Integrative and Comparative Biology Advance Access originally published online on June 6, 2006
Integrative and Comparative Biology 2006 46(5):598-604; doi:10.1093/icb/icl008
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Natural variability in size and condition at settlement of 3 species of marine invertebrates
Department of Ecology, Evolution and Marine Biology, University of California Santa Barbara, CA 9310, USA
Correspondence: 1E-mail: Nicole.Phillips{at}vuw.ac.nz
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Experimental studies have demonstrated that for many marine invertebrate species, variability in larval condition or quality at settlement may have important effects on post-settlement, early juvenile performance. Relatively few studies, however, explicitly examine natural variability in larval condition at settlement. This study examines natural variability in larval attributes (size and lipid index) at settlement for terminal-stage larvae of intertidal mussels (Mytilus sp.) and barnacles (Pollicipes polymerus and Chthamalus dalli) from southern California. Despite significant differences among cohorts in larval attributes, for all 3 species a greater percentage of the variance in larval length (80–100%) and lipids (58–83%) occurred among individuals within a cohort, rather than among cohorts. For all 3 species, coefficients of variation within a cohort for length were much smaller (3–8%) than those for lipid index (30–93%), suggesting that lipid storage is a much more plastic attribute than size for larvae. For mussels, settlement intensity and larval attributes were decoupled, such that average larval condition of a cohort was not related to the number of larvae that settled. At the cohort level, Mytilus and Pollicipes settling together across 3 dates showed similar trends of decreasing lipid index over time, suggesting that environmental conditions may influence co-occurring planktonic larvae similarly across species. This work highlights the need for further experiments in the field on the effects of larval history on recruitment success in natural populations, and further studies to determine what factors influence larval attributes for planktonic larvae in the field.
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Introduction |
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The recruitment of young can play a key role in marine population dynamics (Roughgarden and others 1988
; Gaines and Bertness 1992; Caley and others 1996). Most attention on this issue has focused on the consequences of variation in the number of settling larvae. A growing body of literature, however, demonstrates that substantial variation may also occur in the condition of larval settlers, which can have profound effects on juvenile growth and survival after metamorphosis (reviewed by Pechenik and others 1998; Moran and Emlet 2001; Searcy and Sponaugle 2001; Phillips 2002; Jarrett 2003; Marshall and Keough 2004). For marine invertebrates, the evidence for an important role of larval history on post-settlement success comes largely from studies where larval development is experimentally manipulated. Larval condition or quality (measured as size, lipid storage, and/or organic content) has been influenced by altering egg or hatching size, temperature during development, larval food supply, or by delaying metamorphosis. Subsequent juvenile performance has been followed in both the laboratory and in the field. The presumption of this approach is that factors manipulated in the laboratory during larval development drive similar variability in larval condition in nature. However, other than documenting patterns of size at settlement, few studies have examined larval condition and its natural variability in field populations of marine invertebrates (but see Jarrett and Pechenik 1997; Jarrett 2003).
A number of critical questions can be addressed in field studies of naturally settling larvae. First, how are different metrics of larval condition related, such as morphological measures (for example size) and biochemical indicators (for example lipid stores)? Do different metrics of quality co-vary in space and time, or are they decoupled? Under favorable planktonic conditions throughout development 1 expectation might be that they should co-vary. If food availability is consistently high, larvae may grow large and have plentiful lipid stores by the time they are competent to settle. In laboratory experiments where mussel larvae were fed constant high or low rations until settlement, larvae fed high rations were on average larger and had greater lipid stores than those fed low rations (Phillips 2002). If, however, conditions are variable during larval development, it might be expected that these different components of condition would be decoupled. For example, high-food availability early in development may generate large larvae, but a decline in food availability later may reduce their lipid stores.
Second, can patterns of natural variability in larval quality or condition help us understand the underlying mechanisms that generate them, especially for planktotrophic larvae that are both feeding in, and being transported by, ocean currents? For example, how does larval condition vary within versus among larval cohorts (that is, individuals settling at the same time)? If larvae in a given cohort experience more similar planktonic conditions than larvae settling at different times, there may be less variability among individuals of a cohort than among cohorts. On the other hand, variability among individuals (for example in feeding performance, in genetic or maternal effects, or in tolerance to stress) may generate as much or more variability in condition among individuals within cohorts as among cohorts.
Third, what is the relationship between larval condition and settlement intensity? Are cohorts that settle in higher numbers also of overall higher quality or better condition? If timing of spawning is correlated with favorable planktonic conditions for larval development (for example see Starr and others 1990, 1994), this might be expected. Similarly, conditions conducive to high larval survival may also enhance other aspects of larval condition. If, however, physical features in the ocean that concentrate and deliver larvae to shore are not the same or coincident with those favorable for development, then the number and quality of larvae that settle may be uncoupled or even negatively co-vary. The answer to the question of covariation in larval quality and quantity might help indicate whether potential post-settlement effects of larval history are likely to be complicated by potential density-dependent effects at settlement (for example Shima and Osenberg 2003).
The purpose of this study was to begin to address these questions by examining 2 components of larval condition, size and lipid stores at settlement, of terminal-stage mussel and barnacle larvae collected from the rocky intertidal zone of the southern California coast.
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I placed 20 settlement pads (Tuffy® brand plastic-mesh scrub pads) in the mid-zone of rocky intertidal habitat at 2 sites in southern California, which are
100 km apart: Lompoc Landing (34°43'45'' N, 120°36'56'' W) and Coal Oil Point (COP; 34°24'25'' N, 119°52'41'' W). Pads were collected at bi-weekly to monthly intervals, at which times each site was visited at low tide on 1 of 2 consecutive days, May–September 2001 at Lompoc Landing, and June–September 2001 at COP, all pads remaining were collected on each visit (often some of the pads were lost from the initial deployment) and new pads were deployed as replacements. I brought the pads into the laboratory and, with the aid of a dissecting microscope, removed all the terminal-stage larvae of mussels (Mytilus spp., most likely californianus, see Discussion) and barnacles (Chthamalus dalli and Pollicipes polymerus). Hereafter, terminal-stage barnacle larvae are referred to as cyprids. (Note that cyprids were not collected before July 5 and 6). Tuffies were sorted within 24 h of collection from the field. Larvae were placed in vials and stored at –80°C for later analysis. It is possible for terminal-stage mussel and barnacle larvae to postpone metamorphosis for several days after settlement, although the degree to which this occurs in nature is unknown. I assume therefore, that these collections represent new arrivals, although it is possible that they include individuals that arrived 1 or more days before collection.
Within several weeks of collection, larvae were thawed, and larval size and lipid stores measured. I quantified lipid content using the stain Nile Red. Methods for this technique are detailed elsewhere for bivalve larvae (Castell and Mann 1994; Phillips 2002) and for barnacle cyprids (Hentschel and Emlet 2000). Briefly, larvae are stained with Nile Red, which binds to neutral lipids and fluoresces bright yellow under the appropriate filter on a fluorescent microscope. I took digital images of each larva and performed image analysis using the public domain NIH Image program (developed at the US National Institutes of Health, Bethesda, MD) to calculate the lengths of larvae and the areas of both the larva and its lipid stores. I derived a lipid index for each larva that was standardized by size: lipid area/total larval area. Although this method of measuring lipids is relative, and is based on the 2-dimensional representation of 3-dimensional mass, Hentschel and Emlet (2000) found that lipid content determined using similar image analysis of Nile-Red-stained cyprids had a positive linear relationship with lipid content determined using biochemical extraction.
To examine variability in larval condition, I assayed individuals from different cohorts. I use the term cohort here to refer to a collection of larvae that settled to a single site on 1 date. For 3 cohorts of Pollicipes and 6 cohorts of Mytilus from Lompoc Landing, and 2 cohorts of Chthamalus from COP, I used a single-factor ANOVA to examine the random effect of date on larval length and lipid index. I also examined the relationship between size and lipids (length versus lipid index) at settlement for individuals within each of these cohorts, and coefficients of variation (CVs) for the larval attributes. For the 6 cohorts of Mytilus, I also examined the relationship between settlement intensity and cohort-level lipid index and mean larval size.
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Due to loss of tuffies from the field, between 10 and 20 tuffies were examined for larvae on each date from each site (mean = 15.7, SD = 3.67). For Mytilus and Chthamalus, there was significant variability among cohorts in mean length and lipid index, whereas for Pollicipes, there was significant variability among cohorts in lipid stores, but not in size (Table 1, Fig. 1). Although the barnacles overall had higher lipid indices than Mytilus, on the 3 dates that both Mytilus and Pollicipes settled together at Lompoc Landing, both species showed decreasing lipid indices over time (Fig. 2).
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For the 6 cohorts of mussels examined, there was no relationship at the cohort level between mean length and mean lipid index (P = 0.23), that is, cohorts that were on average larger in size did not necessarily also have greater lipid stores. In fact the trend, if anything, suggests the opposite (Fig. 1). At the level of individuals, there was also no evidence in any of the 3 species for coupling between size and lipid index; larger larvae did not necessarily have a greater proportion of lipid than did smaller larvae.
For all 3 species, although there was significant variability among cohorts in average larval attributes, most of the variation in length (80–100%) and lipids (58–78%) actually occurred among individuals within a cohort rather than among cohorts (Table 2). For all 3 species, however, CVs for each cohort were up to an order of magnitude greater for lipids (30–93%) than for length (3–8%, Table 3). Thus, although there was greater variability in both size and lipids among individuals than among cohorts, variability in size was more constrained than was lipid storage. Lipid storage exhibited much greater plasticity.
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There was no relationship between the size of a pulse, that is the number of larvae that settled, and the average quality of that cohort (for correlations with settlement intensity: length, P = 0.82; lipid index, P = 0.57). Thus, there was no evidence that pulses of mussel larvae that settled in higher numbers settled in overall greater condition as measured by these 2 metrics. There were not enough cohorts of Pollicipes or Chthamalus to ask this question at the species level for barnacles. Taken together, however, there was a positive relationship between settlement intensity and lipid index (r = 0.97; P = 0.004, Fig. 3).
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Discussion |
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Larval history is increasingly being recognized as a potentially important influence on post-metamorphic juvenile performance for a variety of marine invertebrates (reviewed by Pechenik and others 1998
, see articles in this volume). In 1 study, Phillips (2002) manipulated larval food environments for the mussel Mytilus galloprovincialis and found that larvae that had developed in a constant high-food environment were, on average, larger and had greater lipid stores at settlement than did those that had experienced constant low larval food. Moreover, early juveniles from those high-food larvae had higher growth rates and better survival for at least several weeks after metamorphosis. This study demonstrates that natural variability in lipid stores and size at settlement for larval mytilid mussels is substantial and commonly spans most of the range that can be generated in the laboratory. In interpreting these data, 1 caution necessary is the possibility that inter-specific differences may be contributing to the variability as I could not distinguish larvae from the 2 species possibly present for Mytilus (either M. galloprovincialis or M. californianus). I would argue that any inter-specific differences played at most a minor role as M. galloprovincialis is relatively uncommon in the region, especially at exposed sites north of Point Conception (for example Lompoc Landing). Also, genetic identification of larvae to species were made on settlers from several intertidal sites in this region in 2000 and >83% of the settlers were M. californianus (L. Miller and G. Pogson unpublished data).
The specific physiological mechanisms by which differences in larval history operate to affect post-metamorphic performance are not yet known, making the task of choosing the most relevant index of larval condition more challenging. Size at metamorphosis seems to be a good indicator of future performance in some marine invertebrates (Connell 1961; Marshall and Keough 2003, 2004). Other studies have indicated that juvenile performance may be more affected by larval energy stores than by size (Emlet and Hoegh-Guldberg 1997; Ito 1997) and that lipid in particular is important (reviewed by Holland [1978]). Thus, there is evidence that size and energy stores may both be good indicators of larval quality or condition at settlement especially as it is relevant to post-metamorphic juvenile performance.
However, a key result from this study is that lipid stores and size were decoupled for natural populations of mussel and barnacle larvae. These 2 metrics of larval quality did not co-vary. An issue that warrants considerable attention is the role of temporal variability in food supply during larval development. Differential amounts or quality of food available in the plankton during early versus late stages of development may lead to different physiological responses of larvae to either grow or store energy. For example, Phillips (2004) found for M. galloprovincialis that larval food treatments which varied timing and amount of food resulted in different relationships between larval length and lipid index at settlement. For barnacle cyprids, which are non-feeding, the decoupling of size and lipid stores might indicate a delay of metamorphosis after competence in some individuals.
Reports publishing data on multiple indices of larval condition at settlement are rare for marine invertebrates. In part it is difficult owing to the small size of marine invertebrate larvae (especially planktotrophic larvae) and the fact that hundreds or thousands of individuals may be required to generate a signal for analyses of biochemical constituents. For 2 congeners of a tropical reef fish (Pomacentrus), however, Kerrigan (1996) found similar results. In her study, relationships among morphometric measures of condition (for example length) and biochemical components (for example lipid, protein, and carbohydrate composition) of newly settled fish were inconsistent both between species and among seasons. For speckled goatfish, McCormick and Molony (1993) also found that length at settlement was not tightly coupled to biochemical measures of condition or quality at settlement. Taken together these studies suggest that for marine organisms, the relationship between morphological indices of condition (such as length) and biochemical indices are complex and not predictive of each other for natural populations of larvae.
For mussels and barnacles in this study, variability in size was more constrained than was lipid index, which was more plastic. Kerrigan (1996) found a similar pattern for the Pomacentrid fish she studied, where for both species biochemical indices (for example lipid, protein, and carbohydrate content) were much more variable (CVs of 25–49%) than length (CV of 5%) for newly settled fish. This suggests that constraints on variability in size may commonly be greater than those on biochemical components for newly settled marine organisms. This may in part be attributed to the fact that body length is a relatively long-term and integrative measure of growth, and that although growth rates may vary and slow, they do not generally fall below 0. There may also be pressure to attain at least a minimum size at metamorphosis. Biochemical composition, however, may fluctuate more rapidly, via growth or reduction, in response to environmental variability (especially food availability and temperature fluctuations) and larval activity.
The results of this study demonstrate that significant variability in size and lipid stores at settlement can be common among cohorts, even those settling within 2 weeks of each other, but also that most of the variation in both size and lipid storage actually occurred among individuals within cohorts (rather than among cohorts). These findings are consistent with the few other studies that examined temporal variation in some metric of larval condition. For example, Connell (1961) showed extensive variability in size at settlement for cyprids of the barnacle Semibalanus balanoides both among individuals and among cohorts settling weeks apart. Similarly, Jarrett and Pechenik (1997) found substantial variability in organic content of S. balanoides between cohorts settling within a week of each other, and Jarrett (2003) reported decreasing energy content of cyprids of S. balanoides over a settlement season. Kerrigan (1996) found significant variability in size and biochemical composition of the reef fish Pomacentrus amboinensis among settlement pulses, but similar to this study she also found most of the variability occurred among individuals within a pulse (60% for lipid content and length), rather than between pulses. If variability among larvae is an important component of recruitment success, taken together the results from this and other studies suggest that variability in larval condition is commonly present both within cohorts and among cohorts, and that variability among individual larvae (for example in feeding performance or growth, genetic or maternal effects, or tolerance to stress) may be an important driver of natural variability in larval attributes.
For mussels, there was no relationship between the numbers of larvae in a settlement pulse, and the average condition of the larvae in that pulse. This suggests that any post-settlement effects of larval history and density-dependence at settlement are unlikely to act synergistically to alter initial settlement patterns for this species. It also demonstrates that factors controlling the number of larvae in a pulse are different from those influencing the attributes of those larvae, at least in this system. Although there were not enough data to examine this question for each of the barnacle species, taken together, the results were suggestive that for barnacles there may be a positive relationship between lipid index and settlement intensity. This may indicate that the relationship between the number and average condition of larvae across settlement pulses is species- or taxon-specific and warrants further investigation.
An interesting result of this study was that over the 3 dates during which they settled together, mussels and gooseneck barnacle larvae showed a similar temporal trend of decreasing lipid index with time. Although with such limited data it is difficult to say anything conclusive, these results suggest that the planktonic environment experienced by the larvae of those pulses may have affected the energy storage of the 2 species in similar ways. Kerrigan (1996) also found for the 2 species of Pomacentrus that she studied, on 2 successive pulses in which the 2 species settled together, they had similar trends in lipid levels, that is both species increased their lipid levels from the first to second pulse. Again, the data are limited; however, she also suggested that an aspect of the planktonic environment (for example temperature or food availability) could account for this concordance. There are few, if any, other published accounts comparing larval attributes at settlement of different species that potentially develop in the same water mass. Many benthic marine invertebrates that interact as juveniles/adults (for example are space competitors like mussels and barnacles) may have similar reproductive seasons or larval strategies (for example planktotrophic for days to weeks) and thus commonly develop in the same water mass and settle together. If different species have similar responses to the planktonic environment in terms of cohort-level patterns in larval condition, and settle together, this could have important implications for the outcome of species interactions and early community development.
Given the laboratory evidence that larval condition can strongly influence early juvenile performance and the field evidence presented here that such variability in larval condition is widespread at several scales, it is likely that juvenile performance of marine invertebrates is frequently influenced by larval history. Ultimately, we need a mechanistic understanding of how larval number and larval quality interact to affect recruitment rates. The results presented here suggest this synthesis is possible from coordinated studies of the number and quality of larval settlers. This and other issues (that is, how larval history interacts with the severity of the juvenile environment, patterns of variability in larval condition among competitors or predators and prey whose larvae settle together) highlight the necessity of conducting further experiments in the field, to determine the extent to which larval history influences juvenile success of natural populations, and how it operates in conjunction with other processes that together influence recruitment of marine species.
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Many thanks to R. Podolsky and A. Moran for the invitation to participate in the Society of Integrative and Comparative Biology Symposium on Integrating Function Over Marine Life Cycles. This research was supported by funds from a National Science Foundation graduate fellowship to the author, and in part by NSF BIR94-13141 and NSF GER93-54870 to W. Murdoch, and the David and Lucile Packard Foundation to S. Gaines.
Conflict of interest: None declared.
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2Present address: School of Biological Sciences, Victoria University of Wellington, New Zealand
From the symposium "Integrating Function Over Marine Life Cycles" presented at the annual meeting of the Society for Integrative and Comparative Biology, January 4–8, 2006, at Orlando, Florida.
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References |
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