Larvae of humpback grouper show drastic changes in morphology as they develop from hatched larvae to juvenile stage (Mishima and Gonzares 1994). Until larvae complete metamorphosis to juvenile stage, they are very sensitive to environmental conditions and are prone to stress and high mortality rates.
Egg handling should be done very carefully to prevent mechanical shocks. Sensitivity of humpback grouper eggs may be different at each stage of egg development. Eggs are very sensitive before the embryonic stage and just before hatching. Salinity, temperature and egg density are important factors during the incubation of marine fish eggs (Holliday 1988).
The purpose of the present studies was to determine the effect of egg density, water exchange and aeration rate on embryonic development and hatching rate of humpback grouper, Cromileptes altivelis, eggs.
Material and Methods:
Spawners and Eggs
Grouper spawners weighing 1.06–3.16 kg each, total length 41.5–57.0 cm were held in round concrete tanks (75 m 3 ) with a density of 24 fish (16 female and 8 male). Broodfish were fed trash fish and squid that were supplemented with a vitamin mix and vitamin C. Water exchange in the broodstock tanks was 300–500% per day using flow through system.
Humpback grouper spawn at night, mostly around 22:00–24:00 hours. Since fertilised grouper eggs float, spawned eggs flowed directly into the egg collector (300–400 µ m mesh size) that was connected to the tank overflow pipe. Collected eggs were then harvested on the following morning and transferred into a transparent 30-litre polycarbonate tank filled with filtered sea water. Only floating eggs were transferred into the polycarbonate tank for the experiment.
Experiment 1:
Effect of Egg Density
Six treatments of egg density were studied in this experiment: 500, 1000, 1500, 2000, 2500 and 3000 eggs/litre. The experiment was conducted using three litre transparent bottles each filled with two litres of sea water. The experiment was a completely randomised design with three replicates for each treatment. Samples of eggs were collected every hour after stocking for observation of embryonic development.
Experiment 2:
Effect of Water Exchange
This experiment used 12 transparent tanks, 100 litres in volume and filled with sea water (34 ppt). In each tank were stocked newly spawned eggs at a density of 500 eggs/litre. Treatments for this experiment were: without water exchange; 100% water exchange; and 200% water exchange per day.
The experiment was a completely randomised design with three treatments and three replicates per treatment. Samples were taken every hour after stocking for embryonic development and hatching observation.
Experiment 3:
Effect of Aeration Rate
This experiment used 12 transparent bottles, three litres in volume filled with sea water (34 ppt). In each bottle was stocked newly spawned eggs at a density of 500 eggs/litre. The treatments in this experiment were: (A) aeration rate of 0 ml/min, (B) 200 ml/min, (C) 400 ml/min, and (D) 600 ml/min. The experiment was a completely randomised design with four treatments and three replicates per treatment. Samples were taken every hour after stocking for observation of embryonic development and hatching.
Embryonic Development:
Time from spawning to various embryonic stages was recorded for the different treatments. Ten to 30 eggs were analysed each hour and their development was examined by microscope.
Result:
Experiment 1
Egg development from spawning to various embryonic stages at different egg density is summarised in Table 1. Total time required from spawning to hatching ranged from 18 hours and 41 minutes to 20 hours and 29 minutes. The hatching rate was significantly higher for eggs incubated at lower density (Table 2). However, eggs incubated at the density of 2000, 2500 and 3000 eggs per litre resulted in the same hatching rate.
Table 2. Average value of hatching time and hatching rate of humpback grouper eggs incubated at different densities.
Table 3 shows that embryonic development from many cells to hatching was faster at higher water exchange. Almost 70% of eggs had hatched at 17:00 for eggs incubated at a water exchange of 200%/day. On the other hand, eggs incubated at 100%/day water exchange and without water exchange had a hatching rate of only 60% and 50% at 17:30 and 18:00, respectively.
The hatching rate of eggs incubated at 200%/day (71.6%) and 100%/day (57.7%) water exchange was not significantly different, but significantly higher compared with no water exchange (48.3%).
Experiment 3
Hatching time of eggs incubated at different aeration rates are shown in Table 4. Hatching time is significantly different (P < 0.05). All eggs hatched at around 18:00 in the afternoon. Dissolved oxygen increased and ammonia concentration decreased in incubation tanks with increasing aeration rate (Table 4). Hatching rate and survival of 3-day old larvae increased significantly (P < 0.05) with increasing aeration rate (Table 5).
Table 4. Hatching time of humpback grouper eggs incubated at different aeration rates, dissolved oxygen (DO) and ammonia concentration in the incubation water.
Table 5. Hatching rate of humpback grouper eggs and survival of 3-day old larvae after the eggs were incubated at different aeration rates.
After fertilisation, humpback grouper eggs hatched within 18.41–19.51 hours at an egg density of 500–3000/litre; 18–18.38 hours at aeration rate of 0–600 ml/min; 19.50–20.50 hours at water exchange of 0–200%/day at 29–32°C water temperature and salinity 34 ppt. Hussain et al. (1975) (cited in Kawahara et al. 1997) reported that grouper Epinephelus tauvina eggs hatched within 26–35 hours after fertilisation at 27–30°C, much longer than the present observation.
Hatching time of E. fuscoguttatus eggs was 18–19 hours at 28–30°C water temperature (Chao et al. 1993 cited in Kawahara et al. 1997). The hatching time of artificially-fertilised egg of E. striatus was 21.1–22.3 hours for the firs thatching and 23–25.5 hours for the complete hatching at 28°C (Watanabe et al. 1995). However, other environmental conditions, such as light intensity, aeration and salinity could influence the incubation time of grouper eggs.
The average hatching rate success of Cromileptes altivelis eggs was 77%, with an egg density of 500/litre, 71.6% with a water exchange 200%/day, and 78.67% with an aeration rate of 600 ml/minute. Watanabe et al. (1995) reported that hatching success of E. striatus eggs was as high as 82.5% at the water temperature of 26–30°C, but the mortality of yolksac larvae was accelerated at higher temperatures.
The result of survival rate of pre-feeding stage of C. altivelis larvae (day-3) in present study was 62.3% with an aeration rate of 600ml/min. Biochemical quality of eggs that related to the condition of the broodstock can influence larval survival (Watanabe et al. 1984). Other environmental factors such as aeration (water circulation) and light intensity may also influence metabolism and survival of yolksac larvae.
Author:
M.A. Rimmer
