Centre for Sustainable Aquaculture (CSAR)

Swansea University
Singleton Park
Swansea SA2 8PP

Tel. 01792 602030

Fax 01792 295955

Technical Report
submitted to Aker BioMarine

Contract Research
ID # Aker02

Project manager: Ingrid Lupatsch

Performance of juvenile Litopenaeus vannamei fed diets at increasing

levels of krill meal.
Centre for Sustainable Aquaculture (CSAR)
Swansea University

Research Project Identification

Contract ID Aker02

Contractor Aker BioMarine ASA

Fjordallèen 16
POB 1423 Vika, 0115 Oslo
Norway

Contacts Dr. Simon Wadsworth, Dr. Sigve Nordrum

Email: simon.wadsworth@akerbiomarine.com
Email: Sigve.nordrum@akerbiomarine.com
Project Title Performance of juvenile Litopenaeus vannamei
fed diets at increasing levels of krill meal.

Trial Location Centre for Sustainable Aquaculture Research

Swansea University
Singleton Park
Swansea, SA2 8PP

Project manager Dr. Ingrid Lupatsch

Email: i.lupatsch@swansea.ac.uk

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

Performance of juvenile Litopenaeus vannamei fed diets at increasing levels of

krill meal.

The objective of the following trial was to assess the growth performance, feed
efficiency and survival of juvenile Litopenaeus vannamei fed feeds where fish meal
was gradually replaced by full fat krill meal QrillTM.

Material and Methods

Shrimp and rearing conditions

Pacific white shrimp (Litopenaeus vannamei) were obtained from the rearing tanks at
CSAR. These shrimp were bred at CSAR from the original broodstock shipped in
from Bonaire. Shrimp selected for the trial (~ 0.5g) were hand graded to
approximately uniform size. Prior to the trial shrimp had been fed a local shrimp feed
(42% protein, 10% lipid) manufactured by Dragon Feed Ltd (Port Talbot, U.K.)
which specializes in feeds derived from ragworm, Nereis virens.
The experimental system consisted of fifteen identical 20L plastic aquaria. The tanks
were covered with individual plastic mesh covers to prevent shrimp from escaping.
Water temperature was maintained at 27°C ± 0.8 using a thermostatically controlled
water heater within the sump of the recirculation system. The salinity of 32 ppt was
maintained by adding marine grade salt supplied by Tropic Marin®. The light regime
of the room was 16 hr of light (7:00 hours to 23:00 hours) and 8 hr of dark.

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 Centre for Sustainable Aquaculture (CSAR)
Swansea University

Feed preparation

The experimental feeds were formulated and manufactured at CSAR using commonly
available feed ingredients as described in Table 1. All feeds were formulated to
contain 40% protein and 10% lipid and fish meal was gradually replaced by krill meal
QrillTM substituting 0%, 30%, 60% and 100% of fish meal as indicated in Table 2.
The ingredients (Table 1) were combined and thoroughly mixed before the addition of
the fish oil and water. A doughy mixture was produced of approximately 40-50%
moisture content, this dough was then extruded through a meat grinder with
approximately a 1.5mm diameter orifice plate. The resulting extruded spaghetti-like
strands were then dried at 450C for 24 hrs.

Table 1: Composition of feed ingredients (per kg feed).

Ingredient Dry Matter Crude Ash Lipid Gross

(g) Protein (g) (g) (g) Energy (MJ)
Krill meal1 960 549 95 220 23.08
Fish meal2 926 666 167 100 18.95
Wheat gluten3 920 760 60 8.0 21.24
Wheat starch 930 29 - 16.23
Fish oil 1000 38.49
1
Krill meal (QrillTM ) – source Aker BioMarine
2
Fishmeal – source Argentina
3
Wheat gluten – source Italy

After the mixture was hardened the spaghetti-like strands were broken up again to
create pellets of between 5 and 10mm in length. After oven drying and when fed to
the shrimps the feeds maintained a residual moisture content of about 50 g/kg diet.

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 Centre for Sustainable Aquaculture (CSAR)
Swansea University

Table 2: Formulation of feeds (g per kg)

Diet 1 Diet 2 Diet 3 Diet 4 Diet 5

(0%) (15%) (31%) (48%)
Fish meal1 480 320 150 0
Krill meal full fat2 0 150 310 480
Wheat gluten3 120 140 155 150
Wheat starch 290 290 290 280

Closed formula
Vitamin and Mineral4 10 10 10 10
Stay C5 0.8 0.8 0.8 0.8
CaSO4 20 20 20 20
Alginate6 20 20 20 20
KH2PO4 0 10 25 40
Fish oil 60 40 20 0
1
Fish meal - source Argentina, South America.
2
(QrillTM) Aker BioMarine.
3
Source Italy, Europe.
4
Vitamin and Mineral premix for shrimps, Source DSM – nutritional products
5
Phosphorylated ascorbic acid – source DSM nutritional products
6
Binder

Experimental set up

The growth trial was carried out for 53 days in 15 identical 20L plastic aquaria,
providing three replicates per treatment. The internal control was the commercial
shrimp feed (closed formula) provided by Dragon Feeds Ltd. (Port Talbot, U.K.),
Twenty L. vannamei (initial weight ~0.49g) were batch weighed and stocked into
each aquarium. Shrimp were fed manually twice a day to satiation, the total amount of
feed actually consumed was monitored by counting the left over pellets (the average

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

weight of a pellet was determined beforehand and was 15mg / pellet). To ensure that
shrimp were fed to appetite, in tanks where few pellets (<20) were left feed was
increased and in tanks where many pellets (>30) were left, feed was decreased. All
feeds were easily manipulated by the shrimp and once immersed in water remained
intact for up to 24hours. Approximately every 14 days, the shrimp from each tank
were batch weighed and counted. Daily routine - besides feeding - involved recording
of moults and any mortalities.

Sample preparation and chemical analyses

Initial samples of about 25 shrimp were taken and at the end all the shrimp from each
tank were sacrificed and blended to create homogenous samples. Sub-samples for
estimation of dry matter were taken from the sampled shrimp before the remaining
homogenate was oven-dried. The dried samples were once more mixed in a blender
before all remaining analyses. Identical analyses were applied for diets and body
homogenates. Crude protein was measured using the Kjeldahl technique and
multiplying N by 6.25. Ash was calculated from the weight loss after incineration of
the samples for 12 h at 5500C in a muffle furnace. Gross energy content was measured
by combustion in a Parr bomb calorimeter using benzoic acid as the standard.

Results

The results from the feed analysis are presented in Table 3. In general all diets
reflected the intended composition, yet there was a gradual decrease in crude protein
as the krill meal substitution increased, this was due to the fact that Qrill TM contained
actually only 55% protein compared to the 59% protein as stated by the manufacturer.

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 Centre for Sustainable Aquaculture (CSAR)
Swansea University

Table 3: Proximate composition of the feeds (per kg as fed) at increasing inclusion

levels of krill meal.

Dry Matter Crude Protein Ash Gross Energy

(g) ( g) (g) (MJ)
Diet 1 (0 %) 962 424 107.3 19.24
Diet 2 (15%) 955 411 100.1 19.25
Diet 3 (31%) 954 394 94.9 19.24
Diet 4 (48%) 964 388 98.3 19.44
Diet 5 (Commercial) 926 447 92.0 19.63

Table 4 describes the growth performance of shrimp fed the experimental diets.
Superior growth and FCR was found in shrimp fed the commercial feed. Comparison
of weight gain among the shrimp fed the four experimental feeds only showed the
highest weight gain occurred in shrimp fed Diet 2 (15% krill) and the lowest weight
gain in shrimp fed Diet 4 (48% krill). Survival was uniform throughout the trial and
not related to treatment. Analysis of variance however showed that the FCR of the
shrimp fed the different diets was not significantly different among the treatments
(Table 4). The reduced weight gain was strongly correlated to reduced feed intake,
showing a gradually decreasing feed consumption ranging from 9.32% to 6.64% per
day from Diet 1 through to Diet 4.

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 Centre for Sustainable Aquaculture (CSAR)
Swansea University

Table 4: Performance of Pacific white shrimp fed the four experimental

feeds (mean  std of three replicate treatments) after 53 days of growth.

Initial Final Weight gain FCR Feed Survival

BW (g) BW (g) g/shrimp/day intake %
% / day
Diet 1 0.50 3.59 0.058 a,b 2.14 a 9.32 a 65.7 a
0% Krill  0.07 0.03  0.002  0.02  0.80  7.4
Diet 2 0.50 4.22 0.070 a 1.89 a 9.13 a 68.3 a
15% Krill  0.01  0.03  0.001  0.18  0.85  16.1
Diet 3 0.51 3.19 0.051 b 1.84 a 7.29 b 81.7 a
31% Krill  0.02  0.37  0.007  0.19  0.43  2.9
Diet 4 0.46 2.26 0.034 c 2.06 a 6.64 c 70.0 a
48% Krill  0.03  0.57  0.010  0.34  0.27  13.2
Diet 5 0.47 5.71 0.099 1.56 9.40 71.7
Commercial  0.02  0.27  0.005  0.05  0.37  2.9
a,b,c
Values in a column with the same superscript are not significantly different
(p>0.05). Diet 5 was not included in the statistical analysis.

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

Figure 1: Voluntary feed intake in g per shrimp per day estimated for each fortnight
(mean of three replicate treatments).

The voluntary feed intake (g per shrimp per day) as calculated for each weighing
period is depicted in Figure 1. This graph shows that from the start of the trial shrimp
showed a preferential feed intake for Diet 2 and the least feed consumption for Diet 4.
This pattern of feed intake is very well reflected in the subsequent growth
performance (Table 4).

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 Centre for Sustainable Aquaculture (CSAR)
Swansea University

Table 5: Composition of shrimp per g live weight (mean ± SD).

Initial Diet 1 Diet 2 Diet 3 Diet 4 Diet 5

sample (0%) (15%) (31%) (48%)
Dry Matter 234 250 246 243 231 255 ±
(mg) ± 2.4 ± 6.7 ± 2.9 ± 11.3 2.9
Ash 34.7 27.8 28.3 28.9 28.9 28.3 ±
(mg) ± 1.2 ± 1.4 ± 1.4 ± 0.1 1.07
Crude Protein 159 176 172 172 165 181 ±
(mg) ± 2.6 ± 0.3 ± 5.8 ± 6.7 6.1
Gross Energy 4.33 4.89 4.81 4.71 4.33 5.08 ±
(kJ) ± 0.06 ± 0.09 ± 0.07 ± 0.31 0.09

The composition of shrimps (Table 5) reflects the effect of size with shrimps fed Diet
4 being the smallest and having the lowest dry matter and thus energy content.

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

Table 6: Efficiency of energy and protein retention (mean ± SD, n=3) of shrimp fed
krill meal at increasing dietary inclusion levels.

Diet 1 Diet 2 Diet 3 Diet 4 Diet 5

(0%) (15%) (31%) (48%)
Gross Energy intake 2.400 2.553 1.782 1.314 3.037
(kJ / shrimp /day)  0.074  0.261  0.206  0.200  0.215
Energy gained 0.290 0.342 0.242 0.148 0.509
(kJ / shrimp /day)  0.007  0.008  0.036  0.048  0.026
Energy retention 12.10 13.48 13.61 11.05 16.78
efficiency (%)  0.12 a  1.00 a  1.54 a  2.23 a  0.31
Crude Protein 52.9 54.6 36.5 26.3 69.2
intake  1.6  5.6  4.2  4.0  4.9
(mg / shrimp /day)
Protein gained 10.4 12.2 8.8 5.6 18.1
(mg / shrimp /day)  0.2  0.3  1.5  1.7  1.0
Protein retention 19.64 22.54 24.13 21.17 26.17
efficiency (%)  0.32 a  1.70 a  2.73 a  3.46 a  0.54
a
Values in a row with the same superscript are not significantly different (p>0.05)

Discussion

Krill meal has a number of attributes that make it attractive as an aqua-feed. These
include palatability enhancement of feeds for crustacea (Smith et al, 2005), a source
of carotenoid pigments (Floreto et al, 2001), a source of essential fatty acids, a well
balanced amino acid profile and an improvement of larval fish survival.
In several investigations with marine fish species the replacement of fishmeal with
krill meal has had positive affects on growth showing that krill was partially or fully

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

interchangeable with fish meal (Olsen et al, 2006 Suontama et al, 2007) without
effecting or even enhancing growth performance of fish.
In the present study Litopenaeus vannamei only showed superior growth performance
up to a dietary inclusion level of 15% full-fat krill meal. At a 30% krill meal inclusion
(equivalent to 60% of fish meal replacement) performance of shrimp was not
significantly different to the fish meal only feed but fed the all krill meal feed (Diet 4)
growth was significantly inferior. This reduced growth of shrimp fed the high krill
meal feeds was mainly due to decreased feed intake. No significant differences were
found regarding FCR, energy and protein retention efficiency values (Table 6). This
fact supports the concept that whichever feed the shrimp consumed they utilized
equally well. This also suggests, that decreased growth performance might not be due
to lower digestibility of the krill meal or unbalanced amino acid profile, but
palatability might be the cause. This fact is quite surprising as krill meal had been
used in the past as an attractant in aquaculture feeds.
Feed attractants have been characterised and isolated from different marine organisms
such as squid, marine worms, mussels, clam, brine shrimp as well as krill (Smith et al,
2005; Kolkovski et al, 2000). However the use of krill as a feed attractant at higher
inclusion levels has had some interesting results. Kolkovski et al (2000) found that
coating a commercial diet with 10% krill hydrolysate did not improve growth or
survival of yellow perch juveniles compared to the 5% coated diet, thus concluding
that there may be a threshold level of the chemo-receptors of the perch, where a low
inclusion level may be all that’s needed to enhance feed intake.
These results are confirmed in our trial were feed intake was highest in shrimp fed the
15% krill meal feed (Diet 2). However at high substitution levels (shrimp fed Diet 4)
feed intake is markedly reduced throughout the trial.
Krill meal potentially contains a high amount of fluoride, which is mainly derived
from the exoskeleton. Euphausiids are known to have the highest overall fluoride
concentrations of a range of Antarctic marine crustaceans examined; normally around
(1500 mg/kg DM). According to the manufactures’ information Qrill TM in general
contains 1000-2000 (mg/kg DM) with seasonal variations.

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

This relatively high fluoride content has limited the use of krill as a food for fish.
Yoshitomi et al (2006) found that salmon (Salmo salar) showed a decrease in growth
performance as the levels of fluoride were increased. More recently other
investigations have found the use of low fluoride krill (LFK) meal to be successful in
the diets for salmon. LFK has approximately one-fourth the amount of fluoride of krill
meal (Yoshitomi et al, 2007).
Additional research is needed to determine the effects - if any - of increased dietary
fluoride levels on growth performance of shrimp.

Conclusions

The potential use of full fat krill meal as a replacement for fish meal in the diet of L.
vannamei is limited due to a reduced feed intake by the shrimp and resulting in
reduced growth. Based on this trial the highest dietary inclusion level of krill meal
where enhanced growth can be achieved is 15%. Further investigation of the use of
krill meal at substitution levels of up to 30% is needed.

References

Floreto, E., Brown, P. and Bayer, R. (2001). The effects of krill hydrolysate-
supplemented soya-bean based diets on the growth, coloration, amino and
fatty acid profiles of juvenile American lobster, Homarus americanus.
Aquaculture Nutrition 7, 33-43.
Kolkovski, S., Czeny, S. and Dabrowski, K. (2000). Use of krill hydrolysate as feed
attractant for fish larvae and juvenile. Journal of World Aquaculture Society
31, 81–88.

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Centre for Sustainable Aquaculture (CSAR)
Swansea University

Olsen, R., Suontama, J., Langmyhr, E, Mundheim, H., Ringø, E., Melle, W., Malde,
M.K. and Hemre, G.-I. (2006). The replacement of fishmeal with Antarctic
krill, Euphausia superba in diets for Atlantic salmon, Salmo salar.
Aquaculture Nutrition 12, 280-290.
Smith, D., Tabrett, S., Barclay, M. and Irvin, S. (2005). The efficacy of ingredients
included in shrimp feeds to stimulate intake. Aquaculture Nutrition 11, 263–
272.
Suontama, J., Karlsen, Ø., Moren, M., Hemre, G., Melle, W., Langmyhr, E.,
Mundheim, H., Ringo, E. and Olsen, R. (2007). Growth, feed conversion and
chemical composition of Atlantic salmon Salmo salar L. and Atlantic halibut
Hippoglossus hippoglossus L. fed diets supplemented with krill or amphipods.
Aquaculture Nutrition 13, 241-255.
Yoshitomi, B., Aoki, M., Oshima, S. and Hata, K. (2006). Evaluation of krill
(Euphausia superba) meal as a partial replacement for fish meal in rainbow
trout (Oncorhynchus mykiss) diets. Aquaculture 261, 440-446.
Yoshitomi, B., Aoki, M. and Oshima, S. (2007). Effect of total replacement of dietary
fish meal by low fluoride krill Euphausia superba meal on growth
performance of rainbow trout Oncorhynchus mykiss in fresh water.
Aquaculture 266, 219-225

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