Distribution and Dynamics of an Intertidal Ascidian Pseudopopulation

BULLETIN OF MARINE SCIENCE, 45(2): 288-303, 1989

DISTRIBUTION AND DYNAMICS OF AN INTERTIDAL ASCIDIAN PSEUDOPOPULATION

Craig M, Young

ABSTRACT Molgula occidentalis. an abundant ascidian in the shallow subtidal zone of the northern Gulf of Mexico, occurs seasonally on intertidal sandbars. One such population was studied over a 5-year period on Bay Mouth Bar, Franklin County, Florida. Most years, ascidians recruited during the spring and summer months, then died during early morning low tides in January and February. Predation by Fasciolaria hunteria (Gastropoda) accounted for a negligible portion of the mortality. Most mortality occurred when strong north winds caused freezing temperatures and prolonged exposure in the intertidal. Molgula occidentalis aggregate at several spatial scales. They occur most abundantly in areas where seagrasses (predominantly Ruppia maritima) stabilize the sediment. Sand movements and excessive siltation have adverse effects on adults and juveniles. Although larvae are capable of attaching to unconsolidated sand, they prefer to settle on the consolidated sands from the adult habitat. Small- scale distribution is also related to elevation. Few ascidians occur in the bottoms of deep pools or on high sand ridges. Highest densities are attained near the edges of pools, no more than a few em above or below the seawater table at low tide. Because adults can survive in the bottoms of pools, it is suspected that this distribution pattern is established by larval choice andjuvenile mortality. Intertidal Molgula occidenta/is are not true populations because they are not self-sustaining. Recruitment depends entirely on larvae emigrating from subtidal populations.

Individual organisms sometimes colonize regions beyond the usual geographical range of their species. Often, however, such individuals either fail to reproduce because of physiological stress, or die because of inimical biological or physical conditions before attaining reproductive maturity (Thorson, 1946; Mileikovsky, 1961). Thus, the species border may shift slightly with habitat fluctuations from year to year or from season to season, but tends to remain more-or-less constant over the long term (Mayr, 1970). Portions of a population established and sus- tained only by outside propagules are sometimes known as pseudopopulations. Most benthic marine invertebrates with pelagic larvae form "open" populations, meaning that colonizing larvae may originate either from within or from outside of a particular part of the range (Roughgarden et al., 1985). Thus, the distinction between an open population (Roughgarden et al., 1985) and a pseudo population (Mileikovsky, 1961) is tenuous. For the purpose of the present paper, I define pseudopopulations as portions of open marine populations occurring so near the species boundary that individuals either fail to reproduce or persist only tem- porarily. Virtually all individuals in a pseudopopulation are immigrants. The dynamics of pseudo populations are instructive from an evolutionary stand- point; they provide clues as to the factors controlling the species border and thus constraining horizontal gene flow. Nevertheless, most ecological studies focus on the self-perpetuating or otherwise permanent portions of populations geograph- ically removed from the species border. In the marine environment, species have both horizontal and vertical bound- aries. Pseudopopulations frequently occur in deep water, where recruitment depends on larvae washed offshore and reproduction is precluded by cold water temperatures and other environ mental conditions (reviewed by Mileikovsky, 1961). Physiological limits of tolerance to physical factors are generally thought to control

288 YOUNG: ASCI DIAN PSEUDOPOPULATION DYNAMICS 289 the upper limits of distribution for intertidal and shallow subtidal species (Connell, 1972; Underwood, 1985). For example, both hot and cold weather have adverse effects on molluscs occurring high on the shore (reviewed by Newell, 1979). Horizontally, populations of some benthic invertebrates are repopulated from outside sources because of the directions of prevailing currents. One example is in the Oresund of Scandinavia, where many species are replenished by supplies of larvae originating in the Kattegat or in the Baltic Sea (Thorson, 1946). Ascidians, being soft-bodied, are typically subtidal animals. Intertidal ascidians generally occur either in tidepools or in shaded habitats far down on the shore. The most notable exceptions are stolidobranchs in the genus Pyura, which char- acteristically have thick, leathery tunics (Paine and Suchanek, 1983; Stephenson, 1942; Underwood and Fairweather, 1986). Extensive long-term data on the dynamics of natural ascidian populations have only been collected from subtidal Scandinavian fjords (Svane, 1983; Svane and Lundalv, 1982a, 1982b). Many members of the family Molgu1idae occupy soft bottoms. Use of sedimentary habitats by these organisms has been facilitated by apparent adaptations in developmental mode, larval structure, larval behavior, and adult morphology (Ber- rill, 1931; Young et ai., 1988). Specifically, it has been suggested that adaptations for habitat selection have been lost in soft-sediment molgulids because habitat homogeneity renders such traits unnecessary (Berrill, 1931). However, neither population biology nor small-scale distribution of soft-sediment molgulids has been studied. In the northern Gulf of Mexico, Molgula occidentalis and Styela plicata are two dominant ascidians in the shallow subtidal zone. Both species occur in dense aggregations, occasionally attaching to seagrasses, shells, and other ascidians, but more often resting on the sandy or muddy substratum. Of the two species, only Molgula occidentalis is found in large numbers in the intertidal zone. Although it attains local densities as high as several hundred per square meter, intertidal populations are ephemeral. In this paper, I report quantitative and anecdotal observations on the dynamics of Molgula occidentalis at a single intertidal site over a 5 year period. I also report their intertidal spatial pattern on several scales, document behavioral attributes of the larvae, and discuss factors that control patterns of distribution and abundance.

MATERIALS AND METHODS

Study Site. -Pseudopopulations of Molgula occidentalis were studied primarily on Bay Mouth Bar, a large intertidal sandbar, several kilometers long by approximately 400 m wide, located at the mouth of Alligator Harbor, Florida (Fig. ]). The bar stretches north and south; the east side, where most ascidians were located, was on the leeward side of the bar except when winter cold fronts passed through from the north. Population Dynamics and Sources of Mortality. - During the winters of 1982-] 983 and 1983-1984, the corners of permanent sampling plots were marked with wooden stakes near the eastern shore of the sandbar where Molgula occidenta/is were abundant. All individuals were counted on each subsequent visit to the intertidal. Cohort survivorship curves were correlated with low tide predictions taken from NOAA tables and with National Weather Service air temperature data taken at the Tallahassee airport, approximately 50 km north of the study site. Surface seawater temperatures were taken at the Florida State University Marine Lab, located approximately 8 km west of the study site. The potential role of predators was examined by comparing survivorship of caged individuals, individuals surrounded by fences, and uncaged, marked individuals. Circular cages and fences 15 cm in diameter were constructed of I-cm mesh galvanized hardware cloth. Each cage or fence contained only a single individual in order to eliminate any confounding density-dependent effects. One set of treatments was set adjacent to each of the permanent sampling plots. Whenever mortality occurred, the cages were placed over new sets of animals, so data values represent mortality occurring between sampling dates, not cohort survivorship. 290 BULLETIN OF MARINE SCIENCE. VOL. 45. NO.2. 1989

~,r" FS.U. Morine Lob

Figure I. Map of the study area in the northern Gulf of Mexico.

Adult Molgula occidentalis cover themselves with a thick layer of sand, which is held in place by hair-iike extensions of the tunic. I investigated the potential role of this sand covering as a defense mechanism in laboratory experiments. In each of three experimental runs, either 4 or 10 adult ascidians were maintained in running seawater tables with 3 adult Fasciolaria hunteria. Half of the ascidians were scraped gently to remove most of the sand and to expose the bare tunic. After 24 to 48 h, mortality in the two groups was tabulated. Because each run had a small sample size and the results were similar for all runs, the data were pooled for analysis. Three aggregations of adult ascidians were transplanted to the bottoms of deep pools in December 1983 in order to determine if environmental conditions in pools were suitable for adult survival. The ascidian clumps were held in placc with large brass staples and were monitored biweekly until 19 January 1984. A single attempt to monitor early juvenile survivorship and growth by means of field transplants was unsuccessful. Fortuitously, heavy recruitment in a shallow running seawater system during the spring of 1983 enabled me to monitor juvenile mortality under high and low flow conditions. The seawater hose was directed at three marked quadrats, each to em on a side, near the standpipe drain. Individuals in these quadrats were subjected to continous, unidirectional flow and relatively little sedimentation. Mortality and growth of these three sub-cohorts was compared with mortality and growth of sub-cohorts living in other regions of the tank, where flow was lower and up to I cm of fine silt sometimes accumulated. Each individual in the tank was mapped separately and measured with vernier calipers on each sampling date, so growth data represent average increases in individual sizes and arc not biased by size-specific mortality. Measurement of Spatial Distribution. - Distributions ofascidians were characterized by sampling with I m' quadrats, 25 cm x 25 em quadrats, or 100 em x 75 cm rectangular quadrats, depending on the scale of interest. All sampling was contiguous. On the largest scale, transects were delimited at 50-m intervals by laying a 50-m long tape parallel to the north-south edges of the bar. Because tides in this region are heavily dependent on wind direction, nighttime low tides in the spring and summer months were generally not suitable for extensive sampling. Early morning tides from November through February generally occurred in conjunction with north winds that kept the tide out lower and longer than usual, so most sampling was done during this period. General habitat notes were made on all transects. During the 1987 sampling, each individual quadrat was classified into one of several major habitat types. Expected densities for each habitat classification were calculated by multiplying the total number of ascidians counted by the proportions of the total YOUNG: ASCIDIAN PSEUDOPOPULATION DYNAMICS 291 habitat composed of each habitat type. Expected and observed habitat distributions were compared with a goodness of fit test using the G statistic. For these analyses, the transects traversing flat sand plains on the eastern and western shores were not used, as they contained no ascidians. The relationship between elevation (or depth) and density was studied by sampling a grid of 25 x 25 em quadrats that encompassed a deep pool and the exposed substrata surrounding it. Pool depth was measured in the middle of each submerged quadrat with a meter stick; elevation above the seawater table was determined by excavating to the water level, then measuring the depth of the hole. The grid data were also used for analyzing aggregation on spatial scales less than I m. For these latter analyses, the submerged quadrats (which contained no animals) were excluded. Variance/Mean ratios were used as an index of dispersion and were tested statistically by methods outlined in Elliott (1977). Intermediate scales of aggregation (I -10m) were measured by calculating indices of dispersion for data taken along a IOO-m transect positioned in the same general area as the grid. Larval Development and Settlement Behavior. -Gametes for larval cultures were obtained by dissec- tion. Ovaries and testes obtained from hermaphroditic adults were macerated through 253 JLm Nitex mesh into filtered seawater. The embryos were rinsed several times to eliminate supernumerary sperm and cultured in glass dishes of seawater at 25°C, which was ambient seawater temperature during September. After hatching, swimming tadpoles were transferred with pasteur pipettes to the appropriate experimental treatments. Two kinds of experiments were run: "choice" experiments and "delay of settlement" experiments. In the former, small piles of natural substrata (adult tunic; consolidated sand from the adult habitat; unconsolidated sandbar sand), were placed in separate sections of compartmented pyrex petri dishes. The dishes were filled with seawater, larvae were added, and dishes were incubated under fluorescent lights at 25°C for 48 h. Each pile of substratum was then sorted carefully under a dissecting microscope, and the number of larvae selecting each substratum was recorded. Because the proportions of larvae selecting the various substrata in any given dish totaled one, treatments were not independent and could not, therefore, be compared with analysis of variance techniques. Instead, I used the G statistic to test goodness of fit of the rank orders of treatments to the numbers of runs that would be expected to have various rank orders if settlement were random. In "delay of metamorphosis" experiments, 50 larvae were incubated in each of several polystyrene petri dishes with single substrata (i.e., no alternative choices offered). After 12 h, the number of tadpoles undergoing metamorphosis in each dish was counted. Genetic variation in behavior was isolated simultaneously by running the multiple replicates with three different sibling groups originating from self-fertilized eggs.

RESULTS General Habitat Description. - The lowest tides of each year generally occur during the winter months, when strong north winds augment the tidal puIl, pushing the water further out than usual and extending the duration of the low tide for up to several hours longer than normal. On such tides, Bay Mouth Bar is approximately 400 m wide. The lowest regions exposed by winter tides consist of flat, bare sand dominated by enteropneusts and the sand dollar Mellita quinquiesperjorata. This zone is about 80 m wide on the eastern shore and 20 m wide on the western shore. Until 1984, dense beds of the mussel Modiolus americanus occupied a 20-m band, beginning just inshore of the western sand flat. After the winter of 1983-1984, the beds of M. americanus disappeared. By the winter of 1986-1987, shell rubble in this same region had been colonized by the bryozoan Thalamo- porellajloridana. At a comparable elevation on the eastern shore, the substratum was dominated by sparse beds of the tine-bladed seagrass Ruppia maritima. Deep- er pools in this region generally had seagrasses around their margins and loose, unconsolidated sand at greater depths. The highest portions of the sandbar were either bare sand or dominated by Thalassia testudinum. The highest cover of T. testudinum generally occurred in shallow «20 cm deep) tidepools. Sandbar t~- pography changes almost continuously as sand shifts from place to place. Those regions with dense seagrass appear to be the most stable, presumably because of the sediment stabilizing effect of seagrass rhizomes (den Hartog, 1977). Population Dynamics. -Six permanent plots were marked in November 1982 in 292 BULLETIN OF MARINE SCIENCE, VOL. 45, NO, 2, 1989

24 2' - Air - Atr 2D •.• Sea Water 2D e-. Sec Water U 16 16 ~: 12 12 II:j .

-12 c ~ 1 -12

DS D,6 I D' D,' 0 > .3 D,2

-0 >=· .3•

-0.4 .3·• -0.6 IS 22 29 5 12 19 26 2 1 15 22 29 5 12 19 26 JAN FEB DEC JAN 1982 1983 1983 1984

Figure 2. Cohort survivorship curves, water temperatures, daily low air temperatures, and lower low tides during December and January of 1982 through 1984.

the Ruppia zone near the east side of the sandbar. When I returned on 2 December, populations in three of the plots (all located in one area) had declined from an average density of 169 ascidians' m-2 (SD = 40.63) to a density of 0.33· m-2 (SD = 0.58). The remaining three plots experienced no mortality, suggesting that those animals dying in November did so not because of general environmental conditions, but rather because of processes acting locally. Although the causes for the November mortality are not known, it appeared that much sand had shifted in the region where the mortality occurred. I suspect that sand movements either covered or undermined the ascidians. Between 2 December and 11 February all ascidians in the remaining plots died (Fig. 2) and very few living animals were found anywhere on the eastern side of the sandbar. Freezing air temperatures accompanied by strong northerly winds coincided with extremely low early morning tides in December and for several consecutive days in the middle of January (Fig. 2). Because of the unexpectedly rapid decline seen in the 1982-1983 season, I monitored plots more frequently (once per spring tide sequence) during the winter months of 1983-1984. Eight marked plots contained starting densities ranging from 20 to 60 ascidians'm-2 (Fig. 2). Declines of the populations were modest during December despite temperatures as low as -10°C. The lowest temperatures occurred during a neap tide sequence in the latter part of the month. Freezing weather during the subsequent 12 days coincided exactly with a period of very low tides. The entire Molgula occidentalis population crashed over the next 2 weeks (Fig. 2). During this same tide sequence, all of the Modiolus americanus YOUNG: ASCIDIAN PSEUDOPOPULATION DYNAMICS 293

24 20 - Air ..-...... •... ~.•...• •..• Sea Water u 16 .-'-'"L..0 •...... «Q) 12 8 E~::::l .•..• 4 .-E2Q) .~ a.. ~E 0 •....Q) -4 -8 -12

..-... 0.6 E '-'" 0.4 Q)

Q)> -.J 0.2

Q) "U j.:: 0.0 ~ 0 -0.2 -.J L.. Q) -0.4 ~ 0 -.J -0.6 1 8 15 22 29 5 12 19 26 2 DEC JAN FEB 1986 1987

Figure 3. Temperatures and lower low tides during the winter of 1986-1987. living on the western side of the sandbar died. Four years later, mussels still had not recolonized the sandbar (R. Ellington, pers. comm.). Molgula occidentalis did not recruit to the sandbar during 1984 or 1985. In- termediate numbers of individuals appeared by the autumn of 1986. These were sampled quantitatively on 28 January 1987. There was little evidence of mortality at that time. The population survived the remainder of the winter and persisted until at least November 1987 (J. Dalby, pers. comm.). This was the only time intertidal Molgula occidentalis survived the winter during the 5-year study period. This persistence may have been due to a mild winter. During the 1986-1987 winter season, a single day of - 1.O°Ctemperature coincided with the lowest tide of the year in mid December. Other freezing days were isolated, never reached temperatures lower than -4°, and (with a single exception) coincided with tides no lower than -0.2 m. The meteorological data needed to compare the relative severity of the winters (e.g., hourly wind speed for computation of heat loss to convection; actual tidal exposure times) are not available for the study site. How- ever, because water temperature in the nearshore environment tracks consistent air temperature parameters, it provides an indirect, relative measure of the stress ascidians might have encountered during exposure. Water temperatures reached 294 BULLETIN OF MARINE SCIENCE, VOL. 45, NO.2, 1989

Table I. Summary of winter population observations over 5 years

Highest observed density (indi victuals· m-2) Year Oct.-Nov. Jan.-Feb. Observations 1982-1983 215 o Abrupt decline during cold January low tides, 1983-1984 61 o Unusually cold January and December tides eliminated mussel beds as well as ascidians. 1984-1985 0 o No recruitment of either Mo/gu/a or Madia/us during entire year. 1985-1986 0 No Mo/gu/a recruitment, 1986-1987 54 Molgula colonized both sides of sandbar. Modiolus beds re- placed by Thalamoporella jloridana, which provide new ascidian habitat. Mo/gu/a survive winter low tides for first time in five years.

10°C in 1983-1984 and 1982-1983 (Fig. 2), whereas the low was 12.5° in 1987 (Fig. 3). The 12.5° temperature persisted for only 1 week in 1987. By contrast, water temperature remained below 12.5° C for over 3 weeks in December and January of 1983-1984 (Fig. 2). The observed year-to-year variations in winter population phenomena are summarized in Table 1. Predation. -Numerous species of predatory gastropods occur on Bay Mouth Bar (Paine, 1963). One of these, Fasciolaria hunteria, was observed preying upon Molgula occidentalis on numerous occasions in the field. The snail inserts its proboscis into a siphon of the ascidian and consumes the internal organs, leaving the tunic intact. Because all M. occidentalis occurring on the sandbar were completely covered with sand, it seemed possible that sand could deter the predators to some extent, either as a camouflage device or as a direct defense mechanism. In laboratory experiments, adult M. occidentalis with and without attached sand were offered to F. hunteria (Table 2). All animals without sand were consumed, whereas only two of the animals with intact sand coatings were eaten. Mortality in cages and fences was compared with mortality of uncaged, marked control individuals during the population decline of the 1983-1984 season (Fig. 4). All three treatments experienced virtually identical mortality in each run of the experiment, thus demonstrating that predators play an insignificant role in the winter population crashes. Juvenile Mortality and Growth. - Mortality of juveniles in the flow-through seawater system was highest during the first month after settlement (Fig, 5). In low flow (i.e., high silt) regions of the tank, M. occidentalis juveniles all died, whereas in the high flow region, mortality leveled off after the first 1.5 months. Ascidian juveniles demonstrated the following daily growth rates in the high flow quadrats:

Table 2. Pooled data and contingency table analysis of laboratory predation experiments with Fas- ciolaria hunteria

Treatment Eaten Not eaten Total With sand 2 7 9 Without sand 9 0 9 Total II 7 18 G = 7.271, P < 0.01. YOUNG: ASCIDIAN PSEUDOPOPULATION DYNAMICS 295

100 E::22 Cage _ Fence 80 c::J Control

---~ '-" 60 -+-'>- 0 -+-' L- 40 a ~ 20

0 9 to 21 Dec 21 Dec to 5 Jan 5 to 19 Jan 1983 1984 Figure 4. Mortality of caged, fenced and uncaged individuals during three intervals in 1983 and 1984. Sample sizes for each treatment were 16.

100 •....• High Flow ~ ~,~ •..• Low Flow '" '",,,~~ 80 ,,",," " ,," ,"",.... , ,,,,"" ,,,,,, " 0.. 60 , , ,,'',, ..c , ',", Ul ,, ",,, I.- , " o ,, ",,' > , " > 40 , " I.- , " ::J " , ',',.... ,, (f) , , , , ' " ,, '' ", 20 , ' 'e.... , ', , ,, , ,, .' ,, o 1 8 15 22 29 6 13 20 27 APR MAY Figure 5. Survivorship of juvenile cohorts in high flow and low flow conditions in a flow-through laboratory seawater table. 296 BULLETIN OF MARINE SCIENCE, VOL. 45, NO.2, 1989

~ 21 Dec 1983 28 Jan 1987 ~~~ ~- ..

/\ ~

/~ :~ ~~.__ ~~-~.L.·""_'_"~~ __ ._L__ }/

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~o~~'o-~20-~30-~40--so Distance Alang Transect (m) Figure 6. Ascidian densities on nine identical, evenly-spaced quadrats sampled in December 1983 and again in January 1987.

Quadrat A, 0.844 ± 0.465 (SD) mm; Quadrat B, 0.095 ± 0.094 mm; Quadrat C: 0.107 ± 0.152 mm. Growth rates did not differ significantly among quadrats. Distribution. - The large array of 50-m transects was sampled on two low tides, one in December 1983 and the other in January 1987. During both years, most ascidians were concentrated near the eastern and western shores of the sandbar (Fig. 6). The normally subtidal sandflat regions at the edges of the bar were devoid of Molgula occidentalis populations; highest densities were found in the eastern Ruppia maritima zone in 1983 and in the easternmost portion of the Thalassia zone in 1987. In 1983, many M. occidentalis were attached to the mussels near the western shore. In 1987, this same zone contained many ascidians, but the latter were attached to erect, branching bryozoans (Thalamoporella jloridana), which in turn nestled in the Modiolus americanus shell rubble remaining from the former beds. The habitats occupied by Molgula occidentalis during the 1987 sampling period differed significantly from the distribution expected on the basis of habitat availability (Fig. 7). Ascidians occurred less often than expected on exposed sand and slightly more often than expected on submerged sand, on substrata covered with Thalassia testudinum. and on hard surfaces such as shell rubble and sponges. The individuals on submerged sand were seldom anchored; they were mostly loose individuals or clumps that had apparently been washed into pools. In 1987, the distribution of ascidians on the eastern shore was shifted into the T. testudinum zone, whereas in 1983 it had been centered in the Ruppia maritima zone (Fig. 6). This shift apparently occurred because erosion of sand had created small hummocks of T. testudinum with roots and rhizomes exposed around the edges. Most individuals of M. occidenta/is were attached to these exposed rhizomes. Scales of aggregation were investigated in 1983 by counting grids of 25-cm quadrats and also a 100-m long belt transect of I-m quadrats in the high density region of the Ruppia maritima zone. The large grid contained a II-em deep pool in the middle of it. In order not to bias the data by including populations overlying YOUNG: ASCIDIAN PSEUDOPOPULATION DYNAMICS 297

140 E2.:2I Observed c:::::J Expected 120

(f) 100 G = 477.14 0 ::l -0 p < 0.001 > 80 -0 C '+- 60 0

0 Z 40

Figure 7. Expected and observed distributions of Molgula occidentalis on various substrata in 1987. an obvious habitat change, those quadrats falling in the pool were not included in the initial pattern analysis, but will be discussed later. Significant aggregation was detected at 25- and 50-em scales in the grid data and at l-m, 2-m, 5-m and 10-m scales in the transect data (Table 3). Because the grid data were taken in a relatively homogeneous environment, the presence of small-scale aggregation sug- gests the possibility that gregarious larval settlement influences spatial pattern. Substratum elevation differences also appear to explain some of the density variation at small to intermediate scales (Fig. 8). The highest densities occurred at elevations between tidepoollevel and +4 em elevation. Just a few em below the surface of the pool, densities dropped to zero. This abrupt change corresponded approximately with the region where Ruppia rhizomes became less abundant and the sand became more loose. More than 4 em above the level of the pool, population densities were also low, suggesting that either juveniles or adults may require some threshold of sediment moisture in order to survive low tides. Clumps of adult ascidians (N = 3, >20 individuals per clump) transplanted to the bottoms of three deep pools on 10 December 1983 persisted until 10 January 1984, then disappeared near the end of January, when the rest of the population crashed. It is not known whether the ascidians died or were washed out of the pools. Nevertheless, their survival in pools for nearly a month demonstrates that physical and chemical conditions in the pools do not prevent adults from living there. 298 BULLETIN OF MARINE SCIENCE, VOL. 45, NO.2, 1989

••.•...•...... I fT1 E 200 (1) < 0 0 ~ Z 0 '-'" ::J >. ••.•...•... +' () (J) c 100 3 Q) '--" 0

Distance Along Transect (m)

Figure 8. Densities (mean ± SD) of M. occidenta/is in a grid overlying a deep pool of seawater. Elevations and depths are plotted as connected points.

Larval Development and Settlement. - Unlike many stolidobranchs, Molgula OC- cidentalis is completely capable of self-fertilization. At 25°C, which is near ambient temperature during most of the spring and summer recruitment season, cleavage begins only 45 minutes after fertilization and larvae hatch in 12.5 h, at which time they are competent to undergo metamorphosis. Thus, the embryonic period is unusually short in this species, and dispersal potential is more limited than in many other solitary ascidians.

Table 3. Indexes of dispersion (variance/mean) at several spatial scales in the Ruppia zone of Bay Mouth Bar, 21-22 December 1983. Significance of the ratio was tested by direct comparison with a chi-square table for sample sizes less than N = 31. For larger samples, the normal variable "d" was calculated and tested against a Z table (Elliot, 1977). Two different data sets were used. For scales larger than I m-2, samples were drawn from a 100 m long belt transect. Data for smaller scale analyses came from two 6 x 12 grids of contiguous 25 x 25 cm quadrats

Varianeel Spatial scale N Mean Variance mean d P Small scale (grid): 25 x 25 cm 144 5.06 15.15 2.99 12.36 <0.001 SO x SOcm 36 20.25 116.42 5.75 11.75 <0.001 75 x 75 cm 9 44.94 413.71 9.21 n.s. 100 x 100 cm 6 82.50 734.41 8.90 n.S. Large scale (transect): I x 1m 100 13.16 217.86 16.55 43.21 <0.001 Ix2m SO 26.32 651.78 24.76 39.41 <0.001 Ix4m 25 52.65 1,903.60 36.16 36.16 n.s. I x 5 m 10 131.60 8,817.20 67.00 <0.001 1 x 10 m 5 263.20 5,530.90 21.0 I <0.001 YOUNG: ASCIDIAN PSEUDOPOPULATION DYNAMICS 299

100 _ Sand (Adult) ~ Sand (Ambient) 80 mmm Adult Tunic "0 Q) ;; .-Q) 60 (/) c -Q) 0 40 Q)'- a. 20

Figure 9. Percentage of larvae selecting three different habitats in 12 experimental runs.

Larvae do not require rocks or other hard substrata on which to settle. Given only clean sand as a substratum, they attach initially to sand grains by means of a single large primary ampulla which expands shortly after metamorphosis. Sand grains attached to the tunic surrounding this long ampulla appear to weight down the juvenile while additional ampullae are being produced. After several hours, new ampullae expand the juvenile tunic, which adheres to sand grains and other substrata (e.g., seagrass blades) all around the body. Thus, the characteristic sand coating of this species begins from the very moment of settlement. Larvae were offered a choice of clean, unconsolidated sand from Bay Mouth Bar, sand removed from the adults and containing the characteristic Ruppia maritima debris and other material from the adult habitat, and bare pieces of adult tunic. In every trial but one, sand from the adult habitat was selected significantly more often than either of the other two substrata (Fig. 9). This result differs significantly from the expected number of runs with these rank orders of treatments (G = 23.412; P < 0.001). Since adult tunic was less attractive than sand from the adults, it seems unlikely that larvae settle gregariously in response to adult cues alone. If they aggregate using cues from conspecifics, the cues also require the presence of sand grains from the habitat. In a 2-factor experiment designed to test the effects of parentage and substratum availability on delay of metamorphosis, animals settled on all substrata offered (Fig. 10). Two of the sibling groups settled in polystyrene dishes at a lower fre- quency than on natural substrata. As in the choice experiments, a higher percentage oflarvae underwent metamorphosis on adult sand than in other treatments. Both main effects, "sibling group" and "substratum," contributed significantly to the ANOV A model, as did the 2-way interaction term (Table 4). The fact that sibling groups displayed different behaviors is preliminary evidence that behavior of larvae has a genetic component in this species, and may therefore be influenced by natural selection.

DISCUSSION When Molgula occidentalis occur in the sandy intertidal zone, they form pseudopopulations rather than true self-perpetuating populations. Often they grow to reproductive maturity before dying (as evidenced by the presence of fertilizable eggs in intertidal individuals), but any offspring produced and retained locally must die when the parents do. The pseudopopulations can be viewed as temporary 300 BULLETIN OF MARINE SCIENCE, VOL. 45, NO.2, 1989

70 EZa Sand (Ambient) _ Sand (Adult) 60 DIDDDIIAdult Tunic c:J Polystyrene -0 50 Q) -+-' -+-' Q) (f) 40

L- Q) .J:) 30 E :J Z 20

0 A B c Sibling Group Figure 10. Number of larvae settled on various substrata after 12 h. Larvae from three different parents were examined separately. N = 5 replicates per treatment. extensions of the population into a region higher than the upper species boundary (Mayr, 1970). The ephemeral nature of these particular pseudopopulations can be attributed primarily to meteorological events. Although predators are known to control other ascidians (Gulliksen and Skjaeveland, 1973; Young, 1985), they have a negligible effect on M. occidentalis in comparison to the effects of winter exposure. Thus, the upper species boundary is limited not by the absence of suitable habitat or by predators, but by physical factors associated with the ter- restrial environment. Newell (1979) has reviewed the effects offreezing on intertidal organisms. Many temperate bivalves can tolerate ice crystals in their tissues for a period of time, as well as the tissue dehydration that results from ice for- mation. Although air temperatures often fell below O°C on Bay Mouth Bar, sea water ice was never observed there. The high survival of M. occidentalis during the 1986-1987 winter season demonstrates that this species can tolerate short periods of exposure to sub-freezing temperatures. Prolonged exposure to lower temperatures occurred in the years of population crashes. A study of physiological tolerance limits might be warranted to clarify these field data. It is interesting to consider why no populations of Molgula occidentalis colonized the eastern shore of Bay Mouth Bar for two years following the extreme cold of 1983-1984. Perhaps overwintering source populations were also killed that year. Before 1984, many ascidians occupied the interstices of the Modiolus americanus beds in the shallow subtidal portions of the western and southern shores. If some of these individuals survived the winters, they could have repopulated the eastern portion of the sandbar on a regular basis. The dispersal period of M. occidentalis is only 12 h long, and the embryos do not swim actively during this period. Thus, YOUNG: ASCIDIAN PSEUDOPOPULATION DYNAMICS 301

Table 4. Two-way fixed factor ANOYA for larval delay of metamorphosis experiment (Fig. 10)

SOUTce or variation df Sums of squares Mean square F p Sibling group 2 507.4 253.7 7.8 <0.01 Substratum 3 937.5 312.5 9.6 <0.001 Interaction 6 744.4 124.1 3.8 <0.01 Error 48 1,554.8 32.4 dispersal distances are probably quite short. Prevailing winds during the summer and spring recruitment seasons come from the south and west, so they probably generate the kinds of currents that would carry embryos across the sandbar rather than offshore. Besides the former populations in the Modiolus beds, the only large potential source populations that have been found during several years of trawling in the region are located on the west side of Turkey Point, approximately 7 km away. Thus, I speculate that absence of a local source population could have resulted in 2 years with no recruitment.. When the western shore of the sandbar became populated with large, erect bryozoan colonies, a stable substratum once again became available. By January 1987, both the bryozoan assemblage and the eastern shore had once again been colonized by M. occidentalis. Ascidians living on soft sediment often display adaptations for anchoring themselves and resting above the surface. Molgula occidentalis is no different in this respect. Fine projections of tunic anchor the animal on all sides not only to the sand or mud itself, but to any sediment stabilizing structure nearby. Seagrass roots, rhizomes and blades seem to be the most important such stabilizing features in the intertidal sandbar habitat. The ability of seagrasses to hold sand is well known (Wilson, 1949; den Hartog, 1977). Several lines of evidence suggest that sediment stability is an important habitat feature for Molgula occidentalis. First, although juveniles of M. occidentalis tolerate much more overlying sediment than those of a sympatric species, Styela plicata (Young, unpublished data) and of typical hard-bottom ascidians (Young and Chia, 1984), too much sediment ac- cumulation quickly kills them (Fig. 5). Second, large-scale adult mortality occurring during November, 1983 seemed to be due to shifting sand. Finally, very few ascidians occurred on loose sand that is not stabilized by seagrasses or other structures (Fig. 7). Likewise, high densities near the edges oftidepools contrasted markedly with the absence of animals a few centimeters underwater where no stabilizing seagrasses were present. Berrill (1931) outlined developmental modes in the family Molgulidae and demonstrated that many species occupying soft sediments have evolved direct ("anural") development. Moreover, some molgulid tadpoles have a reduced com- plement of sensory structures in the cerebral vesicle. Many are blind. It has been widely presumed (Berrill, 1931; Millar, 1971; Young et al., 1988) that loss ofa swimming tadpole stage in some species and loss of ocelli in others have arisen because neither swimming nor habitat selection is essential for animals occupying homogenous soft-sediment environments. The data on Molgula occidentalis con- tradict these ideas on several counts. Even though this species lives in a sedimentary environment, its habitat is far from homogeneous. Subtle differences in habitat stability appear to be enormously important for survivorship. Under these circumstances, one might expect habitat selection to have evolved. Larvae of Molgula occidentalis lack an ocellus and have very reduced oral papillae (Young, unpublished; Torrence and Cloney, 1981). Nevertheless, tadpoles can distinguish light from dark and are adept at differentiating between loose sand and sand 302 BULLETIN OF MARINE SCIENCE, VOL. 45, NO.2, 1989 associated with the more stable adult habitat, Presence of an adult is not enough to cue settlement; adult tunic alone was less attractive to the tadpoles than sand and associated debris removed from the surface of the adults. These behaviors probably did not evolve in the intertidal zone, since intertidal mortality tends to be cataclismic rather than selective. Nevertheless, the same behaviors are likely to be important in distinguishing stable and unstable habitats in the shallow subtidal zone.

ACKNOWLEDGMENTS

This paper is dedicated to Donald P. Abbott, who encouraged my interest in ascidian ecology early on. J. Rhymer, R. Ellington, P. Cummings, B. Bingham, and J. Dalby assisted with intertidal sampling. I thank L. Cameron, B. Bingham and A. Davis for comments on the manuscript. This work was supported by NSF grant OCE-8400406. Contribution number 1048 from the Florida State University Marine Laboratory and Harbor Branch Contribution No. 672.

LITERATURE CITED Berrill, N. J. 1931. Studies in tunicate development, II. Abbreviation of development in the Mol- gulidae. Phil. Trans. Roy. Soc. B. 219: 281-346. Connell, J. H. 1972. Community interactions on marine rocky intertidal shores. Ann. Rev. Ecol. Syst. 3: 169-192. den Hartog, C. 1977. Structure, function, and classification in seagrass communities. Pages 89-119 in C. P. McRoy and C. Helfferich, eds. Seagrass ecosystems. Marcel Dekker, New York and Basel. Elliott, J. M. 1977. Some methods for the statistical analysis of samples of benthic invertebrates. Freshwater Biological Assoc. Cumbria, England. 160 pp. Gulliksen, B. and S. H. Skjaeve1and. 1973. The sea star, Asterias rubens L., as predator on the ascidian, Ciona intestinalis (L.), in Borgenfjorden, North Trondelag, Norway. Sarsia 52: 15-20. Mayr, E. 1970. Populations, species, and evolution. Harvard University Press, Cambridge and London. 453 pp. Millar, R. H. 1971. The biology ofascidians. Adv. Mar. BioI. 9: 1-100. Mileikovsky, S. A. 1961. Character and nature of deep-water eurybathic forms of invertebrates with pelagic larvae taking as an example the polychaete Euphrosyne borealis Oersted 1843 from the North Atlantic. Okeanologiya 1: 687-697. Newell, R. C. 1979. Biology of intertidal animals. Marine Ecological Surveys, Ltd., London. 781 pp. Paine, R. T. 1963. Trophic relationships of 8 sympatric predatory gastropods. Ecology 44: 63-73. --- and T. H. Suchanek. 1983. Covergence of ecological processes between independently evolved competitive dominants: a tunicate-mussel comparison. Evolution 37: 821-831. Roughgarden, J., Y. Iwasa and C. Baxter. 1985. Demographic theory for an open marine population with space-limited recruitment. Ecology 66: 54-67. Stephenson, T. A. 1942. The causes of the vertical and horizontal distribution of organisms between tidemarks in South Africa. Proc. Linn. Soc. Lond. 154: 219-232. Svane, I. ]983. Ascidian reproductive patterns related to long-term population dynamics. Sarsia 68: 249-255. --- and T. Lundalv. 1982a. Persistence stability in asci dian populations: long-term population dynamics and reproductive pattern of Pyura tesselata (Forbes) in Gullmarfjorden on the Swedish west coast. Sarsia 67: 249-257. --- and ---. 1982b. Population dynamics and reproductive patterns of Boltenia echinata (Ascidiacea) on the Swedish west coast. Neth. J. Sea Res. 16: 105-118. Thorson, G. 1946. Reproduction and larval development of Danish marine bottom invertebrates. Medd. Komm. Danm. Fisk.-og Havunders., ser. Plankton 4: ]-523. Torrence, S. A. and R. A. Cloney. 1981. Rhythmic contractions of the ampullar epidermis during metamorphosis of the ascidian Molgula occidentalis. Cell Tissue Res. 216: 293-312. Underwood, A. J. 1985. Physical factors and biological interactions: the necessity and nature of ecological experiments. Pages 372-390 in P. G. Moore and R. Seed, eds. The ecology of rocky coasts. Hodder and Staughton, London. --- and P. G. Fairweather. 1986. Intertidal communities: do they have different ecologies or different ecologists? Proc. Ecol. Soc. Aust. 14: 7-16. Wilson, D. P. 1949. The decline of Zostera marina L. at Sa1combe and its effects on the shore. J. Mar. BioI. Assoc. U.K. 28: 395-412. YOUNG:ASCIDIANPSEUDOPOPULATIONDYNAMICS 303

Young, C. M. 1985. Abundance patterns of subtidal solitary ascidians in the San Juan Islands, Washington, as influenced by food preferences of the predatory snail Fusitriton oregonensis. Mar. BioI. 91: 513-522. --- and F. S. Chiao 1984. Microhabitat-associated variability in survival and growth of subtidal solitary ascidians during the first 21 days after settlement. Mar. BioI. 81: 61-68. --, R. F. Gowan, J. Dalby, Jr., C. A. Pennachetti and D. Gagliardi. 1988. Distributional consequences of adhesive eggs and anural development in the ascidian Molgula pacifica (Hunts- man, 1912). BioI. Bull. 174: 39-46.

DATEACCEPTED: September 9, 1988.

ADDRESS: Division of Marine Science, Harbor Branch Oceanographic Institution, Inc., 5600 Old Dixie Highway. Ft. Pierce, Florida 34946.