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Distribution of sea cucumbers, Holothuria atra, on reefs in the upper Gulf of Thailand and the effect of their population densities on sediment microalgal productivity
Estuarine, Coastal and Shelf Science 235 (2020) 106514
Contents lists available at ScienceDirect
Estuarine, Coastal and Shelf Science journal homepage: http://www.elsevier.com/locate/ecss
Distribution of sea cucumbers, Holothuria atra, on reefs in the upper Gulf of Thailand and the effect of their population densities on sediment microalgal productivity Voranop Viyakarn *, Suchana Chavanich, Eliza Heery, Chalothon Raksasab Reef Biology Research Group, Department of Marine Science, Faculty of Science, Chulalongkorn University, Bangkok, 10330, Thailand
A R T I C L E I N F O
A B S T R A C T
Keywords: Microalgae Holothurian Abundance Deposit feeder Feeding behavior
Populations of sea cucumbers in Thailand have been declining because of overfishing in recent years; however, little is known about the possible long-term effects of this decline on the sea cucumber population and reef communities. The purpose of this study was to examine the distribution patterns and habitat compositions of Holothuria atra in the upper Gulf of Thailand. In addition, field and laboratory experiments were conducted to investigate the impact of sea cucumber density and feeding activity on the sedimentary microalgal community. The results showed that there was a correlation between the density of H. atra and substrate composition. The density of H. atra increased as the percentage of sand cover in the habitats increased. High concentrations of chlorophyll–a were detected in gut contents. In addition, results from field and laboratory experiments showed that when H. atra was absent, a high concentration of chlorophyll-a in the reef sediment was recorded. The higher the number of H. atra, the greater the capability to reduce the microalgal biomass in the sediment. Thus, H. atra plays an important role in recycling nutrient in reefs in the Gulf of Thailand, and consequently, a decline in the sea cucumber population may alter the reef community.
1. Introduction Sea cucumbers are considered to be one of the most conspicuous species in coral reef communities. Over the past decade, sea cucumbers have been heavily harvested and exported to China and other Asian markets because of high demand (Conand, 2001; Uthicke and Benzie, 2000; Kinch et al., 2008; Purcell, 2010). Pressure on sea cucumbers as a resource has increased in many countries such as Thailand, and global export figures indicate that populations of sea cucumbers are currently overexploited (Conand, 2001; Purcell, 2010). Sea cucumbers recover very slowly, and thus, can take decades to recover from over-exploitation (Battaglene and Bell, 1999; Bruckner et al., 2003; Purcell, 2010). However, in Thailand, little is known about population density, and the possible effects of a decline in sea cucumber numbers (Bussarawit and Thongtham, 1999). In coral reef ecosystems, the distribution of sea cucumbers can in fluence environmental factors such as nutrient concentrations in the sediment (Uthicke, 2001a, 2001b). In contrast, environmental factors such as temperature and salinity can also have an effect on sea cucumber abundance (Uthicke, 1994; Mercier et al., 2000). Different holothurian
species can co-exist and partition resources within the same habitat (Hammond, 1980; Hammond et al., 1985). Holothurians are important recyclers of inorganic nutrients and are believed to have an important role in bioturbation (Uthicke, 1999; Wolfe and Byrne, 2017). Sea cucumbers ingest large amounts of sediments, and their gut contents can be composed of bacteria, diatoms, algae, molluscan shells, and others (Hamel et al., 2001). High holothurian densities may reduce benthic microalgal production and biomass (Moriarty, 1982; Uthicke, 1999). However, Uthicke and Klumpp (1997) and Uthicke (2001a, 2001b) have shown that the feeding activity of holothurians at natural densities increased nutrient levels in the sedi ment, which also benefitted microalgae because of the excretion of ammonium ions by the holothurians. As coral reefs degrade and shift to algal-dominated states, it is important to characterize the key processes and taxa that can mediate benthic algal productivity. It appears that sea cucumbers play an important role in benthic community processes on coral reefs through sediment bioturbation, but empirical evidence in the literature is scarce. In Thailand, sea cucumbers are currently threatened by over exploitation; however, the effect of the declining population on reefs
* Corresponding author. E-mail address: [email protected] (V. Viyakarn). https://doi.org/10.1016/j.ecss.2019.106514 Received 2 May 2019; Received in revised form 18 November 2019; Accepted 29 November 2019 Available online 2 December 2019 0272-7714/© 2019 Published by Elsevier Ltd.
V. Viyakarn et al.
Estuarine, Coastal and Shelf Science 235 (2020) 106514
Fig. 1. Map of study area.
and reef communities was unknown (Bussarawit and Thongtham, 1999). The purpose of this paper was to examine the distribution pattern of Holothuria atra in the upper Gulf of Thailand. Bottom coverage, po tential nutrient content of surrounding sediments, and gut contents were also evaluated and tested to estimate the potential effects of their feeding behavior on the sediment. Field and aquarium experiments were conducted to assess the impact of density and feeding activity on the sedimentary microalgal community.
2.2. Density and habitat structure of sea cucumbers Densities and percent composition of the habitats of Holothuria atra were investigated around the islands located in Chon Buri and Trat Provinces using transect lines and quadrats. A 50-m transect line was extended along the substrate on a flat reef parallel to the shore. A diver then swam along the length of the line and recorded the number of sea cucumbers within 1 m of each side of the line. At least three transect lines were established on each island. Data were then compiled and divided by the total area surveyed (100 m2). The percent composition of sea cucumber habitats was also determined along the transect lines. A quadrat (0.5 � 0.5 m) with string crossed to form 25 points were placed immediately adjacent and on both sides of the transect at 0, 5, 10, 15, 20, 25, 30, 35, 40, 45, and 50 m. In each quadrat, the type of substrate below the points was recorded within the following categories based on Reef Check method: soft coral, hard coral, algae, rock, rubble, sand, or others. Silt was not found in the areas. Once the data were compiled, the percent composition was derived from the fraction of each category over the total 25 points.
2. Methods 2.1. Study sites Data and sample collections took place at 20 islands located in Chon Buri and Trat Provinces in the upper Gulf of Thailand. Most islands located in Chon Buri Province, but Kham and Khram Islands are the most restricted and protected by the Royal Thai Navy. Thus, no harvesting of marine organisms is allowed in these 2 islands. The study sites was shown in Fig. 1. 2
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Estuarine, Coastal and Shelf Science 235 (2020) 106514
2.3. Gut content analysis Thirty sea cucumbers were collected in the sand areas of reef flats at Kham Island. The samples were analyzed to determine the gut contents. The composition of stomach contents and bacterial numbers were examined and counted. To determine the composition in the stomach, all contents of each stomach were removed. Then, they were spread in a petri dish with lines crossed to form 100 points. Type of contents found in each point was recorded. For the bacterial count, the small contents from each sea cucumber’s stomach were spread in a petri dish con taining bacterial agar, and allowed bacteria to grow for a week. After one week, the numbers of bacterial colonies grew in each plate were counted under the microscope. To test whether sea cucumbers consume benthic diatoms, an addi tional 20 sea cucumber gut samples of each sea cucumber size, 20 sediment samples from the surrounding habitat, and 20 sea cucumber feces samples were collected for chlorophyll-a analyses. Three sizes of sea cucumbers were collected: small, medium, and large (<10, 10–25, >25 cm in length respectively). Each sample was collected in separated zip-lock plastic bags by divers. Samples and specimens were immedi ately frozen and protected from light when brought to the surface, and remained so until analyzed in the laboratory. In the laboratory, sea cucumber specimens were defrosted, weighed, measured for length and dissected. The gut contents were removed and again frozen and pro tected from light. Pigment extraction and analysis was performed once the samples had been thawed in a dark location. The method of chlorophyll analysis was adopted from Gilpin (2001). For each sample, 5 g was placed in a 30-ml container with a screw on cap. Then, 20 ml of methanol was added and the container stirred vigorously and allowed to stand for 5 min. A 1-ml aliquot of the methanol containing pigments was then extracted with a pipette and placed into a test tube with a top. Then, 9 ml of 90% acetone was added to the solution and the tube wrapped in foil until analysis in a spectrometer. The methanol-acetone solution was poured into a cuvette with a pathlength of 1 cm. After wiping the cuvette thoroughly, it was placed in a spectrometer for measurement. Absorbency readings were made at 665 nm and 750 nm before 4 drops of HCl were added to the sample and the measurements repeated. With each change in wavelength and acid addition, the spectrometer was calibrated with a blank of 90% acetone, containing or excluding 4 drops of HCl, depending on the constituents of the sample. The compiled data was then calculated with the following formulas for the derivation of chlorophyll-a concentration: Chlorophyll-a
Fig. 2. Average densities of Holothuria atra and the sand compositions on twenty reefs in Chon Buri and Trat Provinces.
A field experiment was conducted at Kham Island, Chon Buri Prov ince, to examine the relationship between sea cucumber population density and the biomass of benthic microalgae using chlorophyll as an indicator of benthic microalgal biomass. The uniqueness of this study site is that sea cucumbers are not overexploited, and the site is protected by the Royal Thai Navy. The study site was manipulated by the authors as follows: 1) an area without sea cucumbers; 2) an area where the density of sea cucumbers equaled the average or natural density (approximately 25 sea cucumbers per 100 m2); 3) an area where the density of sea cucumbers is two times higher than the natural density (approximately 50 sea cucumbers per 100 m2). Each area was caged. All experimented sea cucumbers were collected from the area. Because it was decided to measure the actual concentrations of chlorophyll-a with only the density of sea cucumbers as a fixed factor, the densities of other benthic organisms were not manipulated and remained natural. Ten sediment samples in each area treatment were collected one month after the experiment for chlorophyll-a analysis. A one-way ANOVA followed by a Tukey pairwise comparison test was used to test differences in chlorophyll-a concentrations in each area.
[chl] (μg g 1) ¼ 26.7 x (665b–665a) / L 665b: absorbance at 665 nm before acidification minus absorbance at 750 nm before acidification 665a: absorbance at 665 nm after acidification minus absorbance at 750 nm after acidification L: pathlength of the spectrophotometer cuvette used (cm)
3.2. Laboratory experiment A laboratory experiment was conducted to determine the relation ship between the density of sea cucumbers and the density of benthic microalgae in the sediments. All sea cucumbers used in the experiment were collected from Kham Island, and were between 25 and 30 cm in length. Three treatments with ten replicates each were used. All exper iments were conducted in 20-gallon glass aquaria (76 � 31 � 31 cm) at room temperature (30 � C), a salinity of 30 psμ, and in moderate sunlight. The artificial seawater was used, and the seawater was changed daily. Sand used in the experiment was collected from the habitat where sea cucumbers were actually found at Kham Island. Each aquarium con tained one of the following: 1) sand on the bottom and one individual sea cucumber; 2) sand and three individuals; 3) a control with sand and no sea cucumbers. The chlorophyll-a concentrations of sands from each aquarium were measured at the beginning, middle, and end of the
3. Experiments on the relationship between sea cucumber and benthic microalgal densities 3.1. Field experiment From preliminary surveys, high chlorophyll concentrations were detected from gut content analyses; thus, indicating that H. atra con sumes benthic microalgae. Therefore, sea cucumbers may play a role in controlling the biomass of primary production on coral reefs. The hy pothesis was that the density of sea cucumbers influenced the density of benthic diatoms. In this study, when benthic microalgae were referred, it meant benthic diatoms. 3
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5. Discussion The field surveys showed that the highest density of Holothuria atra was recorded at Kham Island, followed by Khram Island. As these two islands are protected by the Royal Thai Navy, fishermen are not allowed
Fig. 3. Percent composition of Holothuria atra habitats on twenty reefs in Chon Buri and Trat Provinces.
experiment. Each time, five replicates were collected in each aquarium. The experiment was run for four days. A one-way ANOVA followed by a Tukey pairwise comparison test was used to test differences between in chlorophyll-a concentrations in each treatment. Fig. 5. Chlorophyll-a concentration in the gut, feces, and in the surrounding sediment of Holothuria atra.
4. Results The field survey results showed that the density of Holothuria atra at Kham and Khram Islands, areas protected by the Royal Thai Navy, were higher than on other unprotected islands (Fig. 2). In addition, difference of the densities of H. atra in each site depended on the substrate composition. The density of H. atra increased with the percentage sand cover within the habitat (Fig. 3). Regarding gut content analysis, results showed that the gut content of small size class (<10 cm in length) of H. atra was composed of sand only, while in medium (10–25 cm in length) and large (>25 cm in length) size classes, both sand and shell fragments were found in the guts (Fig. 4). The average bacterial number recorded in the gut of H. atra was between 10 1 and 10 2 CFU. A comparisons of chlorophyll-a concen tration between in H. atra gut, feces, and the surrounding sediment showed that chlorophyll-a was higher in the gut than in the feces or sediment (Fig. 5). The results from the field experiment showed that concentrations of chlorophyll-a were higher in the control plot containing no H. atra specimens (Fig. 6). However, the area with double the density of H. atra had a lower chlorophyll-a concentration than the other plots. Results of the laboratory experiment showed that the highest chlorophyll con centrations were in the aquaria with more individuals of H. atra (Fig. 7).
Fig. 6. Chlorophyll-a field experiment.
concentration
Fig. 4. Gut contents in different size classes of Holothuria atra. 4
in
sediments
after
a
1-month
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Estuarine, Coastal and Shelf Science 235 (2020) 106514
1985). In addition, toward the end of our field experiment, it was observed that the sea urchin, Diadema setosum, was moving into the area (approximately 1 individual/1 m2) where the H. atra population was cleared out. The migration of sea urchins could be attracted by the in crease in the benthic microalgal biomass. Sea urchins usually prefer both micro and macroalgae as food sources (Harris et al., 2001). Neverthe less, other studies have found that holothurians induced nutrient regeneration through excretion of ammonium ions (Uthicke, 2001a, 2001b). Further studies are required to investigate in detail the rela tionship between holothurians, benthic algae, and other benthic reef organisms. The results of this study showed that difference of the densities of H. atra in each site depending on the substrate composition. It was also shown that the feeding activity of H. atra has an impact on benthic microalgal production and biomass in reefs in the upper Gulf of Thailand. Holothurians play an important role in recycling nutrients in reef ecosystems, and the decline in sea cucumber numbers may nega tively affect reef communities.
Fig. 7. Changes in chlorophyll-a concentrations after 2 and 4 days in the lab oratory experiment.
to access, in contrast to the situation around other islands. In this study, the density of H. atra was correlated to its habitat composition. The higher the percentage of sand cover in an area, the greater the density of H. atra. It has been shown that holothurians are distributed mainly on a sand substratum (Mercier et al., 1999). However, some studies indicated that other factors, such as organic matter content of the sediment, water movement, etc. may also influence the distribution of sea cucumbers (Kerr et al., 1993; Mercier et al., 1999, 2000). Holothuria atra is categorized as a deposit feeder. In this study, gut content analysis showed that the gut of the small size class was composed solely of sand, while the guts contents of the larger size classes contained a mixture of sand and mollusk shell fragments. The gut con tents of other sea cucumber species, such as H. scabra, comprised bac teria, copepods, diatoms, algae, molluscan shells, foraminiferans, sand, and muds (Wiedemeyer, 1992; Baskar, 1994; Hamel et al., 2001). In this study, although bacteria were found, the numbers were very low. Thus, bacteria may not be a major food source of H. atra. A comparison of chlorophyll-a concentration in H. atra gut, feces, and surrounding sands showed that the highest concentrations were found in the gut. The results suggested that benthic microalgae may be an important food sources for H. atra. It has been suggested that chlo rophyll-a concentration and bacterial content could be used as in dicators of the nutritional value of sediments for holothurians (Moriarty, 1982). In the Solomon Islands, Battaglene and Bell (1999) observed that cultured juveniles of H. scaba fed on epiphytic algae and bacteria on the substrate. Other studies have showed that H. scaba, can display substrate selection, and was able to distinguish between sediments based on grain size (Wiedemeyer, 1992; Baskar, 1994). However, there is also a pos sibility for sea cucumbers to have their own additional gut flora than in the sediment, which led to higher chlorophyll concentrations in the gut than in the habitat. More studies in depth on the feeding behavior of sea cucumber in a relation to gut contents are needed. Gut content analysis suggested that the feeding behavior of H. atra could reduce the concentration of chlorophyll-a in surrounding sedi ments since the chlorophyll concentration in the feces was lower than the surrounding sediment. Yingst (1976) and Hammond (1983) reported that some diatoms were digested by holothurians. Even though the feces had lower chlorophyll as sea cucumbers digested the algal food, this was not necessarily shown that sea cucumbers reduced the chlorophyll concentration in the environment (Uthicke and Klumpp, 1997). The results of field and laboratory experiments demonstrated that H. atra may play an important role in controlling the density of benthic diatom and epiphytic algae in sediments. The higher the number of sea cucumbers, the greater the capability to reduce the algal biomass in the sediment (Fig. 6). Previous studies have shown that benthic microalgal communities have decreased through the feeding activity by holothu rians (Moriarty et al., 1985; Uthicke, 1999). However, when holothu rians were absent, cyanobacterial mats were observed (Moriarty et al.,
6. Conclusions At present, sea cucumbers are overharvested and their populations have been declined dramatically. This study showed that the increase and decrease of sea cucumber populations can have significantly affected to the benthic microalgal production in the sediment and reef ecosystems. Thus, further studies are needed to investigate in details of the linkages between sea cucumber populations and other benthic reef organisms and their in depth roles in reef ecosystems. Acknowledgements This study was part of the Plant Genetic Conservation Project under the Royal Initiative of Her Royal Highness Princess Maha Chakri Sir indhorn. The authors would like to thank members of the Reef Biology Research Group and the Naval Special Warfare Command, Royal Thai Fleet, Royal Thai Navy for assisting in data collections. This work was partially supported TRF (RSA6080087), PADI Project AWARE grant, PADI Foundation grant, and Mubadala Petroleum (Thailand) Limited. Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi. org/10.1016/j.ecss.2019.106514. References Baskar, B.K., 1994. Some observations on the biology of the holothurian Holothuria (Metriatyla) scabra Jaeger. In: Rengarajan, K., James, D.B. (Eds.), Proceedings of the National Workshop on Beche-De-Mer. Bulletin of the Central Marine Fisheries Research Institute 46. Indian Council of Agriculture Research, Cochin, India, pp. 39–43. Battaglene, S.C., Bell, J.D., 1999. Potential of the tropical Indo-Pacific sea cucumber Holothuria scabra for stock enhancement. In: Howell, B.R., Moskness, E., Svasand, T. (Eds.), Stock Enhancement and Sea Ranching. Blackwell Science, Oxford, pp. 478–490. Bruckner, A.W., Johnson, K.A., Field, J.D., 2003. Conservation strategies for sea cucumbers. Can a CITES Appendix II listing promote sustainable international trade? SPC Beche-de mer Inf. Bull. 18, 24–33. Bussarawit, S., Thongtham, N., 1999. Sea cucumber fisheries and trade in Thailand. In: Baine, M. (Ed.), Proceeding of the International Conference “The Conservation of Sea Cucumber in Malaysia, Their Taxonomy, Ecology and Trade”. Hariot-Watt University and Fisheries Research Institute, Kuala Lumpur, pp. 26–36. Conand, C., 2001. Sea cucumber retail market in Singapore. SPC Beche-de-mer Inf. Bull. 14, 12–14. Gilpin, L., 2001. Marine Biology: methods for analysis of benthic phytoplankton pigment. Available at: www.oaerre.napier.ac.uk/users/p.tett/MB/benchl01.html. Hamel, J.F., Conand, C., Pawson, D.L., Mercier, A., 2001. The sea cucumber Holothuria scabra (Holothuroidea: Echinodermata): its biology and exploitation as beche-demer. In: Southward, A.J., Tyler, P.A., Young, C.M., Fuiman, L.A. (Eds.), Adv. Mar. Biol. 41, 129–223.
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Estuarine, Coastal and Shelf Science 235 (2020) 106514 Moriarty, D.J.W., Pollard, P.C., Hunt, W.G., Moriarty, C.M., Wassenberg, T.J., 1985. Productivity and bacteria and micro algae and the effect of grazing by holothurians in sediments on a coral reef flat. Mar. Biol. 85, 293–300. Purcell, S.W., 2010. Managing sea cucumber fisheries with an ecosystem approach. In: Lovatelli, A., Vasconcellos, M., Yimin, Y. (Eds.), FAO Fisheries and Aquaculture Technical Paper No, vol. 520, p. 157pp. Rome. Uthicke, S., 1994. Distribution patterns and growth of two reef flat holothurians, Holothuria atra and Stichopus chloronotus. In: David, B., Guille, A., Feral, J.P., Roux, M. (Eds.), Echinoderms (Dijon), Proc. 8th Int. Echinoderm Conf. Balkema, Rotterdam, pp. 569–576. Uthicke, S., 1999. Sediment bioturbation and impact of feeding activity of Holothuria (Halodeima) atra and Stichopus chloronotus, two sediment feeding holothurians, at Lizard Island, Great Barrier Reef. Bull. Mar. Sci. 64, 129–141. Uthicke, S., 2001. Interactions between sediment-feeders and microalgae on coral reef : grazing losses versus production enhancement. Mar. Ecol. Prog. Ser. 210, 125–138. Uthicke, S., 2001. Nutrient regeneration by abundant coral reef holothurians. J. Exp. Mar. Biol. Ecol. 265, 153–170. Uthicke, S., Benzie, J.A.H., 2000. Effect of beche-de-mer fishing on densities and size structure of Holothuria nobilis (Echinodermata: Holothuroidea) populations on the Great Barrier Reef. Coral Reefs 19, 271–276. Uthicke, S., Klumpp, D.W., 1997. Ammonium excretion by holothurians enhances production and turnover in benthic diatom communities. In: Lessios, H.A., Macintyre, I.G. (Eds.), Proc. 8th Int. Coral Reef Symp., Panama, pp. 873–876. Wiedemeyer, W.L., 1992. Feeding behavior of two tropical holothurians Holothuria (Metriatyla) scabra (Jaeger 1833) and H. (Halodeima) atra (Jaeger 1833), from Okinawa, Japan. In: Proc. 7th Int. Coral Reef Symp., Guam, pp. 853–860. Wolfe, K., Byrne, M., 2017. Biology and ecology of the vulnerable holothuroid, Stichopus herrmanni, on a high-latitude coral reef on the Great Barrier Reef. Corel Reefs 36 (4), 1143–1156. Yingst, J.Y., 1976. The utilization of organic matter in shallow marine sediments by an epibenthic deposit feeding Holothurian. J. Exp. Mar. Biol. Ecol. 23, 55–69.
Hammond, L.S., 1980. Diversity and coexistence in an assemblage of deposit-feeding holothurians and spatangoids in the reef lagoon, Discovery Bay, Jamaica. Proc. Assoc. Is. Mar. Labs. Carib. 15, 1–11. Hammond, L.S., 1983. Nutrition of deposit-feeding holothuroids and echinoids (Echinodermata) from a shallow reef lagoon, Discovery Bay. Jamaica. Mar. Ecol. Prog. Ser. 10, 297–305. Hammond, L.S., Birtles, R.A., Reichelt, R.E., 1985. Holothuroid assemblages on coral reefs across the central section of the Great Barrier reef. Proc. 5th Int. Coral Reef Symp., Tahiti 285–290. Harris, L.G., Tyrrell, M., Williams, C., Sisson, C., Chavanich, S., Chester, C., 2001. Declining sea urchin recruitment in the Gulf of Maine: is overfishing to blame? In: Barker, M. (Ed.), Echinoderm 2000. Balkema, Rotterdam, pp. 439–444. Kerr, A.M., Stoffel, E.M., Yoon, R.L., 1993. Abundance and distribution of holothuroids (Echinodermata: Holothuroidea) on a windward and leeward fringing coral reef, Guam, Mariana Islands. Bull. Mar. Sci. 52, 780–791. Kinch, J., Purcell, S., Uthicke, S., Friedman, K., 2008. Population status, fisheries and trade of sea cucumbers in the Western Central Pacific. In: Toral-Granda, V., Lovatelli, A., Vasconcellos, M. (Eds.), Sea Cucumbers: A Global Review of Fisheries and Trade. FAO Fisheries and Aquaculture Technical Paper, pp. 7–55. No. 516, Rome. Mercier, A., Battaglene, S.C., Hamel, J.F., 1999. Daily burrowing cycle and feeding activity of juvenile sea cucumber Hotothuria scabra in response to environmental factors. J. Exp. Mar. Biol. Ecol. 239, 125–156. Mercier, A., Battaglene, S.C., Hamel, J.F., 2000. Periodic movement, recruitment and size-related distribution of the sea cucumber Holothuria scabra in Solomon Islands. Hydrobiologia 440, 81–100. Moriarty, D.J.W., 1982. Feeding of Holothuria atra and Stichopus chloronotus on bacteria, organic carbon and organic nitrogen in sediments of the Great Barrier Reef. Aust. J. Mar. Freshw. Res. 33, 255–263.
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Contents lists available at ScienceDirect
Estuarine, Coastal and Shelf Science journal homepage: http://www.elsevier.com/locate/ecss
Distribution of sea cucumbers, Holothuria atra, on reefs in the upper Gulf of Thailand and the effect of their population densities on sediment microalgal productivity Voranop Viyakarn *, Suchana Chavanich, Eliza Heery, Chalothon Raksasab Reef Biology Research Group, Department of Marine Science, Faculty of Science, Chulalongkorn University, Bangkok, 10330, Thailand
A R T I C L E I N F O
A B S T R A C T
Keywords: Microalgae Holothurian Abundance Deposit feeder Feeding behavior
Populations of sea cucumbers in Thailand have been declining because of overfishing in recent years; however, little is known about the possible long-term effects of this decline on the sea cucumber population and reef communities. The purpose of this study was to examine the distribution patterns and habitat compositions of Holothuria atra in the upper Gulf of Thailand. In addition, field and laboratory experiments were conducted to investigate the impact of sea cucumber density and feeding activity on the sedimentary microalgal community. The results showed that there was a correlation between the density of H. atra and substrate composition. The density of H. atra increased as the percentage of sand cover in the habitats increased. High concentrations of chlorophyll–a were detected in gut contents. In addition, results from field and laboratory experiments showed that when H. atra was absent, a high concentration of chlorophyll-a in the reef sediment was recorded. The higher the number of H. atra, the greater the capability to reduce the microalgal biomass in the sediment. Thus, H. atra plays an important role in recycling nutrient in reefs in the Gulf of Thailand, and consequently, a decline in the sea cucumber population may alter the reef community.
1. Introduction Sea cucumbers are considered to be one of the most conspicuous species in coral reef communities. Over the past decade, sea cucumbers have been heavily harvested and exported to China and other Asian markets because of high demand (Conand, 2001; Uthicke and Benzie, 2000; Kinch et al., 2008; Purcell, 2010). Pressure on sea cucumbers as a resource has increased in many countries such as Thailand, and global export figures indicate that populations of sea cucumbers are currently overexploited (Conand, 2001; Purcell, 2010). Sea cucumbers recover very slowly, and thus, can take decades to recover from over-exploitation (Battaglene and Bell, 1999; Bruckner et al., 2003; Purcell, 2010). However, in Thailand, little is known about population density, and the possible effects of a decline in sea cucumber numbers (Bussarawit and Thongtham, 1999). In coral reef ecosystems, the distribution of sea cucumbers can in fluence environmental factors such as nutrient concentrations in the sediment (Uthicke, 2001a, 2001b). In contrast, environmental factors such as temperature and salinity can also have an effect on sea cucumber abundance (Uthicke, 1994; Mercier et al., 2000). Different holothurian
species can co-exist and partition resources within the same habitat (Hammond, 1980; Hammond et al., 1985). Holothurians are important recyclers of inorganic nutrients and are believed to have an important role in bioturbation (Uthicke, 1999; Wolfe and Byrne, 2017). Sea cucumbers ingest large amounts of sediments, and their gut contents can be composed of bacteria, diatoms, algae, molluscan shells, and others (Hamel et al., 2001). High holothurian densities may reduce benthic microalgal production and biomass (Moriarty, 1982; Uthicke, 1999). However, Uthicke and Klumpp (1997) and Uthicke (2001a, 2001b) have shown that the feeding activity of holothurians at natural densities increased nutrient levels in the sedi ment, which also benefitted microalgae because of the excretion of ammonium ions by the holothurians. As coral reefs degrade and shift to algal-dominated states, it is important to characterize the key processes and taxa that can mediate benthic algal productivity. It appears that sea cucumbers play an important role in benthic community processes on coral reefs through sediment bioturbation, but empirical evidence in the literature is scarce. In Thailand, sea cucumbers are currently threatened by over exploitation; however, the effect of the declining population on reefs
* Corresponding author. E-mail address: [email protected] (V. Viyakarn). https://doi.org/10.1016/j.ecss.2019.106514 Received 2 May 2019; Received in revised form 18 November 2019; Accepted 29 November 2019 Available online 2 December 2019 0272-7714/© 2019 Published by Elsevier Ltd.
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Estuarine, Coastal and Shelf Science 235 (2020) 106514
Fig. 1. Map of study area.
and reef communities was unknown (Bussarawit and Thongtham, 1999). The purpose of this paper was to examine the distribution pattern of Holothuria atra in the upper Gulf of Thailand. Bottom coverage, po tential nutrient content of surrounding sediments, and gut contents were also evaluated and tested to estimate the potential effects of their feeding behavior on the sediment. Field and aquarium experiments were conducted to assess the impact of density and feeding activity on the sedimentary microalgal community.
2.2. Density and habitat structure of sea cucumbers Densities and percent composition of the habitats of Holothuria atra were investigated around the islands located in Chon Buri and Trat Provinces using transect lines and quadrats. A 50-m transect line was extended along the substrate on a flat reef parallel to the shore. A diver then swam along the length of the line and recorded the number of sea cucumbers within 1 m of each side of the line. At least three transect lines were established on each island. Data were then compiled and divided by the total area surveyed (100 m2). The percent composition of sea cucumber habitats was also determined along the transect lines. A quadrat (0.5 � 0.5 m) with string crossed to form 25 points were placed immediately adjacent and on both sides of the transect at 0, 5, 10, 15, 20, 25, 30, 35, 40, 45, and 50 m. In each quadrat, the type of substrate below the points was recorded within the following categories based on Reef Check method: soft coral, hard coral, algae, rock, rubble, sand, or others. Silt was not found in the areas. Once the data were compiled, the percent composition was derived from the fraction of each category over the total 25 points.
2. Methods 2.1. Study sites Data and sample collections took place at 20 islands located in Chon Buri and Trat Provinces in the upper Gulf of Thailand. Most islands located in Chon Buri Province, but Kham and Khram Islands are the most restricted and protected by the Royal Thai Navy. Thus, no harvesting of marine organisms is allowed in these 2 islands. The study sites was shown in Fig. 1. 2
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2.3. Gut content analysis Thirty sea cucumbers were collected in the sand areas of reef flats at Kham Island. The samples were analyzed to determine the gut contents. The composition of stomach contents and bacterial numbers were examined and counted. To determine the composition in the stomach, all contents of each stomach were removed. Then, they were spread in a petri dish with lines crossed to form 100 points. Type of contents found in each point was recorded. For the bacterial count, the small contents from each sea cucumber’s stomach were spread in a petri dish con taining bacterial agar, and allowed bacteria to grow for a week. After one week, the numbers of bacterial colonies grew in each plate were counted under the microscope. To test whether sea cucumbers consume benthic diatoms, an addi tional 20 sea cucumber gut samples of each sea cucumber size, 20 sediment samples from the surrounding habitat, and 20 sea cucumber feces samples were collected for chlorophyll-a analyses. Three sizes of sea cucumbers were collected: small, medium, and large (<10, 10–25, >25 cm in length respectively). Each sample was collected in separated zip-lock plastic bags by divers. Samples and specimens were immedi ately frozen and protected from light when brought to the surface, and remained so until analyzed in the laboratory. In the laboratory, sea cucumber specimens were defrosted, weighed, measured for length and dissected. The gut contents were removed and again frozen and pro tected from light. Pigment extraction and analysis was performed once the samples had been thawed in a dark location. The method of chlorophyll analysis was adopted from Gilpin (2001). For each sample, 5 g was placed in a 30-ml container with a screw on cap. Then, 20 ml of methanol was added and the container stirred vigorously and allowed to stand for 5 min. A 1-ml aliquot of the methanol containing pigments was then extracted with a pipette and placed into a test tube with a top. Then, 9 ml of 90% acetone was added to the solution and the tube wrapped in foil until analysis in a spectrometer. The methanol-acetone solution was poured into a cuvette with a pathlength of 1 cm. After wiping the cuvette thoroughly, it was placed in a spectrometer for measurement. Absorbency readings were made at 665 nm and 750 nm before 4 drops of HCl were added to the sample and the measurements repeated. With each change in wavelength and acid addition, the spectrometer was calibrated with a blank of 90% acetone, containing or excluding 4 drops of HCl, depending on the constituents of the sample. The compiled data was then calculated with the following formulas for the derivation of chlorophyll-a concentration: Chlorophyll-a
Fig. 2. Average densities of Holothuria atra and the sand compositions on twenty reefs in Chon Buri and Trat Provinces.
A field experiment was conducted at Kham Island, Chon Buri Prov ince, to examine the relationship between sea cucumber population density and the biomass of benthic microalgae using chlorophyll as an indicator of benthic microalgal biomass. The uniqueness of this study site is that sea cucumbers are not overexploited, and the site is protected by the Royal Thai Navy. The study site was manipulated by the authors as follows: 1) an area without sea cucumbers; 2) an area where the density of sea cucumbers equaled the average or natural density (approximately 25 sea cucumbers per 100 m2); 3) an area where the density of sea cucumbers is two times higher than the natural density (approximately 50 sea cucumbers per 100 m2). Each area was caged. All experimented sea cucumbers were collected from the area. Because it was decided to measure the actual concentrations of chlorophyll-a with only the density of sea cucumbers as a fixed factor, the densities of other benthic organisms were not manipulated and remained natural. Ten sediment samples in each area treatment were collected one month after the experiment for chlorophyll-a analysis. A one-way ANOVA followed by a Tukey pairwise comparison test was used to test differences in chlorophyll-a concentrations in each area.
[chl] (μg g 1) ¼ 26.7 x (665b–665a) / L 665b: absorbance at 665 nm before acidification minus absorbance at 750 nm before acidification 665a: absorbance at 665 nm after acidification minus absorbance at 750 nm after acidification L: pathlength of the spectrophotometer cuvette used (cm)
3.2. Laboratory experiment A laboratory experiment was conducted to determine the relation ship between the density of sea cucumbers and the density of benthic microalgae in the sediments. All sea cucumbers used in the experiment were collected from Kham Island, and were between 25 and 30 cm in length. Three treatments with ten replicates each were used. All exper iments were conducted in 20-gallon glass aquaria (76 � 31 � 31 cm) at room temperature (30 � C), a salinity of 30 psμ, and in moderate sunlight. The artificial seawater was used, and the seawater was changed daily. Sand used in the experiment was collected from the habitat where sea cucumbers were actually found at Kham Island. Each aquarium con tained one of the following: 1) sand on the bottom and one individual sea cucumber; 2) sand and three individuals; 3) a control with sand and no sea cucumbers. The chlorophyll-a concentrations of sands from each aquarium were measured at the beginning, middle, and end of the
3. Experiments on the relationship between sea cucumber and benthic microalgal densities 3.1. Field experiment From preliminary surveys, high chlorophyll concentrations were detected from gut content analyses; thus, indicating that H. atra con sumes benthic microalgae. Therefore, sea cucumbers may play a role in controlling the biomass of primary production on coral reefs. The hy pothesis was that the density of sea cucumbers influenced the density of benthic diatoms. In this study, when benthic microalgae were referred, it meant benthic diatoms. 3
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5. Discussion The field surveys showed that the highest density of Holothuria atra was recorded at Kham Island, followed by Khram Island. As these two islands are protected by the Royal Thai Navy, fishermen are not allowed
Fig. 3. Percent composition of Holothuria atra habitats on twenty reefs in Chon Buri and Trat Provinces.
experiment. Each time, five replicates were collected in each aquarium. The experiment was run for four days. A one-way ANOVA followed by a Tukey pairwise comparison test was used to test differences between in chlorophyll-a concentrations in each treatment. Fig. 5. Chlorophyll-a concentration in the gut, feces, and in the surrounding sediment of Holothuria atra.
4. Results The field survey results showed that the density of Holothuria atra at Kham and Khram Islands, areas protected by the Royal Thai Navy, were higher than on other unprotected islands (Fig. 2). In addition, difference of the densities of H. atra in each site depended on the substrate composition. The density of H. atra increased with the percentage sand cover within the habitat (Fig. 3). Regarding gut content analysis, results showed that the gut content of small size class (<10 cm in length) of H. atra was composed of sand only, while in medium (10–25 cm in length) and large (>25 cm in length) size classes, both sand and shell fragments were found in the guts (Fig. 4). The average bacterial number recorded in the gut of H. atra was between 10 1 and 10 2 CFU. A comparisons of chlorophyll-a concen tration between in H. atra gut, feces, and the surrounding sediment showed that chlorophyll-a was higher in the gut than in the feces or sediment (Fig. 5). The results from the field experiment showed that concentrations of chlorophyll-a were higher in the control plot containing no H. atra specimens (Fig. 6). However, the area with double the density of H. atra had a lower chlorophyll-a concentration than the other plots. Results of the laboratory experiment showed that the highest chlorophyll con centrations were in the aquaria with more individuals of H. atra (Fig. 7).
Fig. 6. Chlorophyll-a field experiment.
concentration
Fig. 4. Gut contents in different size classes of Holothuria atra. 4
in
sediments
after
a
1-month
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1985). In addition, toward the end of our field experiment, it was observed that the sea urchin, Diadema setosum, was moving into the area (approximately 1 individual/1 m2) where the H. atra population was cleared out. The migration of sea urchins could be attracted by the in crease in the benthic microalgal biomass. Sea urchins usually prefer both micro and macroalgae as food sources (Harris et al., 2001). Neverthe less, other studies have found that holothurians induced nutrient regeneration through excretion of ammonium ions (Uthicke, 2001a, 2001b). Further studies are required to investigate in detail the rela tionship between holothurians, benthic algae, and other benthic reef organisms. The results of this study showed that difference of the densities of H. atra in each site depending on the substrate composition. It was also shown that the feeding activity of H. atra has an impact on benthic microalgal production and biomass in reefs in the upper Gulf of Thailand. Holothurians play an important role in recycling nutrients in reef ecosystems, and the decline in sea cucumber numbers may nega tively affect reef communities.
Fig. 7. Changes in chlorophyll-a concentrations after 2 and 4 days in the lab oratory experiment.
to access, in contrast to the situation around other islands. In this study, the density of H. atra was correlated to its habitat composition. The higher the percentage of sand cover in an area, the greater the density of H. atra. It has been shown that holothurians are distributed mainly on a sand substratum (Mercier et al., 1999). However, some studies indicated that other factors, such as organic matter content of the sediment, water movement, etc. may also influence the distribution of sea cucumbers (Kerr et al., 1993; Mercier et al., 1999, 2000). Holothuria atra is categorized as a deposit feeder. In this study, gut content analysis showed that the gut of the small size class was composed solely of sand, while the guts contents of the larger size classes contained a mixture of sand and mollusk shell fragments. The gut con tents of other sea cucumber species, such as H. scabra, comprised bac teria, copepods, diatoms, algae, molluscan shells, foraminiferans, sand, and muds (Wiedemeyer, 1992; Baskar, 1994; Hamel et al., 2001). In this study, although bacteria were found, the numbers were very low. Thus, bacteria may not be a major food source of H. atra. A comparison of chlorophyll-a concentration in H. atra gut, feces, and surrounding sands showed that the highest concentrations were found in the gut. The results suggested that benthic microalgae may be an important food sources for H. atra. It has been suggested that chlo rophyll-a concentration and bacterial content could be used as in dicators of the nutritional value of sediments for holothurians (Moriarty, 1982). In the Solomon Islands, Battaglene and Bell (1999) observed that cultured juveniles of H. scaba fed on epiphytic algae and bacteria on the substrate. Other studies have showed that H. scaba, can display substrate selection, and was able to distinguish between sediments based on grain size (Wiedemeyer, 1992; Baskar, 1994). However, there is also a pos sibility for sea cucumbers to have their own additional gut flora than in the sediment, which led to higher chlorophyll concentrations in the gut than in the habitat. More studies in depth on the feeding behavior of sea cucumber in a relation to gut contents are needed. Gut content analysis suggested that the feeding behavior of H. atra could reduce the concentration of chlorophyll-a in surrounding sedi ments since the chlorophyll concentration in the feces was lower than the surrounding sediment. Yingst (1976) and Hammond (1983) reported that some diatoms were digested by holothurians. Even though the feces had lower chlorophyll as sea cucumbers digested the algal food, this was not necessarily shown that sea cucumbers reduced the chlorophyll concentration in the environment (Uthicke and Klumpp, 1997). The results of field and laboratory experiments demonstrated that H. atra may play an important role in controlling the density of benthic diatom and epiphytic algae in sediments. The higher the number of sea cucumbers, the greater the capability to reduce the algal biomass in the sediment (Fig. 6). Previous studies have shown that benthic microalgal communities have decreased through the feeding activity by holothu rians (Moriarty et al., 1985; Uthicke, 1999). However, when holothu rians were absent, cyanobacterial mats were observed (Moriarty et al.,
6. Conclusions At present, sea cucumbers are overharvested and their populations have been declined dramatically. This study showed that the increase and decrease of sea cucumber populations can have significantly affected to the benthic microalgal production in the sediment and reef ecosystems. Thus, further studies are needed to investigate in details of the linkages between sea cucumber populations and other benthic reef organisms and their in depth roles in reef ecosystems. Acknowledgements This study was part of the Plant Genetic Conservation Project under the Royal Initiative of Her Royal Highness Princess Maha Chakri Sir indhorn. The authors would like to thank members of the Reef Biology Research Group and the Naval Special Warfare Command, Royal Thai Fleet, Royal Thai Navy for assisting in data collections. This work was partially supported TRF (RSA6080087), PADI Project AWARE grant, PADI Foundation grant, and Mubadala Petroleum (Thailand) Limited. Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi. org/10.1016/j.ecss.2019.106514. References Baskar, B.K., 1994. Some observations on the biology of the holothurian Holothuria (Metriatyla) scabra Jaeger. In: Rengarajan, K., James, D.B. (Eds.), Proceedings of the National Workshop on Beche-De-Mer. Bulletin of the Central Marine Fisheries Research Institute 46. Indian Council of Agriculture Research, Cochin, India, pp. 39–43. Battaglene, S.C., Bell, J.D., 1999. Potential of the tropical Indo-Pacific sea cucumber Holothuria scabra for stock enhancement. In: Howell, B.R., Moskness, E., Svasand, T. (Eds.), Stock Enhancement and Sea Ranching. Blackwell Science, Oxford, pp. 478–490. Bruckner, A.W., Johnson, K.A., Field, J.D., 2003. Conservation strategies for sea cucumbers. Can a CITES Appendix II listing promote sustainable international trade? SPC Beche-de mer Inf. Bull. 18, 24–33. Bussarawit, S., Thongtham, N., 1999. Sea cucumber fisheries and trade in Thailand. In: Baine, M. (Ed.), Proceeding of the International Conference “The Conservation of Sea Cucumber in Malaysia, Their Taxonomy, Ecology and Trade”. Hariot-Watt University and Fisheries Research Institute, Kuala Lumpur, pp. 26–36. Conand, C., 2001. Sea cucumber retail market in Singapore. SPC Beche-de-mer Inf. Bull. 14, 12–14. Gilpin, L., 2001. Marine Biology: methods for analysis of benthic phytoplankton pigment. Available at: www.oaerre.napier.ac.uk/users/p.tett/MB/benchl01.html. Hamel, J.F., Conand, C., Pawson, D.L., Mercier, A., 2001. The sea cucumber Holothuria scabra (Holothuroidea: Echinodermata): its biology and exploitation as beche-demer. In: Southward, A.J., Tyler, P.A., Young, C.M., Fuiman, L.A. (Eds.), Adv. Mar. Biol. 41, 129–223.
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