1. WALLACE RESOURCE LIBRARY
Module 01 - Ecosystems: Coral Reefs
D04 – Interspecific variation in anemonefish (clownfish)

2. Coral distribution
There are a few key things to note from this map (see if the students can spot any of them – this is good practice for data analysis and interpretation):
That corals are restricted to within the tropics (i.e. close to the equator). This is due to the temperature requirements of reef building corals – they generally can’t survive in temperatures below 19°C.
The three centres of coral diversity are found in the Western areas of the three main oceans: (1) The Caribbean (Western Atlantic), (2) the Indo-Pacific (Western Pacific Ocean), (3) the Red Sea and East Africa (Western Indian Ocean). This is due to the prevailing currents in the tropical oceans being in a western direction.
There are large areas of the tropics with very little or no coral diversite (e.g. West Africa and South America). This is because these regions experience upwelling of cold nutrient rish waters, so you find totally different ecosystems there.
The Caribbean is much less diverse than other areas of the world (see the next slide).
There are three main centres of coral diversity, (1) the Caribbean, (2) the Indo-Pacific and (3) the Western Indian Ocean

3. Anemonefish - Clown fish
Live on their host anemone in a unique symbiotic relationship.
They help each other by providing protection from predators.
28 species and often more than one species will live on the same anemone.
Background to the research:
The lionfishes comprise about 22 species within the scorpion fish family. Like scorpion fish, lionfish are venomous and can deliver a potent sting using venomous spines. The red lionfish, Pterois volitans, is perhaps the best recognized group member. Prized by aquarium hobbyists for their beautiful coloration and hardy nature, the fish are the bane of biologists struggling to manage exotic introductions in the Mediterranean and western Atlantic. The Atlantic introduction is especially troubling as the fish have rapidly established persistent populations from North Carolina on the U.S. eastern seaboard, all the way into the Gulf of Mexico and into the Caribbean Sea. They are voracious predators with few natural enemies and early indications are that they will negatively impact the ecological balance of Florida and Caribbean reefs.
Temperature is an important environmental factor influencing red lionfish ecology and distribution, but at the moment we lack good information on the thermal ecology of lionfish from their native range. The immediate goal of the experiments described below is to model high temperature tolerance responses of lionfish acclimated (acclimatised) to different constant temperatures in the laboratory. In the long term, thermal tolerance data may provide a better understanding of red lionfish thermal biology, eventually leading to more effective management strategies to mitigate the fish’s adverse impacts on western Atlantic ecosystems.

4. Anemonefish -Clownfish
Fish of the same species communicate by unique calls or ‘popping’ sounds.
These sounds are used to establish hierarchies, warn off predators and stop other fish invading their territory.
This study looks at how the calls between species differ and how and why this is achieved.
Additional reading:
For more background on this experimental approach, and to see how these data can be used in larger ecological context, please read the following:
Beitinger, T. L. and W. A. Bennett Quantification of the Role of Acclimation Temperature in Temperature Tolerance of Fishes. Environmental Biology of Fishes, 58:
Dabruzzi, T. F., W. A. Bennett, Rummer, J. L., and Fangue, N. A. In Press. Juvenile Ribbontail Stingray, Taeniura lymma, demonstrate a unique suite of physiological adaptations to survive hyperthermic nursery conditions. Hydrobiologia.
Eme, J. and W. A. Bennett Critical Thermal Tolerance Polygons of Tropical Marine Fishes from Sulawesi, Indonesia. Journal of Thermal Biology. 34,
These documents have been included in this folder and are scientific papers that have been, or are scheduled to be published in peer-reviewed scientific journals.

5. Research questions and methodology
In this physiological investigation 20 lionfish are used to see how they adapt to a number of different sea-water temperatures.
Four different temperatures are investigated and these results are used to understand how they develop thermal tolerance.
Data collection methods:
Five replicate lionfish are selected and acclimated (acclimatised) by placing them individually into special biologically filtered 16-L plastic sea water tanks at 20.9 degrees C for minimum of 12 days. This is repeated for more lionfish at 26.7, 29.9 and 32.7 degrees C making a total of 20 of fish for the trial.
These tanks are initially held at lab temperatures (~ 26°C); however, after the fish begin feeding, water temperature is increased or decreased in each acclimation aquarium by 1°C per day until the appropriate treatment temperature is reached. Lionfish are then held for an additional 12 days before undergoing temperature tolerance trials.
For each Critical Thermal Trial (CTM), one or two lionfish from the chosen acclimation treatment aquarium were placed into an insulated 45 × 30 × 15 cm CTM chamber set at the appropriate treatment temperature. The CTM chamber consists of a 15-L recirculating seawater bath that can be heated at a constant rate of 0.25°C/min. Water temperature increase is then initiated and continues until the fish experiences Loss Of Equilibrium (LOE). When equilibrium loss is observed, the water temperature is recorded and the fish immediately returned to acclimation temperature to recover.
LOE is determined when the fish cannot maintain an upright position (loss of dorso-ventral orientation) for a minute.
The trial process is repeated until all experimental fish have been tested. All fish are released at their site of capture at the end of the study and suffer no ill effects.
Temperature tolerance values are estimated as critical thermal maximum (CTmax), and derived using the critical thermal methodology (CTM) as described by Beitinger and co-workers (2000).

6. Research questions and methodology
A diver makes a 10 minute sound recording for one specific fish and repeats this for another 19 fish of the same species.
This is repeated for another species living on the same anemone.
The results are analyzed using graphs and statistics.

7. Research questions and methodology
Which vocalisation characteristics differ between Amphiprion clarkia and Amphiprion perideraidon?
2. What could be the driving force of differences in vocalisations between species?

8. Discussion
The data suggests that there are clear differences in vocalisation between the two species studied.
Further statistics support the idea that it is the body size of the fish that is related to the vocalization differences between the two species that occupy the same anemone. A discussion can link this idea to your own ideas on the process of evolution.
Answers/discussions to research questions:
1. Our data show that thermal tolerance of the native red lionfish population on Hoga is indeed altered by changes in water temperature. From our results we may infer that lionfish experiencing different thermal regimens would also exhibit different thermal tolerance characteristics. We might go one step further and predict that lionfish in our experiment gained heat tolerance in a predictable manner. We can look to the regression model to help us address this question.
2. The relationship between acclimation temperature and critical thermal maximum suggests that for every increase in acclimation temperature of 1°C, lionfish thermal tolerance increases by 0.16°C. This value is given by the slope of your trend line, and in thermal ecology jargon, we refer to it as the acclimation response ratio or ARR.
Though we have only four points on our graph, the R2 value of suggests the relationship between acclimation temperature and heat tolerance is a strong one. Typically ARR values are low in tropical fish that see little seasonal change in water temperature. So if for example, we found a higher ARR value in the invasive Atlantic lionfish population, it may indicate a high phenotypic plasticity in this thermal tolerance trait. Such a finding may in turn have implications for expansion of the fish into other novel habitats where seasonal water temperature changes are more profound. Of course these findings are tentative and definitive answers will only be forthcoming after many more experiments.