R2 = 0.2096. 0. 50. 100. 150. 200. 250. 300. 350. 0. 1. 2. 3. 4. 5. 6. Sampling Day. T o x ic ity. - u g. S. T. X e q. /1. 0. 0 g. R01 - Starvation- seawater only ...
Concentration of live non toxic algal feed Use of non toxic algal concentrates Starvation Clam density Water quality
3 x Raceways (2m L x 0.4m W x 0.2m D) with buffer tanks Air conditioned, insulated unit 180 litres of water- total volume Water Temperature - 16°C (+/- 1) 900 litres/ hour flow rate 3 - 3.5kg in weight in each raceway 80 clams in each
divided equally into 4 baskets
Fluorescence readings used to monitor algal concentration in raceways
10 clams selected on each sampling day Individual clams tested for PSP toxin levels Level of toxicity was determined using a Saxitoxin enzyme immunoassay method (Ridascreen Fast Saxitoxin Kit from rBiopharm). Ammonia and nitrite levels monitored daily Biodeposits and seston removed twice daily Siphon behaviour observed twice daily
Minimum of 10 days duration Dinoflagellate Alexandrium minutum Cultured 60l bags Conc. of cultures -20,000 -25,000 cells/ml Used at 9-11 days of age Feed concentration at 200-400 cells/ml Continuous pump feed with microalgae plus manual addition Required level of toxicity approx. 200 g/100g STX eq Samples taken at day 0 and regular intervals during toxification
4 - 8 days duration Live non toxic algal cultures Isochrysis galbana (Tahitian strain- T.Iso) 2 x 106 cells/ml culture conc. Pavlova lutheri 3 x 106 cells/ml culture conc. Non Living algal concentrates, BlueBiotek T. Isochrysis 2 x 109 cells/ml culture conc. 400 x 106 cells/ml culture conc. Continuous peristaltic pump feed plus manual addition Samples taken on each day of detoxification (where practicable)
Mean toxicity levels obtained ranged from approximately 30 400 g STX eq./ml Toxification Data Mean Toxicity - ug STX eq /100g
The Effect of Algal Concentration on Detoxification The effect of algal concentration on detoxification kinetics
450
R01-72,000 cells/ml R02 -36,000 cells/ml
400
y = 381.46e-0.1322x R 2 = 0.8339
Toxicity - ug STX eq/100g
350
R03 -12,000 cells/ml
300 250 200 150
y = 300.03e-0.1923x R2 = 0.8706
y = 246.17e-0.2024x R 2 = 0.9907
100 50 0 0
1
2
3
4 Sam pling Day
5
6
7
8
Effect of starvation on detoxification kinetics The effect of starvation on detoxification kinetics R01 - Starvation- seawater only
350
Toxicity - ug STX eq /100g
300 y = 236.57e-0.0654x R2 = 0.2096
250 200 150 100 50 0 0
1
2
3 Sam pling Day
4
5
6
Effect of Algal concentrates
Effect of non toxic algal concentrates R02 -400 x106 cells/ml 350
R03 -2 x109 cells/ml
Toxicity -ug STX/ 100g
300 y = 259.45e-0.0707x R2 = 0.35
250 200 150
y = 272.22e-0.1332x R2 = 0.8963
100 50 0 0
1
2
3 Sam pling Day
4
5
6
The table shows the predicted number of days to reach a safe level of 80 g STX/100g for an initial toxin level of 200 g STX/100g, calculated using the mathematical model t(d) = -0.91/ k, where the k value is taken from each exponential trend. EXPERIMENTAL CONDITIONS
It was observed that feeding was inhibited ( FU did not fall as quickly) when water quality deteriorated. To ensure no inhibition of feeding occurred a complete water change was carried out when the ammonia rose above 0.6ppm (maximum acceptable level 0.9ppm) or nitrite level reached 2.0ppm (maximum acceptable level 2.5ppm) This generally equated to approximately every 2-3 days.
Although ammonia and nitrite levels increased over the 4 days no inhibition of feeding was seen. % Fall at 60 mins - comparison on each day 100
R02
90 80
R03
70 60
% fall
RO1
50 40 30 20 10 0
day 01
day 02
day 03
Time
day 04
60 mins day 01 day 02 day 03 day 04
RO1
R02 70 76.11 75 72.65
R03 58.72 67.35 89.07 56.7
51.23 48.26 50.04 65.36
Effect of Clam density in raceways
R01 R02 R03
Weight of clams (kg) 4 2.5 1
Volume of water (l) 100 100 100
Density ratio 1:25 1:40 1:100
The reduction in algal concentration was not significantly different from one raceway to another. This may have been due to there being insufficient difference in clam density between raceways. Change in algal concentration - Day 4 R01
70
R02
60
R03
50
Expon. (R01)
40
Expon. (R03)
30
Expon. (R02)
20 10 0 0
50
100 Time - mins
150
200
Algal conc. -(x1000) cells/ml
Algal Conc. -(x1000)cells/ml
Change in algal concentration - Day 1 70
R01
60
R02 R03
50
Expon. (R01)
40
Expon. (R03)
30
Expon. (R02)
20 10 0 0
50
100 Time mins
150
200
Siphon behaviour was recorded twice daily during toxification and detoxification. Visible show of mantle shell open was taken as feeding Number of clams showing siphon decreased at high levels of PSP toxin Siphon behaviour of clams 100
R02
90
% of clams siphoning
R01
R03
80 70 60 50 40 30 20
Start of Detoxification
10 0 18/01/04
20/01/04
22/01/04
24/01/04
26/01/04
28/01/04
Date
30/01/04
01/02/04
03/02/04
05/02/04
Safe levels (i.e. 1.5mg/L TPM) Feeding with concentrated algal pastes did not decrease the PSP to a safe level within 4 days of detoxification. The predicted days were more than twice those predicted for live algal feed. Starvation of clams did not give safe levels of PSP within 4 days of detoxification. No significant difference in rate of detoxification was observed between starvation and feeding with the most concentrated algal paste.
A. minutum was not seen to be inhibitory to feeding (at 0.5mg/L TPM). Ammonia and nitrite levels appeared to have an effect on feeding kinetics. As levels in the water increased feeding was inhibited. This was explored further in experiment 4, but was not confirmed. At very high concentration of non toxic feed algae (216,000 cells/ml or > 6mg/L TPM) excessive production of pseudofaeces was observed. Siphoning behaviour was observed to be inhibited at high levels of PSP toxins.
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