Feeding preference of herbivores can influence the structure of communities and alter plant diversity in an area. High grazing pressure can lead to a decrease in overall plant diversity but intermediate pressures can result in an overall increase in diversity. Tripneustes gratilla have strong chemoreceptors that can sense food items from over a distance of 1 metre, making olfactory tests a useful way of determining food preference. Results indicate they preferentially feed on red (Rhodophyte) algae over green (Chlorophyta) (p=0.03).Invasive red algae has proved to be of detriment to tropical reefs worldwide with local herbivorous fishes finding it unpalatable, thereby reducing the biological control effectiveness. Introduction of T. gratilla to these areas can severely reduce the spread of these invasive algaes, complementing the biological control of fishes in the system. 

Feeding preferences of grazers can dramatically change the way an ecosystem looks and functions by interacting with the competitive abilities of plants, influencing their overall survivorship (Vadas 1977). Grazers can preferentially choose to predate on certain types of prey, altering the structure of the system. If a grazer preferentially preys on certain types of algae for example, this can result in new space for other plants to inhabit. An intermediate level of grazing should be present in a system in order to maintain high algal diversity, in turn creating a wider range of food sources that can be utilized by other organisms (Paine & Vadas 1969).

In tropical waters, both red (Rhodophyta) and green (Chlorophyte) algae are abundant and can be found competing for space in some areas. Red algae is generally found at greater depths than green algae due to its absorption of blue high energy wavelengths, although there is a relatively large overlap between the two habitats (Burger et al. 1999). Grazers are commonly found feeding on both types of algae and can alter the ratio of each depending on their preference. Some companies have developed algae sheets for marine aquaria organisms solely using dried algae. Ocean Nutrition™ have both red and green algae sheets which are comprised of either dried Rhodophyta or Chlorophyte algae respectively. These sheets are highly useful in laboratory experiments as they can be easily standardized between tests and nutritional value is very similar. The red algae sheets contain marginally more fat than the green but slightly less fibre. These sheets also break up easily, releasing odours into the water which can be used in olfactory tests. Due to the composition of these sheets, they can be utilized by a range of herbivores from fish through to grazing invertebrates. It is for these reasons that these sheets were used in this experiment instead of live algae.

Collector sea urchins (Tripneustes gratilla) are found at depths ranging from 2 to 30 metres in tropical waters world-wide (Valentine & Edgar 2010). Mature collector urchins are generally found conspicuously along the sea floor and have found to be capable of sensing potential food sources from over 1 metre away using chemoreceptors located within their jaw epithelium (Raible et al. 2006). They are known to be voracious predators on algae and seagrasses and unlike some other urchins, are found to graze continuously day and night (Vadas 1977). Stimson et al. (2007) suggest that T. gratilla are generalist herbivores but are capable of displaying feeding preferences on algae. Shunula & Ndibalema (1986) conducted a feeding preference experiment on a range of sea urchins and found that they overall prefer red algae to green. This is thought to be due to green algae usually being harder for urchins to consume due to the location of the mouth parts on their oral side, acting as a physical deterrent. Although this experiment involves using algae sheets instead of live algae, the test urchins were caught from the wild and therefore could have naturally developed a preference for either.  

Five collector sea urchins (Tripneustes gratilla) of similar sizes were randomly selected from the aquaria at the University of Queensland, having been recently caught in the wild and starved for the week prior to performing the experiment. Urchins were numbered 1 to 5 and identified by specific colouring or spine orientation. A black, 80 litre tub was filled with enough water to ensure urchins were fully submerged and care was taken to ensure tub water conditions matched the tank water which the urchins were kept in. The tub was kept in the aquaria which had soft lighting and easy water access to minimize stress on the urchins. At each end of the tub, a 3x3 cm square of algae was placed and covered with a glass slide to prevent it floating away. At one end was green Ocean Nutrition™ marine algae and the red equivalent was placed at the opposing end. Each piece of algae was rubbed slightly to release its odour into the water. Urchin number one was then placed in the centre of the tub and timed for 5 minutes to see which colour algae it moved towards. A successful choice was counted if the body of the urchin passed the middle marker (Figure 1). If the urchin did not fully cross the marker or spent the time moving in circles, this was counted as a 'no choice' result. Care was taken to ensure the urchin did not actually consume any algae as this could have confounded results.

Once the urchin had successfully chosen a colour or the time was expended, it was placed back into the holding tank and urchin number two was placed in the tub. This was repeated until all five urchins had been tested. Once all were tested, the tub water and algae sheets were emptied and the tub was filled with new water and new algae sheets were placed again at opposing ends. After each water change, the placement of each coloured algae was swapped to ensure urchins were not preferring one side of the tub to the other. Urchins 1 to 5 were then tested again using the above procedure and water was changed after each group of tests. Three replicates of each individual were done each week over three weeks as time allowed with urchins being starved between weeks.

Once results were collected, they were arranged in Microsoft Excel and statistically analysed in R studio using a binomial distribution test. This was done by assuming that there would be an even 50/50 split in preference if H0 were true. 

Of the 50 data points collected, 23 were ‘no choice’ results where the urchin did not move over the marker or moved in circles for the 5 minute duration. However, as the focus of this experiment is which colour algae they prefer, these points were omitted from the data. Urchin 1 chose the red algae 5 times and the green algae twice. Urchin 2 chose red 3 times and green once. Urchin 3 chose the red algae 4 times and the green algae once. Urchin 4 chose the red algae 5 times and the green algae once. Urchin 5 chose the red algae twice and the green algae 3 times. Of the 10 tests conducted on each urchin, the remaining choices were ‘no choice’ results (Figure 2). Results from the binomial distribution test suggest that T. gratilla significantly prefer red algae to green (p=0.03). In the 27 tests where a choice was made, red algae was chosen 19 times (Figure 3). 

Of the 50 tests recorded, 23 times the urchin did not make a choice and therefore these were disregarded in the results. When completing the tests, these ‘no choice’ results tended to occur later in the day after the first few tests had been completed. It is believed that this occurred as a result of being stressed from being handled too often, although care was taken to ensure minimum stressors were present. Over the three weeks of tests, these ‘no choice’ results were also occurring more frequently towards the last week. It was found that the urchins were slowly dying off as others in the holding tank (not ones used in this experiment) had to be discarded. These ‘no choice’ results were mainly from the urchin not moving more than a few centimetres over the five minutes allocated, rather than moving but not making a definitive choice. Future studies should consider using more replicates over a shorter time period to minimize the overall stress that these urchins experienced.

Red algae as a whole generally displays greater levels of herbivore deterrence by being harder to consume, either by being courser or tougher than green algae. However if urchins are preferentially grazing on red algae, this in turn can increase herbivory by other fishes or invertebrates on green algae (Lubchenco 1978). Paine & Vadas (1969) concluded that an intermediate level of urchin grazing can result in a greater diversity of algal species in an area. However, they also found that algal diversity is severely reduced under intense grazing pressures and hypothesized that this could eventually result in a monoculture. Local destruction and possible deterrents could minimize grazing pressure to maintain high algal diversity in an ecosystem.

Urchins prefer different algae to herbivorous fishes and are therefore usually found inhabiting different niches, thereby they can complement the biological control agents utilized by the fishes (Stimson et al. 2007). Invasive red algae has been known to spread rapidly and smother reefs, likely due to its ability to grow from small attachment points on the substrate. Rodgers & Cox (1999) found that red algae has the ability to spread at a rate of over 250 metres per year, a result of accidental aquaculture introductions. Manual removal of this algae requires around two hours to clear a 1 metre squared patch and red algae can also be found at depths that make it hard for humans to remove manually (Conklin & Smith 2005). Due to the low palatability of this algae to native fishes and the reasons above, the introduction of T. gratilla as a biological control agent for invasive red algae has proved highly successful, complementing the niche that native herbivorous fishes inhabit.

In conclusion, this experiment has shown to support the proposed hypothesis with T. gratilla showing a preference for red algae over green. From previously conducted studies, it is believed that this is mainly a result of the higher palatability that this colour displays for urchins, complementing the preferences of most herbivorous fishes. The use of T. gratilla as a biological control agent can severely reduce the spread of invasive red algaes and assist in maintaining algal diversity on reefs. Future studies could focus on the complementary feeding behaviours of herbivorous fishes and urchins to determine a ratio that would provide the highest possible algal diversity in an area, a focus that was beyond the scope of the time restraints of this course.

I would like to thank Bernie and Sandy Degnan for coordinating such a fun and interesting course and giving us the opportunity to further our knowledge of marine invertebrates. I would also like to thank the tutors Tahsha, Dylan and Eunice who shared their time and knowledge to assist in this experiment.

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