Laboratory Culture of the Soft-Coral Carijoa riisei.
In 1972, Carijoa riisei, a shallow-water azooxanthellate octocoral, was discovered invading the fouling community in Pearl Harbor. Invasive populations of C. riisei are now so dominant that they are considered a grave threat to Hawaii's native reef ecosystems and economy. C. riisei's juvenile and reproductive biology is not very well understood because of the difficulty of field observations at the depth that C. riisei inhabits. A laboratory based culture technique would represent a significant advancement in our ability to investigate questions involving the spread and control of C. riisei. Flow-through aquariums were set-up to house captive C. riisei colonies over a period of seven weeks. C. riisei colonies were successfully grown on freeze-dried copepods and exhibited release of gametes. The success of this culturing method bodes well for long-term maintenance of C. riisei and further experiments with C. riisei biology.
IntroductionCarijoa riisei was discovered invading the fouling community in Pearl Harbor in 1972 (Evans et al. 1974). C. riisei, which is native to the tropical Western Atlantic and Caribbean, is described as a fast growing, shallow-water octocoral (Bayer 1961; Rees 1969) (Fig. 1). Since its initial discovery, C. riisei has spread rapidly throughout the main Hawaiian Islands and is now commonly observed on hard substrata in low-light habitat with moderate current flow (Thomas 1979; Devaney and Eldredge 1977; Coles and Eldredge 2002). Invasive populations of C. riisei are now so dominant that it is considered a grave threat to Hawaii's native coral populations (Thomas 1979; Coles and Eldredge 2002; Grigg 2003, 2004; Kahng and Grigg 2004).
The extent of the C. riisei invasion became apparent from 2001-2004 deep water surveys of the Au'au Channel between the islands of Maui and Lanai. These surveys recorded C. riisei overgrowing at least fifty percent of the indigenous black coral colonies (A. dichotoma and A. grandis) at depths below seventy meters (Grigg 2003; Kahng and Grigg 2004) (Fig. 2). C. riisei has also been observed overgrowing large fields of scleractinian plate corals (Leptoseris sp. and Pavona sp.) in the lower photic zone (Grigg 2003, 2004; Kahng and Grigg 2004). This overgrowth has generated great concern because the Hawaiian black corals A. dichotoma and A. grandis are commercially valuable species used for the manufacture of precious coral jewelry. Black coral is the official gemstone for the state of Hawaii and supports a $30 million state-wide precious coral industry (Grigg 2004). Clearly, the growing dominance of C. riisei now threatens the black coral industry in Hawaii, which suffers mortality from both competition and harvesting.
The threat from C. riisei to black coral arises from its ability to attain densities greater than 1600 axial polyps per square meter. This dense growth can saturate the substrata to the exclusion of native fauna. For example, C. riisei routinely smothers cup corals and bivalves in shallow water, which demonstrates a potential for high ecological impact wherever it finds a favorable habitat. Because C. riisei lacks zooxanthellae, it is an obligate predator of zooplankton which it filters from the water column. C. riisei has a voracious appetite which helps fuel a growth rate greater than 1 cm per week (Bayer 1961; Rees 1969). This high growth rate yields densities much higher than the indigenous fauna it replaces, significantly altering the ecosystem in favor of C. riisei.
The invasion of Hawaii's coral reef community by C. riisei is a unique and devastating event. Of the 287 nonindigenous marine invertebrates identified in Hawaii, most occur within the major harbors and with little proliferation on coral reef communities (Coles and Eldridge 2002; Eldredge and Carlton 2002; Demopoulos 2004). In terms of biomass and displacement of native species, C. riisei appears to be the most threatening invasive and nonindigenous marine invertebrate in Hawaii (Kahng and Grigg, in review). The invasion by C. riisei is particularly interesting because in many coral reef ecosystems, especially in the Caribbean and the Indo-Pacific, shallow-water octocorals form a major and sometimes dominant faunistic component (Bayer 1961, Dinesen 1983; Spalding 2001). However, Hawaii's native shallow-water octocoral fauna consists of only four species: Sarcothelia edmonsoni (Family Xeniidae), Sinularia densa (Family Alcyoniide), Sinularia molokaiensis (Family Alcyoniide) and Acabaria bicolor (Family Melithaeidae) (Devaney and Eldredge 1977; Grigg and Bayer 1976; Hoover 1998). Compared with other taxa, the diversity of shallow-water octocorals in Hawaii is anomalously low. This low diversity and abundance of octocorals may contribute to Hawaii's susceptibility to C. riisei. Unfortunately, because of Hawaii's unique environment, the susceptibility may also apply to other nonindigenous octocoral species should they be transported to Hawaii.
The reproductive and juvenile biology of Carijoa riisei is not well understood. A full understanding of the growth and reproduction system of C. riisei is needed before a viable management plan can be developed and implemented. However, it is difficult to monitor these characteristics in situ because of the depth at which C. riisei lives. Consequently, a laboratory based culture technique for C. riisei would represent a significant advancement in our ability to investigate this important invasive species and to address questions that have been difficult to answer from field studies. This work presents the development of one such technique and preliminary data to evaluate its efficacy.
METHODS AND MATERIALS
Collection of Carijoa riiseiC. riisei colonies were collected from three sites. Four colonies were obtained in 1m of water under a bridge at Hawaii Kai, Oahu. Three colonies were collected from the YO-257 shipwreck in 100ft of water at Kewalo Basin, Oahu. The final colony was collected in 0.5m of water from beneath the Sunken City' in Kaneohe Bay, Oahu.
Aquarium Set-upC. riisei were first housed in recirculating aquariums for initial acclimation and observations. After the initial observation period of one week C. riisei were transferred to a 20 gallon flow-through system (Fig. 3). Coarse-filtered seawater pumped from Kaneohe Bay was used to both fill the tanks and for the flow-through system at a rate of 72 gallons per hour. To allow for easy observation and cleaning, the tanks were kept bare-bottomed. Flow was augmented in the aquaria using adjustable powerhead pumps (M1500, SEIO).
Addition of Carijoa riiseiC. riisei was permanently affixed to the skeleton of Fungia scutaria with thick-gel superglue (cyanoacrylate); this creates a base to which C. riisei could adhere, but still allowing movement of the entire colony. Easily identifiable features of the skeleton base were subsequently used for unambiguous colony identification of C. riisei. Because C. riisei is prone to colonization by other organisms, including the predatory nudibranch Phyllodesmium poindimiei (Kahng and Grigg 2004), large fouling organisms were removed to avoid water quality degradation from death or decomposition. A kole tang (Ctenochaetus strigosus) was also added to the tank to help reduce algae growth. Colonies of C. riisei were kept in the recirculating aquarium (as described above) for one week to allow any fouling organisms that were not removed to crawl off or die.
To help acclimate C. riisei to captive conditions, colonies were moved after the first week and re-orientated until full polyp extension and feeding were observed. Individual colonies of C. riisei were then separated and placed into individual 20 gallon flow-through aquaria.
Each C. riisei colony was fed frozen and freeze-dried copepods twice a day at 10 am and 10 pm respectively. Three grams of food per colony were added to each aquarium. Copepods were suspended in sea water for 5 minutes before feedings to allow for rehydration. To avoid mechanical removal of food, the protein skimmer and inflowing seawater were turned off for 45 minutes. The power heads were left on to circulate the food. Ingestion of copepods was confirmed by visual observation using a stereo microscope (SMZ-U, Nikon).
Gamete Bundle Collection
Material that had settled out of the water column was collected from the bottom of aquaria and examined under stereo microscopy for gamete bundles. Bundles were then removed and stored in 5% formalin for sex determination at a later date. A 200 m mesh filter was also placed on the sea water exhaust of each aquarium to collect any gamete bundles that had stayed suspended in the water column (Fig. 4).This filter mesh was removed daily, washed with filtered seawater, and the filtrate was examined under stereo microscopy for gamete bundles. After initial acclimation to the culture system, gamete bundles from each colony were searched daily over a period of four weeks.
Culture of Carijoa riisei
Consumption of Cyclop-eeze (Argent Labs) brand freeze-dried copepods by Carijoa riisei was confirmed visually by stereo microscopic observation of polyp feeding. All C. riisei colonies showed lateral axial polyp growth. In particular, two colonies from the YO-257 shipwreck site grew beyond the Fungia base and made new attachments to aquarium walls. (Fig. 5) For the remaining colonies, it could not be determined if axial polyp growth represented new growth or repositioning of colony resources because biomass could not significantly determined before and after culture trials
Gamete Collection from Carijoa riisei
Collected gamete bundles were approximately 400-500 size m in diameter (Fig. 6). Gametes collected over a four week time period suggest that C. riisei cultures can be maintained in reproductive condition for several weeks at a time. These results suggest that C. riisei cultures maintained as described herein are viable for long-term captive observation and experiments.
Culture of Carijoa riisei
Finding a proper food source for C. riisei is the primary hurdle for the long-term maintenance of C. riisei cultures in the laboratory. Because freeze-dried copepods represent an easily obtainable and relatively cheap source of food, the success with the short-term maintenance and growth of C. riisei cultures using Cyclop-eeze freeze-dried copepods bodes well for long-term maintenance of C. riisei cultures. Having a cheap, easily obtainable food helps maintain laboratory cultures of animals over a long period of time.Carijoa riisei Gamete Collection
The extended period of time over which gametes were collected suggests that C. riisei was sexually reproducing and releasing gametes naturally. Any short-term spawning may be a result of stress-induced spawning, but prolonged release of gametes is much more likely to represent a natural reproductive process. Gamete bundles were not found in the sediment removed from the bottom of the tank throughout the experimental period. Collection of gametes from only the aquarium overflow suggests that gamete bundles from C. riisei stay in the water column and do not settle out rapidly at the flow regime maintained in these cultures.
Cultures of C. riisei can be maintained in artificial lab environments on a commercially available diet of freeze-dried copepods. Colonies in culture fed on a daily basis, and released gametes continuously over a four-week period. The success of this culturing method bodes well for long-term maintenance of C. riisei and further experiments with C. riisei biology in the laboratory setting. This culture method may also be applicable to other zooplankton-feeding soft corals.
The authors gratefully acknowledge support from DOD and NSF through NSF grant #02-43600 and the University of Hawaii Sea Grant College Program. Additional funding came from Hawaii Coral Reef Initiative (HCRI) and Hawaii Invasive Species Council (HISC) research grants. The author also thanks Dr. Michael Cooney, Dr. Brian Bowen, Daniel Wagner, Greg Concepcion, Melissa Skillings and Tracy Campbell for editing and support.
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