Dietary Protein Level and Water Temperature Interactions for Juvenile Greenlip Abalone (Haliotis Laevigata)

Dietary protein level and water temperature interactions for juvenile greenlip abalone (Haliotis laevigata)

David Stone Matthew Bansemer* James Harris Georgia Mercer Hanru Wang This study forms part of an Australian Seafood CRC/AAGA Project (2010/736): Development of formulated diets for cultured abalone Principal Investigator: Assoc. Prof. David Stone

Background - Abalone culture in Australia

• Predominantly in land-based systems in south eastern Australia • Reliant on formulated feeds • Currently, juveniles are weaned off microalgae & on-grown using a single diet feeding strategy

• Pre-2013, crude protein levels ranged from 25 to 30% (Coote et al. 2000)

• Post-2013, crude protein levels increased to 30 to 35% (Stone et al. 2013) Background – Dietary protein

Water Average Optimal dietary Abalone species temperature starting Reference protein level (%) (°C) weight (g)

Greenlip abalone 20 0.81 27 Coote et al. (2000) (Haliotis laevigata) 14 1.8 29 Stone et al. (2013)

22 1.8 35 Stone et al. (2013)

14 22.9 24 Stone et al. (2013)

22 22.9 34 Stone et al. (2013)

South African abalone 22 1.4 47 Britz (1996) (Haliotis midae) 18 0.2-1.0 34 Britz & Hecht (1997)

18 7.0-14.0 44 Britz & Hecht (1997)

Blacklip abalone 17 0.04 45 Dunstan (2010) (Haliotis rubra)

Green Ormer 14 0.18 35 Mai et al. (1995) (Haliotis tuberculata)

Pacific abalone 14 0.38 35 Mai et al. (1995) (Haliotis. discus hannai) Background – Dietary protein

Water Average Optimal dietary Abalone species temperature starting Reference protein level (%) (°C) weight (g)

Greenlip abalone 20 0.81 27 Coote et al. (2000) (Haliotis laevigata) 14 1.8 29 Stone et al. (2013)

22 1.8 35 Stone et al. (2013)

14 22.9 24 Stone et al. (2013)

22 22.9 34 Stone et al. (2013)

South African abalone 22 1.4 47 Britz (1996) (Haliotis midae) 18 0.2-1.0 34 Britz & Hecht (1997)

18 7.0-14.0 44 Britz & Hecht (1997)

Blacklip abalone 17 0.04 45 Dunstan (2010) (Haliotis rubra)

Green Ormer 14 0.18 35 Mai et al. (1995) (Haliotis tuberculata)

Pacific abalone 14 0.38 35 Mai et al. (1995) (Haliotis. discus hannai) Background – Dietary protein

Water Average Optimal dietary Abalone species temperature starting Reference protein level (%) (°C) weight (g)

Greenlip abalone 20 0.81 27 Coote et al. (2000) (Haliotis laevigata) 14 1.8 29 Stone et al. (2013)

22 1.8 35 Stone et al. (2013)

14 22.9 24 Stone et al. (2013)

22 22.9 34 Stone et al. (2013)

South African abalone 22 1.4 47 Britz (1996) (Haliotis midae) 18 0.2-1.0 34 Britz & Hecht (1997)

18 7.0-14.0 44 Britz & Hecht (1997)

Blacklip abalone 17 0.04 45 Dunstan (2010) (Haliotis rubra)

Green Ormer 14 0.18 35 Mai et al. (1995) (Haliotis tuberculata)

Pacific abalone 14 0.38 35 Mai et al. (1995) (Haliotis. discus hannai) Background – Water temperature

• Water temperature affects growth, metabolic rate, feed intake & digestive function of abalone • Water temperature fluctuates during abalone grow-out • 10 to 25°C • Optimum temperature for growth: • Greenlip abalone (H. laevigata) • 18°C for 82 mm • 22°C for 23 mm • Red abalone (H. rufescens) • 18°C for 44 mm • Pacific abalone (H. discus hannai ) • 20°C for 40 mm

Aims

Investigate the interaction of dietary protein level & water temperature on the growth performance & feed utilisation of post-weaned greenlip abalone

This work complements two other studies in the larger project: • The protein requirements of Year 1 & 2 greenlip abalone at 14, 18, 22°C (Stone et al. 2013) • On-farm feed trial for land based abalone culture investigating multi-diet feeding strategies

Experimental design

3 Water Temperatures (ºC) 14 17 20

4 Nominal protein levels (%) 27 30 33 36 27 30 33 36 27 30 33 36

Replicates (n=4) 1 2 3 4 Experimental design

3 Water Temperatures (ºC) 14 17 20

4 Nominal protein levels (%) 27 30 33 36 27 30 33 36 27 30 33 36

Replicates (n=4) 1 2 3 4

Proximate composition Nominal protein (%) (g/100g diet as fed) 27 30 33 36 Moisture 10.35 10.48 10.61 10.30 Crude protein 27.00 31.10 34.30 37.30 Digestible protein (calculated) 20.27 23.54 26.13 28.57 Lipid 3.60 3.60 3.70 3.50 Gross energy (MJ/kg) 17.00 17.25 17.64 17.27 Digestible energy (MJ/Kg) (calculated) 12.24 12.35 12.57 12.53 Ash 5.02 5.31 5.24 8.17 NFE (calculated) 64.38 59.99 56.76 51.03 Digestible CP:GE (g/MJ) 16.57 19.06 20.79 22.80 Experimental design

3 Water Temperatures (ºC) 14 17 20

4 Nominal protein levels (%) 27 30 33 36 27 30 33 36 27 30 33 36

Replicates (n=4) 1 2 3 4

3 Water Temperatures (ºC) 14 17 20

4 Nominal protein levels (%) 27 30 33 36 27 30 33 36 27 30 33 36

Replicates (n=4) 1 2 3 4

• 48 tank flow-through seawater system • 18 post-weaned, 6-month old abalone per tank • Weight: 0.9 g; shell length: 19.5 mm • Selected from heat resistant stock Temperature controlled laboratory • Photoperiod (12 h low light [3.4 lux]: 12 h D) at SARDI ASC, West Beach, Adelaide, South Australia • Fed to excess 4:00 pm daily • Un-eaten feed collected the following day at 8:30am • Water quality monitored daily • Water temperature, DO (mg/L; % sat), pH, salinity • Duration: 91 days 13-L culture unit with bottom surface area of 1,129cm2

Results: Will report on

• Specific growth rate • Apparent feed consumption rate • Apparent feed conversion ratio • Apparent protein efficiency ratio • Apparent energy efficiency ratio • Analysis • Two-factor ANOVA • Regression analysis

Specific growth rate

Two-factor ANOVA

Water temperature (P < 0.001) 14 < 17 < 20°C

Dietary protein level (P > 0.05)

Interaction (P > 0.05)

Regression analysis All water temperature (P > 0.05)

Values (Mean ± SE; n = 4) Apparent feed consumption rate

Two-factor ANOVA

Water temperature (P < 0.001) 14 < 17 < 20°C

Dietary protein level (P > 0.05) Interaction (P > 0.05)

Regression analysis 14°C (R2 = 0.54 ; P < 0.05; y = 0.08x2 - 4.75x + 86.79)

17°C (R2 = 0.30 ; P < 0.05; y = 0.31x + 8.40)

20°C (R2 = 0.50 ; P < 0.05; y = -0.3569x + 37.799)

Values (Mean ± SE; n = 4) Apparent feed conversion ratio

Two-factor ANOVA

Water temperature (P < 0.001) 14 > 17 = 20°C

Dietary protein level (P > 0.05)

Interaction (P > 0.05)

Regression analysis 14°C (R2 = 0.406, P < 0.05 y = 0.01x2 - 0.32x + 6.30)

18 & 22°C (P>0.05)

Values (Mean ± SE; n = 4) Apparent feed conversion ratio

Values (Mean ± SE; n = 4) Apparent protein efficiency ratio

a b b c

Values (Mean ± SE); Two-factor ANOVA Water temperature (P < 0.001; n = 16) Dietary protein level (P < 0.001; n = 12) Interaction (P > 0.05)

Apparent protein efficiency ratio

PER of post-weaned greenlip abalone at 20°C PER of Year 1 greenlip abalone at 22°C (Current study) (Stone et al. 2013) a ab b b

Values (Mean ± SE); Two-factor ANOVA Values (Mean ± SE); One-factor ANOVA Dietary protein level (P > 0.05; n = 4) Dietary protein level (P < 0.001; n = 4)

Apparent energy efficiency ratio

Values (Mean ± SE); Two-factor ANOVA Water temperature (P < 0.001; n = 16) Dietary protein level (P > 0.05; n = 12) Interaction (P > 0.05)

Conclusions and Implications

• Raising water temperature resulted in improved growth rates • Plausible to heat nursery systems → shorter production time • Dietary protein had no significant effect on growth • Faster growing abalone at 20°C fed low protein diets compensated by increasing feed intake • Results from the current study complement Stone et al. (2013) • Based on results at 22°C, Stone et al. (2013) concluded that there is considerable scope for multi-diet feeding strategies • Currently testing & evaluating on-farm to improve production • Plausible to switch to a higher protein diet (~35% CP) at >20°C • These animals were heat tolerant, results may differ between farms

Acknowledgments

• Australian Abalone Growers Association • Abalone feed companies Joel Scanlon (Adam and Amos Abalone Feeds) Dr Tom Coote and Kym Heidenreich (Eyre Peninsula Aquafeeds) Dr Rhys Hauler and Dr Matt Bransden (Skretting Australia). • South Australian Premier’s Science and Research Fund • Marine Innovation South Australia • Australian Seafood CRC • Fisheries Research and Development Corporation • SARDI Aquatic Sciences, Adelaide, Australia Raymond Cultura (SARDI - Australasian Experimental Stockfeed Extrusion Centre (AESEC) Roseworthy) • Flinders University, Adelaide, Australia Elise Schaefer, Duong Duong Ngoc, Thanh Hoang Hai, Krishna-Lee Currie, Brett Lange Diet Ingredients

Ingredient composition Nominal protein (%) (g/100g diet as fed)

27 30 33 36

Salmon fish meal 4.00 4.00 4.00 4.00 Solvent extracted soybean meal 18.90 21.40 23.90 26.47 Lupins (de-hulled) 20.80 23.60 26.40 29.14 Waxy maize starch 30.67 29.07 27.59 19.96 Pregelatinised waxy maize starch 10.00 5.62 1.15 0.00 Wheat gluten meal 5.00 5.00 5.00 5.00 Casein 5.48 6.53 7.59 8.63 Diatomaceous earth 1.76 1.79 1.77 4.60 Fish oil 1.22 0.84 0.46 0.10 EPA Vitamin/mineral premix 0.20 0.20 0.20 0.20 Sodium alginate 0.30 0.30 0.30 0.30 Vitamin E 0.01 0.01 0.01 0.01 Calcium sulphate 0.43 0.36 0.30 0.22 Monosodium phosphate 0.72 0.68 0.65 0.61 Arginine 0.31 0.37 0.41 0.46 Threonine 0.20 0.23 0.27 0.30

Dry matter leaching loss

10.0 10.0 a 8.0 8.0 a a 6.0 b 6.0 b c 4.0 4.0 c

2.0 2.0 d Dry matter leaching loss (%) loss leaching matter Dry (%) loss leaching matter Dry 0.0 0.0 14 18 22 1 2 4 8 16 Water temperature ( C) Time (h)

Stone et al. (2013) Absolute growth rate

Two-factor ANOVA

Water temperature (P < 0.001) 14 < 17 < 20°C

Dietary protein level (P > 0.05)

Interaction (P > 0.05)

Regression analysis All water temperature (P > 0.05)

Values (Mean ± SE; n = 4)