Alberto J.P. Nunes1, Sigve Nordrum, Otávio Serino Castro, Marcelo V.C. Sá

LABOMAR*. Brazil1E-mail: albertojpn@uol.com.br

*Part of Universidade Federal do Ceará

SUPPLEMENTATION OF KRILL OIL IN THE FEEDING OF Litopenaeus vannamei CAN COUTERACT THE DETRIMENTAL EFFECTS OF HIGH SALINITY

WAS 2009 MeetingVeracruz, MexicoShrimp Nutrition Special SessionSeptember 26th, 200903:10 pm

Rationale In shrimp farms water salinity

can vary from less than 1‰ to more than 50‰

Salinity fluctuates mainly as a response to season, pond depth, water exchange rates, pumping site

Ideal salinities for the rearing of L. vannamei is around 20‰

Species can tolerate wide range of salinities, but above 40‰, osmoregulatory ability can be depreciated

High water salinity leads to increased feed intake, poor FCR and slow growth

Grow-out pond in an hypersaline area with salt accumulated on the bottom

Influence of pumping site to riverine water affects water sality

Review Liu et al. (2007) supplementing

vitamin E (dietary tocopheryl acetate) at 600 and 1,000 mg/kg of diet for L. vannameiexperienced an increase in shrimp resistance to acute salinity changes Hurtado et al. (2006) were able

to demonstrate that growth of L. vannamei at high salinities was enhanced when fed on HUFA-enriched diets Hurtado et al. (2007) observed

that a higher proportion of HUFA in gill membrane in shrimp fed the high-HUFA diet counteracted the influence of salinity on water content in gills during a long-term salinity exposure

Outside cell

Inside cell

fatty acid

The cell membrane is formed by lipid bi-layer. Phospholipids are the major lipid group within the membrane and contain a range of fatty acids including eicosapentanoic acid (EPA).

Source: AkerBiomarine ASA, Norway

Objectives1. To evaluate if

supplementation of Krill oil improves the growth performance of juveniles of L. vannamei when reared under hypersaline water conditions

2. To determine optimum inclusion levels of Krill oil in diets for the Pacific white shrimp when exposed to high salinity rearing conditions

Krill oil (Qrill™, AkerBiomarine ASA, Norway) produced from the Antarctic Krill (Euphausia superba)

LABOMAR/UFCEusébio. BRAZIL

Pacoti River Estuary

19/jan/2009

Outdoor tanksIndoor tanks

Rearing System

Clear water500-L volume 0.57 m2 areaXXXX shrimp/m2

INDOOR50 tanks

Experimental Design Experimental diets:

• One diet containing a combination of fish and soybean oil (diet FISH)

• One diet with Krill and soybean oil (diet KRILL)

• One diet with soybean oil alone (diet SOY)

• Two diets containing low and high inclusion levels of Krill oil in combination with soybean oil (diets KRILL- and KRILL+, respectively)

Salinity conditions• IDEAL (21 – 26‰) and HIGH

(40 – 47‰) salinity conditions• KRILL- and KRILL+ tested

under high water salinity alone

DIETS Salinity EPA+DHA Req.FISH Both Satisfied*KRILL Both Satisfied*SOY Both DeficientKRILL- HIGH 50% deficientKRILL+ HIGH 25% above**based on 80% of that required by Penaeus monodon (Glencross et al., 2002)

Formulas and experimental design

FISH FISH KRILL SOY

KRILL SOY KRILL- KRILL+

FJ46

FJ47

FJ48

FJ49

FJ50

FI41

FI42

FI43

FI44

FI45

FH36

FH37

FH38

FH39

FH40

FG31

FG32

FG33

FG34

FG35

FF26

FF27

FF28

FF29

FF30

FE21

FE22

FE23

FE24

FE25

FD16

FD17

FD18

FD19

FD20

FC11

FC12

FC13

FC14

FC15

FB06

FB07

FB08

FB09

FB10

FA01

FA02

FA03

FA04

FA05

FISH

IDEAL water salinity (20 - 26‰) HIGH water salinity (40 - 47‰)

SOY KRILL KRILL- KRILL+

Six replicate tanks were assigned for each diet, except FISH and KRILL which used a total of seven replicate tanks under IDEAL salinity conditions.Allotment of feeds in rearing tanks followed a random block design

Tank Distribution

INGREDIENT

Soybean meal, 46% CP

FISH

350.00

KRILL

350.00

SOY

350.00

KRILL-

350.00

KRILL+

350.00Wheat flour 298.71 299.80 300.00 300.00 291.46Poultry by-product meal 100.00 100.00 105.00 100.00 67.50Fishmeal, Anchovy 60.17 71.58 70.55 75.62 68.90Soybean protein concentrate 32.48 19.10 16.65 16.65 20.00Corn gluten meal, 65% CP 0.00 0.00 0.00 0.00 40.00DL-methionine crystalline, 99% 8.00 8.00 8.00 8.00 4.61Fish oil 26.65 0.00 0.00 0.00 0.00Krill oil (QRILL™) 0.00 48.28 0.00 14.53 55.00Soybean oil 10.00 4.45 34.49 21.21 3.82Lecithin, soybean 15.00 0.00 15.00 15.00 0.00Cholesterol (Solvay) 0.00 0.00 1.31 0.00 0.38Rice, broken 40.00 40.00 40.00 40.00 40.00Phosphate monodicalcium 13.00 13.00 13.00 13.00 13.00Potassium chloride (KCl) 10.00 10.00 10.00 10.00 10.00Squid meal, whole 10.00 10.00 10.00 10.00 10.00Salt common 10.00 10.00 10.00 10.00 10.00Mineral-Vitamin Premix (DSM) 10.00 10.00 10.00 10.00 10.00Synthetic binder (Bentoli) 4.00 4.00 4.00 4.00 4.00Ascorbic acid polyphosphate (DSM) 2.00 1.80 2.00 2.00 1.34

Pro

tein

Ingr

edie

nts

Fixe

d po

rtion

Lipi

d so

urce

sFormulas (g/kg)

Chemical Composition FISH SOY KRILL KRILL+ KRILL-Nutritional Levels

Crude protein¹ 351.8 354.4 353.5 353.1 351.8Crude fiber¹ 12.0 9.0 13.3 12.0 13.3Ash¹ 93.7 89.5 91.0 82.3 91.0Gross Energy (kcal/kg) 4,184 4,273 4,209 4,212 4,306

Inclusion of Lipid SourcesFish oil1 26.6 0.0 0.0 0.0 0.0Krill oil1,2 0.0 0.0 48.3 55 14.5Soybean oil1 10.0 34.5 4.5 3.8 21.2Cholesterol1,3 0.0 1.3 0.0 0.4 0.0Soybean lecithin1 15.0 15.0 0.0 0.0 15.0Ratio Marine:Plant Oil 2.7 0.0 11.0 14.4 0.4

Nutrient LevelsTotal lipid content1 88.80 94.00 80.80 91.30 80.50DHA (C22:6n-3)4 2.54 0.28 1.60 1.37 0.59EPA (C20:5n-3)4 5.10 0.65 5.35 5.07 0.00Ʃ HUFA4,5 7.64 0.93 6.94 6.44 0.59LOA (C18:2n-6)4 28.32 44.71 16.21 17.97 33.66LNA (C18:3n-3)4 3.40 4.91 1.52 2.03 3.94Ʃ EFA4,6 39.36 50.54 24.68 26.44 38.19Astaxanthin (ug/100 ul) 13.80 14.70 21.40 17.30 ---

1in g/kg of diet as wet basis.2QRILL™ oil, Aker Biomarine ASA (Oslo, Norway).3Cholesterol XG, Solvay Pharmaceuticals BV (Weesp, Netherlands). 4in % of total lipid content in the diet.5sum of highly unsaturated fatty acids (DHA+EPA).6sum of essential fatty acids (DHA+EPA+LOA+LNA).

CONDITIONING PERIOD140 shrimp/m2 (80

shrimp/tank) and raised for 22 days for a conditioning period to water salinity IDEAL: from 25 ± 0.9‰ (3.4%

CV) to 24 ± 0.4‰ (2.0% CV)HIGH: from 36 ± 0.8‰ (2.2%

CV) to 40 ± 0.4‰ (1.0% CV)

Study Set-up

PL Stocking

Rearing in 3,000 L tanks

Harvest

Salt dilution

Stocking and acclimation

NURSERY PL12 reared in

nursery tanks of 3,000 L at 2.4 PL/L for 48 days when they reached 0.65 ± 0.28 g (n = 152)

Header tank20,000 L

Data Collection1. After 22 days of acclimation started on

experimental diets at 2.79 ± 0.60 g2. Density reduced to 70 shrimp/m2 or 40

shrimp/tank3. Fed twice daily in feeding trays at

0730 and 1600 h on a consumption basis

4. Daily water analysis• pH, temperature, salinity and

dissolved oxygen5. 22-24 day interval – 10 shrimp/tank

were weighed6. After 64 days shrimp were counted

and individually weighed1. weekly growth rate (g/week)2. Final body weight (g)3. final survival (%)4. yield (g/m2)5. food conversion ratio (FCR)

Feeding protocol used to adjust the amount of feed delivered based estimated consumption from trays% Amount of Feed Left in Trays

Feed Ration Adjustment

< 10% No change in feed ration> 10% Deliver 75% of original feed rationNo feed remains Increase 10% of original feed ration

Water Quality Trend towards increasing

salinity, from 22‰ to 25‰ under IDEAL salinity tanks and from 41‰ to 45‰ under HIGH salinity tanks No differences between

treatments for pH, salinity and temperature Significant differences

between HIGH and IDEALfor water salinity

Salin

ity (p

pt) -

Tem

pera

ture

(oC

)

pH

6.8

7.0

7.2

7.4

7.6

7.8

8.0

8.2

8.4

8.6

20

25

30

35

40

45

50

Days of Rearing

Salinity (‰)Temperature (ƒC)pH

IDEAL water salinity tanks

6.6

6.6

6.8

7.0

7.2

7.4

7.6

7.8

8.0

8.2

8.4

8.6

20

25

30

35

40

45

50

1 3 6 8 11 14 16 18 21 23 27 29 31 34 36 39 42 44 46 50 52 55 57 60 63

Days of Rearing

Salinity (‰)Temperature (ƒC)pH

HIGH water salinity tanks

Salin

ity (p

pt) -

Tem

pera

ture

(oC

)

pH

Par. IDEAL HIGHpH 7.27 ± 0.33 7.35 ± 0.21 Salin. 23 ± 1.2* 44 ± 2.0* Temp. 27.3 ± 0.46 27.5 ± 0.46

Final Shrimp Survival (%)

1. Chronic exposure to high salinity did not deteriorate shrimp survival2. Increasing n-3 HUFA provided no additional benefit to shrimp survival

91.8% 92.5%90.0%

93.8% 96.3% 95.0% 95.0% 94.2%

60.0%

70.0%

80.0%

90.0%

100.0%

FISH SOY KRILL FISH SOY KRILL KRILL - KRILL+

IDEALwater salinity HIGH water salinity

P = 0.896P = 0.720

Shrimp Yield (g/m2)

1. Final shrimp yield did not vary significantly among different diets regardless ofthe salinity concentration

533

555569

529536

598

531

579

400

450

500

550

600

650

FISH SOY KRILL FISH SOY KRILL KRILL- KRILL+

IDEALwater salinity HIGH water salinity

P = 0.550 P = 0.370

Yield for KRILL 12-13% higher compared to FISH, SOY and KRILL-

Shrimp Final Body Weight (g)

11.12

11.52

12.03

10.96 10.86

11.91

10.88

11.79

10.00

10.50

11.00

11.50

12.00

12.50

FISH SOY KRILL FISH SOY KRILL KRILL- KRILL+

HIGH water salinity

P < 0.0001

P < 0.0001

IDEAL water salinity

a

AA

B

A

b

c

B

Factorial analyses:both water salinity and diet type had a significant effect on shrimp body weight

at 1.45% KRILL- could not counterbalance the effects of high salinity

No growth improvements by further increase in KRILL+ at 5.50%

KRILL able to promote a significantly higher shrimp growth compared to FISH and SOY, regardless of salinity

Weekly Growth Rate (%)

8.33%

3.22%2.94%

Y = 1.451X2 - 24.84X + 109.0R² = 1.000

0.0

2.0

4.0

6.0

8.0

10.0

SOY FISH KRILL

WEIGHT GAIN DEPRESSIONAFTER SALINITY STRESS (%)

As water salinity increased from IDEAL to HIGH there was a growth depression, less significant with animals fed the KRILL diet

ConclusionsSALINITY1. The higher the salinity, the more important was n-3 HUFA (DHA + EPA) to

boost shrimp growth2. Under 21 – 26‰ shrimp did not appear to require diets with high n-3 HUFA

(DHA + EPA) levels as they performed well when fed a diet containing only a vegetable oil source (i.e., soybean oil)

SOURCES3. KRILL oil delivered an increased shrimp growth under both regular (21 –

26‰) and hypersaline (40 – 47‰) rearing conditions compared to the other lipid sources tested

LEVELS4. Under persistent hypersaline conditions, daily exposure to KRILL oil

appeared to be more important than an increased inclusion level 5. Under hypersaline water, n-3 HUFA, particularly DHA had the greatest

impact on shrimp growth. Best final body weight was achieved when projected DHA achieved 1.6% of total lipid content

6. No further enhancement in growth was observed when DHA levels exceeded this threshold for a salinity of 44 ± 2.0‰.