초록

1. 능성어 대량 종묘 생산 기반 조성
- 어미관리
- 광주기${\cdot}$수온 조절을 통한 인위적 성성숙 유도
2. 능성어 성성숙 유도 및 산란 그리고 수컷 유도
- 자연산란 및 인공채란
- 수컷유도 및 정자 냉동 보존
- 먹이계열 개발
3. 대량종묘 생산 기술 개발
- 자${\cdot}$치어 행동생태 및 형태적 변태과정 탐색
- 능성어 종묘생산 기술 확립

Abstract ▼

Groupers are protogynous hermaphroditic fish that inhabit coral reefs in the tropics and subtropics. They belong to the family Epinephelinae, which includes 159 marine spe챠es in 15 genera (FAO, 1993). Eleven species of grouper inhabit the Southern Sea near Jeju Island, Korea, including the longtooth

Groupers are protogynous hermaphroditic fish that inhabit coral reefs in the tropics and subtropics. They belong to the family Epinephelinae, which includes 159 marine spe챠es in 15 genera (FAO, 1993). Eleven species of grouper inhabit the Southern Sea near Jeju Island, Korea, including the longtooth grouper, Epinephelus bruneus, the sevenband grouper, E. septemfasciatus, and the red spotted grouper E. akaara, (Kim et aL, 2001). Groupers are highly regarded as food and as ornamental fish in Korea, China, Japan, and Southeast Asia.
Studies on the reproductive biology of groupers have begun (Hwang et aL, 1998; Johnson et aL 1998; Lee et al., 2002b; Song et aL, 2003), including studies on brood stock management for seed production (Kayano, 1996; Duray et al, 1997; Sugama and Ikenoue, 1999), sex inversion (Lee et al., 2000, 2002a; Tsuchihashi et aL, 2003; Yeh et aL, 2003), and sex maturation (Kim et al., 1997; Marino et al., 2003), to support commercial cultivation or conservation of resource programs.
The sevenband grouper is an excellent eating fish. Due to overfishing in Jeju coastal waters, the catch has been decreasing rapidly. To allow the recovery of this species in coastal waters, and to help the aquaculture industry, a larviculture technique must be developed. This paper describes the sex maturation, ovulation, fertilized egg and larval development, malformation, and larval rearing of the sevenband grouper, E. septerrifasciatus, and longtooth grouper, E. bruneus.
1. Natural spawning
During the period 2000 to 2002, the sevenband grouper, E. septerrifasciatus, spawned naturally in a rearing tank. In 2000, the fish spawned naturally from 6 July to 28 August. The total volume of eggs collected was 3,855 ml, and the total volume of floating eggs was 265 m!. The fertilization rate was 0${\sim}$76％. In 2001, the fish spawned naturally from 5 July to 6 August. The total volume of eggs collected was 1,650 ml, and the total volume of floating eggs was 472 ml. The fertilization rate was 0${\sim}$80％. In 2002, the fish spawned naturally from 25 July to 20 August. The total volume of eggs collected was 1,330 ml, and the total volume of floating eggs was 38 ml. The fertilization rate was 0${\sim}$50％. Egg production and quality decreased gradually at the end of the spawning period.
2. Induction of sex reversal
In this species, techniques for accelerating sex reversal to obtain male brooders are required for the success of induced breeding. We examined the induction of sex reversal in immature female E. septerrifasciatus by injecting 17${\alpha}$-methyltestosterone (MT). MT was injected at a dose of 0.5${\sim}$2.0 mg/kg BW. Initial controls had immature ovaries containing primary oocytes in lamellae that extended into the central lumen. At four weeks, the ovaries of control fish were similar to those of the initial control. In contrast, the MT treatment group was undergoing spermatogenesis, although no spermatozoa were observed. Sperm were obtained from gonads from the 1.0 and 2.0 mg/kg MT treatment groups at eight week using the stripping method.
3. Cryoprotectant
We studied the cryopreservation of sperm in E. septemfasciatus. The survival rate and activity were highest using 5％ glucose as the diluent and DMSO and TYB as cryoprotectants for E. septemfasciatus spermatozoa, although the levels were lower than in controls (P<0.05). The fertilization rate of eggs was highest in the GDS group at 97.6${\pm}$1.2％ and next highest in the control group at 94.9${\pm}$2.0％, although the difference wasnot significant (P>0.05). The hatching rate of eggs was 94.2${\sim}$97.0％.
4. Induction of ovulation
Fish were maintained under the ambient photoperiod and temperature in a 50 $m^3$ flow-through tank The ovary maturity of the broodstock was assessed weekly during the spawning season using biopsy methods. A cannula was inserted into the oviduct of female E. septemfasciatus and E. bruneus, and a sample of gonadal tissue was removed by suction.
Hormone-induced spawning was attempted in females with an average vitellogenic oocyte diameter of at least 400 um To induce spawning, HCG (human chorionic gonadotropin) was used. The hormone treatment strategies tested included HCG alone (500 IU/kg).
Females with mean oocyte diameter ranging from 300 to 500 um were suitable for hormone-induced spawning. Oocyte diameter increased to 300∼700 um within 24 h of the injection and to 800 um within 48 h. Eggs were stripped successfully 48 h after the injection. The total volume of stripped eggs was 2,480 ml, and the total volume of floating eggs was 1,360 ml; floating rate was 54.9％.
5. E. septemfasciatus
Egg development
Stripped fertilized eggs of E. septemfasciatus were colorless spheres, 790-890 um (average 821.8${\pm}$2.0 um) in diameter, with a 170${\sim}$230 um oil globule (average 192.9${\pm}$0.93 um). The fertilization and hatching rates were 56.2${\sim}$94.9 and 70.0${\sim}$97.7％, respectively, using the dry fertilized method.At a water temperature of 22.0${\pm}5^{\circ}C$, first cleavage took place about 1.0 h after fertilization. The egg reached the morula, blastula, gastrula, and embryo stages about 5.0, 12.0, 16.5, and 23.0 h after fertilization, respectively. Hatching began about 46.0 h after fertilization.
Yolk sac stage
To examine the time taken for yolk absorption at various water temperatures (22 and $25^{\circ}C$), changes in the absorption rate of the yolk and oil globule in newly hatched larvae were investigated. The rate of yolk absorption at $25^{\circ}C$ in newly hatched larvae was 83.5％ after 24 hand 97.8％ after 48 h. Yolk absorption was completed within 48 h. By contrast, at $22^{\circ}C$, the rate of yolk absorption was 32.7％ after 24 h, 88.2％ after 48 h, and 93.1％ after 72 h.
The rate of oil globule absorption at $25^{\circ}C$ in newly hatched larvae was 34.0％ after 24 h and 80.3％ after 48 h. By contrast, at $22^{\circ}C$, the oil globule absorption was 29.5, 40.2, and 28.2％ after 24, 48, and 96 h, respectively. The oil globule persisted until mouth opening. The elapsed time from hatching to mouth opening was 3 days at $25^{\circ}C$, and 4 days at $22^{\circ}C$.
Upper jaw length (UJL)
The UJL of larvae with newly opened mouths was 0.148${\pm}$0.009 mm, and this increased to 0.169${\pm}$0.006 and 0.230${\pm}$0.005 mm at 4 and 11 DAH (day after hatching), respectively.
We defined 0.75d as 75％ of the mouth opening and 0.5d as 50％ of the mouth opening. At 3${\sim}$11. DAB, when the larvae were first fed, the mouth size (d) of the larvae was 0.20 9${\sim}$0.325mm, 0.75d was 0.157${\sim}$0.244 mm, and 0.5d was 0.105${\sim}$0.162mm.
Morphological development
In newly hatched larvae, the oil globule was situated in the posterior part of the yolk. The body surface, marginal fin, and yolk were covered with colorless granules. Most newly hatched larvae tended to float to the surface. The larvae were suspended head down in the water column, and swam rapidly soon after hatching.
At 2${\sim}$3 DAH, the larvae retained the yolk and oil globule, and the mouth had not opened. The head skeleton formed, and the pectoral fin bud appeared. Most of the larvae tended to settle to the bottom. Most of the yolk and oil globule were absorbed by this time. The larvae began to feed on 3 DAH By 4${\sim}$5 DAH, the yolk and oil globule were absorbed completely, and the mouth and anus had opened. The eyes appeared in the melanophores.
The rudiments of the second dorsal and pelvic spines appeared at 11 DAH. Both spines, a larval characteristic, began to elongate; the abdominal cavity was densely lined with melanophores; and the two spines appeared on the preoperculum of larvae at 17 DAH. Growth
The newly hatched larvae measured 1.75 mm TL. By day 4, the larvae reached 2.72 mm TL and the mouth and anus had opened. By day 11, the larvae reached 3.24 mm TL and began to metamorphose. By day 24, the larvae reached 5.12 mm TL.
6. E. bruneus
In total, 735,000 eggs were stripped from a female E. bruneus and 700,000 (95.2％) were floating eggs. The fertilized eggs were colorless spheres, 830-950 um (average 900${\pm}$2um) in diameter with a 200${\sim}$260um oil globule (average 244${\pm}$2um). The fertilization and hatching rates were 97.7${\pm}$0.6％ and 96.7${\pm}$0.5％, respectively.
At a water temperature of 25.0${\pm}0.5^{\circ}C$, first cleavage took place about 1.0 h after fertilization.The egg reached the morula, blastula, gastrula, and embryo stages about 5.0, 4.5, 10.0, and 13.0 h after fertilization, respectively. Hatching began about 32 h after fertilization.
The newly hatched larvae measured 2.02${\pm}$0.20 mm TL. By day 11, the larvae reached 4.12${\pm}$0.09 mm TL and began to metamorphose. By day 54, the larvae reached 41.12${\pm}$1.20mm TL, and had completed metamorphosis. Thereafter, the larvae grew quickly and reached 93.78${\pm}$0.98 mm TL at 93 DAH.
The AL (anal length) of the newly hatched larvae was 1.22${\pm}$0.02 mm, and the AL at 54 and 72 DAH was 22.53${\pm}$0.51 and 30.57${\pm}$0.81 mm, respectively.
The second dorsal and pelvic spines appeared at 11 DAH. The first and third dorsal spines appeared at 17 DAH.
The second dorsal spine was initially 0.56${\pm}$0.20 mm long, and increased to 5.35${\pm}$0.20 mm at 38 DAH. The pelvic spine was initially 0.83${\pm}$0.23 mm long, and increased to 4.11${\pm}$0.17 mm mm at 33 DAH. The dorsal and pelvic spines differentiated into fins at 48 DAH. At 23 DAH, when TL was 7.62${\pm}$0.33 mm, the size of the second dorsal and pelvic spines relative to TL attained their maximums: 40％ and 35％, respectively.

목차 Contents

• 제 1 장 서론...20
• 제 2 장 재료 및 방법...21
• 제 1 절 어미 관리 및 사육...21
• 제 2 절 성성숙 유도 및 산란...23
• 제 3 절 기능적 수컷유도 및 정자 냉동 보존...28
• 제 4 절 난 및 자ㆍ치어 발달...35
• 제 5 절 먹이계열 개발 및 대량 종묘 생산...38
• 제 6 절 자바리 종묘생산...41
• 제 3 장 결과...44
• 제 1 절 자연산란...44
• 제 2 절 성성숙 유도 및 인공채란...48
• 제 3 절 기능적 수컷유도 및 정자 냉동 보존...57
• 제 4 절 난 및 자ㆍ치어발달...64
• 제 5 절 먹이계열 개발 및 대량 종묘 생산...75
• 제 6 절 자바리 종묘생산...86
• 제 4 장 고찰...96
• 제 5 장 연구개발결과의 활용계획...106
• 제 6 장 참고문헌...109