon tubing attached to the nipple of a 20-50 cc plastic syringe. Initially, the plunger of the syringe is depressed when the Tygon tubing is inserted into the genital opening located just posterior to the anus. Milt is collected by massaging the abdominal area over the testes to force milt into the collecting ducts and slowly withdrawing the plunger while moving the tubing back and forth in the genital opening. This technique allows for the removal of large quantities of milt without contamination by urine, feces or water.

Once eggs are collected, excess ovarian and coelomic fluids are decanted from the eggs as these fluids negatively impact sturgeon sperm motility. Approximately 1,000 eggs (~ 250-375ml) are decanted into a clean 2l stainless Columbia White Sturgeon 2007 Annual Report

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steel bowl. For fertilization of the eggs, a diluted milt solution (1:20 milt:water; 20ml milt to 1l water) is added to the eggs. The mixture is gently and slowly turned over by hand using a feather. Mixing continues until the first few sticky eggs are observed, usually in 2-3 minutes. Excess fluid is again poured off, and a de-adhesion solution is added that contains diatomaceous (Fullers’) earth. This solution coats the adhesive eggs and prevents the eggs from clumping. The deadhesion process of turning eggs over using a feather in a stainless steel bowl continues for an hour or until all of the eggs have lost their stickiness. Total spawned egg numbers are then estimated volumetrically using counts of 3 subsamples aliquots of 25ml of eggs.

Of the five females that were transported and held at the Kootenay Sturgeon hatchery, four were successfully induced to spawn. Each of the five females was checked regularly to determine their ripeness. The procedure for the maturation determinations followed methods described in the Broodstock Evaluation section of the Hatchery Manual for the White Sturgeon, University of California. The female that did not mature and spawn was implanted with a sonic Tag and released so it could be tracked if it subsequently went to the natural spawning area and possibly spawn. Throughout the 2007 spawning season, there were no irregularities seen in the above routine. Once the fish were under care at KSH protocol was followed and the fish responded to treatment as expected.

Total egg volume and number was not recorded for individual females for the 2007 spawning season nor were all ovulated eggs taken from spawning female fish. From the three spawning events on June 12th, 27th and July 4th, a total of 572,000 green eggs were collected. Of these eggs, the TWG decided to cull 316,000 eggs from Families 7, 8 and 9. The FFS used 136,000 eggs in a delayed fertilization study (see Section 14.8 below). Researchers at the UBC and USask Teck Cominco used 30,000 neurulated eggs for further experimental work (see Sections 14.6 and 14.7, respectively). Thus, 90,000 eggs remained to produce juvenile fish for release. The release targets of 4,000 juveniles for release at Columbia White Sturgeon 2007 Annual Report

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Revelstoke and 12,000 below the Hugh L. Keenleyside dam were used as an indication of approximate egg needs to produce enough juvenile fish to meet the demands. MacDonald jar loading was 0.9l of fertilized eggs per jar (one family). As sturgeon eggs have been very close to 23,000 eggs per ml over successive years, approximately 20,000 eggs were loaded, by volume into each MacDonald jar. With the target number set and the family numbers capped (by male and female crosses to make a family), a factor of about 10X is applied to the number of eggs taken and fertilized. This factoring includes fertility rate, neurulation to hatch rate, losses at feeding and juvenile survival rate in addition to excess production estimates of 100% extra juvenile fish.

For all the 2007 spawning events, one MacDonald jar of fertilized eggs per male female cross were incubated and denoted as a family. Thus, there were approximately 20,000 fertilized eggs per family for six families that were incubated separately. Families were kept separate throughout incubation, early rearing and juvenile rearing. Three pairings of male and female gametes where completed, but later discarded. There was a redundancy in spawning numbers that was meant to cover unexpected events such as low fertilization or neurulation rate or an environmental/mechanical event. In short, three pairings were made for insurance purposes and culled after neurulation. These pairings are shown in Table 4, Section 5.1 Spawning Summary.

Eggs are not counted into MacDonald jars for incubation; only an estimate based on sample counts of aliquots are used. Typically about 25ml of eggs are measured out and enumerated in three to four successive samples from one female. An average number of eggs per ml is then determined and that number applied to the volume of eggs placed in each jar. At six to eight hours post fertilization, a subsample of eggs may be removed to check on fertilization rate. In this case, about 25 – 50 eggs may be removed and the number of eggs that have reached the 4-8 cell stage enumerated and applied as a percentage to the calculated egg number. At neurulation, three subsamples of eggs are again Columbia White Sturgeon 2007 Annual Report

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removed and evaluated for normal development or dead. Inventory is likewise adjusted if required. However, all inventory numbers are estimates because no complete inventories are completed until fish are graded some weeks later.

In 2007, observations of fertility rate, neurulation rate, hatch rate and survival to first feeding were not recorded; these data were noted, but not entered in to the permanent record. As in past years, these data were considered secondary to juvenile production. Fish are not completely inventoried and enumerated until grading (see below Section 8.1 Juvenile Rearing).

5.1 2007 SPAWNING SUMMARY The four females that were spawned this year were crossed with five males to create nine families (Table 4). Spawning was conducted using a monogamous (single female with single male) breeding design. This breeding strategy followed the matrix suggested in past practices stemming from information received from the TWG (the Pollard Approach). Of these nine families, six were chosen to be raised and released as juveniles. The surplus fish were later euthanized. The decision on which ones to keep was made by the Columbia Sturgeon TWG (Table 5). Table 4: Male female sturgeon crosses that contributed to the formation of the initial families. Crosses 4, 5 and 6 (denoted by an asterisk after the female ID number) indicate that the family was culled and not released as juveniles. Female Spawn Date

(Capture Location)

Male

1:

1F6C77126C

(Kootenay) 7F7D4F2F69

2:

7F7D186E66

(Kootenay) 7F7D4F2F69

3:

7F7D186E66

4:

7F7D186E66* (Kootenay)

5: 6:

Capture Location (Kootenay)

June 12

(Kootenay)

June 25

4265646344

(Waneta)

June 25

7F7D21605B

(Keenlyside)

June 25

4158696408*

(Kootenay) 7F7D4F2F69

(Kootenay)

June 25

4158696408*

(Kootenay) 4265646344

(Waneta)

June 25

(Kootenay)

Columbia White Sturgeon 2007 Annual Report

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7:

4158696408

8:

985161000057769 (Kootenay) 985120025166158 (Waneta)

July 4

9:

985161000057769 (Kootenay) 7F7D490B02

July 4

(Kootenay) 7F7D21605B

(Keenlyside)

June 25

(Waneta)

Table 5: Final selection of sturgeon families released as juveniles. Family

Female

Male

1

1F6C77126C

7F7D4F2F69

2

7F7D186E66

7F7D4F2F69

3

7F7D186E66

4265646344

4

4158696408

7F7D21605B

5

985161000057769

985120025166158

6

985161000057769

7F7D490B02

6.0 BROODSTOCK RELEASE After hatchery spawning events, which can occur several times for males, fish are held for about three additional days and then returned to the Columbia River. This additional time in captivity is to assure the staff that the fish are recovered fully from the spawning event and that there are no fish health issues that should be addressed prior to release. Before fish are returned to the river a DNA sample (fin clip) is collected. This sample is placed in ethanol, labelled and stored on site in a secure dry area at the KSH. Fish are also re-checked for the presence of a PIT tag to ensure future identification. The same holding and transport equipment used to transport fish from the river to the hatchery is employed to carry them back to the river, where they are released as near as possible to the capture area. For 2007 adults released back into the wild at both the Robson boat launch (Ft. Shepard and Kootenay Eddy captures) and at Beaver Creek (Waneta captures), water and weather conditions were ideal (fair weather) and water temperatures matched hatchery and transport temperatures at 15°C. All

Columbia White Sturgeon 2007 Annual Report

20

adult releases were completed without incident and all fish appeared well at time of release.

7.0 VEMCO TAGGING ADULT BROODSTOCK Thirteen of the mature adults taken to the hatchery were implanted with V16-6HR64K Vemco coded transmitters (Vemco tags;Table 6). These implanted tags allow for fish movements to be tracked when they pass by stationary VR2 receivers that are situated in various locations in the river. These Vemco tags have a life of 3,000 days or nearly 10 years. This will allow movement data to be collected on these adults to determine if or when they return to spawn and if there are additional spawning areas being used in the river. Table 6: Vemco tag and PIT tag number for adults released at the Beaver Creek location after spawning. Tag Serial Number

ID Code

PIT Tag Number

Sex Size Kg

Date

1041266 1041267 1041268 1041269 1041272 1041273 1041274 1041278 1041279 1041280 1041281

5404 5405 5406 5407 5410 5411 5412 5416 5417 5418 5419

42657C7B47 4265646344 7F7D21605B 7F7D4F2F69 985161000065272 7F7D490B02 985120025166158 4158696408 7F7C77126C 7F7D186E66 98516100057769

F M M M M M M F F F F

July 6 July 5 June 29 June 29 July 5 July 6 July 6 July 5 June 25 June 28 July 7

68 39 45 40 51 37 35 120 80 62 73

Release Location Beaver Creek Beaver Creek Beaver Creek Beaver Creek Beaver Creek Beaver Creek Beaver Creek Beaver Creek Beaver Creek Robson Robson

8.0 INCUBATION AND LARVAL DEVELOPMENT Fertilized eggs were placed in MacDonald Jars for incubation with water outflow from the jars directed into aluminum, free-embryo troughs. Jars were positioned over individual troughs that were labelled and segregated by family. FFSBC staff ensured adequate flow to maintain egg separation and oxygenation, while guarding against egg loss from jars as they become more buoyant during Columbia White Sturgeon 2007 Annual Report

21

development. Dead eggs were removed at intervals throughout the day to control the development of fungal infestations. Egg condition and number were monitored to ensure juvenile and larval release goals were met.

Hatch out for eggs was between 8-10 days post fertilization at 15°Cfor all families of the 2007 brood year. Embryos emerged from the MacDonald jars and were flushed into aluminum troughs that contain 10-12 lpm flowing water of about 10cm depth. Water level is controlled by a standpipe and larvae are protected from the exit flows by a stainless mess screen. Water flows are set to exchange water, but not unduly disturb larvae and cause them to swim. Overhead partitions on the troughs provide cover for the larvae.

After about 10 days the feed is introduced into the larval tanks A custom formulation is produced at the hatchery that contains standard Skretting salmon/trout starter feed with small amounts of anchovy oil, freeze dried blood worm (chronomid larvae), dried krill powder and the commercial product Cyclopeze (Argent Chemicals). Proportions of the ingredients varied with the progress of the larvae, but in general, the additives represented one-third of the feed mass at the beginning of feeding and progressed to Skretting feed by about the 1g stage.

Feed is presented to feeding larvae by hand in two methods. At first, feed is continuously (24h) applied to the water surface and pressed to the trough wall at the waterline. Young sturgeon rise to feed on the vertical surfaces after their primary introduction to feed on the bottom of the trough. Feeding is done on an ad libitum basis as directed by the fish culturist. As fish develop, feed is delivered to the tank wall and water interface by a belt feeder in excess of need.

Common fish culture practice of sturgeon mimics that of trout culture in that tanks and screens are cleaned throughout the day on an ‘as needed’ basis. The monitoring of fish health and feeding activity is likewise observed during daily Columbia White Sturgeon 2007 Annual Report

22

routine. In this fashion, the care and culture of sturgeon, especially for the younger, more vulnerable life stages is continuous throughout the working day.

8.1 RECORD KEEPING Sturgeon culture record keeping in 2007 was driven by release numbers. The number of released fish is the main determinant of how many eggs are taken, the intensity of culling fish that are showing signs of malformation, weakness or duress, and the stringency of grading. It is of utmost importance to record inventory, tag numbers and morphological data for release fish in addition to assuring there is a clear health record backed up by Fish Health Lab certification.

Incubation and rearing data for egg, larvae and juvenile sturgeon were not entered into a permanent log for the 2007 brood year as it was seen as superfluous to the overall objective of attaining release numbers. This is not to state that the data were not gathered only that it was not entered into a permanent hatchery log. Decisions as to critical husbandry practices were based on culture data and observations; once developmental milestones were achieved, the information was discarded. Clearly, the goal in 2007 was to produce release numbers that were supported by complete morphometric records at time of release; other prior culture information was not regarded as important to achieving the release objective. With the signing of the BC Hydro/FFSBC Contribution Agreement, more complete fish culture details will be captured.

Over the period of the 2008 – 2011 Contribution Agreement, full data sets will be a matter of permanent record as established in the agreement. 8.2 JUVENILE REARING At grading, fish are hand-picked into either large or small categories and placed into separate tanks. This is the first instance of inventory. Numbers for all prior milestones of development are then back-calculated from this point. The splitting

Columbia White Sturgeon 2007 Annual Report

23

of the two sub-populations has two effects. Firstly, the splitting of the group decreases densities for the two new populations of fish and removing tank effects for growth restraint. Secondly, non-competitive access to feed is important to the smaller, downgraded fish. These fish will recover from any feeding competition and quickly establish a higher growth rate. As post-release survival is putatively thought to be influenced by size, it is commonly practiced to grow the fish as large as possible to time of release. Additionally, the larger graded fish need an appropriate feed size to continue a positive growth pattern.

As well as a size selection during grading, culturists remove malformed or damaged fish in addition to fish with overt fish health signs. The latter fish are examined and if required, sent to the Fish Health Unit for further examination. In the 2007 season, the levels of deformities witnessed in the 2006 were not repeated and the total percentage of deformed fish for all stages was less that 1% of the population as determined by back calculation at the time of grading and inventory.

During grading, smaller fish remain in troughs or smaller circular tanks until they catch up on growth. Further grading and culling may occur, but care is taken to ensure that smaller fish are not excluded from the population that will contribute to the final release numbers. Culls for population density control are from all tanks in equal measure to ensure that artificial genetic selection is minimized. Briefly, fish are randomly selected from rearing containers using small nets and counted out into a vessel containing 500mg/l TMS (tricaine methanesulphonate) according to FFSBC Standard Operation Procedure: Euthanasia. Culling continues until the desired numbers of fish remain in the culture container. 8.3 MONTHLY REPORTS After grading and inventory is complete, the data are entered into the permanent record. The monthly reports for fish culture activities of late summer 2007 to spring 2008 appear in Appendix 1. Columbia White Sturgeon 2007 Annual Report

24

9.0 JUVENILE RELEASES 2007 Sub–yearling sturgeon are typically released annually in the spring of the year at and average size of 20 cm and 50 gms in weight (SE not detemined; Table 7). A portion of the fish to be released were grown to a larger size (150 gms) that permitted them to be implanted with Vemco tags (Table 8). Alternatively, if seasonal release experiments are incorporated into a study design, juvenile releases may occur in the fall. Prior to release, juvenile fish of approximately 50g are individually handled to insert a PIT tag into the dorsal musculature at the midpoint between the dorsal and lateral scute line inferior to the anterior margin of the dorsal fin. The PIT tag number, length and weight data are recorded for each fish. Each individual fish can subsequently be identified to its release location and date of release in addition to family record. Juveniles are transported in FFSBC fish transport vehicles according to UCWSRI TWG transport protocols. Juvenile releases in the mid Columbia near Revelstoke take into consideration the load shaped hydrograph by releasing fish during periods of elevated flows to minimize the likelihood of stranding. Water quality conditions at the time of the release were favourable for water flow (moderate) and temperature (8°C). Weather conditions at the time of release were ideal. In addition, FFSBC staff help coordinate and participate in public awareness and educational activities during the release events. Table 7: Juvenile sturgeon release numbers and total kilograms released Fall 2007 and Spring 2008 for the 2007 year class. 2007 Year Class Releases: Total Fall Release Total Spring Releases

4,029 12,982

148.7 Kgs. 818.4 Kgs.

Combined Releases

17,011

967.1 Kgs.

Release Summary - Fall 2007 BY SITE Date

Family

Columbia White Sturgeon 2007 Annual Report

#

Kgs

Wt g

Average FLcm

25

Beaver Creek

3 4 5 6

400 295 576 579 1850

17.1 10.2 18.8 19.3 65.4

42.7 34.5 32.7 33.4

17.5 16.3 16.1 16.3

HLK

1 2 3 4

870 725 284 300 2179

38.0 22.2 12.7 10.4 83.2

43.7 30.6 44.7 34.5

18.1 15.7 18.2 16.4

4029

148.7

Kgs

Wt g

Total

BY FAMILY Family

Date

#

Mean FLcm

18.1

Beaver Creek HLK

1 Nov. 14/07 Family Total

870 870

38.0 38.0

43.7

Nov. 14/07

725 725

22.2 22.2

30.6

15.7

Beaver Creek HLK

Nov. 15/07 Nov. 14/07

400 284 684

17.1 12.7 29.8

42.7 44.7

17.5 18.2

Beaver Creek HLK

Nov. 15/07 Nov. 14/07

295 300 595

10.2 10.4 20.5

34.5 34.5

16.3 16.4

Beaver Creek HLK

Nov. 15/07

576

18.8

32.7

16.1

Beaver Creek HLK

576

18.8

579

19.3

33.4

16.3

Beaver Creek HLK

579 4029

19.3 148.7

2 Family Total

3 Family Total

4 Family Total

5 Family Total

6 Family Total Total

Site

Nov. 15/07

Columbia White Sturgeon 2007 Annual Report

26

Release Summary -Spring 2008 BY SITE Beaver Creek

HLK

Revelstoke Day

Revelstoke Night

Mean

Family 1 2 3 4 5 6

# 563 515 433 600 554 568 3233

Kgs 46.2 27.9 25.3 37.9 26.3 31.2 194.8

Wt g 82 54 58 63 48 55

FLmm 22 19 20 21 19 19

1 2 3 4 5 6

581 493 453 523 577 588 3215

49.4 30.4 31.8 37.0 35.7 33.1 217.4

85 62 70 71 62 56

22 19 20 21 20 19

1 2 3 4 5 6

100 100 100 100 100 100 600

8.0 5.5 4.5 6.3 5.7 5.1 35.1

80 55 45 63 57 51

22 19 18 20 20 19

1 2 3 4 5 6 Mixed Sonic

1022 913 749 1193 1007 1000

84.5 52.8 37.9 73.7 57.4 53.3

83 58 51 62 57 53

22 19 19 20 20 19

50 5934

11.5 371.1

229

32

12,982

806.9

Kgs 188.1 116.6 99.5 154.9 125.1 122.7 11.5 818.4

Wt g 83 58 57 64 56 54 229

All Locations

BY FAMILY Family 1 2 3 4 5 6 Mixed Sonic Totals

# 2,266 2,021 1,735 2,416 2,238 2,256 50 12,982

Mean FLmm 22 19 20 21 20 19 32

Columbia White Sturgeon 2007 Annual Report

Site

27

Table 8: Physical data for late-release 2007 juvenile sturgeon implanted with Vemco tags.

Vemco-Tagged Juvenile Releases April 29, 2008 -Location- Big Eddy- Revelstoke Fish

PIT Tag#

1

985120029758960 985120029785388 985120029782910 985120029787712 985120029797636 985120029798978 985120029799077 985120029991792 985120030014417 985120031139162 985120032408900 985120032436799 985120032480526 985120032481754 985120032524002 985120032527187 985120032533431 985120032405742 985120032411890 985120032480156 985120032518770 985120032529633 985120032530152 985120032543065 985120032546155 985120032549240 985120032408791 985120032411696 985120032446638 985120032479681 985120032519755 985120032523035 985120032523058 985120032528372 985120032539313 985120032527403 985120032528738 985120032531634 985120032540035 985120032543041

2 3 4 5 6 7 8 9

10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40

Sonic Tags Serial # ID Code

1049959 1049960 1049956 1049955 1049958 1049952 1049957 1049953 1049954 1049965 1049966 1049962 1049964 1049968 1049961 1049967 1049963 1049969 1049971 1049975 1049972 1049970 1049973 1049976 1049974 1049977 1049993 1049984 1049979 1049978 1049980 1049985 1049983 1049981 1049982 1049990 1049988 1049989 1049987 1049991

10087 10088 10084 10083 10086 10080 10085 10081 10082 10093 10094 10090 10092 10096 10089 10095 10091 10097 10099 10103 10100 10098 10101 10104 10102 10105 10121 10112 10107 10106 10108 10113 10111 10109 10110 10118 10116 10117 10115 10119

Columbia White Sturgeon 2007 Annual Report

FLmm

WTg

YrC

StkYr

Family

20 19 19 21 20 21 20 20 20 17 16 17 17 17 17 16 17 21 20 19 21 19 20 19 19 21 18 16 18 17 18 18 18 17 18 18 17 18 17 18

65 50 62 62 55 69 54 56 53 42 40 35 44 40 40 34 30 62 58 57 66 60 64 54 55 68 43 34 43 41 45 47 44 40 48 44 42 48 46 46

2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007 2007

2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008 2008

1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 4 5 5 5 5 5

28

41 42 43 44 45 46 47 48 49 50

985120032546421 985120032547429 985120032409553 985120032519244 985120032519265 985120032526611 985120032527660 985120032528835 985120032545133 985120032550565

1049992 1049986 1049995 1049996 1049997 1049998 1049994 1049999 1050000 1050001

10120 10114 10123 10124 10125 10126 10122 10127 10128 10129

17 18 19 18 18 18 19 18 18 17

47 50 50 42 48 40 50 43 47 41

2007 2007 2007 2007 2007 2007 2007 2007 2007 2007

2008 2008 2008 2008 2008 2008 2008 2008 2008 2008

5 5 6 6 6 6 6 6 6 6

10.0 FISH HEALTH TESTING SUMMARY 10.1 VIRUS SCREENING FOR BROODSTOCK Samples of ovarian fluid and milt were collected from spawners contributing gametes were screened for viruses (IPNV, WSHV1, WSHV2 and WSIV) using standard tissue culture methods as described in Section X: Procedures for the Detection of Viruses as listed in the Canadian Fish Health Protection Regulations Manual of Compliance: Fluids were collected from egg or milt samples and frozen at -20 prior to testing. All results for ovarian fluid and milt samples were negative for viral or bacterial contamination.

10.2 JUVENILE SCREENING HISTORY Subsamples of 5-30 juveniles from all family groups were screened for viruses at 30 days and 60 days post hatch. Fish were processed according to Section X : Procedures for the Detection of Viruses as listed in the Canadian Fish Health Protection Regulations Manual of Compliance. All examined juveniles tested negative for the above listed viruses. Viral culture cell lines used for Broodstock testing were also used for juvenile testing.

10.3 DEFORMITIES In 2007 year class fish, deformities in all families were almost nil. No changes were made in the way of early rearing practices that could account for the low deformity rate. As there were so few deformities seen, monthly deformity checks were discontinued and deformities were then noted when fish were handled Columbia White Sturgeon 2007 Annual Report

29

during the marking process. No particular deformities were seen with the few that did occur being classed as spinal or fin deformities.

11.0 Permits As with other years permits were obtained from the Introductions and Transfer Committee for adults, as well as for fall and spring release of juveniles. CITES permits were obtained including a letter of Non Detrimental Findings in case eggs or larvae were needed to be shipped to the Columbia Basin Hatchery in Washington. The CITES permit was not used as we did not send any eggs or fish to Washington this year. The permit was returned to the DFO.

12.0 PRESENTATIONS AND MEETINGS Ron Ek gave a short presentation to Sturgeon APG group in Castlegar on Oct 18th to update the group on the 2007 activities.

FFSBC staff members Ron Ek and Bryan Ludwig attended the Nov 15th Recovery Team meeting and Action planning group meetings as scheduled.

A fish culture update was made available at each of these meetings. FFSBC staff also participated in monthly UCWSRI conference calls.

Columbia White Sturgeon 2007 Annual Report

30

13.0 RESEARCH PROJECTS 13.1 COLUMBIA STURGEON MILT CRYOPRESERVATION 2007 In contrast from years past, no Cryopreservation experiments were conducted in 2007. In 2008, the plan is to transfer cryo preserved milt to KSH from the University of Southern Idaho and fertilize some small batches of eggs to investigate if the stored milt is viable upon thawing. The purpose of this work is for gene bank efforts and to maximize genetic breeding potentials. 13.2 WASHINGTON HATCHERY The KSH has previously supplied eggs to Washington state White Sturgeon recovery efforts. However, this year, US staff conducted their own brood capture and successfully spawned 2 females for their culture use. No eggs were shipped to Washington this year. 13.3 DNA SAMPLING Thirty DNA samples total of sturgeon fins were taken from members of each family group and preserved in labelled jars containing ethanol. Samples are stored on site at the KSH. DNA samples also removed from the adults. 13.4 Vemco TAGGING OF ADULT BROOD Thirteen of the mature adults taken to the hatchery were implanted with V16-6HR64K-Coded Vemco tags. These implanted tags allow for these adults movements to be tracked when they pass by stationary VR2 receivers that are situated in various locations in the river. The Vemco tags have a life of 3000 days or nearly 10 years. This tagging should allow adults to be tracked to see if and when they return to spawning areas and if there are additional spawning areas being used in the river. 13.4 Vemco TAGGING OF JUVENILES Fifty juveniles that had been raised to 150 - 200g each were implanted with V92L-R64K coded Vemco tags and released at Revelstoke on April 29th. The purpose of this work was to explore juvenile fish movements after release. 13.6 Substrate Preferences of Sturgeon Fry Fertilized eggs were provided to Steve McAdam at UBC for research use. He is working on sturgeon fry substrate preference and utilization. McAdam submitted the following description (in italics): 2007 Larval needs for Upper Columbia River white sturgeon Project name: White sturgeon larval responses to substrate Investigator: Steve McAdam (604 222-6760 or 604 908-3474) Brief description: Projects being undertaken in 2007 will continue to focus on larval reactions to varying substrate conditions as in past years. In particular this year’s work will look at behavioural reactions to substrate immediately post hatch, as well as the effects of photoperiod on larval reactions to different substrates. Columbia White Sturgeon 2007 Annual Report

31

Project location: UBC, Vancouver Larval needs: Last years work demonstrated that receiving larvae at about 6 days, but still prior to hatch was the best protocol (highest survival). Number of larvae required is approximately 1000 per family. As in previous years I would like to get eggs from each of the expected 6 unique families. Total number of eggs required is therefore 6 x 1,000 = 6,000. 13.7 TOXICOLOGY RESEARCH, UNIVERSITY OF SASKATCHEWAN & TECK COMINCO TRAIL. Fertilized eggs and larvae were supplied to researchers from University of Saskatchewan to be used in toxicology studies both at the Teck Cominco site in Trail and at the Toxicology Centre, University of Saskatchewan, Saskatoon. 13.8 MULTIPLE SPAWNING EXPERIMENT (IN HOUSE) The purpose of this experiment was to see if the fertilization rate changes depending on the time from when eggs are first released by the female until a controlled number of hours later If the fertilization rates stayed the same for a long period of time after initial ovulation, there would not be such a rush to conduct the egg collections and fertilization after seeing the eggs on the bottom of the tank. The reasoning here is that sturgeon often ovulate around dawn, which can make it difficult to collect volunteers to assist with spawning activities. The study is detailed in Appendix 2.

Columbia White Sturgeon 2007 Annual Report

32

Appendix 1: Monthly Hatchery Reports September 2007 Monthly Report

After grading and inventory, fish obtained the size whereby they could be weaned from the custom-prepared starter diet and were weaned onto Skretting Biodiet Grower during the month of September. Once inventory numbers post grading were determined and reported, the TWG directed the culling of three sibling families and these fish were euthanized.

In fish health, Family 1 witnessed higher than normal losses compared to other groups and were administered a 14 treatment of Oxysol 1000 by veterinary order. Rearing densities for the affected group were also reduced and a 5 day NaCl treatment (5‰ for 1h per day) was done. Losses dropped off dramatically as a result of treatment. Pathology results from the testing of samples sent to the Fish Health Unit were as follows: All Adults tested : Results Negative 2007-1091 Columbia sturgeon 126C x 2F69 Bact results:

15 to virus 16 plated on TSA and HS

6/16 possible Aeromonas spp infection 1/16 Flavobacterium psychrophilium

2007-1092 consisted of family crossings: C4 E66 x 05B C6 769 x 158 C1 408 x 05B

Bact results:

inoc Sep 6

inoc Sep 6

30 to virus 30 to bact T

27/30 presumptive for Aeromonas spp HS results not complete

Remaining to be tested: E66 x F69 E66 X 344 &69 X B02

report not complete report not complete remaining to test

Fish culture activities included grading and inventory with the placement of different families and split families in separate rearing units. These rearing units, called ‘ponds’ are in the Columbia Sturgeon Rearing Area and are designated by number according to their location in the hatchery. The prefix ‘C’ denotes the Columbia White Sturgeon 2007 Annual Report

33

‘Columbia’ side of the hatchery building and the numeral the pond designation. All ponds in this instance are 1m circular fibreglass tanks. Table 9 details the results of inventory, fish culture activity and size of the families. Table 9: Fish culture activities and data for Columbia juvenile sturgeon in the months of August and September, 2007. Family (code)

Pond

#1

C-10

Number Last Report 5679

Loss

Culls

1679

C-11

#/kg

Size

# KGs

Comments

100

20

2000

100

10 gms 10 gms

Graded & downsized in August

3851

228

4.3 gms

17

2718

382

2.6 gms

7.1

Inventoried

20

#2

C-7

6501

1375

#3

C-2

3531

813

#4

C-8

5970

103

2000

3867

250

4 gms

15.5

Graded & Inventoried

#5

C-12

5477

88

1425

3964

266

3.8 gms

15

Graded & Inventoried

#6

C-3

5916

304

1575

4037

306

3.3 gms

13

Graded & Inventoried

33,074

4362

6275

22,437

Totals:

Culled half sib Families

1275

Number On Hand 2000

14,000

Graded & Inventoried

107

1-10 gms.

October 2007 Monthly Report Grading, scute marking and PIT tagging was completed for all families. Beaver Creek (Keenleyside) fall releases of 4,000 juveniles planned for Nov 14/15 were marked and ready for release. Thirty fish from each family were set aside to grow in size to accept a Vemco sonic tag in the spring for release into the Revelstoke area. These fish are identified by PIT tag for family tracking. Only 50 fish will be selected to receive a sonic tag and the selection of fish, based on size, will be Columbia White Sturgeon 2007 Annual Report

34

done to ensure that each family is represented equally by 8 fish for 4 families and 9 fish from two families.

To scute mark and PIT tag fish, the fish are removed in small groups of less than 10 fish per group from the tank using a dip net and placed in aerated anaesthetic (75ppm TMS or clove oil in ethanol/water; 30-60mg/l) until mobility stops. Fish are removed from the anaesthetic bath by hand and the scutes on the left hand side of the fish according to a numerical pattern, in this instance scutes 1 and 8 on the left-hand side. This denotes a seven-scute presence between missing scutes indicating the brood year of 2007; seven complete scutes between missing scutes indicates the brood year.

To PIT tag fish, anaesthetized fish are placed ventral side down on a clean immovable surface and the trocar of the PIT tagger inserted into the dorsal musculature dorsal to the lateral line and anterior to the leading margin of the dorsal fin. The trocar and the PIT tag are disinfected in ethanol prior to use. Insertion is subcutaneous and/or intramuscular, but care must be taken to ensure the trocar and inserted PIT tags do not penetrate the body cavity and damage internal organs. The tag is then read by the PIT tag reader, the number recorded and the fish released to fresh water to recover. Pit tag retention and fish survival is generally in the 95+ percentage. Any mortality as a result of the procedures is noted, the PIT tag removed.

In fish health activities, 40 fish samples from each of Families 2, 3 & 6 were sent to the Fish Health Unit for routine testing. Table 10 details the fish culture activities and records for the month of October, 2007 at the KSH. The numbers reported include grading and culls for the ponds indicated.

Columbia White Sturgeon 2007 Annual Report

35

Table 10: Monthly fish culture activities and data for the KSH October 2007. Family (code)

Pond

#1

C-8

Number Last Report 4000

C-17

#2

C-2

110

3851

C-15

#3

C-3

Loss

715

122 40

2776

Culls

864

158

C-16

Number On Hand 950

#/kg

Size

# KGs

28

34

2225

38

36 gms 26 gms

772

44

2053

64

740

34

1838

76

638

45

40 #4

C-4

3865

C-13

#5

C-5

3964

135

256

2836

69

17

727

625

48

2595

71

650

48 69

C-18

#6

C-6

4037

C-14

Totals

22,493

151 40

606

2590

813

3,168

18,512

23 gms 16 gms 29 gms 13 gms 22 gms 15 gms 21 gms 14 gms 21 gms 14 gms

58

18 32

22 24

14 42

13 37

13 37

Comments

Scuted & Pit Tagged Graded & Downsized Scuted & Pit Tagged Graded & Downsized Scuted & Pit Tagged Graded & Downsized Scuted & Pit Tagged Graded & Downsized Scuted & Pit Tagged Graded & Downsized Scuted & Pit Tagged Graded & Downsized

344

November 2007 Monthly Report Releases began November 15 and 16th at Keenlyside 2179 and Beaver Creek 1850. A total of 4,029 fish were released (Table 11). DNA samples were obtained from representatives of each family by fin clip, placed in ethanol in individually labelled jars and stored at the KSH for future investigation.

Columbia White Sturgeon 2007 Annual Report

36

Table 11: Monthly fish culture activities and data for the KSH November 2007. Family (code)

Pond

#1

C-8

Number Last Report 950

Loss

Culls

Number On Hand

#/kg

Size

# KGs

Comments

22

44 gms 52 gms 66 gms 31 gms 33 gms 66 gms 45 gms 43 gms 31 gms 66 gms 35 gms 35 gms 34 gms 66 gms 33 gms 30 gms 66 gms 33 gms 30 gms 66 gms

40

Released Nov 15 HLK

*870* C-17

2225

sonic

C-1

#2

C-7

772

C-15

2053

sonic

C-1

#3

C-9

C-16

25

2250

19

30

15

*725* 8

740

32 2062

30

30

15

*284*

22

*400*

23

1838

1840

32

30

15

17 sonic

C-1

#4

C-10

C-13

638

2836

sonic

C-1

#5

C-5

625

C-18

2595

sonic

C-1

#6

C-6

650

C-14

2590

sonic Totals

*300*

28

*295*

28

39

2810

29

30

15

*576*

31 2605

9

33 15

*579* 11

C-1 18,512

109

4,029

30 2620

33

30

15

14,367

118 2

Hold for Sonic Tag

23

Released Nov 15 HLK

68 2

Hold for Sonic Tag

13

Released Nov 15 HLK Released Nov 15 HLK

17 58 2

Hold for Sonic Tag

11

Released Nov 15 HLK Released Nov 16 BC

10 97 2

Hold for Sonic Tag

18.5

Released Nov 16 BC

79 2

Hold for Sonic Tag

19.3

Released Nov 16 BC Graded & Downsized Hold for Sonic Tag

79 2 511

December, 2007 & January, 2008 Report Routine fish culture activities were continued during the winter months a KSH. Table 12 reports the details for fish culture activities. At this stage of Columbia White Sturgeon 2007 Annual Report

37

development and rearing, there is little remarkable activity as the fish continue to grow and thrive. Mortality rates are typically low and growth is consistent. Table 12: Fish culture activities and data for the KSH December 2007 and January, 2008. Family (code)

Pond

#1

C-11

Number Last Report

C-17

2250

sonic

C-1

30

#2

C-9

Loss

8

0

Culls

25

15

Number On Hand 700

#/kg

Size

# KGs

15

45

1517

16

30

10

700

24

64 gms 62 gms 100 gms 42 gms 33 gms 100 gms 43 gms 40 gms 100 gms 55 gms 50 gms 100 gms 43 gms 40 gms 100 gms 45 gms 40 gms 100 gms

C-15

2062

1347

26

sonic

C-1

30

30

10

#3

C-10

700

23

9

29

C-16

1840

1102

25

sonic

C-1

30

30

10

#4

C-7

700

18

2

136

C-13

2810

1974

20

sonic

C-1

39

30

10

#5

C-12

700

23

1678

25

30

10

700

22

1700

25

30

10

C-18 sonic

C-1

#6

C-8 C-14

sonic Totals

6

221

2605

5

215

2620

C-1 14,360

30

641

13,698

Comments

Marked & Tagged MC

95 3

Hold for Sonic Tag

29

Marked & Tagged MC

52 3

Hold for Sonic Tag

30

Marked & Tagged MC

44 3

Hold for Sonic Tag

39

Marked & Tagged MC

99 3

Hold for Sonic Tag

30

Marked & Tagged MC

67 3

Hold for Sonic Tag

32

Marked & Tagged MC

68 3

Hold for Sonic Tag

648

February 2008 to end March 2008 Report Pathology samples were submitted to the Fish Health Unit in February for release fish. No diseases of note were found. All scute marking and Pit tagging of fish Columbia White Sturgeon 2007 Annual Report

38

was completed as downgraded fish made acceptable weight for tagging. Further details of fish culture activities appear in Table 13. Table 13: Fish culture activities and data for the KSH February and March, 2008. Family (code)

Pond

#1

C-11

sonic #2

Number Last Report

C-1-4

2281

C-19

30

2084

sonic

C-19

30

#3

C-10 C-15

1777

sonic

C-19

30

#4

C-7 C5,6,17

2515

sonic

C-19

30

#5

C-12 C-16

2444

sonic

C-19

30

#6

C-8

Totals

Culls

31

C-9 C-14

sonic

Loss

C-13

2401

C-19

30 13,682

24

41

109

228

129

562

Number On Hand 700

#/kg

Size

# KGs

14

50

1550

16

30

5

700

20

1360

20

30

5

700

20

1036

19

30

5

700

18

1706

17

30

5

700

23

1516

25

30

5

700

22

1572

22

30

5

70 gms 79 gms 200 gms 50 gms 50 gms 200 gms 50 gms 53 gms 200 gms 55 gms 59 gms 200 gms 43 gms 40 gms 200 gms 46 gms 46 gms 200 gms

13,120

Columbia White Sturgeon 2007 Annual Report

95 6 35 68 6 35 55 6 39 100 6 30 61 6 32 71 6

Comments

Marked & Tagged Marked & Tagged Hold for Sonic Tag Marked & Tagged MC Marked & Tagged Hold for Sonic Tag Marked & Tagged MC Marked & Tagged Hold for Sonic Tag Marked & Tagged MC Marked & Tagged Hold for Sonic Tag Marked & Tagged MC Marked & Tagged Hold for Sonic Tag Marked & Tagged MC Marked & Tagged Hold for Sonic Tag

669

39

Appendix 2: Columbia Sturgeon Multiple Spawning Experiment 2007 Background The purpose of this experiment was to see if the fertilization rate changes depending on the time from when eggs are first released by the female until a controlled number of hours later. To spawn sturgeon successfully in captivity the brood stock are induced to ovulate, or spermiate, with exogenous hormone injections. Spawning induction includes injections of the selected hormone preparation. The initial injection is separated from the second, or resolving, injection by 12 hours. Ovulation can be expected within 20 to 40 hours after administration of the resolving injection. Variation in response depends on factors such as the size, age, condition of the fish and water temperature. For the convenience of daytime spawning initial injections are given at 8pm to 9pm and then resolving injections, 12 hours later at 8am or 9am the following morning. Ovulation and spawning are then anticipated during the morning hours of the following day. As this ovulation and spawning can hardly ever be predicted 100% accurately the fish are checked every hour or so starting from 20 hours post resolving injection until eggs are released by the female and egg collection can begin. We always attempt to do the egg collection and fertilization within an hour or so from the first eggs being released. We wanted to see if fertilization rates decreased in accordance with the increased time from when the first eggs were released. If the fertilization rates stayed the same for a long period of time then there would not be such a rush to conduct the egg collections and fertilization. Methods White sturgeon used in the Upper Columbia White Sturgeon Conservation Aquaculture Program were captured by using set lines during the June brood capture session on the Columbia River from Castlegar to the USA border. We used 2 females that were captured on June 22 and biopsied to determine sex and maturation stage. The 2 males used for the experiment were both captured on June 7th and both of these did not need a biopsy as they had flowing milt. (Adult Sturgeon Information is provided in Brood Info Chart.) Every fish collected was weighed and measured, checked for the presence of a PIT tag and once sex and maturity was determined they were directly transferred from a water-filled stretcher to an oxygenated transfer tank and hauled to the Kootenay Sturgeon Hatchery where they were placed in circular holding ponds with water temperature for the Females set to match the river and the males were held at 10’C until they were scheduled to be spawned. Eggs were collected, boiled and fixed in formalin on the boat from both females and the PI (Polarization index) calculation was conducted back at the hatchery.

Columbia White Sturgeon 2007 Annual Report

40

Both of the captured females PI’s were calculated to be under 0.10 thus showing that they were ready for final maturation and ovulation. The selected two female brood stock were placed in individual circular ponds and received two injections of synthetic gonadotropin- releasing hormone LHRHa at a total dose of 50ug/kg body weight: an initial dose (10%) at 8:30 pm June 25th and a resolving dose (90% of total dose) 12 hours later at 8:30 am June 26th. The water temperature in the female holding ponds was maintained at 15’C for the entire period prior to injections and through spawning completion. The two males were together in another circular pond and received single injections of LHRHa at a dose of 10ug/kg body weight on June 25th , with the water temperature also being increased to 15’C at this time. The water appeared to be slightly cloudy on June 26pm so males were checked and both found to be spermiating. The circular tanks holding the females were checked hourly starting at 20 hours after the resolving injections and at 5:30 am June 27th (or 21 hours post injection) the first few eggs were seen in the tank containing Female 7F7D186E66. At 6:15 am ( or 21.75 hours post injection) eggs were seen in the tank containing Female 4158696408. The spawning crew was then called and in place by 6:30 am. At 7:15 am milt was collected from both males by placing the male on its back in the stretcher. Milt is collected with a 5.0 cm length of tygon tubing (1/8” inside diameter;3/16” outside diameter) attached to a 50 ml plastic syringe. Initially, the plunger of the syringe is depressed. The tygon tubing is then inserted into the urogenital opening just posterior to the anus. Milt is collected by massaging the abdominal area over the testes to force milt into the collecting ducts and slowly withdrawing the plunger until the syringe is full. This allows for collecting a fairly large amount of milt without contaminating it with feces or water. The milt is then placed in a small zip lock bag identified with the males pit tag number, filled with oxygen and placed in a temperature controlled incubator at 5’C. A small amount of milt was removed from each of the first collected milt bags and checked for water activated motility using a dissecting microscope. Once the milt collection was completed each female in turn was placed on her back on a stretcher and 500 mls (approximately 17,000 eggs) of eggs were collected by hand expression and placed in a marked stainless steel bowl floating in a trough of 15’C flowing water to maintain the constant temperature. (The spawning times are shown in the spawning time chart.) 50 mls of milt was added to 1 liter of water to make a 1/20 dilution and this was quickly added to the bowl of eggs. Eggs and milt were gently stirred for 3 minutes and then the milt solution was gently poured off the eggs. At this point the eggs were de-adheased by adding a solution of water and Fullers earth to the eggs and gently stirring them with a feather for 1 hour. The Fullers earth solution was then gently rinsed from the bowl and the eggs were poured into a MacDonald jar incubator with adequate water flow through the hatching jar to provide a gentle rolling of the eggs.

Columbia White Sturgeon 2007 Annual Report

41

For this experiment this spawning cycle process was completed 4 times with 2 ½ hours between each cycle. The same male being used to fertilizes the same female each time. ( Refer to Spawning Chart.) Milt was checked again for motility during the last cycle of spawning at 3:30 pm and 4:00pm respectively and found to be as motile as prior to the first spawning in the morning. The MacDonald Jar Incubators were all provided with a flow of 5 liters/minute of 15’C water. Once the eggs reached nerulation on July 1st ( Day 4) four random 200 egg samples were checked from each upweller and the fertilization % calculated. Broodstock Capture History PIT Tag Number 7F7D186E66

Sex F

4158696408

F

7F7D4F2F69

M

7F7D21605B

M

Capture Date 07-Jul-92 07-Nov-92 09-May-03 22-Jun-07 26-Jun-92 18-Dec-97 16-Apr-98 16-Apr-02 22-Jun-07 11-Aug-93 08-Jun-07 04-Oct-96 19-Jun-07

Fork Weight Length kg 142 28.2 142 168.5 52.3 181.5 62 185 59.1 199 65.5 194 64.1 226.5 86.2 225.5 120 123 18.2 170 40 170 45 181 45

Spawning Crosses , Timing and Fertilization % Female # Male # Female # Male # 7F7D186E66 7F7D4F2F69 4158696408 7F7D21605B Time after Time after first eggs Fertilization first eggs observed Spawn Time % observed Spawn Time 2.5 hrs 8:15 AM 87% 2.5 hrs 9:00 AM 5 hrs 10:45 AM 73% 5 hrs 11:30 AM 7.5 hrs 1:15 PM 77% 7.5 hrs 2:00 PM 10 hrs 3:45 PM 78% 10 hrs 4:30 PM

Columbia White Sturgeon 2007 Annual Report

Fertilization % 85% 84% 73% 47%

42

Results We went into this experiment anticipating that there would be a prime time for collecting eggs from female sturgeon after she had started dropping eggs in the tank. This seemed to be true for Female 4158696408 as the highest Fertilization % was in the first spawning and the fertilization rate decreased in each of the following 3 spawnings. This was not the case with Female 7F7D186E66 as her highest fertilization rate was once again in the first spawning, but without an appreciable drop in fertilization rates in any of the last 3 spawnings. It looks like there may not be quite as much rush to spawn sturgeon broodstock after they have started dropping their first eggs, but with that being said it does look like it is slightly better to spawn them within the first few hours. Upon Discussion with Joel Van Eenennaam he brought up the following points -there are many factors that could affect this type of experiment: 1. The PI of the female, as a lower pi female may not be able to maintain egg quality longer than a slightly higher pi. 2. Water temperature is a very major factor. 3. Body size ( we have found large females at cooler water temps can have good quality eggs 4-5 hours after first release, no problem. 4 Female condition in terms of stress factors 5. Time when females actually release the first eggs (as some females seem to “hold back” and are ovulated but delay or do not release eggs at all). This experiment supports results observed in California and Idaho: a good quality female, at good water temps can still have good quality eggs 2-3 hours after first observation of released eggs. Our average time of egg collection is about 1.5 hours after the first eggs, with many fish not being spawned until 2.5-3.5 hours. Therefore, there is no need to rush on observing eggs in the tank, although current thinking and practice is to take eggs when the first egg release is seen. If we were to conduct this experiment again we would try to do the first spawn as soon as possible to when the first eggs are released and then 2.5 hrs ect. It would also be best to have a better experimental design like having 3 (bowls) jars per stripping with 200-300 mils eggs each, for statistical analyses to ensure there was no “bowl or fertilization techniques affect” and then do the 3 samples from each jar for fertility rates.

Columbia White Sturgeon 2007 Annual Report

43