Host selection by shrimps symbiotic with sea anemones: A field survey and experimental laboratory analysis

Journal of Experimental Marine Biology 202 (1996) 165-176

ELSEVIER

JOURNAL OF EXPERIMENTAL MARINE BIOLOGY AND ECOLOGY

and Ecology.

Host selection by shrimps symbiotic with sea anemones: a field survey and experimental laboratory analysis Chau-Chih ‘Instituteof

Guo”, Jiang-Shiou

Marine Biology, College of Fisheries

20224, hDepartmeru

of Systematics

and Ecology,

Received 21 July 1995; revised II

Hwang”, Daphne Gail Fautinb’* Science, National Taiwan,

Tuiwm

Ocrm

fJnivrr.vif~.

Krrlrrr~g.

ROC

6604-r. USA

University

of Km.ws,

December

1995: accepted 29 January 1996

Luwrrtx~,

KS

Abstract In three coastal areas of the Republic of China, we found six species of anemoneshrimps and tive of host sea anemones in 13 symbiotic combinations; four of the I3 combinations were previously documented. In the laboratory, we tested host preference of the three common shrimps, Periclimenes ornutus Bruce, P. brevicarpalis (Schenkel), and Thor umboinensis (De Man), fat anemones of three species. Periclimenes ornatus unequivocally preferred Entucmaea quudricoior (Riippell and Leuckart), the only anemone with which it occurs in northern Taiwan and from which the experimental specimens had been collected. We therefore consider P. ornatus a specialist on E. quadricolor. Results of experiments with P. brevicarpalis and T. umboinensis were ambiguous: for both, outcome of some experiments was affected by identity of the anemone from which the shrimp had been collected, and in other experiments, T. amboinerzsis exhibited no preference. We therefore consider Thor a generalist symbiont, and P. brerkurpalis intermediate in speciticity. In experiments to determine sensory cue(s) used by the shrimps to locate hosts, none could locate an anemone by vision alone. A significant proportion of shrimp of all three species was chemically attracted to anemones of at least one species; as in the preference experiments. source of the shrimp affected the results for P. bre\icarprclis and T. cmzboinensis. The proportion of shrimp chemically attracted to an anemone in the experiments on sensory cues was lower than the proportion attracted in the host preference experiments. We, therefore, infer that shrimp use more than one sensory modality to locate a host anemone. Kepvords:

Actiniaria;

*Corresponding

SOO22-098

Caridea; Host specificity;

author.

0022-098 I /96/$15.00 PI/

Anemoneshrimp;

0

1996 Elsevier Science B.V. All rights reserved

I (96)00020-2

Taiwan

C. Guo et al. I J. Exp. Mar. Biol. Ecol. 202 (1996) 165-176

1. Introduction Symbiotic associations between some shrimps and sea anemones are common throughout the tropics (Bruce, 1976a,b). (The term “symbiotic” is used literally, in the sense of living together, without implication of cost or benefit.) Most research on non-anomuran crustaceans symbiotic with anemones has dealt with caridean shrimps (primarily of families Palaemonidae, Alpheidae, and Hippolytidae). Despite there being many more species of anemoneshrimps in the Indo-Pacific (Bruce, 1976a,b), studies on this symbiosis have been mainly in the Caribbean (e.g. Herrnkind et al., 1976; Smith, 1977; Knowlton and Keller, 1985; Nizinski, 1989). Most shrimps are found with anemones of only some of the species sympatric with them (e.g. Herrnkind et al., 1976; Suzuki and Hayashi, 1977; Knowlton and Keller, 1985; Nizinski, 1989; Gwaltney and Brooks, 1994). This specificity implies the shrimps are capable of distinguishing potential hosts (Bruce, 1976b), but little research has been done on this subject. From host choice trials in the laboratory, Knowlton and Keller (1985) concluded that shrimp of each of four species of Alpheus prefer anemones of the species with which it naturally occurs to an alternative host. However, the distribution of shrimp in the held may not reflect only preferences of the shrimp for particular anemones. Host preference can be modified in some species (Gwaltney and Brooks, 1994), and distribution in the field may be affected by ecological factors such as abundance of hosts and presence of potential competitors (by analogy with anemonefishes [F/ 11). After surveying anemoneshrimps and their hosts in three areas of the Republic of China (ROC), we did laboratory experiments to determine whether shrimp prefer the anemones with which they naturally occur. Of the three most common species of anemoneshrimps and two of host anemones around northeastern Taiwan, we found individuals of Periclimenes ornatus Bruce symbiotic with the sea anemone Entacmaea quadricolor (Riippell and Leuckert), whereas those of P. brevicarpalis (Schenkel) and Thor amboinensis (De Man) occurred with hosts of both E. quadricolor and Stichodactyla tapetum (Ehrenberg). Based on our laboratory results, the three species of shrimp differ in degree of host specificity, P. ornatus being most specific, which is consistent with its host distribution in nature, and T. amboinensis least, which is consistent with its natural occurrence on a variety of cnidarian and non-cnidarian hosts (e.g. Bruce, 1976a,b; Criales, 1984). Also examined was whether chemical or visual cues could explain how anemoneshrimps locate their hosts. We assumed that the association between some species pairs may be chemically mediated whereas those between other pairs may not be, by analogy with the anemonefish symbiosis (Fautin, 1991): e.g. some anemonefish reportedly locate their host visually (Davenport and Norris, 19.58; Mariscal, 1972; Fukui, 1973) and others do so chemically (Miyagawa and Hidaka, 1980; Murata et al., 1986; Miyagawa, 1989). We infer from our results that, for locating a host from a distance, vision plays no role and olfaction may be used by shrimp of some species but not others. Visual, tactile, or gustatory cues may be important at short range.

C. Guo et al. I J. Exp. Mar.

2. Materials

Bid.

Ed.

202 (1996)

16S

176

167

and methods

Surveys to establish geographical distribution and symbiotic associations were made in coastal waters no more than 20 m deep near Keelung in northeastern Taiwan, in Ken-Ting National Park at the southern end of Taiwan, and in the Penghu (Pescadore) Islands, ROC. The sea floor in surveyed areas was rock, coral, or firm sediment. A transect to determine density of shrimp and proportion of anemones occupied was made on Ho-ping Island near Keelung in clonal beds of Entacmaea quadricolor from which no shrimp had been collected. Animals for laboratory experiments were collected by hand, generally while scuba diving; most came from the vicinity of Keelung but some came from the Penghu Islands. Laboratory studies were conducted on the most abundant species found in the held surveys near Keelung: three anemoneshrimps, the palaemonids Periclimenes ornatus and P. brevicarpalis, and the hippolytid Thor amboinensis, and two host anemones, E. quadricolor (family Actiniidae) and Stichodactyla tap&urn (family Stichodactylidae). We also used the non-host anemone Actinia equina (L.) (family Actiniidae) in some experiments. Identification of the shrimps was made or confirmed by A.J. Bruce (Northern Territory Museum, Australia), T.Y. Chan (National Taiwan Ocean University [NTOU]), and M.K. Wicksten (Texas A and M University, USA). The sea anemones were identified by D.G.F. When not part of an experiment, specimens were held in aquaria 30 X 20 X 20 cm containing 12 I of aerated seawater from NTOU’s sea water system, and fed Artemia nauplii daily. Aquaria were kept and experiments were done in the NTOU aquarium room at ambient temperature, which varied from 23 to 28°C depending on outside air temperature, and under artificial lighting, which was on an approximately 12: 12 h cycle. Experiments were begun within 12 d of an animal’s collection; thereafter, surviving animals were returned to the field. Animals were not fed during experiments. For an experiment, a shrimp was dipped from the aquarium in which it had been held using a small glass beaker that was then inverted in the experimental tank; the beaker was removed after 5 min. Each shrimp was used in only one replicate of each type of experiment, although most were used in experiments of more than one type. For choices between anemones of two species, size of anemones was matched as well as possible, but we often used two individuals of A. equina to compensate for their small size. Size of shrimp was not considered in the experiments. Standard length of T. amboinensis and P. ornatus was estimated to be 15-20 mm; in another study, Fautin et al. (1995) found P. brevicarpalis in Taiwan to average 20 mm, with a range of 1 l-28 mm. Each experimental run, which began in the afternoon, lasted 24 h; at its end, the position of the shrimp was recorded (i.e. its movements during the 24 h were unknown). Sixteen replicates were done for most experiments. We discarded data from trials in which animals died and those in which results were ambiguous (e.g. because an anemone moved into the arm of the Y maze occupied by an anemone of another species). The apparatus was thoroughly cleaned after each experiment. Experiments on host choice were done in apparatus of two types. One was a Y maze

168

C. Guo et al. I J. Exp. Mar.

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202 (1996)

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in which the three arms, each 30 cm long and 10 cm wide and high, were set at 120” angles. An experimental shrimp was introduced into the far end of one arm; placement of the two alternative choices at the distal end of the other arms was determined randomly. Water was motionless during the experiment and was not aerated. A trial was scored as “no choice” if the shrimp was in the arm into which it had been placed; a choice was scored if the shrimp was in one of the other arms. In another set of experiments, an individual of E. quadricolor was placed at one end of a rectangular aquarium (30 X 20 X 20 cm), an individual of S. fupetum was placed at the other, and a shrimp was placed on one of the anemones. Air was bubbled into water at one corner of the aquarium during the experiment. Experiments to investigate sensory modalities used by shrimps to make their choices were done in rectangular aquaria. Water was aerated during the trials. To determine the importance of visual cues, a glass container 15 X 15 X 15 cm was placed at each end of an aquarium 90 X 20 X 20 cm holding 22 1 of water to a depth of 12 cm. One container held an anemone and the other a black plastic cylinder of similar size and shape; the placement of the anemone for each trial was determined randomly. A shrimp was introduced to the aquarium halfway between the containers. We assumed the shrimp could see the objects in the containers but could not detect them chemically. Experiments to determine the importance of olfactory cues were conducted in 30 X 20 X 20 cm aquaria. A cylindrical frame made by removing the base and upper portion from a l-l transparent plastic soft drink container in which vertical slits had been cut was set into a Petri dish of slightly larger diameter. Each frame-and-dish structure was wrapped with a single layer of gauze that presumably allowed the passage of diffusible chemicals. For the first series of experiments, a shrimp was given a choice between a specimen of an anemone in a wrapped frame-and-dish structure at one end of the aquarium and an identical wrapped structure without an anemone at the other end, their positions being determined randomly for each trial. A series of experiments in which the frames and Petri dishes were not wrapped with gauze tested if the structure affected a shrimp’s ability to locate an anemone. Experimental results were analyzed by binomial and x2 statistics (Zar, 1984). In Tables 2-6, “source host” refers to the species of anemone from which the shrimp used in the experiment were collected, “non-source host” is another species of anemone with which shrimp of that species occur, and “non-host” is a species of anemone with which shrimp of that species are not known to be symbiotic.

3. Results 3.1. Field survey In this first study of the anemoneshrimp symbiosis in ROC, we found shrimps of six species associated with sea anemones of five species (Table 1). One of us (C.-C.G.) also found Thor amboinensis with an unidentified cerianthid in northern Taiwan. Commonly, more than one species of anemone and shrimp occurred at a site. Most shrimp of the species Periclimenes ornatus, P. brevicarpalis, and Hamopontonia

C. Guo et al. I J. Exp. Mar. Biol. Ecol. 202 (1996) 115-176

169

Table I Symbiotic associations between anemoneshrimps and their host sea anemones in coastal waters of northeastern Taiwan (N), southern Taiwan (S) and the Penghu Islands (P). 0 indicates a previously recorded combination Anemone

species

Entacmaea quadricolor

Macrodactyla doreensis

Stichodactylo tcrpetum

Phymanthus SP.

N, S, P(0) N, P (0)

P S, p

N

N

Heteractis aurora

Shrimp species Periclimenes ornatus Periclimenes hrevicarpalis Periclimmes venustus Periclimenes sp. Hamopontonia corallicola Thor amboinensis

S P N, P (0) N, P (0)

P

N

N

corallicola occurred singly or in pairs. When more than two individuals inhabited one anemone, they were usually a male, a female and young shrimp. We observed at least 25 individuals of Periclimenes venustus on one specimen of Heteractis aurora and as many as 11 of T. amboinensis on Stichodactyla tapetum. The largest shrimp in these groups tended to be near the center of the oral disc, with smaller individuals at the periphery. Although most individual anemones contained shrimp of only one species, we found individuals of Entacmaea quadricolor occupied by shrimp of all three of the most abundant species (P. ornatus, P. brevicarpalis, and T. amboinensis) and by shrimp of two of these species in all three possible combinations. The anemone Macrodactyla doreensis was abundant only in the Penghu Islands and was not found in northern Taiwan. Individuals of P. ornatus were associated with it only where it was not sympatric with E. quadricolor; at a site where both anemones occurred, shrimp were with E. quadricolor. An individual of S. tapetum was occupied by shrimp of P. brevicarpalis and T. amboinensis, either singly or together. Most individuals of T. amboinensis associated with E. quadricolor were beside the anemone whereas individuals associated with S. tapetum were on the host’s oral disc. Table 2 Choice by anemoneshrimps

in Y maze: source host anemone

Shrimp (source host)

Control

Periclimenes ornatus (Entacmaea quadricolor) Periclimenes brevicarpalis (Entacmaea quadricolor) Periclimenes brevicarpalis (Stichodactyla tapeturn) Thor amboinensis (Entacmaea quadricolor) Thor amboinensis (Stichodactyla tapetum)

Empty arm Stone Empty arm Stone Empty arm

N = 16 for all experiments.

Choice host/no

versus control

choice/control

P choice/no

choice

P host/control

16/O/O 15/l/O 16/O/O 16/O/O 16/O/O

0.00002 0.00026 0.00002 0.00002 0.00002

0.00002 0.00003 0.00002 0.00002 0.00002

Stone

15/l/O

0.00026

0.00003

Empty arm

61812 6/9/l 11/5/O 11/2/3

0.59818 0.401x0 0.10505 0.00209

0.14454 0.06250 0.00049 0.02860

Stone

Empty arm Stone

Probabilities

were calculated

with a one-tailed

binomial

test.

host)

ornatus

brevicarpalis

tapetum)

Periclimenes

(Stichodactyla

q./S. q./A. q./S. q./A. LIE. t./A. q./A. q./S. q./A. t./E. t./A. q./A.

t. e. t. e. q. e. e. t. e. q. e. e.

host, and non-host

81612 11/l/4 51912 81315

21717

14/l/l 15/o/ 1 16/O/O 15/l/O

81216

16/O/O 14/O/2

2

anemones

Choice species 1 /no choice/species

N = 16 for all experiments. Probabilities were calculated with a one-tailed binomial test except those indicated ’ E. q. = Entacmaea yuadricolor; S. t. = Stichodactyla tapeturn; A. e. = Actinia equina.

E.

S.

tapetum)

(Stichodactyla

E. E. E.

S.

S.

E. E. E. E.

S.

quadricolor)

source host, non-source

Anemones tested” species I /species 2

in Y maze: among

Thor amboinensis

(Entacmaea

Thor amboinensis

brevicarpalis

quadricolor)

Periclimenes

(Entacmaea

quadricolor)

(Entacmaea

Periclimenes

(source

Shrimp

Table 3 Choice by anemoneshrimps

choice

by ’ were two-tailed.

0.22724 0.00026 0.40180 0.01063

0.40180

0.00026 0.00002 0.00002 O.OGQ26

0.00209

0.00002 0.00002

choice/no

P

0.05470 0.1184S2 0.22626 0.29052

0.1796@

0.00049 0.00052’ 0.00002 o.OOOQ3

0.79052’

0.00002 0.00209

species 1 /species

P

2

C. Guo et al. / .I. Exp. Mar. Biol. Ecol. 202 (1996) 165-l 76 Table 4 Choice by anemoneshrimps

in rectangular

tank: source host anemone

171

versus non-source

host or non-host

anemone Non-source

Shrimp (source host)

or non-host anemone tested

On non-source host or non-host

On source host

Moved

Stayed

Moved

Stayed

Periclimenes ornatus (Entacmaea quadricolor) Periclimenes hrevicarpalis

Stichodactyla tapetum Actinia equina Stichodactyla tapetum

0 0 4

16 16 12

16 16 3

0 0 13

(Entacmaea quadricolor) Periclimenes hrevicarpalis

Entacmaea

0

16

13

3

(Stichodactyla tapeturn) Thor amhoinensis

Stichodactyla

tapetum

12

5

2

14

(Entacmaea quadricolor) Thor amboinensis

Entacmaea

quadricolor

4

12

14

2

(Stichodactyla

quadricolor

tapeturn)

For each trial, a shrimp was placed on the oral disc of an anemone. were all
Probabilities,

calculated

with a x2 test,

of the 100 specimens of E. quadricolor in the census, 42 contained a total of 5.5 shrimp (for an average of 1.3 per occupied anemone). Although as many as four shrimp occupied one individual, most anemones had but one.

3.2. Laboratory

experiments

Given a choice in the Y maze between a specimen of the anemone from which they had been collected and an empty arm or a stone (Table 2), shrimp of the species l? omatus and P. brevicurpalis overwhelmingly chose the anemone. Behavior of specimens of T. amboinensis depended on the species of anemone from which they had been collected. Results of Y maze experiments in which shrimps chose between anemones of two species are shown in Table 3. Specimens of P. ornatus preferred E. quadricolor, preference of individuals of P. brevicarpalis depended on their source, and shrimp of T. amboinensis exhibited no preference. In the other test of host preference (Table 4)

Table 5 Experiments

to determine

if anemoneshrimps

Shrimp (source host)

Anemone

Periclimenes ornatus (Entacmaea quadricolor) Periclimenes brevicarpalis (Entacmaea yuadricolor) Thor amboinensis (Entacmaea quadricolor)

can locate hosts using visual cues tested

Choice (anemone/control)

P

Entacmaea quadricolor Actinia equina Entacmaea yuadricolor

917 5/11 6110

0.40180 0.10505 0.22724

Entacmaea quadricolor

818

0.598 18

Source host Entacmaea quadricolor or non-host Actinia equina versus control (black plastic cylinder). for all experiments. Probabilities were calculated with a one-tailed binomial test.

N = 16

172

Table 6 Experiments

C. Guo

to determine

et al. I J. Exp. Mar. Biol. Ecol. 202 (1996) 165-176

if anemoneshrimps

can locate hosts using chemical

Shrimp (source host)

Anemone

Gauze wrapping

N

Choice (anemone/control)

P

Periclimenes ornatus (Entucmuea quadricolor)

Entacmnea quadricolor Entacmuea quadricolor Stichodactyla taprtum Actinia equincr Entacmaea quadricolor Entucmaeu quadricolor Stichoductylrr tapetum Actinia equina Entacmaea quadricolor Stichodactyla tupetum Entacmaea quadricolor Entacmaea quadricolor Stichodactyla tupetum Actiniu equina Entucmuea quadricolor Stichodactyla tapetum

Yes

19 8 16 19 19 8 16 17 16 16 18 8 16 16 16 17

14/s 810 6/10 IO/9 141.5 8/O 3113 6/11 1214 1313 IO/X 7/l 917 8/8 818 14/3

0.03 I79 0.0039 1 0.22724 0.5000 I 0.03179 0.0039 I 0.01063 0.16616 0.03840 0.01063 0.40725 0.035 I6 0.40 180 0.598 18 0.59818 0.00637

Periclimenes brevicarpalis (Entacmaea quadricolor)

Periclimenes brevicurpalis (Stichodactyla tapetum) Thor amboinensis (Entacmaea quadricolor)

Thor amboinensis (Stichodactyla tapeturn)

tested

Anemone (in Petri dish, surrounded by gauze-wrapped frame in Petri dish). A trial without gauze wrapping one-tailed binomial test.

No

Yes Yes Yes NO

Yes Yes Yes

Yes Yes No

Yes Yes Yes

Yes

cues

plastic frame) versus control (gauze-wrapped was also done. Probabilities were calculated

plastic with a

specimens of P. ornatus unequivocally chose E. quadricolor, behavior of individuals of P. brevicarpalis depended on their source, and individuals of T. amboinensis favored S. tapetum. None of the shrimps located an anemone by visual cues alone (Table 5). A significant proportion of shrimp of all species was attracted chemically to one or both of the host anemones but none was attracted to specimens of A. equina (Table 6).

4. Discussion Of the 13 symbiotic combinations (Table 1) that were found, the four involving Entacmaea quadricolor had been recorded previously. At least six of the others are new records; if the unidentified species of Phymanthus and Periclimenes are new, the three symbioses in which they are involved would also be new. The three most common species of anemoneshrimps, which we used in our laboratory experiments, occur throughout the tropical Indo-Pacific as obligate associates of invertebrates (although all can be kept in the laboratory without anemones). None of them is a cleaner. Suzuki and Hayashi (1977) identified two hosts of P. ornatus in Japan: Parasicyonis actinostroides and Pa. maxima. The correct identification of both is E. quadricolor (see Dunn, 1981; Bruce and Svoboda, 1983; Fautin and Allen, 1992; Fautin et al., 1995). Bruce and Svoboda (1983) mentioned in the text that P. ornatus occurs with “G. helianthus" , a species absent in their table. Presumably this was Gyrostoma helianthus, also a synonym of E. quadricolor (see Dunn, 1981). In addition to E. quadricolor, P.

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173

ornutus has been recorded with anemones of two other species (Bruce and Svoboda, 1983); we add a fourth. Periclimenes brevicarpalis had been recorded with at least eight species of sea anemones (Bruce and Svoboda, 1983); we add three. Entucmueu quadricolor, the host with which the shrimp is most often found in Taiwan, and with which it occurs elsewhere (Bruce and Svoboda, 1983) is also apparently the most common host in Japan, misidentified by Suzuki and Hayashi (1977) as the two species of Parusicyonis. A third host species in Japan, identified by Suzuki and Hayashi (1977) as Radianthus maculata, is unidentifiable, that name being unknown for an anemone (Bruce and Svoboda, 1983; Fautin et al., 1995). Thor amboinensis occurs in the Caribbean as well as the Indo-Pacific (e.g. Herrnkind et al., 1976; Criales, 1984; Nizinski, 1989). Its hosts include a range of cnidarians aside from anemones (Bruce, 1976a,b) and even crinoids (Criales, 1984); we add three anemone hosts and the first report of this shrimp with a cerianthid. Many symbiotic shrimps occur in pairs (Bruce, 1976b), although large groups of Thor (e.g. Suzuki and Hayashi, 1977) and P. anthophilus (see Nizinski, 1989) have been documented. The presence of shrimp of more than one species on a single host individual is not uncommon (Bruce, 1976b). In all our experiments, P. ornatus overwhelmingly preferred E. quadricolor to anemones of the other two species. This is not surprising, E. quadricolor being the only one of the three with which P. ornatus naturally occurs. Omori et al. (1994) associated the specificity of P. ornatus for E. quadricolor (which they referred to as Pa. maxima) with length of its host’s tentacles, because individuals of this species eat their host’s tentacles (Suzuki and Hayashi, 1977) as do those of P. brevicarpulis living with E. quadricolor (see Suzuki and Hayashi, 1977; Bruce and Svoboda, 1983) which enhances the shrimp’s growth, reproduction, and survival (Fautin et al., 1995). However, Omori et al. (1994) provided no information on sympatric potential hosts or on experiments to test this idea. We infer that differences in tentacle length are only coincidental, and that shrimp choice is based on other factors. Although we found this shrimp also with the long-tentacled Macrodactyla doreensis, where M. doreensis and E. quadricolor were sympatric, P. ornatus preferred the latter; and although we did not find it with the short-tentacled S. tapetum, it occurs with the anemone Cryptodendrum adhaesivum, the short tentacles of which it eats (Bruce and Svoboda, 1983). Individual l? brevicarpalis can clearly detect host anemones, selecting in significant proportion an anemone of either host species over nothing, a stone, or a specimen of a non-host anemone (Tables 2 and 3). The lack of choice between anemones of the two species with which P. brevicarpalis naturally occurs by individuals collected from E. quadricolor is presumably not due to an inability to make a distinction between them. for individuals collected from S. tapetum exhibited a strong preference (Tables 3 and 4). These results indicate that prior experience does not necessarily imprint a shrimp. A similar asymmetry occurs in P. yucatanicus (see Gwaltney and Brooks, 1994). Although we found P. brevicarpalis more commonly with E. quadricolor in the field, we conclude it prefers S. tapetum. Because S. tupetum is less abundant than E. quadricolor, we infer that there may be a shortage of the preferred anemones, forcing most shrimp to associate with a less preferred host.

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The least host specific of the three shrimps studied, T. amboinensis, had a weak preference for S. tapetum. Source of the shrimp affected results of some experiments but not others: in the Y maze, only those collected from S. tapeturn showed a preference of an anemone to a stone or nothing (Table 2), which is consistent with results from the chemical cue assay (Table 6). But source was irrelevant in the experiment in which a shrimp was placed on an anemone: a significant proportion of those collected from either host remained on or moved to S. tapetum (Table 4). By contrast, in a direct choice between anemones of the two host species, shrimp from neither host exhibited a preference (Table 3). Given the broad geographic and host distributions reported for T. amboinensis, it is possible that the animals identified as such may actually be sibling species, of which many remain undiscovered among marine organisms (Knowlton, 1993). Knowlton and Keller (1985), for example, resolved four sibling species of anemoneshrimps belonging to a complex that had been previously identified as a single species. The lack of consistency in our results may be evidence of more than one taxon; alternatively, it could be evidence that this shrimp is, indeed, a generalist, with wide host and geographic distributions (although not necessarily as wide as implied by application of the name). We found too few shrimp in the field to draw a meaningful conclusion about whether their distribution is consistent with the results of our experiments. This may be because we, like Herrnkind et al. (1976), missed many of them in our searches. Individuals of Thor typically occur as we saw them with E. quadricolor, under or beside their anthozoan host (Suzuki and Hayashi, 1977; Nizinski, 1989). However, the small, white-spotted dark shrimp (Suzuki and Hayashi, 1977) live on the oral disc of S. tapetum, where they are well camouflaged (personal observation); this may relate to the shrimp’s preference for that anemone. The inability of shrimp to locate an anemone visually (Table 5) may be related to the fact that they are more active at night than during the day (C-C.G., J.-S.H. personal observation on P. brevicarpalis using infrared video recordings). By contrast, shrimp of all three species could locate anemones of some species by olfaction (Table 6). Although distance from the center of the aquarium, where the shrimp was placed, to an anemone was greater in the experiments on vision (about 35 cm; results in Table 5) than in those on olfaction ( lo- 12 cm; results in Table 6), that difference cannot account for the lack of visual attraction because shrimp travelled about 50 cm to an anemone in the Y maze. Moreover, the pattern of data in Table 6 is consistent with that of Tables 2-4, indicating that type of apparatus and differences in aeration did not affect our results. The lower proportion of animals locating E. quadricolor when it was within a gauze-wrapped frame than when it was within the frame without wrapping may be because diffusion was reduced by the gauze. However, diffusion was sufficient that there was a difference in results with symbiotic and non-symbiotic anemones, and, paradoxically, the only choice based solely on olfaction that was significant at the 1% level was that of the extreme generalist, T. amboinensis. We therefore infer that cues in addition to olfaction are used by shrimp to locate an anemone. Whereas in the experiments without gauze, at the end of 24 h a shrimp was commonly on or right beside the anemone, in experiments with gauze, few shrimp were actually on the gauze. If diffusion were being blocked, or molecules were being adsorbed onto the gauze, the concentration gradient would be abrupt at the gauze so the shrimp that had

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managed to locate the gauze-wrapped frame would be expected to cling to it. Our observation is consistent with shrimp detecting an anemone from a distance by diffusing compounds, then switching to cues such as vision, touch, or contact chemosensation (gustation) near the anemone. Unable to confirm the presence of an anemone by vision, touch, or gustation (all of which would be blocked by the gauze), the shrimp would not cling to the gauze. Multiple modalities are important for insects locating plants on which to feed, an activity once attributed entirely to olfaction (Miller and Strickler, 1984). Among marine arthropods, the caridean shrimp Betueus macginitieae uses both olfaction and vision to locate its echinoid host (Ache and Davenport, 1972), and males of the isopod Serolis politu use olfaction and touch to locate females (Michel, 1987). There are separate receptors for chemosensation at a distance (olfaction) and contact chemosensation (e.g. Schmidt and Gnatzy, 1987). Miller and Harris (1985) argued convincingly that multiple modalities probably influence many behaviors, so providing single stimuli is unlikely to elicit realistic responses. Olfaction may not function in all anemoneshrimp symbioses, by analogy with the clownfish symbiosis in which it cannot explain all partnerships. Murata et al. (1986) and Miyagawa (1989) showed that post-larval clownfish of each of two species were attracted chemically by one of two host anemones but not the other. Although fish of each species occur in nature with the anemone to which they were attracted, fish of one of the species also occur naturally with the anemone to which they were not attracted (Fautin, 1991). Studies on the anemoneshrimp symbiosis have shown the associations to be varied (Bruce, 1976a), so findings from one partnership may not apply to another.

Acknowledgments This research was supported by grant number NSC 83-0418-B-019-008 BH from the National Science Council of Taiwan, ROC, and by a grant from the Ministry of Education, ROC, both to J.-S.H. It is adapted from the Masters Thesis of C.X.G., submitted to National Taiwan Ocean University, Keelung, Taiwan, ROC. We are grateful for accommodation and research space at the Ken-Ting Field Station, National Sun Yat-Sen University, and to Paul Barlow for help in preparation of the manuscript. Comments on earlier versions of the manuscript were made by J. Atema, W.J. Bell, R.W. Buddemeier, C.P. Chen, M.D. Greenfield, W.N. Hamner, N. Knowlton, K.Y. Soong, and R. Zimmer-Faust. This research was conducted at the National Taiwan Ocean University, Keelung, Taiwan, ROC.

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