Comparing the Relative Importance of Water-Borne Cues and Direct Grazing for the Induction of Defenses in the Brown Seaweed Fucus vesiculosus

Some seaweed species have been shown to release water-borne cues after herbivore attack, for example, to attract natural enemies of the herbivore. These cues may also be sensed by neighboring seaweeds and used to adjust their defenses in anticipation of a possible herbivore attack. Several studies indicated information transfer between seaweed individuals in the past, including the brown seaweed Fucus vesiculosus. Previous work showed induction of defenses in F. vesiculosus in response to water-borne cues released by isopod-grazed conspecifics. In contrast, another study on induced responses after exposure to cues from isopod-grazed neighbors using the same seaweed species yielded contradictory results. This study reassessed the ability of F. vesiculosus individuals to sense water-borne cues released by isopod-grazed neighbors in a series of experiments that monitored F. vesiculosus palatability in response to direct grazing by Idotea baltica and water-borne cues from isopod-grazed neighbors relative to unmanipulated seaweed pieces. Two-choice feeding assays were conducted with both fresh and reconstituted seaweed pieces. Direct grazing by I. baltica induced a chemical defense in F. vesiculosus, confirming results of previous studies. In contrast, evidence for increased herbivore resistance in seaweed pieces that were located downstream of isopod-grazed F. vesiculosus could not be provided. The lack of defense induction in response to grazing of conspecific neighbors may be explained by the environmental conditions and the scattered distribution of F. vesiculosus individuals in the intertidal zone of Helgoland, which may render resource investment in the emission and/or response to water-borne cues at this site unprofitable.     

Polar extracts of the brown alga Fucus vesiculosus (L.) reduce assimilation efficiency but do not deter the herbivorous isopod Idotea baltica (Pallas)

Plant secondary metabolites provide resistance against herbivory if they are capable of decreasing herbivore performance or deterring feeding. Here we test the effects of phlorotannin-containing polar extracts from the brown alga Fucus vesiculosus on the assimilation efficiency and feeding preferences of the herbivorous isopod Idotea baltica. We test the effects separately for each sex of the isopod to test the hypothesis that the digestive efficiencies of the sexes may have diverged. Phlorotannin-containing polar extracts were incorporated into an agar-based artificial food in varying amounts. We also varied the quality of the food, using high-quality food containing animal proteins and low-quality food containing only algal material. The total assimilation efficiency as well as the assimilation efficiencies of carbon and nitrogen decreased with an increasing concentration of phlorotannin-containing extracts. The negative effects on carbon and nitrogen assimilation were higher when extracts were incorporated in the high-quality food. Females were consistently more efficient than males in all assimilation measures; there were no interactions between sex and food quality or phlorotannin concentration, suggesting that the digestive efficiency of the sexes had diverged but that the two sexes responded similarly to phlorotannin enriched artificial food. Although phlorotannin-containing extracts were harmful to performance, they did not deter feeding in preference tests. On the contrary: the isopods preferred the food containing more phlorotannins, suggesting that phlorotannins may act as cues in host recognition. Thus phlorotannins were inefficient deterrents, and I. baltica seems to be adapted to utilizing phlorotannin-containing foods despite their detrimental effects on assimilation. We suggest that F. vesiculosus provides a predation refuge and temporal constancy in availability for I. baltica, both of which select for the ability to utilize it as a food. Phlorotannin-containing polar extracts from F. vesiculosus are still harmful digestibility reducers, and may thereby prevent complete specialization and generate selection for utilization of a wider host range.     

LACK OF DEFENSE OR PHLOROTANNIN INDUCTION BY UV RADIATION OR MESOGRAZERS IN DESMARESTIA ANCEPS AND D. MENZIESII (PHAEOPHYCEAE)1

Phlorotannins are polyphenoloic metabolites occurring only in the Phaeophyceae that have numerous putative primary roles (e.g. cell‐wall construction and storage) as well as secondary metabolic roles, which include herbivore feeding deterrence and protection from UV radiation. The proposed role of phlorotannins in the defense against UV radiation is of particular importance in the Antarctic due to depletion of the stratospheric ozone layer in that area. Several studies of brown algae have found evidence of an induction response (the production of defensive metabolites, including phlorotannins) after grazing by various mesograzers, after simulated grazing/wounding, and after exposure to increases in UV radiation. This study aimed to determine if phlorotannin production or other defenses in two dominant, endemic Antarctic species (Desmarestia menziesii Montagne and Desmarestia anceps J. Agardh) could be induced by an increase in exposure to UV radiation or by natural and artificial grazing. An in situ experiment failed to detect any effect of UV radiation on phlorotannin concentrations in either species or on subsequent palatability in feeding bioassays. A laboratory‐based experiment did not detect any effect of mesoherbivore grazing or simulated grazing (wounding) on palatability or the concentration of phlorotannins in D. menziesii. Instead, phlorotannin concentrations increased in all treatments in both experiments, consistent with an increase in overall resource availability due to an increase in available PAR compared with the in situ irradiance at the algal collection sites.     

Resistance of the brown alga Fucus vesiculosus to herbivory

Herbivory is typically intense in marine littoral environments; thus, macrophytes are expected to evolve defenses against grazing. Although putative defenses of macrophytes are widely studied, there is lack of studies demonstrating the main premises of defense adaptations: the consequences of herbivory to macrophytes, genetic variation of defense traits and the costs and benefits of defenses in natural environment. We conducted a factorial experiment, where we manipulated amount of herbivory, growing depth and nutrient availability, and measured resistance to herbivory as well as genetic variation and costs of phlorotannins, putative defensive secondary metabolites, in the brown alga Fucus vesiculosus . Herbivory on algae varied with depth: grazing did not cause losses close to the surface, but, most of the algal production was consumed at the deeper end of the algal belt. The higher the genotypic phlorotannin content the less damage the genotype received implying that phlorotannins acted as a resistance trait. Production of phlorotannins was associated with costs for growth. Consistent with the prediction that the cost of defense will be greatest when resources are limiting, the cost appeared only in the deep end of the algal belt where growth was slowed down. Phlorotannins displayed phenotypic plasticity; the three factors influenced phlorotannins interactively, with the main tendencies of nutrient enrichment decreasing and herbivory and increasing depth increasing phlorotannins. Despite this plasticity, variation of phlorotannins was mainly due to the genotype of algae. These results emphasize the role of herbivory as a selective agent for algal defenses and the importance of genetic variation in the constitutive level of phlorotannins in interactions with natural enemies. The cost of phlorotannins may constrain the evolution of resistance in environments where growth is limited by light availability.     

Influence of light and nitrogen on the phlorotannin content of the brown seaweeds Ascophyllum nodosum and Fucus vesiculosus

Phlorotannins, C-based defence compounds in brown seaweeds, show a high degree of spatial and temporal variation within seaweed species. One important model explaining this variation is the Carbon Nutrient Balance Model (CNBM), which states that the relative supply of carbon and limiting nutrients will determine the level of defence compounds in plants. Nitrogen is often considered to be the limiting nutrient for marine macroalgal growth and the CNBM thus predicts that when the carbon:nitrogen ratio is high, photosynthetically fixed carbon will be allocated to production of phlorotannins. In the present study, we evaluated the effects of light (i.e. carbon) and nitrogen on the phlorotannin content of two intertidal brown seaweeds, Ascophyllum nodosum and Fucus vesiculosus. This was done in an observational field study, as well as in a manipulative experiment where plants from habitats with different light regimes were subjected to different nitrogen and light treatments, and their phlorotannin content was measured after 14 days. The results showed that there was a negative relationship between tissue nitrogen and phlorotannin content in natural populations of F. vesiculosus, but not in A. nodosum. In the short term, the phlorotannin content in both algal species was not affected by changes in nitrogen availability. Exposure to sunlight had a positive effect on the phlorotannin content in natural populations of both algal species but, in the manipulative experiment, only F. vesiculosus showed a rapid response to changes in light intensities. Plants subjected to sunlight contained higher phlorotannin content than shaded plants. In conclusion, the results imply that nitrogen availability explains some of the natural variation in the phlorotannin content of F. vesiculosus, but the light environment has greater importance than nitrogen availability in predicting the phlorotannin content of each species.     

Testing for the induction of anti-herbivory defences in four Portuguese macroalgae by direct and water-borne cues of grazing amphipods

Herbivory is a key factor in regulating plant biomass, thereby driving ecosystem performance. Algae have developed multiple adaptations to cope with grazers, including morphological and chemical defences. In a series of experiments we investigated whether several species of macroalgae possess anti-herbivore defences and whether these could be regulated to demand, i.e. grazing events. The potential of direct grazing on defence induction was assessed for two brown (Dictyopteris membranacea, Fucus vesiculosus) and two red seaweeds (Gelidium sesquipedale, Sphaerococcus coronopifolius) from São Rafael and Ria Formosa, Portugal. Bioassays conducted with live algal pieces and agar-based food containing lipophilic algal extracts were used to detect changes in palatability after exposure to amphipod attacks (=treatment phase). Fucus vesiculosus was the only species significantly reducing palatability in response to direct amphipod-attacks. This pattern was observed in live F. vesiculosus pieces and agar-based food containing a lipophilic extract, suggesting that lipophilic compounds produced during the treatment phase were responsible for the repulsion of grazers. Water-borne cues of grazed F. vesiculosus as well as non-grazing amphipods also reduced palatability of neighbouring conspecifics. However, this effect was only observed in live tissues of F. vesiculosus. This study is the first to show that amphipods, like isopods, are capable to induce anti-herbivory defences in F. vesiculosus and that a seasonally variable effectiveness of chemical defences might serve as a dynamic control in alga–herbivore interactions.     

Genotypic variation in tolerance and resistance to fouling in the brown alga <Emphasis Type="Italic">Fucus vesiculosus</Emphasis>

In this study, we examined genetic variation in resistance and tolerance to fouling organisms in the brown alga Fucus vesiculosus. We first grew 30 algal genotypes in the field, where we allowed fouling organisms to colonise the genotypes at natural levels. We then conducted a manipulative experiment, where we grew 20 genotypes of algae in aquaria with or without fouling organisms. We measured host resistance as the load of fouling organisms and tolerance as the slope of the regression of algal performance on fouling level. Fouling organisms decreased host growth and contents of phlorotannins and thus have the potential to act as selective agents on algal defenses. We found significant among-genotype variation in both resistance and tolerance to fouling. We did not find a trade-off between resistance and tolerance. We found a marginally significant cost of resistance, but no cost of tolerance. Our results thus indicate that both the tolerance and resistance of F. vesiculosus can evolve as a response to fouling and that the costs of resistance may maintain genetic variation in resistance.     

Fouling mediates grazing: intertwining of resistances to multiple enemies in the brown alga Fucus vesiculosus

Macroalgae have to cope with multiple natural enemies, such as herbivores and epibionts. As these are harmful for the host, the host is expected to show resistance to them. Evolution of resistance is complicated by the interactions among the enemies and the genetic correlations among resistances to different enemies. Here, we explored genetic variation in resistance to epibiosis and herbivory in the brown alga Fucus vesiculosus, both under conditions where the enemies coexisted and where they were isolated. F. vesiculosus showed substantial genetic variation in the resistance to both epibiosis and grazing. Grazing pressure on the alga was generally lower in the presence than in the absence of epibiota. Furthermore, epibiosis modified the susceptibility of different algal genotypes to grazing. Resistances to epibiosis and grazing were independent when measured separately for both enemies but positively correlated when both these enemies coexisted. Thus, when the enemies coexisted, the fate of genotypes with respect to these enemies was intertwined. Genotypic correlation between phlorotannins, brown-algal phenolic secondary metabolites, and the amount of epibiota was negative, indicating that these compounds contribute to resistance to epibiosis. In addition, phlorotannins correlated also with the resistance to grazing, but this correlation disappeared when grazing occurred in the absence of epibiota. This indicates that the patterns of selection for the type of the resistance as well as for the resistance traits vary with the occurrence patterns of the enemies.     

Food and habitat choice of the isopod Idotea baltica in the northeastern Baltic Sea

The isopod Idotea baltica is the most important benthic herbivore in the Baltic Sea. There exists a significant correlation between the distribution of the adult isopod and the belts of bladder wrack Fucus vesiculosus. However, following the eutrophication induced blooms of the filamentous macroalga Pilayella littoralis and the disappearance of F. vesiculosus a notable increase in idoteid abundances has been observed. The aim of this paper was (1) to evaluate experimentally whether F. vesiculosus provides either food, shelter or both to the isopods and (2) to estimate the role of associated filamentous algae in the habitat selection process. Amongst six abundant macroalgal species, about 50% of isopod population was attracted to F. vesiculosuscovered with the filamentous algae P. littoralis. The majority of the remaining part of the population was either swimming freely or attracted to non-epiphytic P. littoralis. When both live algae and artificial substrata were provided, P. littoralis growing on artificial substrata was clearly preferred by the isopods over epiphyte-free F. vesiculosus. In the grazing experiment where I. baltica was allowed to choose between F. vesiculosus and P. littoralis the latter contributed practically 100% of the diet of the isopod. The results indicate the importance of P. littoralis as a food item and F. vesiculosus as a shelter for I. baltica.     

In situ tests on inducible defenses in Dictyota kunthii and Macrocystis integrifolia (Phaeophyceae) from the Chilean coast

Numerous experimental studies have reported inducible defenses in macroalgae, but most of them have been conducted in laboratory environments where algae were maintained detached from the substratum and in artificial flow regimes. The results of those experiments might not reflect the natural situation, which can only be studied in situ. We examined whether the brown macroalgae Dictyota kunthii (C. Agardh) Greville and Macrocystis integrifolia (Bory) show inducible defenses following exposure to different grazing levels (direct, water-borne cues from nearby grazed conspecifics, presence of a non-grazing herbivore and natural grazing) in field experiments, striving to maintain natural conditions as much as possible. We measured palatability of algae after exposure to different grazing levels by using live algae and agar-based food containing non-polar extracts. M. integrifolia showed no induction of defenses (at least not of non-polar compounds), suggesting constitutive defenses, absence of defenses (tolerance) or use of another strategy to avoid herbivory. These results are similar to those from previous laboratory experiments. In D. kunthii, defense was induced after two weeks of direct grazing by amphipods under field conditions. Water-borne cues from nearby grazed conspecifics, presence of a non-grazing herbivore and natural grazing did not induce defenses. Induction of defense in response to direct grazing agrees with results from a previous laboratory study, but while indirect cues induced defenses in the laboratory, there was no measurable induced defense in the field. Probably chemical cues from grazers are diluted quickly in the field, not reaching concentrations that cause induction of defenses. This might be the reason why in some algae induction by direct grazing is a more important defensive strategy than induction by water-borne cues. The results from our study also suggest that laboratory experiments showing induced defenses in response to grazed neighbours or mere grazer presence need to be interpreted with caution.     

Geographical divergence in host use ability of a marine herbivore in alga–grazer interaction

When the selective environment differs geographically, local herbivore populations may diverge in their host use ability and adapt locally to exploit the sympatric host population. We tested whether populations of the marine generalist herbivore Idotea baltica have diverged in host us ability and whether they locally adapted to exploit the sympatric population of their main host species, the bladderwrack Fucus vesiculosus. We fed isopods from three local populations reciprocally with the sympatric and two allopatric populations of the host. The bladderwrack populations varied in their quality as food for isopods suggesting variation in the selective environment. The ability to exploit the main host showed considerable divergence among the isopod populations. There was no significant interaction between host and isopod origin, indicating that the patterns observed in the reciprocal feeding experiment could be explained by differences in overall suitability of the hosts and differences in overall performance of the isopod populations. Isopod population that was sympatric to a bladderwrack population with low phlorotannin content showed high performance on the algae from the sympatric but low performance on the algae from the two allopatric populations. Performance of isopods, especially in this population, decreased quickly with the increasing phlorotannin content of food algae. We therefore hypothesize that the isopods adapted to a low phlorotannin content were unable to utilize high-phlorotannin algae efficiently. Isopod populations sympatric to the high-phlorotannin bladderwrack populations may be generally better adapted to deal with phlorotannins, being thereby able to utilize a range of bladderwrack populations.     

Seasonal changes in situ grazing of the mesoherbivores Idotea baltica and Gammarus oceanicus on the brown algae Fucus vesiculosus and Pylaiella littoralis in the central Gulf of Finland, Baltic Sea

The in situ grazing experiments were performed in the shallow water rocky habitat of the northern Baltic Sea during ice-free season 2002. In the experiments the effects of algal species and choice on the grazing of the mesoherbivores Idotea baltica (Pallas) and Gammarus oceanicus Segerstråle were tested. Salinity, temperature, concentration of nutrients in water and macroalgae and net production of macroalgae were considered as random effects in the analysis. The invertebrate feeding rate was mainly a function of the net photosynthetic activity of Pylaiella littoralis (L.) Kjellman and Fucus vesiculosus L. Feeding rate increased significantly with decreasing algal photosynthetic activity. When the two algal species were incubated together invertebrates fed primarily on P. littoralis. Low selectivity towards P. littoralis coincided with its high photosynthetic activity. The presence of F. vesiculosus did not modify the invertebrate feeding on P. littoralis. The results indicated that (1) the grazing on F. vesiculosus depended on the availability of P. littoralis, (2) the photosynthetic activity of algae explained the best the variation in grazing rate and (3) the grazers are not likely to control the early outbreak of filamentous algae in the northern Baltic Sea by avoiding young and photosynthetically active algae. The likely mechanism behind the relationship is that the increased photosynthetic activity of macroalgae coincides with their higher resistance to herbivory.     

Galactolipids rather than phlorotannins as herbivore deterrents in the brown seaweed<Emphasis Type="Italic"> Fucus vesiculosus</Emphasis>

The first investigation of seaweed chemical defense against herbivores involved the brown seaweed Fucus vesiculosus and suggested defense via phlorotannins. The first demonstration of seaweed induction of secondary metabolites in response to herbivory also involved the genus Fucus and assumed a defensive function for phlorotannins. Many other investigations correlate herbivore feeding preference with changing levels of phlorotannins in this genus and others, but few directly test the effects of phlorotannins. No studies have assessed Fucus chemical defenses using bioassay-guided separation to investigate the complete complement of compounds deterring herbivores. We investigated the deterrence of F. vesiculosus chemical extracts using herbivore bioassays to guide our chemical investigations. Although crude extracts from F. vesiculosus strongly deterred feeding by the sea urchin Arbacia punctulata, phlorotannins from this extract did not deter feeding at 2x or 4x natural concentration by dry mass. Feeding deterrence was due to: (1) a polar galactolipid in the ethyl acetate-soluble extract, and (2) a non-phenolic compound, or compounds, in the water-soluble extract. Although this is the first evidence of galactolipids deterring herbivores, such defenses could be geographically and taxonomically widespread. The galactolipid we discovered in Fucus occurs in marine dinoflagellates, and a related metabolite that deters herbivory has recently been discovered in a tropical green seaweed. We were unable to identify the second deterrent compound, but deterrence occurred in a fraction containing carbohydrates, including sulfated sugars, but no phlorotannins. Given the polarity of these chemical deterrents, they could co-occur with and confound bioassays of phlorotannins if investigators test phlorotannin-containing algal extracts without further purification.     

To grow and defend: lack of tradeoffs for brown algal phlorotannins

The concept of cost is an integral element of ecological theories, including optimal defense theory, resource availability theory, and growth-differentiation balance theory. Indeed most frameworks that attempt to explain within-plant patterns of secondary metabolites, as well as account for the evolution of induced defenses, presume that defenses are 'costly'. One way in which investigators have sought to quantify the cost of secondary metabolites is to examine growth/defense tradeoffs, which are predicted to occur wherever resources cannot be simultaneously allocated to both growth and defense. However, emerging evidence suggests that these critical assumptions may not be valid for brown algal phlorotannins, compounds that occur throughout the division Phaeophyta and have served as analogs to vascular plant tannins in numerous tests of terrestrial-derived ecological theories in the marine environment. Here we present a model of phlorotannins as metabolites with both primary and secondary roles and argue that apparent trade-offs between algal growth and phlorotannin content are not a reliable indicator for establishing a cost of defense. We suggest the ecological theories which presume that defenses are costly because resources allocated to defense cannot also be allocated to other 'primary' functions are unlikely to accurately predict the striking variations in algal phlorotannin concentrations that are observed in nature.     

Induction of phlorotannin production in a brown alga: defense or resource dynamics?

Increase of phenolic secondary metabolites, phlorotannins, in brown algae due to gastropod grazing has been interpreted as an anti-herbivore adaptation. Here we tested whether such a response could be due to changes in truly available resources for the alga, not by the grazing activity of snails as such. We allowed two species of snails, Theodoxus fluviatilis and Physa fontinalis to graze on Fucus vesiculosus . These species feed on epibiota and particulate matter on the thallus but do not eat the thallus of F. vesiculosus . We further simulated snail grazing by nutrient enhancement, removal of epibiota and by a combination of the two. Manipulations of nutrient and light availability revealed the crucial role of epibiota in mediating resource availability for F. vesiculosus . Nutrient enhancement alone increased epibiota and decreased phlorotannins. Cleaning the thallus resulted in increased growth, and together with nutrient enhancement also in a trade-off with phlorotannins. Presence of T. fluviatilis on the thallus induced phlorotannin production, a response differing from the simulations of snail grazing. However, we suggest that the increase in phlorotannins may not be an induced defense but rather a consequence of a specific way of resource manipulation by this snail species. T. fluviatilis removes hyaline hairs that facilitate nutrient uptake. P. fontinalis did not remove hyaline hairs and the response of the alga to its grazing was similar to the treatment where we mechanically removed epibiota suggesting that cleaning of the thallus is the major mechanism how this snail species affects F. vesiculosus . Genetic variation in phlorotannin concentrations highly exceeded the induced responses of simulated or real snail grazing. This casts doubt for the efficiency of induced phlorotannin production to act as a defense, but is not contradictory with the interpretation of phlorotannins responding to variation in resource availability.     

Density dependent grazing effects of the isopod Idotea baltica Pallas on Fucus vesiculosus L in the Baltic Sea

In the Baltic Sea, abiotic factors are often supposed to explain the distribution of the key species Fucus vesiculosus. Still, in many areas, decline of F. vesiculosus has coincided with mass occurrence of the herbivorous isopod Idotea baltica. The aim of this work was to examine whether, how and at what densities I. baltica can affect the distribution of F. vesiculosus in the central Baltic proper. Both large-scale field surveys and a two-week grazing experiment have been performed.In the field survey there was a correlation between density of I. baltica and reduction in depth penetration of F. vesiculosus. At 80 animals per 100 g F. vesiculosus wet weight, the depth penetration of the F. vesiculosus belt was reduced by 2.5 m within a year. In the grazing experiment there was a correlation between density of I. baltica and loss of F. vesiculosus biomass and meristems. In the controls biomass and number of meristems increased by 50%, while at 20 animals per 100 g of F. vesiculosus there was no net growth of F. vesiculosus. Intensity of grazing did not differ between isopod densities of 20, 40 and 60/100 g. At isopod densities of 80 and 100/100 g though, biomass and meristems decreased by 50%, indicating a threshold for the survival of F. vesiculosus in the experiment. At all densities the isopods preferred younger tissue to older.Our results indicate that grazing by Idotea baltica is an important structuring factor in the Baltic Fucus vesiculosus populations.     

Interaction between isopod grazing and wave action: a structuring force in macroalgal communities in the southern Baltic Sea

The macroalgal belt in the southern Baltic Sea may be partly structured by the interaction of physical and biological factors. A field study, spanning the 1990s, describes a rapid decline of the Fucus spp. stands along the wave-exposed Swedish southeast coast. During this period, a relative dominance of Fucus vesiculosus L. shifted to a relative dominance of Fucus serratus L. The decline of F. vesiculosus coincided with observations of large numbers of the grazing isopods Idotea baltica (Pallas) and Idotea granulosa Rathke, or with field observations of frequent grazing marks on Fucus fronds. I. baltica, but not I. granulosa, tended to aggregate in the declining Fucus spp. stands, indicating a strong preference for Fucus spp. In a mesocosm experiment I. baltica, when given a choice, grazed both Fucus species at weak water motion. At strong water motion grazing was concentrated on F. vesiculosus. It is hypothesized that one of the reasons I. baltica preferred F. vesiculosus to F. serratus at strong water motion may have been differences in habitat quality, like width of thallus, influencing the ability to cling to the plant. Smaller thallus, as in F. vesiculosus, thus is the preferred habitat for grazing of I. blatica. We postulate that the existence of F. serratus in the area may be favoured by strong wave action and moderate but not strong grazing by I. baltica, relaxing the interspecific competition from F. vesiculosus.