International Summer Water Resources Research School · 2012. 11. 13. · Therese Areskoug...

2012

International Summer Water Resources Research School Dept. of Water Resources Engineering, Lund University

Clearance rates for different phytoplankton grazed

by the jellyfish Aurelia aurita in polyp and ephyrae stage

By

Therese Areskoug

Source: www.jellyfishfacts.net

Therese Areskoug 200120712

Clearance rates for different phytoplankton grazed by the jellyfish Aurelia aurita in polyp and ephyrae

stage

Source: www.jellyfishfacts.net

Acknowledgements: This project is part of the Lingfeng Summer Research School 2012 at Xiamen University, China. Apart from thanking the organizers for a succesful project, I would like to extend a sincere gratitude towards the people who have helped me complete this project; professor Huang Banquin, assistant Huang Xuguang and last but not least Ge Muyang who have all helped me in various ways to complete this project. I would also very much like to thank the sponsors Xylem and Tyréns for making the trip to Xiamen, China financially possible.


AbstractJellyfish blooms are becoming an increasing liability for swimmers, fishermen and powerplant intakes all over the world. In order to be able to come to terms with these problems, the ecological impact of these creatures must be determined. This study is focused on the cnidarian scyphozoan Aurelia aurita, and its feeding habits in the ephyrae stage. The ephyrae were measured and fed for seven days. Three different types of food were used; the dinoflagellate Alexandrium tamarense, the zooplankton Artemia franciscana, a combination of both and in addition a control only containing seawater. We have found that on a diet of A. tamarense, the ephyrae will decrease in size over time. And even when fed with the combination of both zooplankton and dinoflagellate, the ephyrae will not grow even though A. franciscana is a suitable food source to grow on. The most probable reason is thought to be that as the dinoflagellates move, the ephyrae will use energy to capture it. However, it cannot digest the dinoflagellate and therefore cannot make use of the food to grow.

By counting how many contractions the ephyrae make per minute in the presence of different high concentrations of A. tamarense, their ability to swim has been evaluated. The samples were first incubated for 4 hours to evaluate inhibition, and then for 1 hour to evaluate recovery. The results show that an increasing amount of algae cause a stronger swimming inhibition. However, if the algae are removed, they can recover to some extent. For further confirmation of this, two sample plates were prepared, one with algae, and one with algae filtered away. Without the physical presence of A. tamarense, the swimming was not inhibited at all. This indicates that it is the physical prescence of the dinoflagellates that cause the inhibition rather than the algae releasing toxic compounds.

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Table of contents

Abstract .................................................................................................................................................11. Introduction.......................................................................................................................................3

1.1 Aim........................................................................................................................................32. Background........................................................................................................................................3

2.1 Feeding habits of A. aurita....................................................................................................32.2 Project restritions...................................................................................................................4

3. Method............................................................................................................................................... 43.1 Materials................................................................................................................................43.2 Calculating the correct dilution.............................................................................................43.3 Toxicity experiment...............................................................................................................43.4 Algae/no algae experiment....................................................................................................53.5 Recovery experiment.............................................................................................................53.6 Feeding experiment...............................................................................................................6

4. Results and Discussion......................................................................................................................64.1 Toxicity experiment...............................................................................................................64.2 Algae/no algae experiment....................................................................................................74.3 Recovery experiment.............................................................................................................84.4 Feeding experiment.............................................................................................................10

5. Conclusions.......................................................................................................................................116. References.........................................................................................................................................12

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1. IntroductionAurelia aurita or moon jellyfish (English common name) are present in many coastal regions around the world. They can tolerate a wide range of environmental conditions which allows them to thrive from 70 o N to 40 o S and their abundance is generally higher in shallow enclosed waters (Cathy H. Lucas, 2001). There have been five published accounts for blooms of Aurelia aurita in China since 2004. In 2008, over 4000 tons of jellyfish were cleaned up in Qinhuangdao which gives some perspective as to the scale of the problem (Dong, Liu, Keesing, 2010). Some probable causes for these jellyfish blooms are eutrophication, overfishing, restricted water exchange (Cathy H. Lucas, 2001) and manmade coastal structures (Lo et al. 2007) such as aquaculture rafts and harbours which provide excellent settling places for the polyps.

Jellyfish blooms have increased worldwide in the last 10-15 years and it is becoming a liability to swimmers, fish populations, fishing industry and power plant water intakes. In 2003 a species of jellyfish called Nemopilema nomurai had a large bloom in the East China Sea, which resulted in a certain species of fish declining by 20% during the bloom (Dong, Liu, Keesing, 2010).

Even though it is the medusae that cause the problems, it is the previous stages that determine the population size (Kamiyama, 2011). If the polyps thrive in this stage, they can produce numerous ephyrae and in the end cause massive blooms. In order to do that, a number of environmental factors influence strongly, whereof one is access to suitable food.

1.1 AimThe initial aim of this project is to examine whether or not the dinoflagellate Alexandrium tamarense could be used as a food source for the ephyrae stage of A. Aurita. It will be measured as the ability of the ephyrae to grow when fed with A. tamarense. More specifically it is interesting to see if there is any difference if the ephyrae are fed with a toxic or a non-toxic strain.

The project will also try to find out if the ability of the ephyrae to swim is inhibited by high concetrations of algae, or algae toxin. If so, it will also be investigated if the ephyrae can recover from any possible inhibition by some means.

2. Background

2.1 Feeding habits of A. auritaA. aurita as well as most other jellyfish has a life cycle with two main stages, one benthic and one pelagial. As a polyp, the jellyfish is attached to a surface reproducing asexually. Ephyrae are then formed as a middle stage or a larvae stage between the polyp and the fully grown medusa. A. aurita medusae feed on zooplankton and some fish larvae, basically the same diet as predator fish, meaning that if jellyfish blooms are becoming more common, it can add to overfishing, causing fish populations to crash (Cathy H. Lucas, 2001).

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Adult medusae feed on zooplankton and fish larvae but planktonic algae have been found in the gastric pouch. However, there was no indication that the diatoms found were important as a food source (Båmstedt, 1990). In 2001 Båmstedt et al. showed that phytoplankton may be important as a food source in the very beginning of the ephyrae stage, especially if coincided with a phytoplankton bloom (Båmstedt, Wild, Martinussen, 2001). A study has also been made on whether the polyps of A. aurita feed on ciliates or not with a control containing phytoplankton. In this study, it was actually found that the polyps were actively feeding on both nano- and mictorphytoplankton. However, compared to the ciliates of similar sizes the feeding rates were significantly lower (Kamiyama, 2011). The current research indicate that both ephyrae and polyp stages of A. aurita can feed on phytoplankton to some extent, but it does not seem to be very important as a food source.

2.2 Project restrictionsThe original aim was to find out the clearance rates for different phytoplankton in A. aurita ephyrae and polyp stage. Because of the time limitation, we have focused on ephyrae feeding patterns in connection to the dinoflagellate A. tamarense. Because the connection between algae and ephyrae has been little studied, we ran in to some problems during the first days in the lab that required us to change direction a little bit. We discovered that we could not calculate the clearande rate because the ephyrae died or were strongly inhibited very soon after the initial exposure to the dinoflagellate. Also, we had trouble counting the algal cells with enough accuracy to be able to tell if the ephyrae had digested any or not. These two factors led to a small shift in our project aim.

3. Method

3.1 MaterialAll experiements were performed on a 12 well plate, where each well has a volume of 6 ml with a lid for protection during incubation. Filtered seawater, a beaker containing ephyrae, algal solutions of both toxic and non-toxic Alexandrium tamarense and a beaker containing Artemia franciscana were also included in the materials used. The lab equipment consisted of microscope, pipettes, counting chamber, miroscope slide, cover glass, potassium iodine and a filtration syringe.

3.2 Calculating the correct dilutionFor all the experiments, the initial concentration of the algal solution is required. Therefore, a sample of 200 μl is taken from the well mixed beaker containing A. tamarense solution and put in a small eppendorf tube. It is mixed with 20 μl potassium iodine in order to kill the algae and stop them from moving around during counting. 100 μl of this solution was then taken out and put on a microscope slide with a counting chamber and covered with a cover glass. The algae were then counted using a microscope with 5x magnification and then the algae concentration is known for calculating the right dilution in the experiments.

3.3 Toxicity experimentTwo 12-well plates were prepared with the same dilution series of algae but one was prepared with a toxic strain and one with a non- toxic strain of A. tamarense. Row one

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was used as a control so it contained no algae. The second row had a concentration of 125 cells/ml, the third had 250 cells/ml and the fourth row contained a concentration of 750 cells/ml. Then, one ephyrae was inserted into each well. The samples were incubated at 20°C in natural light for 16 hours and then visually examined.

3.4 Algae/no algae experimentIn order to see if it is the algae itself or if they release some toxin into the water, two 12 well plates were prepared with the same dilution series. The first row of the first plate was a control and only contained filtered seawater. The second row had a concentration of 125 per ml, the third 250 per ml and the fourth 750 per ml. On the second plate however, the algae samples were filtered with a filtration syringe, before they were inserted into the wells. The same amount of algal solution was used in each row as in plate one.

Once the dilution series were done, the individual amount of contractions made by each ephyrae during 1 minute were counted. Then both plates were incubated for 4.5 hours in 20°C under natural light. After incubation, another round of counting was performed.

3.5 Recovery experimentA dilution series was made with duplicates of three on three different plates. Each column of the plate was given a different concentration of A. tamarense. The different concentrations in the columns can be seen in Table 1.

Table 1: The different algae concentrations prepared for each plate

Conc. col 1 (cells/ml) Conc. col 2 (cells/ml) Conc. col 3 (cells/ml) Conc. col 4 (cells/ml)

200 400 800 1200

After the wells had been prepared with the right mixture of seawater and algae solution, ephyrae were randomly selected from the ephyrae beaker and one was inserted into each well. Then the amount of contractions made by each ephyrae were counted during one minute in order to observe any difference connected to the different concentrations of algae. The ephyrae were not removed from its well at any time in order to disturb it as little as possible. Only contractions made with all arms were counted. After counting, all plates were incubated in in natural light and 20°C for 4 hours. After this incubation period, the amount of contractions made by each ephyrae were counted during one minute.

To see if the ephyrae were able to recover from any inhibition that might be occurring,In order to cover any differences along the dilution gradient, each of the three rows of wells were treated with different methods. On the first row, half of the original amount of water was taken out after mixing carefully and was replaced by 2.5 ml seawater containing a few A. franciscana. Both row two and row three were completely emptied on water. In row 2 it was replaced by 5 ml of the same A. franciscana solution as in row 1. Row 3 was replaced with 5 ml pure filtered seawater. The samples were then incubated for 1 hour, and the amount of contractions were counted once again during 1 minute each.

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3.6 Feeding experimentIn order to see if ephyrae of A. aurita can grow on a diet of A. tamarense, a new 12- well plate was prepared. The first column was filled with 5 ml filtered seawater in each well, the second with an algae concentration of 250 cells/ml, the third with two A. franciscana and the fourth with the same algae concentrations as in colum 2 and two A. franciscana. Then 12 ephyrae were randomly selected and distributed among the wells.

Between the first and the second measurement, two days of no measuring occured. Then the ephyrae were measured each day for five days in succession. Between Day 6 and Day 7 there was also a day with no measuring. A total of seven measurements were made during a period of ten days. The ephyrae were measured using a microscope with 5x magnification and a computor. A picture was taken of each of the ephyrae with the program Q-capture and the radius of the central disc was measured using a program called Simple PCI. The arms of the ephyrae were not included because it was too difficult to measure them while the ephyrae were alive and moving. The diameter of the central disk was measured if possible, but if the ephyrae was too big, only the radius was measured. An accuracy of 0,1 mm was used. The ephyrae were never removed from their wells in order to disturb them as little as possible. After each measurement, the two rows containing A. franciscana were supplied with 2 new ones regardless of how many was left since the day before. The algae were not renewed.

4. Results and Discussion

4.1 Toxicity experimentThe plates were examined visually and they both showed the same pattern. The last row is completely curled up as can be seen in Figure 1, and are not moving at all.The third row has two curled up ephyrae and one feebly moving. This result together with some other early experiments where all ephyrae died at high concentrations of algae (no matter if they were toxic or not) indicates that it is not the toxin of the A. tamarense that cause the ephyrae to die. It rather seems like it is a physical process where the ephyrae is worn down by either having too much food to eat so that it gets tired, or the algae simply sticks to its surface and wears it down. At least, there seem to be extremely little or no difference in the effect on ephyrae between the toxic and the non toxic strain.

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4.2 Algae/no algae experimentThe results of this experiment clearly showed that there is of course some variability in the activity amongst the ephyrae. Still, it is very clear from Figure 1 that the activity of the samples with algae after incubation and at high concentration is clearly more inhibited than the other samples. Mean activity drops to almost 10 contractions/min, which is significantly lower than the sample without algae after incubation, which has a mean activity of over 70 contractions/min.

From this experiment, it is clear that it is the algae themselves that cause the inhibition and not some toxin released into the water. This only reinforces the results obtained from the Toxicity experiment.

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0 125 250 750

05

1015202530354045505560657075

Average activity with/without algae

Algae, before incubation

Algae, after incubation

No algae, before incubation

No algae, after incubation

Algae concentration (cells/ml)

Ave

rag

e a

ctiv

ity (

con

tract

ions/

min

)

Figure 2: Average activity at different concentrations of the two plates in the Algae/No algae experiment

Figure 1: Typical appearance of phyrae before (left) and after incubation (right) with high concentrations of A. tamarense


4.3 Recovery experimentThe results from the recovery experiment are quite uncertain because of the high standard deviation, which is caused by the high individual and time dependent variation in the activity of the ephyrae.

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Figure 4: Mean activity of the ephyrae at different dinoflagellate concentrations and recovery with zooplankton

Figure 3: Mean activity of the ephyrae at different dinoflagellate concentrations and recovery with 50% zooplankton and 50% algae


However, Figure 4 and 5 show the same pattern before incubation as was indicated by the results of the toxicity and the algae/no algae experiment. Both show a decrease in activity as a result of the increasing concentration. Figure 3 does not show the same connection, which could be due to the uncertainty of the graphs, but could also mean that there is no connection between activity and algae concentration. But since different experiments have all indicated the same thing, the most probable reason is that there is actually a connection although perhaps not always an evident one.

It can also be seen that almost without exception, the activity has decreased after incubation and some recovery seems possible. This is interesting as it may mean that both the algae concentration and the time of exposure influence activity. No difference in recovery can be distinguished between the different treatment methods.

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Figure 5: Mean activity of the ephyrae at different dinoflagellate concentrations and recovery with seawater


4.4 Feeding experiment

Figure 6: The mean radius of the central disk of the ephyrae fed with zooplankton, algae, a combination and the control respectively.

As shown in Figure 6, the ephyrae fed with the zooplankton A. franciscana grew a considerable amount compared to the other samples. The control and the combination of zooplankton and dinoflagellate did not change significantly in size, and the sample containing only A. tamarense seems to have decreased in size. The figure shows a clear result, but the reason behind the behaviour needs some discussion. it is not surprising that the ephyrae can grow well when fed with the zooplankton since it is a previously known food source for A. aurita ephyrae. However, it is very interesting that the ephyrae in the combination samples cannot grow even though they are presented with suitable food in the form of zooplankton when A. tamarense is also present.

Both the control and the zooplankton samples indicate that the experiment has a high validity since they both show the expected results. It is also very interesting that the ephyrae seem to decrease in size when fed only with the dinoflagellate. The most probable explaination is that as the dinoflagellates can move, they trigger the feeding mechanism much like a swimming zooplankton would do. However, if the ephyrae cannot digest A. tamarense, they will not be able to extract any engery from the hunt. This might be the explaination as to why the ephyrae are not able to grow, even when presented with suitable food in combination. The energy obtained from the few zooplankton they catch is not enough to make sustain growth in this environment. Though it can keep them from decreasing in size which seems to be the case when only A. tamarense is present.

To our knowledge, the only previous study made on A. aurita ephyrae and phytoplankton showed that the ephyrae could in fact grow on a phytoplankton diet, at least during the first 10 days of so after strobilation (Båmstedt et al. 2001). The ephyrae used for this experiment were alredy 10 days when the experiment started and had previously been fed A. franciscana. Båmstedt et. al made a very similar experiment, but instead of A. tamarense, they used the cryptophyte Rhodomnas baltica. Or the time limitation for using

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phytoplankton as food source is very specific, so our ephyrae were too old to be able to feed on it. It seems as though our results show that some algae can be used as a food source but not all, so further investigation is required.

5. Conclusion

The swimming ability of the ephyrae is inhibited at high concentrations of A. tamarense. There is no effect if the phytoplankton are filtered away or is present only at low concentrations. No difference can be distinguished between toxic and non toxic A. tamarense indicating it is not the toxin of the dinoflagellates but their physical presence. Ehyrae might be able to recover to some extent from theswimming inhibition if the dinoflagellates are removed.

Ephyrae can't grow when eating the dinoflagellate A. tamarense, in fact, the results indicate that they decrease in size. The dinoflagellate also inhibits growth even if in combination, the ephyrae are supplied with more suitable food such as A. franciscana.

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6. ReferencesBåmstedt U, 1990, ”Trophodynamics of the scyphomedusae Aurelia aurita. Predation rate in relation to abundance”. J Plankton Res.

Båmstedt U, Wild B, Martinussen M.B, 2001, ”Significance of food type for growth of ephyrae Aurelia aurita (Scyphozoa)”. Marine Biology.

Dong Z, Liu D, Keesing J K, 2010, ”Jellyfish bloomsin China: Dominant species, causes and consequences”. Marine Pollution Bulletin.

Kamiyama T, 2011, ”Planktonic ciliates as food source for the scyphozoan Aurelia aurita (s.l.): Feeding activity and assimilation of the polyp stage”. Journal of experimental Marine biology and Ecology.

Lo W T et. al, 2007, ”Enhancement of jellyfish (Aurelia aurita) populations by extensive aquaculture rafts in a coastal lagoon in Taiwan”. ICES Journal of Marine Science.

Lo W T, Chen I L, 2007, “Population succession and feeding of scyphomedusae, Aurelia aurita, in a eutrophic tropical lagoon in Taiwan”. Estuarine, Coastal and Shelf science.

Lucas C H, 2001, ”Reproduction and life history strategies of the common jellyfish,Aurelia aurita, in relation to its ambient environment”. Hydrobiologica.

Riisgård H U, Madsen C V, 2010, ”Clearance rates of ephyrae and small medusae of the common jellyfish Aurelia aurita offered different types of prey”. Journal of Sea Research.

Titelman J, Hansson L J, 2005, “Feeding rates of the jellyfish Aurelia aurita on fish larvae”. Marine Biology.

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International Summer Water Resources Research School · 2012. 11. 13. · Therese Areskoug...

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