Chrysomelid Beetle Herbivores and Salix Phenolglycosides
Amy L. Seidl
Colorado State University
Fort Collins, Colorado 80523
Chemical Ecology Review Paper
EN 570
Spring 1994
Abstract
Salix spp. are known to contain phenolglycosides as main secondary
compounds (Palo 1984). Salicortin and salicin, the primary
phenolglycosides, are found in the bark and leaves of Salix spp.
respectively. Plants with high levels of these compounds tend to be
eaten by specialized herbivores, including chrysomelid beetles in the
genus Phratora and Chrysomela. Closely related chrysomelid beetles
synthesize chemical defenses both autogenously and through the use of
plant precursors. In Ph. vitellinae, salicin is the plant precursor used
to synthesize the volatile irritant salicylaldehyde. Researchers have
speculated that beetles have evolved the use of host plant precursors
from de novo biosynthesis due to (1) a host plant shift from herbaceous
plants to trees, and (2) the superior metabolic efficiency of using a
plant precursor. Coevolution of chrysomelid beetles and Salix toxins has
been aided by a unique exaptation: enzymes used in autogenous production
of chemical defenses are the same as those used in the synthesis of
chemical defenses with a plant precursor.
Introduction
Ehrlich and Raven (1965) defined the term coevolution as "the
patterns of interaction in between two major groups of organisms with a
close and evident ecological relationship, such as plants and
herbivores." In their seminal paper, they identified five steps of
coevolution between plants and herbivores. Briefly, they are (1) plants
make secondary compounds through mutation and recombination, compounds
which are not necessary for primary metabolism, (2) by chance, secondary
compounds alter plant suitability for herbivores, (3) plants, freed from
herbivory, adaptively radiate, (4) herbivores evolve mechanisms of
resistance by mutation and recombination, and (5) herbivores exploit
previously unsuitable resources and adaptively radiate (Ehrlich and
Raven 1965). Berenbaum (1983) has elegantly shown this sequence of
coevolution in her study of coumarins and caterpillars, where toxic and
phototoxic coumarins in species of the plant family Apiaceae make them
unpalatable to generalist herbivores. Only specialized herbivores,
including Papilio polyxenes, are unaffected by the plants' toxicity
(Berenbaum 1983).
This paper reviews a similar system where coevolution has occurred
between plant and herbivore. Interestingly however, the plant's toxic
compound, salicin, enhances the survivorship of the specialist
herbivore, via its chemical defense, a mechanism of resistance not due
to mutation or recombination but by an exapted trait. In turn, the
herbivores have a negative effect, through herbivory, on the plant and
are defended from their predators as well (Smiley et al. 1985).
Chemical Ecology of Willows
Willows in the genus Salix (Salicaceae) contain phenolglycosides,
phenolic compounds which are bitter tasting to herbivores (Palo 1984).
Salicin and salicortin are the identified toxic compounds found in
willow, birch and poplar species, all in the family Salicaceae.
Salicortin is found in the bark of almost all species of Salix, and
contains higher levels of phenolic concentration and glucoside diversity
than salicin in leaves (Palo 1984). In Finnish Salix species, Julkunen-
Tiitto (1989) found the amount and composition of phenolgylcosides in
each willow was moderately species-specific. Additionally, the
magnitude of concentration varied considerably in twigs and leaves,
between 0.05% to 12.0%, as did glucoside levels in buds and flowers
(Julkunen-Tiitto 1989). Salix spp. of Central and Northern Europe are
known to have differing levels of phenolglycosides as well, from none as
in S. alba to high levels as in S. nigricans (Rowell-Rahier 1984b).
Salix not defended by phenolglycosides are often defended by
proanthocyanidins or by morphological structures such as trichomes that
deter herbivores (Rowell-Rahier 1984a, Pasteels et al. 1989). Glucoside
content in willow parts has been reported to be dependent on the
physiological activity of willow tissue, including diurnal, seasonal and
intra-species fluctuations (Julkunen-Tiitto 1989). Altitudinal gradients
may also affect glucoside content (Smiley et al. 1985).
Because phenolglycosides are carbon-based secondary compounds
(Price et al. 1989), willow plants are hypothesized to maintain a
balance between nutrient and carbohydrate concentrations in the tissues
of their leaves and twigs (Price et al. 1989). Bryant et al. (1983)
proposed a carbon-nutrient balance hypothesis wherein plants, which are
under nutrient stress, will continue to photosynthesize such that
carbohydrates and carbon-based compounds like phenolgylcosides
accumulate. Interestingly, in Salix lasiolepis male plants have a
different carbon balance than female willow plants and have lower
concentrations of phenolglycosides, especially salicortin and tremulacin
(Price et al. 1989). Similar differences in male and female
phenolglycoside levels were found in S. caprea, where females had two
times the concentration of males (Palo 1984). Differences in plant
toxin between sexes represents differences in plant quality as well:
female willows are better defended and therefore less palatable (Price
et al. 1989). Female clones invest more resources into sexual
reproduction and defense and less into vegetative growth whereas male
clones invest more into growth, and thereby may be prone to greater
herbivory (Price et al. 1989). This is most clearly seen in S.
lasiolepis where shoot length is longer in males than females of the
same species (Price et al. 1989). How these differences translate into
sex-biased herbivory is largely correlative (Boecklen et al. 1990).
However bud-galling sawflies Euura mucronata (Hymenoptera:
Tentredinidae) oviposit preferentially on the more vigorously growing
male clones of S. cinerea (Roimnen 1991), a fact which may be
phenologically based with a secondary result being lower phenol levels
(Roimnen 1991). Alternatively, females may be responding to the rapidly
growing longer shoots, rather than to low phenolglycoside content, since
vigorous growth often signals high sugar and photosynthate resources
(Roimnen 1991).
Studies of the effects of salicortin and salicin on herbivores
include (1) digestibility of phenols in apical parts of Salix twigs on
ungulate browsers (Palo et al. 1992), (2) premature leaf abscission and
subsequent loss of host plant resource for Phyllonorycter spp.
(Lepidoptera) (Preszler et al. 1993), (3) sexually dimorpic leaf
chemistry and phenology on sawfly oviposition sites (Boecklen et al.
1990, Roimnen 1991), and (4) coevolution of chrysomelid beetles to
salicin and salicortin and subsequent production of the defensive
compound salicylaldehyde (Eggenberger and Rowell-Rahier 1993, Pasteels
et al. 1983, Pasteels et al. 1989, Pasteels et al. 1990, Rowell-Rahier
1984a, 1984b, 1984c, Tahvanainen et al. 1985). The remainder of this
paper will focus on how some chrysomelid beetles have become specialists
on Salix spp. and what adaptations and/or exaptations exist for them to
detoxify phenolglycosides for proximate use in their own defense against
predators.
Chemical Defense in Chrysomelid Beetles
Chrysomelid beetles feed in large aggregations on patchy food
plants in open habitats, and are likely prey to predators such as birds
(Pasteels et al. 1989). Chrysomelid (leaf) beetles are known for their
aposematic coloration, they often have bright metallic markings which
serve as intra- or interspecific signals to other individuals signaling
their toxicity (Pasteels et al. 1989). Visual aposematism works as a
rapid and powerful negative reinforcement and serves to protect the
species as a whole. Like other aposematic species, chrysomelid beetles
have evolved non-cryptic behaviors, for instance they feed on young
nutritious leaves that are often on the exposed tops of plants, and
their aggregation behavior makes them conspicuous. Specialization on
plants which live in open habitats may have supported the adaptation of
aposematism and defense by autogenously produced toxins (Pasteels et al.
1989).
The larvae of the chrysomelid beetles Chrysomela tremulae and
Phratora vitellinae use salicin as the precursor for salicylaldehyde
(Pasteels et al. 1983). Salicylaldehyde is a volatile irritant and is
frequently used as a defensive secretion by arthropods, especially
against parasitoids and small predators such as ants (Pasteels and
Gregoire 1983). In Ph. vitellinae there is no autogenous source of
salicylaldehyde. This was shown by feeding Ph. vitellinae a salicin-free
diet and then examining its excretion, which did not contain
salicylaldehyde (Pasteels et al. 1983). When fed its host plant
however, S. nigricans, salicylaldehyde was produced (Pasteels et al.
1983). The use of salicin as a biosynthetic
precursor is advantageous to the larvae because (1) larvae do not need
to make the initial compound, making the production of salicylaldehyde
less metabolically expensive, (2) larvae are able to continue to feed on
willow and sequester salicin, an ability which would otherwise not allow
them to exploit a nutritional resource, (3) salicylaldehyde deters ants,
the main predator of chrysomelid beetles, better than salicin (Pasteels
et al. 1983), and (4) larvae are able to mobilize the glucose released
through the hydrolysis of salicin (Soetens et al. 1993).
The evolution of salicylaldehyde has evolved three times in
chrysomelid beetles, in the genera Chrysomela and Phratora and in the
species Plagiodera versicolora (Pasteels et al. 1983). Shift in host
plant affinities and the adaptation by chrysomelid beetles to Salix
toxins, changed the physiological and morphological structure of
defensive glands (Pasteels et al. 1983, Pasteels et al. 1989). The
beetles became much more efficient in not only breaking down toxins but
in restructuring defensive glands so that new and less costly compounds
could be made, ones that were chemically based on the plant's toxin
(Pasteels et al. 1989).
Although identical defensive allomones have been observed in
unrelated species, convergence of chemical defenses in sympatric or
allopatric species is unknown (Pasteels and Gregoire 1983). In
chrysomelid beetle larvae, distinctly different mixtures of chemical
defenses are used (Pasteels and Gregoire 1983). There are two
hypotheses why closely related species may have different chemical
defenses: (1) predators may differ (Pasteels and Gregoire 1983), and (2)
a community of arthropods with a diverse chemical defense system is more
likely to limit the ability of predators to overcome chemical defenses,
i.e. a generalist predator could not detoxify a number of different
chemical defenses simultaneously (Pasteels and Gregoire 1983). However,
Mullerian mimicry of odors is frequent among aposematic beetles,
including aposematic lycids, cerambycids and coccinellids (Rothschild
1961). Similar defensive odors, but different main toxins, send a strong
"community" signal to predators and more rapidly reinforces the
ecological message of warning coloration and unpalatability (Pasteels
and Gregoire 1983).
Phratora vitellinae (Coleoptera:Chrysomelidae): A Case Study
Phratora vitellinae, a leaf beetle, specializes on Salix spp. with
high levels of salicin (S. nigricans, S. purpurea, S. repens) which it
then converts into salicylaldehyde (Pasteels et al. 1983). Ph.
vitellinae overwinter in sheltered spaces and emerge in the spring to
colonize Salicaceous trees and to feed, mate, and reproduce on them
(Rowell-Rahier 1984b). Ph. vitellinae has nine pairs of eversible glands
through which they secrete an aqueous emulsion containing
salicylaldehyde. The secretion accumulates in vacuoles that converge on
a linear duct and ends in an opening onto the body surface (Pasteels et
al. 1989) (Fig. 1). Openings exist all over the pronotum and the elytra
(Pasteels et al. 1989). Defensive glands (whose parts include vacuoles,
linear ducts, and openings) are hypothesized to be the derived state of
the primitive dermal glands and are most abundant along the lateral
portion of the body (Pasteels et al. 1989).
To derive salicylaldehyde from salicin two enzymes are needed (1)
B-glucosidase and (2) oxidase (Soetens et al. 1993). B-glucosidase
hydrolyses the salicin into saligenin and glucose. Glucose is later
recovered by the beetle larvae. Pasteels et al.(1983) suggested that
the reabsorbed glucose may cover as much as 31.6% of the daily caloric
requirement. The second enzyme, oxidase, is used to transform saligenin
into salicylaldehyde (Soetens et al. 1993). Interestingly, the walnut-
feeding chrysomelid Gastrolina depressa utilizes the same two classes of
enzymes in its secretion of the toxin juglone (Matsuda and Sugawara
1980).
Closely related species to Ph. vitellinae biosynthesize iridoid
monoterpenes de novo, suggesting that the primitive condition for
Phratora spp. is autogenous production of chemical defenses (Pasteels et
al. 1990). The shift made by Ph. vitellinae to derived chemical
defenses is probably due to a shift in host plant, i.e. to Salix spp.
In their review of toxins in chrysomelid beetles, Pasteels et al. (1990)
illustrate how the enzymes used for de novo synthesis are those that
would permit the evolution and synthesis of plant-derived glucosides for
defense. This evolutionary sequence, wherein a previous adaptation is
used secondarily for a present benefit is termed "exaptation", sensu
Gould and Vrba (1983).
Pasteels et al. (1990) explain that species which secrete iridoid
monoterpenes do so because they are relatively stable and less toxic and
are easy to transfer across membranes, i.e. from internal glands and
cells to the external openings which release the poison to the outside.
Additionally, because so much glucose is recovered by the larvae after
the transformation of saligenin to salicylaldehyde, and because glucose
increases as the quantity of hydrolyzed salicin increases, sequestration
and subsequent use of plant toxins has become adaptive. Chemical
defenses replaced autogenous synthesis because they are less costly and
yield a net energetic advantage to the beetles (Pasteels et al. 1990).
Conclusion
What is unique about the coevolution scenario of chrysomelid
beetles and Salix spp. is that the adaptation of the beetles to the
plant toxins is not due to the evolution of resistance mechanisms
"through mutation and recombination" as Ehrlich and Raven (1965)
proscribed. Instead, resistance is due to exaptations, the presence of
key enzymes and defensive gland physiology both of which aid in
detoxifing and sequestering plant toxins. A similar scenario of
exaptation and insect resistance to plant toxins was described by Brower
et al. (1988) in their study of monarch sequestering of cardenolides.
Therefore, although chrysomelid beetles use "preadaptive" traits to
convert salicin to salicylaldehyde, thereby not evolving specific
adaptive traits of resistance, their specialist nature has allowed them
to adaptively radiate onto, and coevolve with, Salicaceous plants which
other herbivores find unpalatable.
Finally, chrysomelid aposematism has been suggested as an
exaptation as well (Pasteels et al. 1989). Aposematism in beetles is
hypothesized to be as ancestral a trait as de novo biosynthesis, and the
two are in fact coupled. Aposematic coloration reinforced beetle
toxicity to predators in their primitive state since autogenously
produced toxins make the beetles as unpalatable as using plant
precursors (Pasteels et al. 1989). Although intuitively this hypothesis
appears reasonable there is no conclusive evidence for it. More
taxonomic and ecological research is needed to ascertain if changes in
coloration have occurred simultaneously with changes in the synthesis of
chemical defenses.
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