Abstract In the rocky intertidal of the Monterey region, the slipper limpet Crepidula adunca is à common inhabitant of the shells of Tegula funebralis. In the Hopkins Marine Life Refuge, 13% of Tegula funebralis are host to Crepidula adunca. Sixty-three percent of Crepidula are found alone on Tegula shells, while 37% of Crepidula are on Tegula that are host to at least one other Crepidula as well. Initial lab experiments to determine movement rate of Crepidula between hosts showed that 16% of Crepidula moved at least once, 100% of whom were male. Experiments were then set up to determine what motivated male Crepidula to move. Males were removed from their original hosts and reattached on new hosts, then placed in replicate tanks containing Tegula with either no Crepidula or specific combinations of male and/or female Crepidula. Results of 19 replicates show that on average, 48% of Crepidula movements were to Tegula containing female Crepidula, 33% were to Tegula containing no Crepidula, and only 12% were to Tegula containing only male Crepidula. Results suggest that movement of male Crepidula was influenced by the presence of female Crepidula on the host snail, but this can not be isolated as the sole cause of movement. Other factors in Crepidula movement, such as overall abundance of male Crepidula on a host snail and chance events (such as rate of Tegula movement and Crepidula encounter) remain to be investigated. Introduction It is not uncommon in the marine world to find organisms living in host-guest relations with one another. One such example in the rocky intertidal zone of Monterey Bay is the interaction between Crepidula adunca (commonly known as the slipper snail) and its snail host, Tegula fune bralis (hereaster they will be referred to as Crepidula and Tegula respectively). This study examined why and how Crepidula chose their host snail. Crepiduld are sequential hermaphrodites, which means they reproduce as one sex (in this case males) early in life, then change to the other sex for the remainder of their life span (Hobday 1997). Crepidula shell size increases as a function of time, the sex of an individual is correlated with its size. Males are often found stacked on top of females - this is the position necessary for copulation (Charnov 1982). Hobday (1997) studied the movement of Crepidula based on reproductive potentials, hypothesizing that social situation can affect the time of male-female transition. That is, if a male is in the presence of females, it may change to a female later than if it were alone, in order to maximize its reproductive potential. This means that the sex of a Crepidula cannot be determined solely by its size. Later in this paper à precise way of determining the sex of a Crepidula will be explained in further detail Crepidula are filter feeders and can live in a sedentary state indefinitely. Vermeii et al. (1987) studied the interaction between Crepidula adunca on the host snail Calliostoma ligatum and showed that Crepidula who had been moved from their host shell and reattached were more likely to be attacked by predators since they no longer had as close a fit. Crepidula shell size increases as a function of time. Therefore the fit of a Crepidula on à host improves with time. Movement therefore makes them more vulnerable to predators as they grow larger and it would seem logical were they to move solely for reproduction Putnam (1964) studied the dispersal of young Crepidula adunca from Tegula funebralis and found that while 150-200 young are released from a female on one host, eight is the masimum number of Crepidula ever seen on one legula shell. She concludes that young must therefore be moving to neu hosts. In preliminary experiments to determine impetus for movement, Putnam (1964) discovered that uhen removed from their hosts, "adult males are atracted by legula sunebralis, but even more strongly by female C. adanco. In contrast. tne newly hatched spat do not apear to be strongly attracted to either T. funebralis or to older C. adumncd, male or female" She concluded that juvenies move about at random and afe only " weakly attacted to either T funebrals or older C. adunca, his attraction is clearby present in older individuals. Though Putnam (1964) states that atraction of males to females is clearly present in older individuals, she fails to give substantial proof to support this claim. This study was conducted to determine if and how osten Crepidula adunca moved fom its host snail, and the impetus for movement. Materials and Methods. Teguld and Crepidula distribution in the intertidal. In order to determine abundance and distribution of Crepidula in the intertidal. I sampled organisms from a transect perpendicular to Agasiz beach in the Hopkins Marine Lite Refuge. Along this transect I collected organisms from 10 quadrats. Tidal heights for each quadrat were measured with a surveyor's transit and stadia rod, relative to the predicted level of low tide. Osing a 23m? quadrat I collected all of the Tegula located within the quadrat and took them back to the lab. Ithen categorized them according to shell size and noted whether they had Crepidula present, the size of the Crepidula, whether the Crepidula were present in multiples or stacked, whether there were other limpets present on the snail, and whether the shell contained a hermit crab. These data provided a picture of the organisms abundance and patterns of their behavior and location within the intertidal. Shell size of Tegula was measured with calipers and the shell was measured by as maximum basal diameter. Crepidula shell size was measured by basal length as it sat on its host Tegula. Initial Movement Experiments These experiments tested if Crepidula were capable of moving from host to host, and ir so, with what frequency they did so. Tegula with and without Crepidula were collected from mid-intertidal zones and placed in tanks. The experiment was run twice, once over six days, and once over five, with four tanks present in each run. Tanks contained Tegula with Crepidula in the following ratios: Ismall (l-3mm), 3 medium (4-Imm), and 2 large CSmm) Crepidula, though absolute numbers differed among tanks. Each tank always contained as many or more Tegula as Crepidula. Tanks were 32 x 16 x 8 cm and contained between 11 and 24 snails. These numbers were approximately equal to the number of organisms found in an area in the intertidal of the same dimensions. Tanks were provided with running seawater and snails were disturbed once a day while data were recorded, but were otherwise lest alone. They received no food during the experiment. Crepidula were color-coded to match their host snail in order to track movement. Organisms were painted with nail polish on the tip of their shell where Crepidula could not attach to. At the completion of the experiment, Crepidula were removed from their hosts and placed under a microscope (in water) to determine their sex (so as to categorize their movement by sex). Male Crepidula have a penis protruding immediately right of the right cephalic tentacle. Females lack a penis, and Crepidula in the transition stage of becoming a female from a male have a stunted penis, which can very in length according to their stage in the transition process. Secondary Movement Experiments These experiments were set up to determine what factors lead to movement of Crepidula between hosts. Experiments were run for 24 hours with a total of 19 experiments. Experiments were run in tanks identical to the previous set of experiments. Tanks were placed in different positions on the sea table every day to eliminate variation between tanks. Each day they were run, the tanks were subdivided into two groups based on the source of male Crepidula. Two tanks contained males that had been collected from hosts which also contained female Crepidula. Two tanks contained males collected from hosts where no females had been present (i.e. they were alone on the snail or in the company of other male Crepidula). Ten tanks were run with males that had been found without females in the environment; nine tanks contained males that had formerly been on snails with female Crepidula present. Results are presented as aggregate totals, as well as in subdivisions based on the source of the males. Each tank contained the same number of Crepidula and Tegula. Male Crepidula were removed from their hosts and reattached to new hosts prior to placement in a tank. Females were not removed so that movement of males would not be influenced by poor fit or injury of a female that had been displaced. Removal of females increases their mortality. as females tend to die more quickly when removed from their original host (Hobday 1997). Females were identified by their large size (generally greater than 9mm) and all males were examined under a microscope to ensure that they were full males, and not juveniles or males in transition to becoming females. Each tank contained 15 male Crepidula and 5 female Crepidula in the following arrangement on host snails: 3 Tegula contained one solitary male Crepidula each. 2 Tegula contained 3 male Crepidula each. 2 Tegula contained 1 female Crepidula each. I Tegula contained 1 female Crepidula and 1 male Crepidula stacked on top of the female. 1 Tegula contained 1 female Crepidula and 2 males stacked on top if it. 1 Tegula contained 1 female, 1 male stacked on the female, and 2 additional males on the Tegula. 10 Tegula contained no Crepidula. At the end of 24 hours, the position of all males was recorded. These combinations provided a mix of situations based on gender of Crepidula and abundance (density) of Crepidula on each snail in order to observe effects that reproduction and density had on the impetus to switch hosts. There were 20 Crepidula and 20 Tegula present in the tank, so that if preference to be alone was a cause of movement, each Crepidula would have its own snail to move onto. Results and Discussion Intertidal Distributions Data from the Hopkins transect were categorized according to intertidal height (Table 1). Both Tegula and Crepidula occur in greater abundance in the lower intertidal zones and the number of Crepidula is proportional to the number of Tegula at any given height (Fig.1). Out of the 2455 Tegula that I collected from the intertidal, 312 (13%) were host to at least one Crepidula. Sixty three percent of Crepidula were found alone while 37% shared their host snail with at least one additional Crepidula. Further breakdown shows that of these, 22% were stacked and 15% were not (Fig. 2). This raised the question as to why a particular Crepidula chose its host snail and what the rate of movement was between shells. Initial Movement Experiments Sixteen percent (15/96) of Crepidula in the tank moved at least once during the experiment (Table 2). One-hundred percent of Crepidula that moved were male. Possible reasons for this include the role of the male as the mobile partner for reproduction, or that only the male is able to move because the female is too large and fits too tightly to its host snail. Among the 15 Crepidula that moved, there were 26 movements: 12 (46%) to Tegula containing female Crepidula, 11 (42%) to bare Tegula, and 3 (12%) to Tegula whose shells were already inhabited by other male Crepidula (Table 2). These results led to two hypotheses as to why male Crepidula are moving. The first is for reproductive reasons - nearly half of all males moved to host Tegula with females. The second hypothesis is that Crepidula prefer not to be in the company of other Crepidula and move to shells where they are alone. This hypothesis is supported by the data that 42% of movements resulted in a Crepidula being alone on a host snail. These hypotheses were tested in the next set of experiments. Secondary (choice) Movement Experiments Results from these experiments are presented in three ways: total movement of all experimental Crepidula, movement of males that had been collected in the field from hosts with female, and movement of males that had been collected alone or with other males (but not females) in the intertidal. This was done to determine whether males that had begun with females were more likely to move back with females than those that had no immediate prior association with females. The percent movement from any given starting position to a particular end position is shown as a row percentage (Table 3). Those Crepidula that were least likely to move began alone on females, which is expected based on the reproduction hypothesis. In order to better interpret movement patterns related to the reproduction hypothesis. I combined data from the five begin categories into 3 starting positions: alone, with females (males could have been present so long as females were present as well), and with only males (Table 4). The table shows the percentage from each category that moved (those that did not move are not included) and whether they moved to be alone, with females, or with only males. Approximately the same number of males moved regardless of their begin position (Table 4). Several movement percentages show patterns which support the reproduction hypothesis. Among all the Crepidula, males that began alone or with other males were the most likely to move to be with females. This holds true regardless of their source in the intertidal. Male Crepidula that were originally collected with females and that started the experiment with males were the most likely to move to be with females, as well as male Crepidula that were originally collected alone or with other males and that began alone or with other males (Table 4). These results are consistent with the reproduction hypothesis since males showed an inclination to be with females. Looking at the table, there are also patterns of movement which are contradictory to the reproduction hypothesis. Among all males, Crepidula starting alone or with females were most likely to move to be alone, which rather than supporting the reproduction hypothesis, supports the hypothesis of movement for reasons of density (i.e. to be alone) general, Crepidula that began with females had the most even distribution of movement among the three choices (Table 4). That they moved almost as much as males that began with other males or alone questions the reproduction hypothesis because we would expect them to stay with the females more than those from the other staring positions. The results tend to support the reproduction hypothesis, but there are enough contradictory patterns to suggest that additional factors may be involved. Total density of Crepidula on a host appears to have little effect: there is little increase in percent movement as density increases (column three in Table 5). A noteworthy entry is that of males who began with one female. These males were significantly less likely to move, which rather than support a hypothesis of density (in which case they would have moved to be alone) supports that of reproduction. However, the data for two males beginning with a female shows that they were as likely to move as those males who began with two other males, which suggests that some factor besides reproduction, whether density or otherwise, may play a role here. The results from these experiments suggest that movement of Crepidula adunca between Tegula funebralis is influenced by the presence of female Crepidula. These results support by Putnam (1964) and her studies on dispersal of young, and studies by Hobday (1997) of social situation as it affects movement and sex change. However, contradictions in the data suggest that movement is influenced by other factors as well, possibly including (but not limited to) density and chance events. Chance events include factors such as the density of Tegula and Crepidula in a certain area of the intertidal and the rate at which Tegula come in contact with one another. Further studies are needed to determine how reproduction and other factors interact to cause movement of Crepidula between Tegula shells. Acknowledgements I would like to thank Hopkins Marine Station and especially my advisor Jim Watanabe for all his help and early mornings in the tidal pools. I would also like to thank Jess Davy and other spring students for their help with data collection and for making it a great spring quarter. Literature Cited Hobday, Alistair J. and Riser, Kristin L., 1997. The role of movement by Crepidula norrisiarum Williamson in altering reproductive potential. Journal of Experimental Marine Biology and Ecology 225, 139-154. Charnov, E.L., 1982. The theory of sex allocation. Princeton University Press, Princeton, NJ. Putnam, Deborah A., 1964. The dispersal of young of the commensal gastropod Crepidula adunca from its host, Tegula funebralis. Veliger 6 (supplement), 63-66. Vermeij, G.J., Lowell, R.B., Walters, L.J., Marks, J.A., 1987. Good hosts and their guests: the relationship between Trocdid gastropods and the epizoic limpet Crepidula adunca. Nautilus 101 (2), 69-74. Table 1: Intertidal Distribution of Crepidula adunca and Tegula funebralis. Data are from one 25 m' quadrat at each tidal height. No. of Abs. Tidal No. of No. of Tegula WII No. of Tegula No. of No. of Crepidula No. of wi multiple Crepidula solitary w/ other stacked Ht.(ft) Tegula Crepidula Crepidula Crepidula Crepidula Crepidula 19 —0.22 56 486 25 0.24 115 455 92 28 0.30 1.30 236 34 25 22 35 1.50 366 10 219 1.92 2.14 132 210 15 2.52 4.18 208 5.4 247 64 390 86 2455 143 TOTALS 312 Table 2. Initial movement experiments. Two experiments were run over 6 days and 5 days respectively. Results are aggregate totals. + Tegula: 150 + Crepidula: 96 + Crepidula who moved: 15 + Total Movements (some Crepidula may have moved more than once): 26 + Movements onto Tegula with female Crepidula present: 12 + Movements onto Tegula with only male Crepidula present: 3 + Movements onto Tegula with no Crepidula present: 11 % of moving Crepidula who were male: 100% Table 3: Percent movement (Row totals add up to 100). Begin' refers to starting condition of male Crepidula on host Tegula. End position' refers to final position after 24 hours. End position Didn't Alone Off w/ females w/ females Move Begin no males and males Source: Total Crepidula Alone 10.5 Alone on female Multiple males on female w/males and females Wonly males TOTAL Source: Crepidula collected from hosts with female Crepidula present Alone Alone on female Multiple males 17 on female w/ males and females Wonly males 13 TOTAL Source: Crepidula collected from hosts with no female Crepidula present Alone Alone on females Multiple males on females w/ males and females w only males TOTAL —N 112 13 w/ males no females TOTAL 100 100 100 100 100 100 285 100 100 100 100 100 100 135 100 100 100 100 100 150 114 Table 4: Begin and end positions of Crepidula adunca on Tegula funebralis. *Numbers are percentages of all experimental animals that began in a given category. *a, b, and c are subsets of moved; numbers are percentages of Crepidula that ended in a given association. End Position % Moved b. % w/ only c. % Alone a. % with females males Begin Source: Total Crepidula 2990 With Females 33% 21%0 42%0 With Males 33 Alone Source: Crepidula collected from hosts with female Crepidula present With Females 11%0 33%0 33% With Males 38 Alone Source: Crepidula collected from hosts with no female Crepidula present With Females 3390 33% 27% 259 With Males Alone Table 5: Movement of Crepidula due to density Percentage Moved Begin (density) formerly w/female not formerly w/female 30% 30% 1 Male 1 Male + 1 Female 1 Male + 2 Females 3 Males 3 Males + 1 Female Total 30% 32 Figure legends Fig. 1. Intertidal Distribution of Tegula funebralis vs. Crepidula adunca in 25m? quadrats in the Hopkins Marine Life Refuge Fig. 2. Crepidula Distribution in the Intertidal No. Crepidula No. Tegula L Percent