The relationship of size and biomass to fission rate in a clone of the sea anemone, Haliplanella luciae (Verrill)

J. Exp. Mar. Biol. Ecol., 1982, Vol. 58, pp. 151-162

Elsevier Biomedical Press

THE RELATIONSHIP

OF SIZE AND BIOMASS

FISSION RATE IN A CLONE

HALIPLANELLA

TO

OF THE SEA ANEMONE,

LUCIAE

(Verrill)’

LEO L. MINASIAN, JR.~ Department of Biological Science, Florida State University, Tallahassee, FL 32306, U.S.A

Abstract:Genetically identical clones of Haliplanella luciae (Verrill) were reared in vitro? and the rate of asexual reproduction (fission), individual dry weight and clonal biomass was determined. When temperature increases, the size of individual anemones decreases due to an increased fission rate (k) and increased catabolism. At 26 “C increased food availability caused decreased individual size, due to increased k; at 21 and 16 “C, no feeding effect was evident. Mean individual size is a linear function of k; the resulting regression equation is given, and its use for prediction of k in natural populations is discussed. At low values of k (i.e., low temperature), clonal biomass increases in proportion to feeding frequency. At high values of k, biomass decreases, indicating that k is maximized at considerable cost in terms of tissue growth. This suggests that for H. luciae, maximizing k provides a benefit other than that of maximizing net energetic input and growth efficiency.

INTRODUCTION

Past literature has demonstrated that certain species of sea anemones rely primarily upon asexual reproduction for recruitment (e.g., Davis, 1919; Stephenson, 1929; Shick & Lamb, 1977; Bucklin, 1979). Several investigators have suggested possible advantages accrued from asexual reproduction in sea anemones. These include : reduced desiccation and mechanical drag within clonal groups (as opposed to individual anemones) living in exposed, intertidal habitats (Francis, 1976); increased competitive capability through clonal expansion, and recognition and destruction of genetically distinct sea-anemone tissue (Bonnin, 1964; Francis, 1973a, b, 1976); increased clonal nutrition through increasing the clonal feedingsurface area (i.e., tentacular surface) available for prey capture (Smith & Lenhoff, 1976; Shick & Lamb, 1977; Sebens, 1979); and selective proliferation of those genotypes which have high fitness in specific locales (Shick & Lamb, 1977; Shick et al., 1979). One of the most widely distributed asexually reproducing species is an intertidal sea anemone, Haliplanella ( = Diadumene) luciae (Verrill) (e.g., Stephenson, 1935 ;

’ Contribution No. 130 of the Tallahassee. Sopchoppy and Gulf Coast Marine Biological Association. ’ Present address: University of Houston, Marine Science Program, 4700 Avenue U. Bldg. 305, Galveston, TX 77550, U.S.A. 0022-0981/82/0000-0000/$02.75 0 1982 Elsevier Biomedical Press

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LEOL.MINASIAN,JR.

Minasian & Mariscal, 1979). All observed reproduction in H. luciue has been asexual, usually through binary, longitudinal fission (Davis, 1919; Minasian, 1976). Minasian (1976, 1979) developed laboratory culture methods which revealed that fission occurs not so much in response to the size of individual anemones, but primarily in response to exogenous factors. This led to the inference that both fission rate (k) and its response to exogenous factors would determine the success of H. Zuciae in natural populations. Moreover, the observation that exogenous factors supersede endogenous factors as regulators of fission activity implied that individual sizes and morphological parameters within sea-anemone populations depend upon k and the environmental factors which control it (Francis, 1976; Johnson & Shick, 1977; Minasian, 1976, 1979). However, that size per se may have an effect upon k is suggested by restriction of fission activity to those anemones which attain a minimal size (Sebens, 1980). Minasian (1979) quantified some morphological parameters of in vitro populations of H. luciae which were sensitive to changes in k, which in turn responds to temperature and feeding frequency (Minasian & Mariscal, 1979). The present investigation ascertains the relationship between size (dry wt) and k in identical, clonal cultures of H. luciue, thus eliminating genetic differences as a source of variation. These data quantify the effects of environmental parameters on individual size and clonal biomass in relation to k. It is concluded that longitudinal fission imparts a marked cost on clonal biomass when k is large. This indicates that maximization of k as a means of achieving increased biomass through increased food intake (Sebens, 1979) is only applicable to H. luciae over a limited range of k values. The present study also indicates limits to the size range within which fission occurs in H. luciae. Conditions under which size may serve for the prediction of k are suggested.

MATERIALSAND

METHODS

ESTABLISHMENTOFCLONALANDEXPERIMENTALCULTURES

The procedures by which stock clonal cultures of H. luciue were established and maintained have been described in Minasian & Mariscal (1979). All anemones are probably from the same clone since they are identical in color pattern and sex (male). Electrophoretic analysis of individuals from the present clone (McCommas, pers. comm.) revealed that they are genotypically identical, being heterozygous at the hexokinase locus, which distinguishes them from two other clones at the collection site (Shick & Lamb, 1977). To summarize, three anemones were isolated from the same collection site on the Florida Gulf coast (29”54.8’N : 84”3O’W) in October, 1976. In the laboratory, they were fed Artemia nauplii, and cloned in small glass culture vessels. Once established, cloned “stock cultures” were maintained for several months under constant temperature (16 to 18 “C) and photoperiod

FISSION, SIZE AND BIOMASS

13

(14 h/l 0 h : light/dark). This stock of acclimated, cloned anemones provided animals for subsequent culture experiments. Expe~ments consisted of six replicate cultures which were reared under three different combinations of temperature (1621, and 26 “C) and three different feeding regimens (fed once every 2 days, once every 4 days, and starved) for 30 days, not including a 2-day acclimation period. Fission rate (k) is defined as the instantaneous rate of population increase attributable solely to iongitudinal fission: dN/dt = kN, where N is the population size. Since this rate is an exponential constant characteristic of each individual culture, k was calculated as the slope of the least-squares regression of the natural log number of anemones (In Ni) versus time (Minasian, 1979; Minasian & Mariscal, 1979). Only some of these experimental cultures showed sufficient, exponential increases to permit calculation of k: fed once every 2 days at 16, 21, and 26°C; fed once every 4 days at 21 and 26°C; fed twice per week at 17°C for 92 days (stock cultures). Thus, six values of k were obtainable from each of six different culture regimens, providing a total of 36 k values. Results of these culture data are illustrated and tabulated in Minasian & Mariscal (1979).

DETERMINATION

OF INDIVIDUAL

DRY WEIGHTS AND STATISTICAL ANALYSIS

In order to determine the functional relationship between k and the mean individual dry weight (w) for each culture, 15 individual anemones, or half of the total culture population (whichever was larger) were randomly selected for dryweight determinations. Anemones were fixed in 5% formalin in sea water for 2 days prior to weighing, then placed indi~dualIy in small, tared, glass shell vials, and dried for 24 h at 105 “C. A mean value of w was then determined for each value of k. I assume w to be the response variable dependent upon k (Minasian, 1976). Thus, a least-squares linear regression of w on k was computed, as well as the correlation coefficient. Since this regression equation was used to predict k from known values of w, it was necessary to utilize the reversed form of the regression equation (Winsor, 1946). Statistical methods permit calculation of confidence limits for the inverse prediction, and are based upon either the F- (Seber, 1977) or t-distribution (Sokal & Rohlf, 1969). I applied the latter method. A two-way analysis of variance (ANOVA) (temperature vs. feeding) was performed on square-root transformed data for individual dry weights from all culture regimens: a 3 x 3 factorial design. The 50/;,significance level was employed for all statistical tests.

DETERMINATION

OF CULTURE CLONAL BIOMASS AND STATISTICAL ANALYSIS

The biomass of each culture was calculated in order to determine temperature and feeding effects on clonal biomass. Since 50% or less of the culture populations were used in the dry-weight analysis, clonal biomass was calculated as the product

LEO L. MINASIAN, JR

154

of the mean individual dry weight (w) and the total number of individuals (N) in each culture. After six such determinations were made, means and SDS for clonal biomass were calculated for all temperature-feeding combinations. Environmental effects on biomass were also analyzed by a two-way ANOVA on square-root transformed data in a 3 x 3 matrix, as done for individual dry weights.

RESULTS EFFECTS OF TEMPERATURE

AND FEEDING ON INDIVIDUAL

SIZE AND CLONAL

BIOMASS

The statistical analysis in Table I of individual mean dry weights demonstrated that only temperature had a significant effect upon size. This effect was due to an increase in k over the 16 to 26 “C temperature range (Minasian & Mariscal, 1979). For example, an increase of 10°C resulted in an average decrease in size of 77% among culture populations fed once every 2 days, after 30 days (Table I). Similarly, anemones fed once every 4 days showed a 69% decrease in size when exposed to the same increase in temperature. Among starved individuals fission activity occurred only at 21 “C and 26 “C, and was limited to less than one population doubling (Minasian & Mariscal, 1979). At 26 “C starved anemones were 44% smaller than those starved at 16 “C. Some of this size difference was probably due to accelerated catabolism at the higher temperatures, in addition to weight loss resulting from fission. TABLEI Dry weights (grand mean f SD) of individual H. luciae reared for 30 days under different combinations of temperature and feeding frequency: each grand mean is based on means from six replicate clonal cultures; the total number of anemones included in each determination of a grand mean is given in parentheses; a two-way ANOVA, examining temperature and feeding effects on mean dry weights among experimental cultures, was performed on square-root transformed data; results indicate if exogenous effects were significant (+) or not significant ( -) at P < 0.05. Temperature (“C) Feeding frequency

16

21

26

2 days

2.550 + 0.588 (50)

f ,204 f 0.284 (90)

0.597 f 0.068 (90)

4 days

2.564 f 0.585 (39)

1.313f 0.633 (90)

0.790 f 0.142 (90)

Starved

1.767 f 0.493 (30)

1.077 f 0.279 (39)

0.990 f 0.219 (47)

Feeding frequency (-)

Interaction (-)

ANOVA effects

Temperature (+)

FISSION,

SIZE AND

155

BIOMASS

Table I indicates that feeding may have a significant effect on size at some temperatures (e.g., 26°C) but not at others (e.g., 21 “C). Thus, overall magnitude of the feeding effects is not significant in the ANOVA. At 26 “C, increasing food availability increased k, and tended to decrease size. At 21 “C, the decreased size resulting from stimulation of k in fed anemones was similar to the loss of size due to catabolism in the starved group (Table I). Although fission did not occur among starved anemones at 16 “C, fed anemones at 16 “C tended to be larger, notwithstanding the occurrence of limited fission. This can be attributed to net anabolism due to the availability of food at 16 “C. TABLE II

Biomass (mean *SD) of six clonal cultures of H. luciae reared under different combinations of temperature and feeding frequency for 30 days; biomass for each replicate culture was calculated from the product of the mean dry weight (w) and the total number of anemones (N,) present at the time of the weight determination; total number of anemones included in each biomass determination is in parentheses; a two-way ANOVA, performed on square root-transformed data, indicates if effects of temperature or feeding frequency on biomass are significant (+ ) or not significant ( -) at P < 0.05. Temperature

(“C)

16

21

26

42.36 +- 7.51

57.61 f 13.69

50.50 + 6.47

(50)

(90)

(90)

4 days

30.84 f 5.37

17.25 + 12.32

49.98 +- 7.76

Starved

17.67 k 4.93

Feeding

frequency

2 days

(39)

(90) 13.50 f

(30) ANOVA

effects

Temperature (+)

(90) 4.07

(39) Feeding

frequency (+)

14.93 f 3.10 (47) Interaction (-)

Table II shows that both temperature and feeding had significant effects upon clonal biomass. The maximal clonal biomass over the 30-day culture period was accrued at 21 “C. At 26 “C, clonal biomass, like mean individual size, was smaller, presumably due to enhancement of k by the higher temperature. At 16 “C, clonal biomass was also less than that in cultures reared at 21 “C, although individual size was greater at the lower temperature (Table I). If anemones were not fed, clonal biomass varied inversely with temperature, indicating that increased temperature reduces biomass by increasing catabolism and fission activity (Table II). Fed clones accrued much more biomass than starved clones at all temperatures (Table II). However, Table II shows that biomass was not necessarily maximized in clones receiving the most food, but was equally as high (26 “C) or higher (21 “C) in clones fed once in 4 days. At 16°C doubling the feeding frequency increased clonal biomass by 29%. The opposite effect occurred at 21 “C: doubling feeding

LEO L. MINASIAN,

156

JR

frequency reduced clonal biomass by 34%. Thus, the benefit of increased clonal biomass did not result from increased food availability at higher temperatures.

Fed

every

Fed every

second

day

fourth

day

starved 518

rl -

o-

Culture

Temperature

Fig. 1. Clonal biomass of H. luciue reared at different combinations of temperature and food availability for 30 days: biomass values represent a mean of six cultures; error lines are SDS; values of k, where calculable, are given atop histograms, and consist of a mean of six cultures.

This is clarified by Fig. 1, which demonstrates that biomass depended upon not only temperature and nutrition, but also upon k. When k was low (16 “C), clonal biomass was minimal, and increased in proportion to food availability. At temperatures above 20 “C, where k was moderate to large (i.e., k > 0.05), biomass did not increase in relation to food availability. Instead, biomass changed inversely with k. Thus, k is maximized at the expense of biomass. FUNCTIONAL

RELATIONSHIP

BETWEEN

DRY WEIGHT

AND

FISSION

RATE

Mean dry weight per anemone (w) was plotted as a function of k for 36 clonal cultures of H. luciae (Fig. 2). This function is given by the regression equation: w = -37.1447k + 3.2985. The correlation coefficient (r = -0.9332) indicates the

FISSION,

SIZE AND

157

BIOMASS

strong, inverse relationship between variables. In order to use w as a means of predicting k, the above function was algebraically rearranged into the form appropriate for the inverse prediction : t%= (3.2985 - w)/37.1447. Here, the estimated fission rate (l) is expressed as a function of w.

.

1

3.6

‘s r= -0.9332

32-

1.6-

Oi-

0.4-

O.o-. 0.010

,

.,

, 0.020

1,.

I,,

0.030

1

I,,

I

0.040 Fission

I,

0.050 Rate

I

I,,

,

0.060

,

,

,

,

,

0.070

(k)

Fig. 2. Least-squares regression of w on k: each point is a mean dry weight of 15 anemones from the same clonal culture, exhibiting a single value of k; numbers are next to those points for which sample sizes were c 15 anemones; the equation given is that used for the inverse prediction of k based on W.

Several previously unknown characteristics of the size-fission rate relationship were revealed by the distribution of points in Fig. 2. First, the spread of points was greater at low values of k. Thus, the predictability of k based on w may be better for smaller sized anemones. Accordingly, lower values of k cause a population of H. luciue to exhibit greater variability in size. Second, the distribution of data points approached a minimal limit (i.e., “bottomed out”) at an approximate dry weight of 0.5 mg (Fig. 2). This suggests that there is a minimal mean size, below which fission activity is inhibited. However, some anemones were smaller, with dry weights as low as 0.1 mg due to the occurrence of unequal fissions (Minasian, 1976). The apparent limit of mean dry weight to 0.5 mg indicates that parent anemones must usually exceed this minimal size in order to execute fission. Likewise, the plot of w vs k indicates the existence of a maximal value of k. The inter-fission time limits the

158

LEO L. MINASIAN, JR.

fission rate, and may represent the time necessary for anemones smaller than the size necessary for fission to grow back to that minimal size. Thus, a maximal value of k is logically linked to a minimal size for the occurrence of fission. Fig. 2 reveals that the maximal value of k for this clone is approximately 0.08, corresponding to a maximal population doubling time of 8.7 days.

DISCUSSION

The pronounced effect of temperature on the size distribution of H. luciae supports laboratory data suggesting that temperature is the primary environmental factor regulating both k (Ninasian & Mariscal, 1979) and the observed size variability in H. hciae (Hand, 1955; Minasian, 1979). Since increased food availability increases individual size through tissue growth, and reduces individual size through enhancement of k, the effects of feeding on individual size depend upon whether or not the temperature permits fission, The tendency of increased temperature and food availability to maximize biomass is balanced by the catabolic effects of increased k. Thus, biomass was optimized at reduced temperatures and levels of food availability. Sebens (1979) constructed a model which predicts optimal polyp size, type of clonal existence, and occurrence of asexual reproduction, given prey items of specified size and availability. He applied this model to all benthic marine invertebrates which exhibit indeterminate growth, but focused on anthozoan polyps, parti~ularIy sea anemones of the genus A~~~op~eu~a. The basic rationale for the model is as follows: if food is limiting, increasing tentacular surface area provides sufftcient energetic benefit to select for specific polyp sizes and kinds of clonal (= colonial in terms of the model) relationships, in order to maximize food intake. Detailed conclusions of this model apply well to H. Iuciae. Under favorable habitat conditions (e.g., warm temperatures, abundant food), the model predicts that k should increase, and polyp size should decrease as previously demonstrated (Minasian, 1976; Minasian & Mariscal, 1979). However, this prediction is contradicted by data from Smith & Lenhoff (1976) showing that abundant food minimizes asexual reproduction (i.e., pedal laceration) in an anemone recently identified as Aiptasiogeton coma&s (Seaton, pers. comm.). Chia (1976) likewise stated that actiniarian asexual reproduction is generally maximized under conditions of inadequate nutrition, or in otherwise unfavorable environments. Sebens’s model predicts that individuals of H. luciae with sizes >0.0023 mg dry wt should undergo fission, assuming that prey is abundant and small (0.5 mm diameter). This prediction is reasonably accurate, since H. luciae always ,exceeds this size value, and can divide continuously (Minasian, 1976). Furthermore, culture data on Ii. fuciae have proved that polyps of all sizes thrive on planktonic prey (viz., newly hatched Artemiu nauplii, average diameter x0.9 mm). Sebens’s model

FISSION, SIZE AND BIOMASS

159

also predicts that polyps feeding upon prey 0.5 mm in diameter should be colonial (or clonal), and exhibit determinate unit size. When small prey are abundant, k is maximized, and the variance in polyp size decreases (Table I; 26 “C, fed every 2 days), thus approaching a constant unit size, in agreement with his model. Sebens (1979) regarded H. luciue as consuming different, larger prey (diameter = 15 mm) than I have assumed for the above application of his model. This is in agreement with Hausman’s (1919) observations that H. luciue feeds on small benthic invertebrates (viz., amphipods, etc.). Based upon prey of this diameter Sebens’s model predicts that polyps should be clonal or colonial, and of indeterminate unit size. This better fits the characteristics of H. luciue than the above prediction, based upon smaller prey size. However, Sebens’s assignment of H. Zuciue as a predator on intermediate-sized prey results in quantitative predictions of polyp size at fission (150 mg) and diameter at fission (2.0 cm) which are in excess of those actually observed either in the laboratory or in the field. Notwithstanding the decreased accuracy of this second prediction, Sebens’s model does generate a set of values within the limits of which the parameters of H. luciue appear to be accommodated. This is the model’s strength. However, the rationale for his model does not apply well to H. Zuczizewhen k is optimized. The purpose of the model was to support the hypothesis that asexual reproduction is a means for maximizing energetic surplus as biomass, including tissue growth and gonad production (Sebens, 1979,198O). The model has apparently been successful in this regard for some species (Sebens, 1979). Fig. 1 showed that a clone of H. luciue amassed optimum energetic benefit when k was close to 0.05. It was at low temperatures, where k approximated 0.02, and both k and biomass increased in proportion to food availability, that Sebens’s model applies. The reduction in biomass at higher temperatures was not due to physiologically stressful culture conditions per se. Indeed, all anemones remained active and healthy in these cultures (Minasian & Mariscal, 1979). Anemones receiving food every second day at 21 “C showed higher k and lower biomass than those receiving half as much food (i.e., fed every fourth day) (Fig. 1). This occurred despite the fact that cultures were fed to repletion with Artemiu nauplii, evident from remaining nauplii at the end of the feeding period and numerous boli egested a day or two after feeding. Thus, maximizing k at both 21 and 26°C caused an apparent reduction in feeding or conversion efficiency (i.e., [increase in clonal biomass]/[no. of feedings per 30-day culture period]). When k ~0.05, asexual reproduction in H. luciue is an energetic liability rather than an asset, being advantageous only in environments or during seasons when k is small. Thus, in environments where k is moderate to large, k is advantageous for reasons other than the strategy proposed by Sebens (1979). Alternative strategies which may account for the observed success of H. luciue have been discussed by Shick & Lamb (1977) and Shick et al. (1979). Since sexual production of larvae is so infrequent as not to have yet been confirmed, much

160

LEO L. MINASIAN,

JR.

dispersal and colonization must be achieved by the usually attached polyp. Minasian (unpubl. data) has suggested that the high probability of reproductive success (i.e., population growth) afforded by maximizing k permits clonal survival in environments which are unpredictable or characterized by frequent disturbance. Data on H. luciue from Johnson & Shick (1977) also imply a negative effect of k on biomass. These authors correctly reasoned that a clone consisting of more individuals per unit area would receive less food per individual than a clone consisting of fewer individuals per unit area, given that both clones received equal aliquots of food. However, if a consequence of more individuals per clone is increased prey capture by a clone (Johnson & Shick, 1977; Sebens, 1979) then increasing k should still result in greater biomass. Since high values of k cause decreased clonal biomass, catabolism associated with frequent fission activity must be great. This supports the conclusions of Calow (1977) and Shick et al. (1979) that partitioning of a clone into a larger number of smaller units causes greater catabolic output per unit weight. Zamer & Mangum (1979) have provided empirical evidence for this: H. luciue shows long-term increases in the rate of oxygen uptake after exposure to a high “developmental” temperature (i.e., high k). I believe that such increased catabolism results from extensive regeneration associated with high k values. Regenerative processes of sea anemones, including both morphallactic regulation of structure (e.g., Cary, 1911; Atoda, 1954) and increased cell proliferation (Singer, 1970, 1971) increase catabolic cost in excess of any anabolic benefit of increased prey capture. The suitability of size as an index of k requires several assumptions. The first is that the relationship between size and k is the same in the field population as in the culture population. If a field population consists of more than one clone (e.g., Uchida, 1932; Hand, 1955) it must be assumed that the response of size to k is the same in all clones. Second, the range of sizes used for the prediction of k in field samples must occur within the range of w and k values obtained in the laboratory, and from which the prediction equation was derived. Third, all individuals in the field population are assumed to originate through fission. Fourth, the correlation between w and k must be sufficiently strong for w to be of predictive value. The size of individual anemones results from the equilibrium between organismal growth and k in fissiparous species (Francis, 1976; Minasian, 1979). Temperature and feeding frequency affects both tissue growth and k simultaneously. Thus, the body size (Minasian, unpubl. data) and morphology (Minasian, 1979) of individuals will always reflect the rate of tissue growth, k and the environmental factors which regulate those rates. The functional relationship between w and k also indicates constraints upon the size at which fission may occur. In previous samples of H. luciae, k was neither responsive to nor dependent upon size (Minasian, 1976, 1979); other studies supported this conclusion (Francis, 1976; Johnson & Shick, 1977). However, the

FISSION,SIZEAND BIOMASS

161

present study, involving a wider range of k values than previous studies, showed that as k approaches a maximum possible value of 0.08, mean dry wt does not drop appreciably below 0.5 mg. Likewise, increased variability in size at lower values of k indicates that some large specimens of H. luciue may still divide several times a year (0.005 < k < 0.01) and the w vs k function may change for many of the larger anemones. There are two inherent consequences of this. First, the probability of a fission event may decrease in large individuals (i.e. > 7.0 mg; Minasian, 1976), although I find no size so large that fission is permanently inhibited. Thus, a population of homogeneously large H. luciae would be expected to exhibit lower values of k than a population of smaller clonemates, when under identical environmental conditions. Second, since sexual maturity accompanies attainment of a relatively large size (assuming prior sterility) (Chia, 1976; Francis, 1976), sexual maturity may accompany a decreased response of k to exogenous factors. Such a decrease in k would then represent the physiological correlate of a transition in reproductive strategy from primarily asexual to primarily sexual reproduction (Minasian & Mariscal, 1979). This is why natural populations of H. luciae which exhibit fission often consist of small, sterile individuals (e.g., Shick & Lamb, 1977; Minasian, 1979, unpubl. data) after several months of high seasonal temperatures.

ACKNOWLEDGEMENTS

I wish to thank Dr. R. N. Mar&al for his support during the course of this study, which was also aided by an award from the Lerner Fund for Marine Research. Information provided by R. W. Seaton (F.S.U.) and S. A. McCommas (University of Houston) was also helpful.

REFERENCES ATODA,

K., 1954. The development of the sea anemone, Diudumene luciue, reproduced by the pedal laceration. Sci. Rep. Tohoku Imp. Univ., Ser. 4, Vol. 20, pp. 123-129. BONNIN,J. P., 1964. Recherches sur la “reaction d’agression” et sur le functionnement des acrorhages d’dcrinia equim L. Bull. Biol. Fr. Belg., Vol. 98, pp. 225-250. BUCKLI~T, A., 1979. Sexual and asexual reproduction of the sea anemone ~e~rid~~~ senile in intertidal populations. Am. Zoof., Vol. 19, p. 926 (abstract). CALOW, P., 1977. Ecology, evolution and energetic+. Adv. Ecol. Rex., Vol. 10, pp. l-62. CARY, L. R., 1911. A study of pedal laceration in actinians. Biol. Bull. (Woods Hole, Muss.), Vol. 20, pp. 81-108. CHIA, F. S., 1976. Sea anemone reproduction: patterns and adaptive radiations. In, Coelenterate ecology and behavior, edited by G. 0. Mackie, Plenum Press, New York, pp. 261-270. DAVIS, D. W., 1919. Asexual multiplication and regeneration in &rgurriu luciue Verrill. J. Exp. Zoo/., Vol. 28, pp. 161-263. FRANCIS, L.. 1973a. Clone specific segregation in the sea anemone Anthopiatra efeganrissima. Biof. B&i. (Woods Hole, Mass.), Vol. 144, pp. 6472.

162 FRANCIS, L., 1973b. Intraspecific

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JR

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