E Behavioral Interactions Between Copepods and Six Species of Sea Anemones Found in the Monterey Bay Region Mark R. Colville Hopkins Marine Station of Stanford University Pacific Grove, California Running Title: Behavioral Interactions Between Copepods and Sea Anemones Send all proofs and correspondence to: Behavioral Interactions Between Copepods and Sea Anemones. Abstract The behavioral interactions between copepods (species?) and six species of anemones (Tealia lofotensis, Tealia crassicornis, Tealia coriacea, Anthopleura elegantissima, Anthopleura xantho- grammica, and Epiactus prolifera) were examined. Lab and field studies over a six week period showed that the copepods feed on anemone mucus and are dependant on the anemone for survival. Mature copepods were found to inhabit anemone species specifically. It was also found that copepods showed species specific zonation on the anemone; occupying areas where their color matched that the anemone. Adult copepods failed to visually or chemically cue to anemones. Introduction Copepods were observed to be ectocommensal with six species of anemones; Tealia lofotensis, Tealia crassicornis, Tealia cori- acea, Anthopleura elegantissima, Anthopleura xanthogrammica, and Epiactus prolifera. (Hand, 11956). Observations of copepods in the field and in the lab raised several questions concerning anemone- copepod behavioral relationships. What are the advantages that may accrue to the host and 1) symbiont from this relationship? 2) Are copepods protected against nematocyst discharge from anemones of a species other than their host; and are they protected from other anemones of the same species as their host? shavioral Interactions Between Copepods and Sea Anemones Are there particular zones of the anemone that the copepods inhabit species specifically? 4) Do the copepods display any host recognition through chemical or visual cues? Each question was approached in a series of experiments. The results of all these experiments will be discussed together in an attempt to present a broad overview of the behavioral rela- tionship. Methods and Materials All animals were collected in the Monterey bay area during the month of May, 1975. The locations of the collection sites are shown in figure 1. Anemones were removed from the substrate with a scraper, and sealed in seperate jars to prevent the escape of copepods during transport back to the lab. All anemones were kept in acquaria in the lab, with a constant flow of fresh seawater. Only anemones that were attached, expanded and sensitive to stimuli were used in lab experiments. Field Observations able 2 lists the different types and locations of anemones collected with copepods. Copepods generally were more abundant on subtidal anemones, and were never found on anemones above mean lower low water mark. Table 2 also describes the types of copepods found on the different anemones. The species of the copepods are not known, but are designated as distinct by size and color. Table 2 shows the percentage of anemones with copepods at the different locations. Behavioral Interactions Between Copepods and Sea Anemones The average number of copepods observed on inhabited anemones is also shoun. Lab Observations All copepods observed appeared to be cyclopoids (Brady, 1880). The copepods appeared fully developed and no larvae were observed during the course of the six week experiment. The copepods are poor swimmers with legs adapted mainly for crawling on the surface of the anemone. As copepods crawl over the surface of the anemone, the tentacles move slightly in response to the tactile stimulus, but no feeding response is evoked. The large 2mm copepods are capable of rapid movement, but move infrequently; remaining stationary for up to 2 hours. The 1 mm copepods are characterized by rapid movements over the entire anemone surface, rarely remaining stationary for more than a minute. Experiments: Advantages to Host and Copepod To determine if the copepods eat anemone tissue or mucus, excised tentacles were stained in methylene blue for 40 minutes until both the tentacle and surrounding mucys were dark blue. A white copepod placed on the stained preparation in fresh seawater for 24 hours. was At the end of the 24 hour period, the copepod's gut had turned a pale blue, indicating stained material had been injested. Squash preparations copepods revealed no signs of nematocysts in the copepods. To defermine if copepods are dependent upon the anemones for sur¬ vival, six copepods were kept isolated from anemones in fresh seawater. All six copepods died within 10 days. Two of the six died within three days. Dead copepods were never observed on anemones or in tanks con¬ taining commensal anemones. Other data indicate dead copepods are not eaten by the host. Behavioral Interactions between Copepods and Sea Anemones Lab and field observations were conducted in an attempt to determine if the anemones benefited or were harmed by the presence of copepods. tab observations were made for capture and injesting of copepods by anemones, and dead copepods were fed to anemones to see if they would pe eaten. Visual comparisons were made in the field between anemones with and anemones without copepods. There were no observed differences in appearance between commensal and noncommensal anemones. Anemones were never observed to capture their copepods, or copepods that had died and were fed to the anemone. Copepods that had been crushed were quickly captured and injested. Feeding experiments were conducted to determine if the anemone's feeding behavior was modified by the presence of copepods, and to ob- serve copepod behavior during anemone feeding. It was found that the presence of copepods did not hinder the anemone's ability to capture other prey the size of copepods. Tigriopus were instantly captured and injested by the anemones. When anemones feed of large prey, copepods are observed to move away from the area where the prey is captured. At no time were copepods seen on the captured food, or going into or out of the coelenteron. Copepods that had been coated with limpet juices were captured and injested by the anemone. 2) Extent of Copepod Immunity Copepods found on the same anemone species appeared to be species specific. Several transfers of copepods among anemones of the same species as the host and among anemones of different species were conducted. It was found that copepods from all six different species of anemones were able to be transferred to other anemones of the Behavioral Interactions Between Copepods and Sea Anemones same species without harm to the copepods. Anemones from the same or different areas did not react to intraspecific transfers of copepods, even if they were previously noncommensal. Anemones did react to interspecific transfers of copepods, with the excep- tion of copepod transfers made between Anthopleura elegantissima and Anthropleura xanthogrammica. In all other transfers, the anemone elicited a vigorous feeding response toward the copepod, and the cope- pod displayed escape behavior. Species Specific Zonation Preliminary field observation of Tedia lofotensis and T. crassi- cornis indicated that copepod species were located where their color matched that of the aneomone. Seven T. lofotensis were collected and observed in the lab for a period of three weeks. The anemones were divided into five zones (actinopharynx, oral disc, tentacle base, tentacles, and column) and copepods of each type were counted and their locations recorded. Because T. crassicornis did poorly in the lab, zonation studies were done in the field. Twenty T. crassicornis were observed on two separate dives. Because of inaccuracy in counting copepods while diving, the anemones were divided into only three zones (oral disc, tentacles, and column). The results are summarized in Tables 3 and 4. They show that T. lofotensis, the red copepods are found primarily on the tentacles, were they are the same color as the anemone. White copepods show a preference for the tentacle base area and the tentacles, but show no preference between the two areas, which are of different colors. In the lab, seven red copepods were placed on the oral disc of a lofotensis, which is a brownish-green in color. Within 30 minutes five had moved to the tentacles, which are red, and two had moved onto Behavioral Interactions Between Copepods and Sea Anemones the pharynx, which is also red. The copepods maintained these positions for two hours, when the experiment was terminated. 4) Cueing Towards the Anemone In the field it was noted that often an anemone with few or copepods could be found less than three feet away from an anemone with over 50 copepods. This observation led me to ques¬ tion the method by which copepods spread to new anemones, and how they come to inhabit anemones only of a particular species. Four T. lofotensis were placed in seperate tanks that were entirely white so any movements of copepods off the anemones could be easily observed. Copepods were then placed in the tank at distances from one to seven centimeters away. The time taken for copepods to move to the anemones was recorded. When placed in the tanks, copepods appeared to swim in a random manner, or else to merely settle to the bottom of the tank. there appeared to be no orienting behavior unless the copepod was brought within .5 cm. of the anemone or actually touched to the anemone. If brought this close, the copepod would quickly move to the anemone. In another experiment, a commensal T. lofotensis was placed cm. away from a. noncomménsal. Observations were made daily for three weeks to determine if any copepods had moved to the copepodless anemone. It was found that copepods never spread to the uninhabited anemone, nor were copepods ever observed swimming off of the comménsal anemone. Behavioral Interactions Between Copepods and Sea Anemor Discussi Copepods appear to be commensal on the anemones. Experiments finding that copepods injest anemone mucus, and die when isolated suggest that the copepods could be dependent upon the anemone mucus for sur¬ vival. The anemones, on the other hand, do not appear to benefit nor be harmed in any way by the presence of copepods. Being unable to capture and eat these copepods, the only benefit the anemone might derive from the relationship would be a removal or cleaning of mucus. It appears that copepods do not directly eat the food an anemone captures. Copepods do not travel into the coelenteron, and if they become coated with the fluids of a captured prey, they may be stung and captured by the anemone. The behavior of the anemone is not appreciably modified by the presence of copepods. Although sensitive to the tactile stimulus of the copepod, the anemone appears unable to discharge nematocysts, and does not display the vigorous movements characteristic of a feeding response. The anemone is not, however, unable to capture all animals the size of copepods; quickly siezing and injecting animals of similar or even smaller size. These copepods show both species specific habitation and immunit, except when associated with the genus Anthopleura, where the same copepods are immune to both species. It seems interesting that the two species of Anthopleura seem more alike in terms of copepod immunity than the three species of Tealia. The copepods found on A.elegantissima and A. xanthogrammica were indistinguishable from each other and were immune on either Anthopleura species. The three Tealia species, however, Behavioral Relationships Between Copepods and Sea Anemones were highly specific to seperate types of copepods. Comparisons of mucus from the different anemones could shed further light on this problem. It was also interesting to note that copepods were immune to all anemones of the same species, even when collected from widely different areas. (Carton, 1963. Bouligand, 1966) t was discovered that interspecific transfers of copepods elicit dual recognition of foreignness by both the anemone and the copepod. An anemone, when touched by a foreign copepod, immediately exhibits a vigorous feeding response, yet nematocysts do not discharge. The anemone somehow 'recognizes" the copepod as prey, yet the cue causing nematocyst discharge is lacking. The copepod also exhibits a swimming avoidance behavior away from the anemone. This swimming behavior was never seen by copepods on their host anemone. Experiments using ex¬ cised anemone tentacles show that copepods will cling tenaceously to tentacles of their host, yet will remain indifferent towards tentacles of a different anemone species. It appears then, that the copepod's avoidance behavior is a response to the anemone's sudden movements, and not avoidance of the anemone tissue itself. Copepods living on Tealia lofotensis and Tealia crassicornis showed a preference for zones in which their color matched that of the anemone. jecause white copopods inhabiting T. lofotensis are not camouflaged anywhere on the anemone, one would not expect these copepods to prefer any particular zone. Table 3 shows that the white copepods show an equal preference for either the tentacle bases or the tentacles of the of the anemone, which are brown-green and red respectively. This color blending of copepods to anemones was shown in the other anemones as well. Copepods on all six species of anemone were generally found to be the same color as the anemone. Copepods with blue egg sacs were found on blue Epiactus prolifera, light green copepods were Sehavioral Interactions Between Copepods and Sea Anemones found on subtidal A. elegantissima, and white copepods were found at the white bases of T. coriacea tentacles. The manner in which the copepods assumed the color of the anemone was not determined. No larvae were observed in lab, so their color is not known. It was shoun, however, that the copepods cannot change color within one week. Red copepods that were either isolated or placed on white tentacles never showed a color change. Copepod predation was never observed in the field but certain fish and crustacea are often in close association with anemones. This color matching might afford the copepods some prote tion against these predators. (Ross, 1974) It is intriguing that none of the copepods worked with display visual or chemical cueing towards an anemone at a distance greater than .5 cm. This would indicate that the spread of copepods to new anemones is accomplished during another stage in the copepod's life cycle. (Dudley, 1966). It would be interesting to know the method in which the copepod larvae seek out specific species of anemones. Sehavioral Interactions Between Copepods and Sea Anemones Acknowledgements wish to thank Chuck Baxter for his helpful criticism during the writing of this paper and a special thanks to all the were so quick to offer help when it was needed. Behavioral Relationships Between Copepods and Sea Anemones Literature Cited Recherches Recentes sur les Associes aux Bouligand, Y. 1966. Anthozoaire. p.267-306. In Rees, W. J. (ed.) The Cnidaria and their Evolution. Proc. Symp. Zool. Soc. Lond. 16. Academic Press, N.Y. Brady, George Stewardson. 1880. A Monograph of the Free and Semi-parasitic Copepods of the British Islands. Roy. Soc. London. Carton, Y. 1963. Etude de la Specificite Parasitaire chez 1. Actinae (copepoda cyclopoida). C. R. Sebd. Seanc. Acad. Sci. Paris. P. 256. Dudley, P. 1. 1966. Developement and Systematics of Some Pacific Marine Symbiotic Copepods. The Biology of the Notodelphy- idae. Univ. Wash. Publ. Biol. 21: vt. 288p. Hand, Cadet H. 1956. The Sea Anemones of Central California. Wasserman Journal of Biology, v.12. no.3. v.13 no. 1-2. Ross, D. M. 1974. Behavior Patterns in Associations with Other Animals. p.281-312. In Muscatine and Lenhoff (eds.). Coelenterate Biology: Reviews and New Perspective. Academic Press, N. Y. c ure Legend 1) A map of the Monterey bay region, showing the five collection sites. 3) 4) Table Legend Table showing zonation behavior of red and white copopods on lealia lofotensis. When the anemones were fully expanded no copepods were observed on the column. No counts were made when the anemones were closed. Zonation behavior of copepods on Tealia crassicornis. Only anemones that were fully expanded were counted. 5. Pescadero Pt. Pacific Ocean S MONTEREY P t. Pinos .Hopkins Marine Station P.Whari Carmel FIGUR ILA 1./ Sewer Outfall Monterey 5 80 98 e a08 DO O 0 0 .9 0 oco 801 60 50 0 aaaaaaa- a kaaaaaaa H O 9 P 8 O d 9 O3o F 100 Copepod I red pe Location pharynx as entacl base tentacle colum tota. pharynx oral disc tentacle base tentacles column total Observe Cope red HP Location pharynx and oral disc tentacles column total No. Observed 294 of Total 81.2