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Article history: Monitoring programmes have demonstrated seasonal and short term variation of inlet water quality in
Received 25 July 2013 Chilean salmon hatcheries. Potentially toxic levels of copper (Cu) and aluminium (Al) in many cases may
Accepted 3 April 2014 explain observed increased mortalities during the early life-stages of Atlantic salmon. Therefore, there is
a need for efficient water treatments methods. Two alternative water treatment methods were tested to
Keywords: reduce the toxicity of Cu and Al in fresh water production systems (egg to first feeding): crushed marble
Atlantic salmon
filtration treatment (MF) and sodium silicate addition (SA) treatment. To mimic the observed natural
Water quality
conditions an experiment was set up with short term (2–3 days) pulses of elevated Cu (20 and 30 g L−1 )
Metal toxicity
Fish welfare
and Al (80 and 130 g L−1 ) concentrations. The effect of the two treatments was tested during 3 distinct
Water stabilization developmental periods of the salmon, from 401 to 424, 551 to 567, and 729 to 736 accumulated thermal
Chilean aquaculture units (ATU). Treatments were compared with a positive control group with metal addition (PC), and a
negative control group (NC) exposed to untreated water with no metal addition. Compared to the groups
that received metal pulses with no treatment (PC), mortalities were significantly reduced by both silicate
addition and marble filtration. Both at 896 and 1013 ATU marble filtration (MF) group showed the lowest
mortality. The present study suggests that crushed marble filtration and sodium silicate addition is well
suited to treat Al and Cu pulses in the relevant concentration range tested. This constitutes a low cost
and robust alternative for freshwater aquaculture facilities that need to improve the quality of their inlet
water, and may consequently decrease mortality and improve fish welfare for Atlantic salmon.
© 2014 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.aquaeng.2014.04.001
0144-8609/© 2014 Elsevier B.V. All rights reserved.
78 C.A. Pessot et al. / Aquacultural Engineering 60 (2014) 77–83
Fig. 2. (A) Tray 6 was moved to position of tray 3 and tray 3 to position of tray 6 in order to obtain populations with one and two pulses respectively. (B) After Episode 3,
tray 10 from MF was set on the line of SA obtaining a new treatment group (tray 10).
of modules: Module NC, trays 1–3; module PC, trays 4–6 (from PC1 11–12) with those that finished marble treatment after the first
and PC2 were obtained later); module SA, trays 7–9 and module feeding (group 10).
MF, trays 10–12). Each module consisted of 3 trays (30 cm × 30 cm)
containing 1680 eggs each, with a total of 5040 eggs per test. The 2.4. Sampling
eggs were received at 363 accumulated thermal units (ATU).
As described previously, modules PC, SA and MF (Fig. 1) received Water samples were taken from inlet waters (after treatments
a total of 3 different Al/Cu pulses. Al and Cu were episodically added MF and SA when correspond) from day 0 (370 ATU, after 1 day eggs
to the 3 treatment groups at days 4–7 (Episode 1, 401–424 ATU), were received) until day 67 (912 ATU). No artificial heating of water
24–26 (Episode 2, 551–567 ATU) and 47–48 (Episode 3, 729–736 was applied during the experiment, so ATUs is increased based only
ATU) to nominal concentrations (i.e. concentrations in the modules the natural temperatures and time. Control module samples were
after adding pulses) of 80 g L−1 Al/20 g L−1 Cu for Episodes 1 and taken every 3rd day and every day during Cu/Al pulses.
2, and 130 g L−1 Al/30 g L−1 Cu for Episode 3. Water samples taken from the NC included metal analyses (Al
After the first pulse, tray number 3 from module NC was moved and Cu) and also basic water variables related to metal toxicity
to module PC, and tray number 6 from module PC was moved to modulation: pH, turbidity, total organic carbon (TOC), Ca and Mg.
module NC. Trays 4 and 5 received pulses 1, 2 and 3. Tray 3 received Water samples from the 3 different treatments (PC, SA and MF)
pulses 2 and 3, and tray 6 received pulse 1 (Fig. 2A). were only tested for Al and Cu.
In order to compare the differential effect over mortality of Biological samples (3 replicates of 2 g each) were taken to ana-
crushed marble versus silicate treatment on biological material, lyse total Al and Cu concentrations in complete specimens (eggs
once 1st feeding was starting after day 66 (corresponding to 896 and fry). Samples of eggs at Day 0 and Day 5 were analysed at
ATU) specimens (fry) were rearranged into the following groups in the laboratory of the Norwegian University of Life Sciences, Oslo,
each module: Module NC (group 1–2 from trays 1 and 2, group 6 Norway, with parallel analyses at the Department of chemistry and
from tray 6); module PC (group 4–5 from trays 4 and 5, group 3 from biochemistry at the University of Valparaiso, Chile. Samples from
tray 3); module SA (group 7–8–9 from trays 7, 8 and 9; group 10 Day 17 and onwards were analysed at the University of Valparaiso.
from tray 10 coming from marble treatment); module MF (group Metal determination in biological samples consisted of acid, oxi-
11–12 from trays 11 and 12) (Fig. 2B). This way group 10 from orig- dant digestion at 200 ◦ C for 2 h and detection by Inductive Coupled
inal marble treatment (MF) was exposed to silicate (SA) in order to Plasma (ICP-AES). Certified reference material DOGFISH DORM-2
compare marble treated specimens (fry) after first feeding (group from National Research Council Canada was used as reference.
80 C.A. Pessot et al. / Aquacultural Engineering 60 (2014) 77–83
2.6. Chemical analysis Fig. 3A and B shows the Al and Cu concentrations in the water
samples. Al and Cu analyses were done at the dates of the metal
Water temperature was measured daily. Upon sampling of pulses. Average concentrations of Al and Cu in 1st, 2nd, and 3rd
organisms, water samples for analysis of total organic carbon (TOC), pulse in the PC were 128, 140 and 207 g L−1 for Al and 12.0,
pH and turbidity of the water were taken. Analyses of Al, Cu, Ca and 15.5 and 26.0 g L−1 for Cu respectively. For Al, the first and sec-
Mg concentrations were done by ICP-MS (Induced Coupled Plasma ond pulse the target (nominal values) was 80 g L−1 and for third
with mass detector). pulse the target (nominal value) was 130 g L−1 . For Cu, first and
second pulse target (nominal values) were 20 g L−1 and for the
third pulse the target (nominal value) was 30 g L−1 . In the case of
Al, differences between the nominal values and the obtained con-
2.7. Data analysis centrations are explained by the natural background levels of Al
in the water. The average concentration of Al in the water source
Difference between the different treatments in metal and Ca was 11 ± 6.84 g L−1 whereas maximum concentration recorded
concentration in water was tested by Analysis of variance (one-way was 31 g L−1 . Also some loss of water coordinated molecules
ANOVA). from (Al2 (SO4 ) × 18H2 O) could have affected this. Both conditions
To determine if there were significant differences between explain why the final concentrations were somewhat higher than
treatments after 912 ATU (from first feeding), the Homogeneity calculated.
Test for Regression Coefficients was applied (Gerome C.R. Li, 1964).
The test establishes differences between treatments based on the
3.3. Al and Cu in biological material
slopes presented by different treatments on a defined range of time
(from 912 to 1013) for the dependent variable (mortality).
Samples showed similar trends for both the metals Al and Cu. An
increase in metal accumulation was observed until 522 ATU (initial
sac fry stage) after the 1st metal pulse, whereas a decrease of the
3. Results metal concentrations to background levels (average and standard
deviations were 1.21 ± 0.95 g g−1 ; for Al and 1.01 ± 0.17 g g−1 for
3.1. Background water quality in NC and water Ca concentrations Cu, wet weight), was observed after 559 ATU even though a 2nd and
in treatments 3rd metal pulse were applied at 559 and 736 ATU.
Background water quality monitored during the study 3.4. Accumulated mortalities in the different treatments
showed stable conditions. Average and standard deviations
were, for pH (7.64 ± 0.09), Turbidity as NTU (0.46 ± 0.22), TOC Fig. 4 shows mortality trends in the 6 different treatments (NC,
(0.21 ± 0.06 mg C L−1 ) and Hardness, based on Ca and Mg con- PC, MF, SA, PC1 and PC2). The actual timing of the Al and Cu pulses
centration (16.9 ± 0.59 as mg L−1 CaCO3 ). When comparing Ca is indicated in the graph, described as “1st Pulse”, “2nd Pulse” and
concentrations in water for the different modules, no differ- “3rd Pulse”. The figure also shows the time period when hatch-
ences were detected between PC, NC and SA (4.4 ± 0.04 mg L−1 , ing was recorded at 522 ATU. No significant difference in mortality
4.4 ± 0.09 mg L−1 and 4.4 ± 0.11 mg L−1 , respectively). However, was observed between the different treatments as a response to
treatment with marble showed an increased in Ca concentration to the 1st pulse of Al and Cu before hatching (corresponding to 401
7.1 ± 0.37 mg L−1 compared to PC, SA and NC. Results from the MF ATU) (F0.5 > F under Test for Regression Coefficients). However after
module show that crushed marble increased the Ca concentration hatching and the 2nd pulse of Al and Cu at 567 ATU, it is possible to
in water. This difference was highly significant (ANOVA, P < 0.05). observe a first significant difference (F0.5 < F under Test for Regres-
The Ca concentration over a period of 31 days showed no trends sion Coefficients) in mortality between the eggs that received Al/Cu
and stayed stable. pulse without water pre-treatment and those that received water
C.A. Pessot et al. / Aquacultural Engineering 60 (2014) 77–83 81
4. Discussion
concentrations were lower than Al, SA treatment was not able to 5. Conclusion
reduce the accumulation of this metal in the eggs before hatching
(from 501 to 522 ATU). This could be related to a low affinity The present study confirmed that marble filtration as well as
constant of this metal to silicate and higher relative affinity sodium silicate water treatment can be an effective, low cost and
constant to biological moieties in eggs or fish. An important factor robust alternative for hatcheries facing elevated levels of Al and Cu
to be considered is the high pH this trial was carried out (which is in their inlet waters. The water requirement during early life stages
representative for Chilean salmon hatcheries). in salmonid aquaculture is relatively low, making this technology
MF treatment when compared to PC, showed lower concentra- feasible at this life-stage. The experiment is actually carried out in
tions for Al or Cu in the period from 501 to 522 ATU, and higher a commercial facility, albeit not on a full scale egg group. A 10–20
Cu concentrations than NC in the same period. Biological tissues times up-scaling of the marble filter or sodium silicate dosing is
(complete specimens) from MF treatment showed concentrations quite feasible in terms of costs and/or space requirements.
of Al and Cu lower than those observed for the silicate treatment. There was an effective decrease of Al and Cu levels in the marble
These results are also in accordance with Al and Cu concentration filtrated water which in part can explain the better survival of eggs
in water samples, showing lower concentrations for MF than for PC and fry exposed to this treatment, but it is also shown that increas-
treatment. Results from the MF treatment demonstrated the capac- ing the concentration of Ca by marble treatment can decrease the
ity of the system to significantly decrease metal concentrations in background levels of mortality. This effect could be responsible
water (ANOVA, P < 0.05) but also significantly increasing the con- for the better performance after first feeding, regardless of the
centration of Ca (ANOVA, P < 0.05) compared to the inlet water (see presence of toxic concentrations of Al and Cu. After hatching, as
Fig. 3A and B). These results (Fig. 3A) show that water treated with described previously, no accumulation was observed, but the toxic
crushed marble (MF) caused a decrease in the actual concentration effect was stronger clearly suggesting that this is a more sensitive
of Al. This difference was statistically significant under a P < 0.05 by state possibly given by the Al and Cu targets in biological tissue.
ANOVA when compared with PC treatment. The final concentra- Sodium silicate treatment also confirmed to be an effective tech-
tion of Al in water treated with silicate (SA) showed even a slightly nology to reduce adverse effect driven by Al and Cu in water. Low
higher concentration than NC when no pulse where added and decrease in water Al concentration by MF, suggests that a com-
also higher when pulses were added compared to trays exposed to bined application of marble and silicate should be studied in order
pulses and no silicate or marble treatment. This is explained at some to validate this as an option to control combined metal toxicity.
extent by the presence of Al (38 mg L−1 ) in the original stock solu-
tion of silicate. In the case of Cu, MF treatments showed, the same Acknowledgments
as for Al, it decreased the actual concentration of Cu. This difference
was statistically significant (P < 0.05, ANOVA) when compared with Special thanks to Granja Marina Tornagaleones represented by
PC. Ignacio Ochagavía whom allowed the information obtained dur-
This suggests that the positive effect of marble in terms of reduc- ing this project to be published in this article. Also thanks the
ing the toxic effect of Cu and Al from 501 to 522 ATU, is given by company for facilitating its installations and valuable time that
a dual effect, including adsorbing Al and Cu plus liberation of Ca Andres Streuly, Mauricio Schulz, Cristián San Martin, Beatriz Gar-
to the media. This dual effect becomes more evident starting from rido, Roberto Michell and Claudio Bernales dedicated to this project.
676 ATU and afterwards being the MF treated group the one with Thanks to research assistant Anne May Ilestad for her help in run-
lowest mortalities, even lower than NC. ning the experiment, and to Rafael Pessot for his support in the
At 567 ATU, after the second pulse of Al and Cu was applied, statistical analysis of the data. This research was supported by funds
it was possible to see that NC and pre-treated groups (MF and SA) received from CORFO (Corporación de Fomento para la Producción),
showed lower mortalities than to Cu Al treated groups (PC, PC1 and Ministry of Economy, Chile, Reference No. O9IERI-5623 and Granja
PC2). The rates of mortality allow us to distinguish a separation of Marina Tornagaleones.
trends in the range from 676 to 736 ATU for the different treat-
ments: lower mortalities are shown by NC, pre-treated (MF and
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