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1. National Marine Environmental Monitoring Center, Dalian 116023, China
2. Dalian Ocean University, Dalian 116023, China
Received: December 19, 2011 / Accepted: January 9, 2012 / Published: February 20, 2012.
Abstract: Using a PSP (paralytic shellfish poisoning) toxin-producing strain of Alexandrium tamarense, it studied the timing of toxin accumulation and elimination of PSP toxins in Argopectens irradias. The PSP toxicity was studied by following the standard PSP mouse bioassay developed by the Association of Official Analytical Chemists (AOAC). Alexandrium tamarense was cultured to an average density of 1.26 × 104/mL for a total of about 50 L culture. The toxicity of the alga was 2.18 × 10-6 MU/cell. The results show that PSP content increased with time in both visceral and muscle tissue during a two-week accumulation period during which scallops were fed with A. tamarense. The average toxin level in scallop’s viscera was 49.4 MU/g, with an average of 10.0 MU/g in muscle tissue. This level is 2.5 times higher than the sanitation standard (4.0 MU/g of muscles). The highest value was 61.0 MU/g in the viscera. In summary, the viscera accumulated greater concentrations of toxin than muscle tissue. Scallops that had accumulated toxins were transplanted for two weeks into a field environment containing no toxic algae. The PSP content of the scallops decreased to 7.9 MU/g viscera and 1.6 MU/g muscles two weeks after being transplanted, but did not reach the sanitation standard. Under the experimental conditions, the toxin depuration rate of shellfish toxin was 12% daily. This study worked toward the development of a sanitary shellfish industry and better management of PSP toxin-impacted shellfish in China.
Key words: PSP, argopectens irradias, accumulation and elimination time.
1. Introduction
Countries around the world nowadays take to prevent shell poisonous toxins, reduce shell poisonous loss research, attend shellfish’s healthy aquaculture, establish shellfish purifying center, as well as accelerate shell toxins purification research and so on. Our country is a great economical nation of marine products, and the shellfish culture is an important cornerstone. It does not allow neglect the loss, for the harmful and virulent red tide. Therefore, it is extremely essential to study the algae toxin with the time of in vivo accumulation and elimination in the sea shellfish, and is important to control the sanitary quality of shellfish.
2. Materials and Methods
2.1 Methods of Algae Culture
The experimental algae were Qingdao strain of Alexandrium tamarense. It was cultured from March 2003. The algae were cultured in 30 mL, 15-18 °C. The illumination cycle was the natural photoperiod, and the intensity of illumination was 3,000-5,000 lux. Cell density was counted with blood counting chamber every 5-7 days. When the density was about 1 × 104 cells/mL, it can add the boiling disinfection sea water to culture expansively, with the proportion 1:1 of sea water to the algae. The salinity was 30.1. It should be shook 4 times every day and shook 2 minutes every time. 5 L glass Erlenmeyer flask was used for mass cultured, putted on wooden multi-layered shelf. The culture medium was filtered seawater with f/2 medium which is a nutrient salt
formula [1].
2.2 Determination of Algae Toxin
The densities of Alexandrium tamarense cell was counted on August 1, 2003. The density was 1.38 ×104 cells/mL. The sample of 500 mL was centrifuged at 2000 r/min for 10 min, and 5 mL algae cell was obtained at the bottom of the tube. The algae toxicity was assayed following the standard method of PSP mouse bioassay developed by the Association of Official Analytical Chemists (AOAC).
The densities of Alexandrium tamarens cell and Tetraselmis cell was counted with blood counting chamber.
2.3 Experimental Methods
Started from July 21, 2003, the experiment was divided into four groups of 100 Argopectens irradias, each of which was bred in the 100 L plastic water troughs filled with 80 L sea water and gasified continuously with sufficient air pump. The illumination was about 200 lux, constant temperature 20 °C, salinity 30.1, sea water used was “no organism sea water” after dark deposition for 2 months.
No. 1, 2 and 3 groups were fed with poisonous algae of Alexandrium tamarense. The feeding amount was 1 L every day and average density was 1.26 × 104 cells/mL. The fourth group was fed with nontoxic algae of Tetraselmis (average density was 1.27 × 106 cells/mL), with the amount of 1 L every day from July 21 to July 27. The volume amount of Tetraselmis was corresponding to the volume of Alexandrium tamarense from July 28 to August 3. The length was 48 μm, and the width was 52 μm for Alexandrium tamarense cell (according to sphericity to calculate the volume), and the length and width were 24 μm and 15μm, for Tetraselmis cell (according to cylinder to calculate the volume) respectively. The ratio of these two algae volume was 15:1. Sampling and observation the algae cells was to investigate whether they were taken in food completely after breeding 24 hours. Change the water one time every 24 hours. Every 7 days measure the shell height completely one time, and determine the weight, simultaneously. Take 25 A. irradias to freeze at -20 °C into the plastic bag for determining the shell poison. The experiment altogether carried on for 28 days. The 1st, 2nd and 3rd groups were feed with A. tamarense from July 21 to August 3, but from August 4 to August 18, 4 groups were feed Tetraselmis (average density 1.24 × 106 cells/mL).
2.4 Preparation of Shellfish
The shellfish samples were preserved into -20 °C refrigerator, and opened the shell, took out the soft tissue and the body fluid, cut the liver pancreas carefully with the scissors, weighted, and moved it into the test tube, using the organization refiner mix round. Then add the same weight of HCl or NaOH to adjust the pH to 3-4. The test tube was placed in the water bath to boil 5 min, then cooled to the room temperature and adjusted pH to 2-4 again. Subsequently, the test tube was metered volume according to the rule of 2 mL solution being obtained from 1 g tissue.
The shell poisonous extraction fluid was got by centrifugation at 3,000 r/min for 5 min, and then put the fluid into the refrigerator. 2.5 Calculation of Shell Poison
According to the mouse bioassay developed by the AOAC:ρ = V × D × A × B
ρ: the content of shell poison;
V: the volume of shell poison samples;
A: mouse units (MU) that corresponds to the average dead time of small white mouse;
B: the correction factor of the average weight of small white mouse in experimental group which corresponds to mouse unit in the table;
D: dilution multiple of extraction.
3. Results
3.1 Single Pure Cultivation of Alexandrium tamarense
From March 1, 2003 to August 20, 2003, we cultured Alexandrium tamarense from 30 mL algae fluid to 50 L algae fluid of the average density 1.26 ×104 cells/mL.
3.2 Poison Content of Alexandrium tamarense
Table 1 shows the assay results of the algae cell toxin. The content of the algae liquid toxin was 15.04(MU) after adding to 10 mL, and the algae cell toxic was 2.18 × 10-6 MU/cell. Similar result was showed in Zheng’s study [2] which was 2.21 × 10-6-4.11 × 10-6 MU/cell. So A. tamarense belongs to low poison algae.
3.3 The Effects of Alexandrium tamarense on Argopectens irradias’s Ingestion
By dissection observation, Alexandrium tamarense stayed in Argopectens irradias’s stomach. Cell ruptured easily. And the excrement also became brown. Complete cell of Alexandrium tamarense was not picked out in the excrement, but the obvious Tetraselmis cell was found in the control group’s excrement. Table 2 shows the chlorophyll-a before and after one hour feeding in July 30, 2003. From chlorophyll-a, it knew the algae cells almost all were feed by filtering after one hour feeding, and this result was proved by the result of electron microscope.
4. Discussion
4.1 Average Poison Content in Shellfish
The average algae cells and shellfish toxin contents in Argopectens irradias were examined after feeding Alexandrium tamarense 7 days and 14 days (Fig. 1). So, it is known that 1.4 × 105 cells (Table 3) of Alexandrium tamarense were ingested by Argopectens irradias every day. Feed was continuous for 7ds, the poison content was 19.9 MU/g in visceral, corresponding to 3.7 MU/g in muscle, which were similar to the sanitation standard (4.0 MU/g in muscle). From the eighth day, Argopectens irradias was fed with 3.1 × 105 Alexandrium tamarense cells(Table 3) every day continuously. At the 14th day, the average toxicity level in visceral of the scallops was 49.4 MU/g, corresponding to 10.0 MU/g in muscle. It was 2.5 times higher than the sanitation standard.
4.2 Transformation and Elimination of Shellfish Poisoning
Tetraselmis was fed to Argopectens irradias after continuously feeding for 14 days. Average algae density was 1.72 × 104 cells/mL during the first seven days, while 1.56 × 104 cells/mL from the eighth day to the fourteenth day. The shellfish poison was showed in Fig. 1 at the seventh and fourteenth day.
The poisoning content was 21.3 MU/g in viscera, corresponding to 3.5 MU/g in muscle. And the average poisoning content was 7.9 MU/g in visceral, corresponding to 1.6 MU/g in muscle.
4.3 Time of Accumulation and Elimination of Shellfish Poisoning
The algae toxin was transformed through food chain and accumulated in shellfish in vivo, called poisoning. Shellfish poisoning accumulated to 10.0 MU/g in muscle 14 days at the experimental condition. Under the food satisfied condition, after the shell poisoning transformation and elimination for 14 days, the shellfish poisoning reduced to the 7.9 MU/g in visceral. It equaled to 1.6 MU/g in muscle, whch achieved the edible standard. The eliminating rate of shellfish poisoning was 12% daily.
4.4 Transformation between Algae Toxin and Shellfish Toxin
In the experiment the algae toxin and the shell toxin were not same amount. It was possible that the toxin ingredient had changed and gotten more strongly after shellfish’s metabolism.
Traditionally, the algae toxin did not affect on the shellfish. The shellfish only functioned as a middle medium. The fish was more sensitive to the paralysis algae toxin than the shellfish. The fish was caused to death often at low algae toxin concentration. Therefore it was not possible for fish to take the transmission medium. But after 1980s, studies showed that the toxin algae could affect on the shellfish according to the shell species [3]. The accumulation and distribution of the toxin was related to shellfish’s species, and bio-transformation happened in vivo. This study showed that the shellfish toxin was very different from the algae toxin in component after the shell grazed Prorocentrum sp.
This transformation has affected on the shellfish
toxicity. When low N-sulfuryl ammonia formyl toxoid transformed into high carbamyl toxoid, the shellfish body toxicity would increase. It showed that it must be an enzyme to catalyze bio-transformation in shell body [4].
After common mussel ingested Alexandrium tamarense algae, the relative scale of various toxin components in shellfish in vivo and the algae cells would change strongly [5].
4.5 The Unbalance between Algae Toxin and Shellfish Toxin
Centrifugal concentration method was used to assay algae toxin. This result was the same as the data obtained half a year ago, and corresponding to the results of low value in Zheng [2]. The long-time cultured virulent algae in laboratory would possibly reduce the toxicity [6].
Total algae toxin amount absorbed by average per shell was 2.18 × 10-6 MU/cell × 2.18 × 106 cells = 4.8 MU. However, shellfish toxin accumulated was up to 21.7 MU. The reasons why the toxins were not the same may be that: (1) error of assay method; (2) toxin from dead corrupt algae cells in the algae culture medium; (3) toxin excluded by lived algae cells. These would induce a lot of algae toxins in the algae liquid. It needs further study in the future.
5. Conclusions
5.1 Transformation and Elimination Time of Shellfish Poisoning
Japanese researcher [7] put Patinopecten yessoensis and Ostrea gigas, whose shellfish poisoning amount was 0.02 MU/g, into the red tide zone where the phytoplankton flora contained the toxin algae Dinophysis acuminate. The DSP (diarrheic shellfish poisoning) content reached 0.08 MU/g in these two kinds of shellfish after a week. The toxin algae cells reduced too suddenly to examine nearly after passing through 2 weeks, the DSP content reduced to 0.02 MU/g, which was same as the starting value when the shellfish was just put in the sea area, and was lower than the standard of DSP edible (0.05 MU/g).
In the experiment, when the amount of shellfish poisoning was to 49.4 MU/g in visceral, it was same with that of 10 MU/g in muscle. After two weeks experiment for transformation and elimination, the shellfish poisoning amount reduced to 7.9 MU/g in viscera, corresponding to 1.6 MU/g in muscle. This time of the shellfish poisoning transformation and elimination gotten from the experiment was the same as the one from the natural sea, which told the reason why the shellfish should be collected past though 2 weeks after the harmful algal blooms disappeared in Japan. At the same time, it would provide proof for reducing and preventing the harmful algal blooms in China.
5.2 Elimination Rate of Shellfish Poisoning
The initial shellfish toxin amount of A. irradians was 10.0 MU/g, and the daily elimination rate was 12%. The initial shellfish toxin amount of Mytilus Edulis was 0.61 MU/g, and the daily elimination rate was 9% [5]. The elimination rate was related to species, types and quantities of food. In addition, it was likely that when shellfish was at a high level of toxicity, the elimination rate would still remain at a relatively high level.
Acknowledgments
This study was financed by a grant from the Youth Marine Science Foundation of State Oceanic Administration of China under contract No. 99612 and the National Investigation Research (“908”) Program of China under contract No. 908-02-03-01.
References
[1] R.C. Wang, K.K. Yu, S.C. Yao, G.R. Miu, Z.P. Wang, Handbook of Marine Aquaculture Technology, Science and Technology Press, Shanghai, 2001, pp. 533-534.
[2] S.Z. Zheng, X. Lin, H.Z. Lin, H.S Chen. Studies on paralytic shellfish poisoning of Alexandrium tamarense from Hong Kong waters, Oceano Limnol Sinica 29 (1998) 477-481. (in Chinese)
[3] L.F. Gauney, S.E. Shumway, A compendium of responses of bivalve mollusca to toxic dinoflgellates, J. Shellf 7 (1988) 623-629.
[4] Y. Oshima, Chemical and enzymatic transformation of paralytic shellfish toxins in marine organisms, in: Harmful Marine Algal Blooms; Technique et Documentation—Lavoisier, Intercept Ltd, Paris, 1995, pp. 475-481.
[5] T. Yan, M. Fu, J. Li, R.C. Yu, M.J. Zhou, Accumulation, transformation and elimination of PSP in mytilus edulis, Oceano Limnol Sinica 32 (2001) 420-427.(in Chinese)
[6] T.J. Jiang, W.J. Huang, Z.H. Wang, Y.M. Luo, Y.W. Yin, S. Qi, Effects of water, temperature, salinity and pH on growth and toxicity of Alexandrium tamarense (Lebour) balech (Dapeng strain), Chin. J. Appl. Environ. 6 (2000) 151-154. (in Chinese)
[7] S. Taguchi, G. Hoshiai, A. Ito, Shellfish poisoning in the northeastern cost of Japan, Bulletin on Coas Oceano 32(1994) 55-67.
2. Dalian Ocean University, Dalian 116023, China
Received: December 19, 2011 / Accepted: January 9, 2012 / Published: February 20, 2012.
Abstract: Using a PSP (paralytic shellfish poisoning) toxin-producing strain of Alexandrium tamarense, it studied the timing of toxin accumulation and elimination of PSP toxins in Argopectens irradias. The PSP toxicity was studied by following the standard PSP mouse bioassay developed by the Association of Official Analytical Chemists (AOAC). Alexandrium tamarense was cultured to an average density of 1.26 × 104/mL for a total of about 50 L culture. The toxicity of the alga was 2.18 × 10-6 MU/cell. The results show that PSP content increased with time in both visceral and muscle tissue during a two-week accumulation period during which scallops were fed with A. tamarense. The average toxin level in scallop’s viscera was 49.4 MU/g, with an average of 10.0 MU/g in muscle tissue. This level is 2.5 times higher than the sanitation standard (4.0 MU/g of muscles). The highest value was 61.0 MU/g in the viscera. In summary, the viscera accumulated greater concentrations of toxin than muscle tissue. Scallops that had accumulated toxins were transplanted for two weeks into a field environment containing no toxic algae. The PSP content of the scallops decreased to 7.9 MU/g viscera and 1.6 MU/g muscles two weeks after being transplanted, but did not reach the sanitation standard. Under the experimental conditions, the toxin depuration rate of shellfish toxin was 12% daily. This study worked toward the development of a sanitary shellfish industry and better management of PSP toxin-impacted shellfish in China.
Key words: PSP, argopectens irradias, accumulation and elimination time.
1. Introduction
Countries around the world nowadays take to prevent shell poisonous toxins, reduce shell poisonous loss research, attend shellfish’s healthy aquaculture, establish shellfish purifying center, as well as accelerate shell toxins purification research and so on. Our country is a great economical nation of marine products, and the shellfish culture is an important cornerstone. It does not allow neglect the loss, for the harmful and virulent red tide. Therefore, it is extremely essential to study the algae toxin with the time of in vivo accumulation and elimination in the sea shellfish, and is important to control the sanitary quality of shellfish.
2. Materials and Methods
2.1 Methods of Algae Culture
The experimental algae were Qingdao strain of Alexandrium tamarense. It was cultured from March 2003. The algae were cultured in 30 mL, 15-18 °C. The illumination cycle was the natural photoperiod, and the intensity of illumination was 3,000-5,000 lux. Cell density was counted with blood counting chamber every 5-7 days. When the density was about 1 × 104 cells/mL, it can add the boiling disinfection sea water to culture expansively, with the proportion 1:1 of sea water to the algae. The salinity was 30.1. It should be shook 4 times every day and shook 2 minutes every time. 5 L glass Erlenmeyer flask was used for mass cultured, putted on wooden multi-layered shelf. The culture medium was filtered seawater with f/2 medium which is a nutrient salt
formula [1].
2.2 Determination of Algae Toxin
The densities of Alexandrium tamarense cell was counted on August 1, 2003. The density was 1.38 ×104 cells/mL. The sample of 500 mL was centrifuged at 2000 r/min for 10 min, and 5 mL algae cell was obtained at the bottom of the tube. The algae toxicity was assayed following the standard method of PSP mouse bioassay developed by the Association of Official Analytical Chemists (AOAC).
The densities of Alexandrium tamarens cell and Tetraselmis cell was counted with blood counting chamber.
2.3 Experimental Methods
Started from July 21, 2003, the experiment was divided into four groups of 100 Argopectens irradias, each of which was bred in the 100 L plastic water troughs filled with 80 L sea water and gasified continuously with sufficient air pump. The illumination was about 200 lux, constant temperature 20 °C, salinity 30.1, sea water used was “no organism sea water” after dark deposition for 2 months.
No. 1, 2 and 3 groups were fed with poisonous algae of Alexandrium tamarense. The feeding amount was 1 L every day and average density was 1.26 × 104 cells/mL. The fourth group was fed with nontoxic algae of Tetraselmis (average density was 1.27 × 106 cells/mL), with the amount of 1 L every day from July 21 to July 27. The volume amount of Tetraselmis was corresponding to the volume of Alexandrium tamarense from July 28 to August 3. The length was 48 μm, and the width was 52 μm for Alexandrium tamarense cell (according to sphericity to calculate the volume), and the length and width were 24 μm and 15μm, for Tetraselmis cell (according to cylinder to calculate the volume) respectively. The ratio of these two algae volume was 15:1. Sampling and observation the algae cells was to investigate whether they were taken in food completely after breeding 24 hours. Change the water one time every 24 hours. Every 7 days measure the shell height completely one time, and determine the weight, simultaneously. Take 25 A. irradias to freeze at -20 °C into the plastic bag for determining the shell poison. The experiment altogether carried on for 28 days. The 1st, 2nd and 3rd groups were feed with A. tamarense from July 21 to August 3, but from August 4 to August 18, 4 groups were feed Tetraselmis (average density 1.24 × 106 cells/mL).
2.4 Preparation of Shellfish
The shellfish samples were preserved into -20 °C refrigerator, and opened the shell, took out the soft tissue and the body fluid, cut the liver pancreas carefully with the scissors, weighted, and moved it into the test tube, using the organization refiner mix round. Then add the same weight of HCl or NaOH to adjust the pH to 3-4. The test tube was placed in the water bath to boil 5 min, then cooled to the room temperature and adjusted pH to 2-4 again. Subsequently, the test tube was metered volume according to the rule of 2 mL solution being obtained from 1 g tissue.
The shell poisonous extraction fluid was got by centrifugation at 3,000 r/min for 5 min, and then put the fluid into the refrigerator. 2.5 Calculation of Shell Poison
According to the mouse bioassay developed by the AOAC:ρ = V × D × A × B
ρ: the content of shell poison;
V: the volume of shell poison samples;
A: mouse units (MU) that corresponds to the average dead time of small white mouse;
B: the correction factor of the average weight of small white mouse in experimental group which corresponds to mouse unit in the table;
D: dilution multiple of extraction.
3. Results
3.1 Single Pure Cultivation of Alexandrium tamarense
From March 1, 2003 to August 20, 2003, we cultured Alexandrium tamarense from 30 mL algae fluid to 50 L algae fluid of the average density 1.26 ×104 cells/mL.
3.2 Poison Content of Alexandrium tamarense
Table 1 shows the assay results of the algae cell toxin. The content of the algae liquid toxin was 15.04(MU) after adding to 10 mL, and the algae cell toxic was 2.18 × 10-6 MU/cell. Similar result was showed in Zheng’s study [2] which was 2.21 × 10-6-4.11 × 10-6 MU/cell. So A. tamarense belongs to low poison algae.
3.3 The Effects of Alexandrium tamarense on Argopectens irradias’s Ingestion
By dissection observation, Alexandrium tamarense stayed in Argopectens irradias’s stomach. Cell ruptured easily. And the excrement also became brown. Complete cell of Alexandrium tamarense was not picked out in the excrement, but the obvious Tetraselmis cell was found in the control group’s excrement. Table 2 shows the chlorophyll-a before and after one hour feeding in July 30, 2003. From chlorophyll-a, it knew the algae cells almost all were feed by filtering after one hour feeding, and this result was proved by the result of electron microscope.
4. Discussion
4.1 Average Poison Content in Shellfish
The average algae cells and shellfish toxin contents in Argopectens irradias were examined after feeding Alexandrium tamarense 7 days and 14 days (Fig. 1). So, it is known that 1.4 × 105 cells (Table 3) of Alexandrium tamarense were ingested by Argopectens irradias every day. Feed was continuous for 7ds, the poison content was 19.9 MU/g in visceral, corresponding to 3.7 MU/g in muscle, which were similar to the sanitation standard (4.0 MU/g in muscle). From the eighth day, Argopectens irradias was fed with 3.1 × 105 Alexandrium tamarense cells(Table 3) every day continuously. At the 14th day, the average toxicity level in visceral of the scallops was 49.4 MU/g, corresponding to 10.0 MU/g in muscle. It was 2.5 times higher than the sanitation standard.
4.2 Transformation and Elimination of Shellfish Poisoning
Tetraselmis was fed to Argopectens irradias after continuously feeding for 14 days. Average algae density was 1.72 × 104 cells/mL during the first seven days, while 1.56 × 104 cells/mL from the eighth day to the fourteenth day. The shellfish poison was showed in Fig. 1 at the seventh and fourteenth day.
The poisoning content was 21.3 MU/g in viscera, corresponding to 3.5 MU/g in muscle. And the average poisoning content was 7.9 MU/g in visceral, corresponding to 1.6 MU/g in muscle.
4.3 Time of Accumulation and Elimination of Shellfish Poisoning
The algae toxin was transformed through food chain and accumulated in shellfish in vivo, called poisoning. Shellfish poisoning accumulated to 10.0 MU/g in muscle 14 days at the experimental condition. Under the food satisfied condition, after the shell poisoning transformation and elimination for 14 days, the shellfish poisoning reduced to the 7.9 MU/g in visceral. It equaled to 1.6 MU/g in muscle, whch achieved the edible standard. The eliminating rate of shellfish poisoning was 12% daily.
4.4 Transformation between Algae Toxin and Shellfish Toxin
In the experiment the algae toxin and the shell toxin were not same amount. It was possible that the toxin ingredient had changed and gotten more strongly after shellfish’s metabolism.
Traditionally, the algae toxin did not affect on the shellfish. The shellfish only functioned as a middle medium. The fish was more sensitive to the paralysis algae toxin than the shellfish. The fish was caused to death often at low algae toxin concentration. Therefore it was not possible for fish to take the transmission medium. But after 1980s, studies showed that the toxin algae could affect on the shellfish according to the shell species [3]. The accumulation and distribution of the toxin was related to shellfish’s species, and bio-transformation happened in vivo. This study showed that the shellfish toxin was very different from the algae toxin in component after the shell grazed Prorocentrum sp.
This transformation has affected on the shellfish
toxicity. When low N-sulfuryl ammonia formyl toxoid transformed into high carbamyl toxoid, the shellfish body toxicity would increase. It showed that it must be an enzyme to catalyze bio-transformation in shell body [4].
After common mussel ingested Alexandrium tamarense algae, the relative scale of various toxin components in shellfish in vivo and the algae cells would change strongly [5].
4.5 The Unbalance between Algae Toxin and Shellfish Toxin
Centrifugal concentration method was used to assay algae toxin. This result was the same as the data obtained half a year ago, and corresponding to the results of low value in Zheng [2]. The long-time cultured virulent algae in laboratory would possibly reduce the toxicity [6].
Total algae toxin amount absorbed by average per shell was 2.18 × 10-6 MU/cell × 2.18 × 106 cells = 4.8 MU. However, shellfish toxin accumulated was up to 21.7 MU. The reasons why the toxins were not the same may be that: (1) error of assay method; (2) toxin from dead corrupt algae cells in the algae culture medium; (3) toxin excluded by lived algae cells. These would induce a lot of algae toxins in the algae liquid. It needs further study in the future.
5. Conclusions
5.1 Transformation and Elimination Time of Shellfish Poisoning
Japanese researcher [7] put Patinopecten yessoensis and Ostrea gigas, whose shellfish poisoning amount was 0.02 MU/g, into the red tide zone where the phytoplankton flora contained the toxin algae Dinophysis acuminate. The DSP (diarrheic shellfish poisoning) content reached 0.08 MU/g in these two kinds of shellfish after a week. The toxin algae cells reduced too suddenly to examine nearly after passing through 2 weeks, the DSP content reduced to 0.02 MU/g, which was same as the starting value when the shellfish was just put in the sea area, and was lower than the standard of DSP edible (0.05 MU/g).
In the experiment, when the amount of shellfish poisoning was to 49.4 MU/g in visceral, it was same with that of 10 MU/g in muscle. After two weeks experiment for transformation and elimination, the shellfish poisoning amount reduced to 7.9 MU/g in viscera, corresponding to 1.6 MU/g in muscle. This time of the shellfish poisoning transformation and elimination gotten from the experiment was the same as the one from the natural sea, which told the reason why the shellfish should be collected past though 2 weeks after the harmful algal blooms disappeared in Japan. At the same time, it would provide proof for reducing and preventing the harmful algal blooms in China.
5.2 Elimination Rate of Shellfish Poisoning
The initial shellfish toxin amount of A. irradians was 10.0 MU/g, and the daily elimination rate was 12%. The initial shellfish toxin amount of Mytilus Edulis was 0.61 MU/g, and the daily elimination rate was 9% [5]. The elimination rate was related to species, types and quantities of food. In addition, it was likely that when shellfish was at a high level of toxicity, the elimination rate would still remain at a relatively high level.
Acknowledgments
This study was financed by a grant from the Youth Marine Science Foundation of State Oceanic Administration of China under contract No. 99612 and the National Investigation Research (“908”) Program of China under contract No. 908-02-03-01.
References
[1] R.C. Wang, K.K. Yu, S.C. Yao, G.R. Miu, Z.P. Wang, Handbook of Marine Aquaculture Technology, Science and Technology Press, Shanghai, 2001, pp. 533-534.
[2] S.Z. Zheng, X. Lin, H.Z. Lin, H.S Chen. Studies on paralytic shellfish poisoning of Alexandrium tamarense from Hong Kong waters, Oceano Limnol Sinica 29 (1998) 477-481. (in Chinese)
[3] L.F. Gauney, S.E. Shumway, A compendium of responses of bivalve mollusca to toxic dinoflgellates, J. Shellf 7 (1988) 623-629.
[4] Y. Oshima, Chemical and enzymatic transformation of paralytic shellfish toxins in marine organisms, in: Harmful Marine Algal Blooms; Technique et Documentation—Lavoisier, Intercept Ltd, Paris, 1995, pp. 475-481.
[5] T. Yan, M. Fu, J. Li, R.C. Yu, M.J. Zhou, Accumulation, transformation and elimination of PSP in mytilus edulis, Oceano Limnol Sinica 32 (2001) 420-427.(in Chinese)
[6] T.J. Jiang, W.J. Huang, Z.H. Wang, Y.M. Luo, Y.W. Yin, S. Qi, Effects of water, temperature, salinity and pH on growth and toxicity of Alexandrium tamarense (Lebour) balech (Dapeng strain), Chin. J. Appl. Environ. 6 (2000) 151-154. (in Chinese)
[7] S. Taguchi, G. Hoshiai, A. Ito, Shellfish poisoning in the northeastern cost of Japan, Bulletin on Coas Oceano 32(1994) 55-67.