2. Predators of the High Intertidal Harpacticoid Copepod Tigriopus californicus Tigriopus californicus, a harpacticoid oopepod, occurs in great numbers in high rocky tide pools along the Pacific coast from Vancouver Island, British Columbia, to Baja California (Vittor, 1971). Predation on Tigriopus has not been previously studied, and as of this date no important predators are known (Vittor, 1971). The present paper presents an investigation of some possible predators of Tigriopus. A preliminary survey of tide pools in the vicinity of the Hopkins Marine Station of Stanford University at Pacific Grove, California and Natural Bridges State Park, Santa Cruz, California, beginning in the highest pools and working down to the lower pools, revealed the following list of possible predators: the waterboatman Trichocorixia sp. (Insecta, Hemiptera), the crab Pachygrapsus crassipes, the hermit crab Pagurus samuelis, the barnacle Balanus glandula, Gammarid amphipods, isopods, the bivalve Mytilus californianus, the sea anemone Anthopleura elegatissima, the cottid fish Clinocottus analis, and the shrimp Heptacarpus paludicola. Materials and Methods Possible predators and Tigriopus were collected in April and May, 1977, at Mussel Point, Point Pinos, and Asilomar State Beach, Pacific Grove, Califor- nia, and maintained in either running sea water or in pans of standing sea water at temperatures f approximately 15 to 22 90 until they were used in experiments. Each potential predator species was tested to determine if it fed on adult Tigriopus or its larvae. The basic experimental procedure for testing predation on adult Tigriopus involved counting out 75 Tigriopus and placing them in filtered sea water in a four inch fingerbowl. No gravid females were used, so no hatching of larvae could occur. One to four specimens of the possible predator species being tested, starved for one to two days, were added to each bowl. Observations were then made on the feeding methods of the predators. At the end of 24 hours the predators were removed, the number of living and dead Tigriopus remaining were counted, and the predator's gut contents and fecal pellets were examined. The control for each set of experiments was a fingerbowl with 75 Tigriopus in filtered sea water. Tigriopus larvae were obtained by removing the egg sacs from gravid females and allowing them to hatch in filtered sea water. Larvae were then added to fingerbowls containing possible predators and observations were made to determine the methods, if any, used to capture Tigriopus larvae. After 21 hours the gut contents and fecal pellets of the predators under study were examined for Tigriopus fragments. Results and Discussion The control populations for the earlier experiments occassionally showed an increase in number. This is due to the separation of copulating pairs which in these earlier tests were counted as one individual, or possibly due to inaccurate counting. The predators are considered below in the order of their distribution in the intertidal zone, starting with those found in the highest pools. Trichocorixia The waterboatman Trichocorixia was found in the flocculant detrital material of the very high tide pools. In natural situations Trichocorixia can cling to the bottom. To simulatethis natural condition I placed sterile sand or cotton in the bottom of the fingerbowl; these materials could be grasped by the corixids with the tarsal claws of their second legs (Pennak, 1953). In all of the runs there was a small decline in the number of Tigriopus at the end of the experiment (Table I). While under observation the corixids did not feed. Before feeding, the guts of the corixids did not appear red, while after feeding the guts of some did appear red. An examination of the reddish gut contents did not reveal any recognizable Tigriopus parts, However, the reddish-orange oil droplets in the corixids did look similiar to those in Tigriopus. It is possible that the corixids only suck the internal contents out of Tigriopus with their stylets. However, no empty Tigriopus exoskeletons were found at the ends of experiments. At Hopkins Marine Station Tigriopus are maintained in an outdoor con- crete pool about h.5 meters in diameter. They are fed "TetraMin" fish food. In the flocculant material on the bottom of this Tigriopus pool some red decomposing pieces of the "TetraMin" fish food were found. The corixids used in most of the feeding experiments were captured in this pool. The gut contents of many freshly captured specimens was red. Corixids starved for several days were fed fresh red "TetraMin", or red "TetraMin" which had been allowed to sit in filtered sea water for two days, to determine if the fish food was responsible for their red gut coloration instead of Tigriopus. The results of these experi¬ ments (Table II) indicate that the fresh "TetraMin" is not responsible for the red gut coloration; only one of five corixids tested with the decomposing "TetraMin" had gut contents similiar to those of corixids just taken from the concrete pool. The reddish-orange oil droplets were also present in the freshly collected corixids' guts. Usinger (1956) reports that corixids do not subsist on the flocculant bottom deposits but eat animal food such as larvae; the corixids sweep larvae into their mouths with their first tarsi (Pennak, 1953). Trichocorixia were not observed eating any larvae, but their gut contents were occassionally red after feeding. No larval fragments were found in the corixids' guts. The results of these experiments do not clearly indicate that Trichocorixia either is or is not a predator of Tigriopus. 5 Pachygrapsus crassipes This crab is a common inhabitant of high tide pools and one of Tigriopus's main predators. The crab ate from 2.6 to 36% of the adult Tigriopus presented to it in feeding experiments (Table III). Fragments of adult copepod exoske¬ letons were found in the crab's feces several hours after the feeding experi¬ ment. Pachygrapsus uses its chelae to pick up Tigriopus and bring them to its mouth. It also uses its chelae to scoop dead Tigriopus into its mouth. In its natural environment Pachygrapsus is an omnivore (MacGinitie and MacGinitie, 1968). Its main food source is matted and encrusting algae which it scrapes from rocks with its chelae; detrital material left by the receding tides, and living intertidal organisms are its secondary food sources (Hiatt, 1948). Pachygrapsus is able to ingest some Tigriopus larvae. Fragments of larval exoskeletons were found in the feces. However, the larger the crab gets, the more difficult it is for the crab to pick up the larvae. The reverse respiratory current created by the bailers on the second maxillae and used to clean the gills may carry the larvae directly to the mouth, allowing the crab to feed without the use of its chelae. After a feeding experiment the crabs involved were placed in 702 alcohol and their bodies and the alcohol searched for adult Tigriopus. Of those copepods found, some appeared to have been inside the gill chamber, current. possibly sucked in with the respiratory/ Others were apparently clinging to the hairs on the legs of the crabs. Pachygrapsus often visits high tide pools and is responsible for transportingat least a few Tigriopus from one pool to another. Pagurus samuelis This hermit crab is a scavenger of the high tide pools (MacGinitie and MacGinitie, 1968) and is another predator of Tigriopus. (Table IV). Adult Tigriopus exoskeletons and fragments were found in the feces after the feeding experiments. Pagurus was seen to pick Tigriopus off the bottom and the sides of the fingerbowls with its left cheliped and then bring them to its mouth. Here they are passed to the maxillipeds (Makaro, 1938). Pagurus is unable to pick up the larvae with either cheliped; no larval exoskeleton fragments are found in either the fecal pellets or the gut. Tigriopus was observed to crawl all over Pagurus's body and sometimes to go under the shell covering the gill chamber of the crab. In some cases Tigriopus came out after the hermit crab was placed in 70% alcohol. Pagurus could provide another means of transport for Tigriopus between tide pools, but it does not move between high tide pools as much as Pachygrapsus. Balanus glandula This barnacle was collected on the shells of the mussel, Mytilus californianus. Preliminary experiments showed that adult Tigriopus are too large for the barnacle to catch with its branched cirri. The barnacles were observed while in a fingerbowl with larvae, or with a mixture of larvae, the diatoms, Phaeodactylum tricornatum and Nitzschia longissima, and the flagellates, Monochrysis lutheni and Isochrysis galbara. The phytoplankters were added to stimulate filter feeding. The barnacles used in feeding experiments were placed in fingerbowls after they had been out of water for 18 hours. A small current was created with a pipette to induce them to feed. The cirri occassion¬ ally caught a Tigriopus larvae and retracted into the shell. However, examin- ation of the gut contents of barnacles exposed to copepod larvae did not reveal any larval Tigriopus exoskeleton fragments. Possibly the larvae are rejected at the mouth and released. At any rate, Balanus glandula is not a predator of Tigriopus. Gammarid Amphipods and Isopods Preliminary observations showed that gammarid amphipods could be predators of Tigriopus while the isopods are not. The amphipods and isopods were too scarce in the high tide pools to use in feeding experiments, and are not common enough to be major predators of Tigriopus. Mytilus californianus Preliminary experiments with Mytilus californianus revealed that it is incapable of capturing adult Tigriopus (Table V). Mussels feed by a ciliary- mucoid mechanism on the gills. The small frontal cilia on the gills move the food particles and mucus down the lamelleof the gills bringing them to the ciliated food grooves where both the food and the mucus are moved foward to the palps for sorting. The palps reject the larger and heavier particles which go to form pseudo-feces, while they accept the smaller or lighter parti¬ cles and pass them to the mouth (Meglitsch, 1972). Dissected living specimens of Mytilus with one valve removed were made to determine if the gills could pick up and transport the larvae. The larvae were pipetted directly onto the gill surface and observed to be transported down the gill lamella to the food groove and then carried to the palps. Following this, the same preparation of diatoms, flagellates and Tigriopus larvae that was used to feed Balanus was presented to several healthy Mytilus from 2.6 cm to 7.6 cm long. After the mussels had been allowed to feed for at least 24 hours, their pseudo-feces, fecal pellets, and gut contents were all examined for Tigriopus fragments. The pseudo-feces contained both live Tigriopus larvae (struggling in the mucus), and some empty larval exoskeletons which could have been molted exoskeletons. Larval exoskeletons were found in the stomach and feces of Mytilus. When Mytilus was fed only Tigriopus larvae, live larvae were found in the psuedo-feces and larval parts were found in the feces. Anthopleura elegatissima Anthopleura elegatissima captures Tigriopus with the nematocysts of its tentacles. Once the Tigriopus are stung with the nematocysts, they stick firmly to the tentacles or the tentacles wrap around them. The anemone slowly transports the tentacles with the Tigriopus to the mouth where they are ingested. Anthopleura consumed from 5 to 87% of the Tigriopus presented to it. Tigriopus exoskeletons were found in the anemone feces. Once Tigriopus became acclimated to the experimental environment with the anemones, they generally avoided contact with the anemone tentacles; they climbed around the base and the mouth areas of the anemones without being caught by the tentacles. Their behavior suggested they were receiving chemical cues from the anemone. Anthopleura is able to catch larvae only if they are suspended in the water column. The anemone itself does not create a current that suspends the Tigriopus larvae in the water. The larvae are small and the anemone is not able to sting and grasp them as well as they catch the adult Tigriopus. Clinocottus analis These cottid fish are voracious predators on Tigriopus. They fed rapidly by sighting, moving towards, and snapping at Tigriopus as soon as the latter were introduced to the fingerbowl. One fish ate all 75 Tigriopus within 3.75 hours (Table VII). The fish fecal pellets were red and contained Tigriopus fragments. The fish do not actively seek Tigriopus larvae, but they eat them if they are sucked in with the respiratory current. The fecal pellets contained Tigriopus larval fragments at the end of the experiment. Heptacarpus paludicola This carnivore (MacGinitie and MacGinitie, 1968) is found at the very lower limit of Tigriopus's intertidal range. It is an active predator of Tigriopus. The shrimp ate an average of 5.6% of the 75 Tigriopus in each experiment (Table VIII). Heptacarpus uses its first and second chelipeds to pick up Tigriopus. It holds the copepod with its legs and chelipeds while it brings them to its mouth. The shrimp fecal pellets were red and contained Tigriopus parts. The shrimp capture and eat the Tigriopus larvae using the same method. In early experiments it appeared that the smaller Heptacarpus ate more Tigriopus than the larger ones. An experiment was set up to test the relation- ship between shrimp size and its feeding rate on Tigriopus. Heptacarpus were measured from the tip of the tail to the end of the rostrum and divided into eight size classes over the body length range of 1.2 to 2.7 cm. One to three predators were placed in a fingerbowl with 75 Tigriopus for 24 hours. The smallest shrimp which had no experience catching Tigriopus ate more copepods than the large inexperienced shrimp. However, the large shrimp which had previously fed on Tigriopus ate more than most of the smaller inexperienced shrimp (Figure I). The larger experienced shrimp also ate more than the inexperienced shrimp of comparable size. It appears that the shrimp have the ability to learn how to capture Tigriopus. Summary Under laboratory conditions the major predators of Tigriopus californicus, in decreasing importance, are Clinocottus analis, Pachygrapsus crassipes, Anthopleura elegatissima, Pagurus samuelis, Heptacarpus paludicola, and Trichocorixia (Figure II). However, in natural situations Pachygrapsus and Pagurus are probably the primary predators in the higher pools. Trichcorixia is not as common and if it is a predator, it is not as effective as the two crabs. Anthopleura is an effective predator in the lower high tide pools which receive some fresh sea water at high tides. Even lower in the intertidal are the cottid fish; these and wave action could efficiently keep the Tigriopus population to a minimum down here. Tigriopus were not normally found in the same tide pools as the shrimps. However, heavy wave action could wash Tigriopus down to the lower pools where the shrimp are found, allowing the shrimp to act as occassional predator of Tigriopus. (( Acknowledgements I would like to thank Dr. Don Abbott for his never ending patience and guidance as well as Dr. Robin Burnett, Chuck Baxter and Lynn Hodgson for their help and suggestions. Literature Cited 1. Hiatt, Robert W., 1918, "The Biology of the Lined Shore Crab, Pachygrapsus crassipes Randall", Pacific Science, 2: 13-213 MacGinitie, G. E., and Nettie MacGinitie, 1968, Natural History of Marine 2. Animals, MCGraw-Hill Book Co., New York, pp. 273, 297, 317 Makarov, V. V., 1938, Anomura, Israel Program for Scientific Translation, Jerusalem, translated 1962, p. 168 Meglitsch, Paul A., 1972, Invertebrate Zoology, Oxford University Press, New York, p. 327 5. Pennak, Robert, 1953, Fresh Water Invertebrates of the United States, Ronald Press, New York, pp. 551-559 6. Usinger, Robert, 1956, Aquatic Insects of California with Keys to North American Genera and California Species, University of California Press, Berkeley, pp. 188-196 Vittor, Barry A., 1971, "Effects of the Environment on Fitness-Related Life History Characters in Tigriopus californicus", Phd. Dissertation, University of Oregon 20 0 4 2 uoD Du o - - o 0= - — o - - OO +4 D D0 DDOT O0 -- Oos d 0- D 0 — D 20 0- 0 22 820 ooo OC DT C oo D 0 O O 41-- 29 vaaa oo D1 a — — O Table II Trichocorixia- red colored TetraMin feeding experiments Starting Dura- TetraMin No. Notes date tion condition of Tri (hrs) corixia used Gut contents not red after 4-28 24 fresh feeding 5-19 fresh Cotton put in fingerbowl; 24 gut contents gold in all individuals after feeding 2 days Cotton put in fingerbowl; 5-19 24 gut contents gold in 3, red old in l and slightly red inl after feeding D 0 - 0 D0 D 0 - 205 0-0 1 OCO DT OC 1 — 0 9 3.9 D O OO 0 O -o o 0 oo O ox Ooco o O O E 2 0 N NN. 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O ooaaaaaaaaaggggo ededenass aaaaayaaaagaago o u 100 90 80 —70 50 30 20 10 Figure I 1.2.1.4 Heptacarpus Feeding Experiment with Tigriopus +Experienced shrimp Inexperiencedshrimp + — 1 1.6 1.8 2.0 2.2 2.4 2.6 Heptacarpus size (cm) O Figure II 9. 8.8 —1-— Number of: predators per trial trials Predators of Tigriopus in the Laboratory 100 80 50 40 + -30 20 10 I — o 2 0 1-4 1-3 30 — Range Key: Standard deviation Mean — 21