Oecologia (2004) 138: 223–230 DOI 10.1007/s00442-003-1427-9
PO PULATI ON ECOLOG Y
Sofia A. Wikström . Henrik Pavia
Chemical settlement inhibition versus post-settlement mortality as an explanation for differential fouling of two congeneric seaweeds Received: 18 February 2003 / Accepted: 7 October 2003 / Published online: 13 November 2003 # Springer-Verlag 2003
Abstract It has been proposed that seaweed secondary metabolites, e.g. brown algal phlorotannins, may have an ecologically important function as a chemical defence against epiphytes, by acting against colonisation of epiphytic organisms. We tested whether the low epiphytic abundance on the invasive brown seaweed Fucus evanescens, compared to the congeneric F. vesiculosus, is due to a more effective chemical defence against epiphyte colonisation. A field survey of the distribution of the common fouling organism Balanus improvisus (Cirripedia) showed that the abundance was consistently lower on F. evanescens than on F. vesiculosus. However, contrary to expectations, results from experimental studies indicated that F. vesiculosus has a more effective antisettlement defence than F. evanescens. In settlement experiments with intact fronds of the two Fucus species, both species deterred settlement by barnacle larvae, but settlement was lower on F. vesiculosus both in choice and no-choice experiments. Phlorotannins from F. vesiculosus also had a stronger negative effect on larval settlement and were active at a lower concentration than those from F. evanescens. The results show that Fucus phlorotannins have the potential to inhibit settlement of invertebrate larvae, but that settlement inhibition cannot explain the lower abundance of the barnacle Balanus improvisus on F. evanescens compared to F. vesiculosus. Assessment of barnacle survival in the laboratory and in the field showed that this pattern could instead be attributed to a higher mortality of newly settled barnacles. Observation suggests that the increased mortality was due to detachment of
S. A. Wikström (*) Department of Botany, Stockholm University, 106 91 Stockholm, Sweden e-mail: [email protected]
Fax: +46-8-162268 H. Pavia Tjärnö Marine Biological Laboratory, Department of Marine Ecology, Göteborg University, 452 96 Strömstad, Sweden
young barnacles from the seaweed surface. This shows that the antifouling mechanism of F. evanescens acts on post-settlement stages of B. improvisus. Keywords Antifouling . Balanus improvisus . Epiphytes . Fucus . Phlorotannins
Introduction In marine systems, hard substratum to colonise is often a limiting resource for benthic organisms and many species have adopted the strategy to live as epiphytes on the surface of other organisms. Epiphytes may have a positive effect on the host species, e.g. by protecting the host from predators or other stress factors, but they may also decrease the fitness of the host by increasing mortality or competing for the same resources (reviewed by Wahl 1989). Conversely, many host species are able to affect the epiphytic community through a chemical or mechanical defence (Davis et al. 1989; Wahl 1989), potentially influencing populations of epiphytic species. However, although epiphytism is common in marine communities, the ecology of marine host-epiphyte interactions has not been widely studied, and we still lack understanding of many of the processes involved. Large, long-lived seaweeds provide extensive hard substratum, which is relatively stable in relation to the longevity of most epiphytic species, and many seaweeds host a diverse epiphytic community. Several studies have documented a largely negative effect of epiphytes on their seaweed host. An extensive epiphytic load may decrease seaweed productivity (Littler and Littler 1999), growth (Buschmann and Gómez 1993; Worm and Sommer 2000) and survival (D’Antonio 1985), and the negative effect of fast-growing epiphytic algae has been proposed as one factor explaining the decline of perennial macroalgae in eutrophicated areas (Vogt and Schramm 1991; Worm and Sommer 2000). These disadvantages to the host could be expected to promote the evolution of effective antifouling defences, and the presence of such a defence has been
proposed to explain why some macroalgal species are conspicuously devoid of epiphytes (e.g. de Nys et al. 1995; Schmitt et al. 1995; Littler and Littler 1999). The host seaweed may affect epiphyte abundance by inhibiting colonisation or by decreasing survival of established epiphytes. Extensive shedding of vegetative tissue in some seaweeds has been proposed to act as a defence against survival of epiphytic organisms (e.g. Johnson and Mann 1986; Littler and Littler 1999). However, most studies of seaweed antifouling defences have focused on anti-settlement mechanisms, in particular chemical compounds deterring epiphytic colonisation. Marine seaweeds produce a wide diversity of secondary metabolites, that are thought to function primarily as a defence against natural enemies, e.g. epiphytes (Hay 1996). Chemical mediation of host-epiphyte interactions has been proposed to be a common phenomenon in marine systems (Steinberg and de Nys 2002) and several studies indicate that seaweed secondary metabolites may affect settlement of protists, algal spores and invertebrate larvae (reviewed by Clare 1996; Steinberg et al. 2001). However, to date there are few rigorous studies showing chemical inhibition of epiphyte colonisation at ecologically realistic concentrations (but see de Nys et al. 1995; Schmitt et al. 1995). Brown seaweeds produce polyphenolic metabolites called phlorotannins, which are polymers of phloroglucinol (1,3,5-trihydroxybenzene) (Ragan and Glombitza 1986). Phlorotannins are often present in high concentrations, up to almost 20% of the algal dry weight (Ragan and Glombitza 1986). Several studies have demonstrated that phlorotannins can function as a chemical defence against a wide diversity of herbivores (see Ragan and Glombitza 1986; Targett and Arnold 1998; Pavia and Toth 2000 and references therein). They have however also been proposed to be active against other groups of potential enemies, including epiphytes (Sieburth and Conover 1965; Langlois 1975). While the deterrent effect of phlorotannins on marine herbivores is well documented, the role of these compounds in the interaction between algae and their epiphytes is still much discussed. Phlorotannins have been shown to inhibit settlement of protists and invertebrate larvae (Langlois 1975; Lau and Qian 1997; Lau and Qian 2000) as well as germination of epiphytic algae (Jennings and Steinberg 1997). Despite these findings, the ecological importance of phlorotannins as inhibitors of fouling organisms has been questioned. Since phlorotannins are water-soluble and do not stay at the algal surface, a considerable rate of phlorotannin exudation is needed to maintain a sufficiently high concentration around the plant to interfere with the establishment or survival of epiphytic organisms (Jennings and Steinberg 1997; Steinberg et al. 2001). Natural phlorotannin exudation rates have proved difficult to measure reliably, not least because exudation increases in response to handling of the algae (see Ragan and Glombitza 1986). However, there is evidence that also healthy and unstressed plants in their natural growing sites exude phlorotannins (Carlson and Carlson 1984; Jennings and Steinberg 1994). The high exudation rate observed
immediately after immersion of algae in the tidal cycle (Carlson and Carlson 1984) indicates that the phlorotannin concentration in algal beds may temporarily be high, at least in the intertidal zone. On northern European shores, the invasive brown alga Fucus evanescens C. Ag. is consistently very little fouled by algae and sessile invertebrates, compared to the native congeneric F. vesiculosus L. (Schueller and Peters 1994; Wikström and Kautsky, in press). The large difference in fouling, despite the ecological and morphological similarity of the two species, suggests that the invader has a more effective antifouling defence than the native species. The ability of F. evanescens to deter epiphytes may have facilitated its invasion, since the species has often most successfully invaded eutrophicated areas, where the native fucoids are heavily overgrown by algal epiphytes (Powell 1957; Bokn et al. 1992; Schueller and Peters 1994). An effective defence against potential enemies has been proposed as an important factor in explaining the successful invasion of other exotic seaweeds, e.g. the rapid invasion of Caulerpa taxifolia in the Mediterranean (Lemée et al. 1993). In such cases, knowledge of the defence mechanism may be important for understanding and possibly predicting seaweed invasions. The aim of this study was to investigate if the pattern of epiphytic abundance on F. vesiculosus and F. evanescens could be explained by differences in algal chemical inhibitors of epiphyte settlement, and to assess the potential role of phlorotannins as such inhibitors. An important fouling organism in this system is the barnacle Balanus improvisus L., which is commonly found in the shallow littoral zone on rocks, algae or anthropogenic constructions (Barnes and Barnes 1962). To begin with, we investigated the natural distribution of B. improvisus on F. evanescens and F. vesiculosus at two different sites on the Swedish west coast, in two different seasons. Based on the results from this survey, we performed both laboratory and field experiments where we tested the hypothesis that the larvae of B. improvisus are deterred from settling on F. evanescens, in comparison to F. vesiculosus. The results from these experiments then led us to test the hypotheses: (1) that juvenile mortality of settled B. improvisus is higher on F. evanescens than on F. vesiculosus, and (2) that phlorotannins from both algal species are effective in inhibiting larval settlement but that phlorotannins from F. vesiculosus are more effective than those from F. evanescens.
Materials and methods Study area and organisms The study was performed at Tjärnö Marine Biological Laboratory on the Swedish west coast (58°52′N, 11°09’E) between 2000 and 2002. In this area, shallow hard bottoms are dominated by perennial brown seaweeds of the family Fucaceae. A recent addition to this community is Fucus evanescens, that was first found in the area in 1960 and is still in the process of expanding its range. Where it is
225 found, it often grows mixed with the native fucoids, e.g. F. vesiculosus (Wikström et al. 2002). The barnacle Balanus improvisus is commonly found in the shallow littoral zone attached to stones, algae or man-made constructions (Barnes and Barnes 1962). The species has a planktotrophic larval development consisting of six naupliar stages and one nonfeeding cyprid stage, the latter being responsible for finding and attaching to a suitable substratum. The processes of attachment and metamorphosis of the cyprid larva into an adult barnacle are collectively referred to as “settlement”. The cyprids are capable of exploring surfaces actively and the site of settlement is determined using physical and biochemical properties of the substratum (e.g. Berntsson et al. 2000). On the Swedish west coast, B. improvisus reproduces during the summer and the planktonic cyprid larvae settle during June–October (Berntsson et al. 2000; Berntsson and Jonsson 2003).
Field distribution of B. improvisus To study the natural distribution and abundance of B. improvisus on F. evanescens and F. vesiculosus, 20 plants (>10 cm long) of each algal species were collected randomly from 0.5±0.1 m below mean water level (MWL) at each of two sites. Both sites were artificial stone reefs close to the harbour of Strömstad. Sampling was done on two occasions, in October 2000, at the end of the settlement period, and in April 2001. On each plant, the number of B. improvisus barnacles was counted under a dissection microscope. The algal fronds were then scraped clean of epiphytes and dried in 70°C to constant weight, for measurement of the dry weight. Surface area of the fronds was calculated from an empiric weight/area ratio, obtained by measuring dry weight and area of 20 frond pieces from each Fucus species. The area was analysed by image analysis (software, Scion Image, Beta 4.0.2), after scanning the fresh pieces with a flatbed scanner. The abundance of B. improvisus could then be related to the surface area of the host.
Settlement rate of cyprid larvae on natural algal surfaces Settlement of cyprid larvae of B. improvisus on fronds of the two Fucus species was tested in choice as well as no-choice experiments on two occasions (October and July) during the settlement period of B. improvisus. The same experimental setup was used on both occasions. Plants for this and the other experiments were randomly collected from 0.3–1 m below the MWL in a mixed stand of F. vesiculosus and F. evanescens in a sheltered bay near the laboratory. From each Fucus plant, one frond tip of about 6 cm length (wet weight 10±1 g) was cut off and used in the experiments. To prevent the larvae from settling on the walls of the experimental container, the experiments were performed in bags of plankton net (140 μm mesh size), with an approximate volume of 100 ml. Cyprids of B. improvisus avoid settling on micro-textured surfaces and a surface prepared with 170-μm plankton net has been shown to inhibit settlement completely in laboratory experiments (Berntsson et al. 2000). Each net bag was placed in a 400-ml plastic bottle with through-flowing sterile-filtered seawater (30‰S) heated to 20±0.5° C. In the no-choice experiments, one piece of frond of either F. evanescens or F. vesiculosus was put into each of six parallel net bags. As controls we used 2×8-cm panels of plexiglas, a material on which B. improvisus cyprids are known to settle readily (Berntsson et al. 2000). In the laboratory, cyprid larvae tend to settle near the water surface. To promote settlement, the Fucus fronds and control panels were therefore placed in contact with the surface. To prevent the algae from drying they were sprayed regularly with filtered sea water. The choice experiments were set up in the same way, but with one piece of F. evanescens and one of F. vesiculosus in each of six parallel net bags. Cyprids of B. improvisus were reared in a laboratory cultivating system, as described by Berntsson et al. (2000). Newly moulted
cyprids were collected from the larval cultures and kept at 15°C for 5 days in a 200-ml glass bottle, to age the larvae before starting the experiment. A total of 100 aged larvae were added to each net bag. After 7 days the algal fronds and control panels were removed from the containers and the number of metamorphosed barnacles were counted under a dissection microscope. The net bags were also examined for metamorphosed barnacles using the same method.
B. improvisus colonisation in situ To determine the colonisation rate of B. improvisus in situ, we transplanted Fucus fronds without epiphytes to a site with high barnacle abundance. Epiphyte-free tips of 10 cm length were cut off from newly collected fronds of F. evanescens and F. vesiculosus and attached to a rope with plastic cable stripes. We used 2×8-cm plexiglas panels as controls, as in the laboratory settlement experiment. The rope with frond pieces and control panels was anchored horizontally on the bottom, at 0.5 m depth, in a sheltered site in July 2001. After 2 weeks the number of metamorphosed B. improvisus was counted under a dissection microscope. The area of the algal fronds was measured with image analysis, as described above, to calculate settlement per unit area.
Phlorotannin analyses and inhibition of larval settlement In order to compare natural concentrations of phlorotannins in F. evanescens and F. vesiculosus, five plants of each species were randomly collected from 0.5 m depth in August 2000. The plants were brought to the laboratory and immediately frozen at −70°C and freeze-dried. Tissue samples for phlorotannin analyses were ground to a fine powder, placed in aqueous acetone and shaken under nitrogen in the dark at 4°C for 24 h. After extraction the samples were centrifuged, evaporated in vacuo at