A free lunch? No cost for acquiring defensive plant pyrrolizidine alkaloids in a specialist arctiid moth (Utetheisa ornatrix).

Many herbivorous insects sequester defensive chemicals from their host plants. We tested sequestration fitness costs in the specialist moth Utetheisa ornatrix (Lepidoptera: Arctiidae). We added pyrrolizidine alkaloids (PAs) to an artificial diet at different concentrations. Of all the larval and adult fitness components measured, only development time was negatively affected by PA concentration. These results were repeated under stressful laboratory conditions. On the other hand, the amount of PAs sequestered greatly increased with the diet PA concentration. Absence of a detectable negative effect does not necessarily imply a lack of costs if all individuals express the biochemical machinery of detoxification and sequestration constitutively. Therefore, we used qPCR to show that expression of the gene used to detoxify PAs, pyrrolizidine-alkaloid-N-oxygenase (pno), increased 41-fold in our highest PA treatment. Nevertheless, fitness components were affected only slightly or not at all, suggesting that sequestration in this species does not incur a strong cost. The apparent lack of costs has important implications for our understanding of the evolution of ecological interactions; for example, it implies that selection by specialist herbivores may decrease the levels of certain chemical defences in plant populations.

Macroevolution and the biological diversity of plants and herbivores.

Terrestrial biodiversity is dominated by plants and the herbivores that consume them, and they are one of the major conduits of energy flow up to higher trophic levels. Here, we address the processes that have generated the spectacular diversity of flowering plants (>300,000 species) and insect herbivores (likely >1 million species). Long-standing macroevolutionary hypotheses have postulated that reciprocal evolution of adaptations and subsequent bursts of speciation have given rise to much of this biodiversity. We critically evaluate various predictions based on this coevolutionary theory. Phylogenetic reconstruction of ancestral states has revealed evidence for escalation in the potency or variety of plant lineages' chemical defenses; however, escalation of defense has been moderated by tradeoffs and alternative strategies (e.g., tolerance or defense by biotic agents). There is still surprisingly scant evidence that novel defense traits reduce herbivory and that such evolutionary novelty spurs diversification. Consistent with the coevolutionary hypothesis, there is some evidence that diversification of herbivores has lagged behind, but has nevertheless been temporally correlated with that of their host-plant clades, indicating colonization and radiation of insects on diversifying plants. However, there is still limited support for the role of host-plant shifts in insect diversification. Finally, a frontier area of research, and a general conclusion of our review, is that community ecology and the long-term evolutionary history of plant and insect diversification are inexorably intertwined.

Evolutionary biology today and the call for an extended synthesis.

Evolutionary theory has been extended almost continually since the evolutionary synthesis (ES), but except for the much greater importance afforded genetic drift, the principal tenets of the ES have been strongly supported. Adaptations are attributable to the sorting of genetic variation by natural selection, which remains the only known cause of increase in fitness. Mutations are not adaptively directed, but as principal authors of the ES recognized, the material (structural) bases of biochemistry and development affect the variety of phenotypic variations that arise by mutation and recombination. Against this historical background, I analyse major propositions in the movement for an 'extended evolutionary synthesis'. 'Niche construction' is a new label for a wide variety of well-known phenomena, many of which have been extensively studied, but (as with every topic in evolutionary biology) some aspects may have been understudied. There is no reason to consider it a neglected 'process' of evolution. The proposition that phenotypic plasticity may engender new adaptive phenotypes that are later genetically assimilated or accommodated is theoretically plausible; it may be most likely when the new phenotype is not truly novel, but is instead a slight extension of a reaction norm already shaped by natural selection in similar environments. However, evolution in new environments often compensates for maladaptive plastic phenotypic responses. The union of population genetic theory with mechanistic understanding of developmental processes enables more complete understanding by joining ultimate and proximate causation; but the latter does not replace or invalidate the former. Newly discovered molecular phenomena have been easily accommodated in the past by elaborating orthodox evolutionary theory, and it appears that the same holds today for phenomena such as epigenetic inheritance. In several of these areas, empirical evidence is needed to evaluate enthusiastic speculation. Evolutionary theory will continue to be extended, but there is no sign that it requires emendation.

On the Measurement of Niche Breadth and Overlap.

Measures of niche breadth and overlap that depend on the distribution of individual among resource states (ecological categories) should be independent of the relative abundance of the species and of the number of resource states considered. Such measures should also take into account the degree of distinctness of the resource states from the point of view of the organisms concerned. An ecoassay of the distinctness of resource states may well be easier and more meaningful than measurements of physical and chemical factors. We propose that the species composition of communities utilizing different resource states may be used to develop weighting factors with which each state may be weighted in proportion to its degree of distinctness. The weighting factors are used in the development of indices of niche breadth and overlap that correct for variation in the range and distinctness of resource states and that suffer less from human subjectivity than do the measures used to date. The use of such indices and the relationship of niche overlap to competition are discussed.

Different effects of variation in Xanthium strumarium L. (Compositae) on two insect seed predators.

To determine the relative importance of variation in several plant characters on susceptibility to herbivores, we examined patterns of seed predation by two monophagous insect species and patterns of variation in ten populations of the cocklebur, Xanthium strumarium. Multiple regression analysis disclosed that one seed predator was most influenced by plant chemical variation, the other was significantly influenced by both chemical and morphological variation, but variation in yet another character, general burr size, was most important in conferring resistance to both insects simultaneously. The plant populations differed most in this character. Although many of the plant characters were correlated with each other, those important in determining susceptibility to each insect species were uncorrelated and independent of those conferring resistance to both insects simultaneously.These results imply that ecological similar herbivores may be influenced by different aspects of plant variation, and that predictions of evolutionary responses of local plant populations to herbivory may require knowledge of the structure of local herbivore communities and the dynamics of their establishment.

Apparent transgenerational effects of host plant in the leaf beetle Ophraella notulata (Coleoptera: Chrysomelidae).

The natural host of Ophraella notulata is Iva frutescens (Asteraceae); its close relative feeds on a related plant, Ambrosia artemisiifolia. We reared beetles on both plants, obtained progeny from the four possible crosses (two sexes X two parental hosts), and reared the progeny on both plant species. Survival to the imaginal stage of progeny reared on Iva varied with both maternal and paternal host. Hatchling feeding response to both plants showed a maternal host X paternal host interaction. Consumption of Ambrosia by adult beetles was, counter to expectation, higher for progeny of Iva-reared males than Ambrosia-reared males. Oviposition response, although based on too few data to be definitive, was peculiar: parental host did not affect oviposition on Ambrosia; on Iva daughters of Iva-reared males laid significantly more eggs than did daughters of Ambrosia-reared males, but only if they had been reared on Iva; those reared on Ambrosia displayed the reverse pattern. We discuss the possibility that nongenetic paternal transmission of host plant effects may explain these results, but offer a somewhat uncomfortable hypothesis of selection as a preferable explanation. An important outcome of the experiment is that it provided no evidence of maternal effects of host plant on offspring feeding or oviposition.

Varying herbivore population structure correlates with lack of local adaptation in a geographic variable plant-herbivore interaction.

Local adaptation of parasites to their hosts due to coevolution is a central prediction of many theories in evolutionary biology. However, empirical studies looking for parasite local adaptation show great variation in outcomes, and the reasons for such variation are largely unknown. In a previous study, we showed adaptive differentiation in the arctiid moth Utetheisa ornatrix to its host plant, the pyrrolizidine alkaloid-bearing legume Crotalaria pallida, at the continental scale, but found no differentiation at the regional scale. In the present study, we sampled the same sites to investigate factors that may contribute to the lack of differentiation at the regional scale. We performed field observations that show that specialist and non-specialist polyphagous herbivore incidence varies among populations at both scales. With a series of common-garden experiments we show that some plant traits that may affect herbivory (pyrrolizidine alkaloids and extrafloral nectaries) vary at the regional scale, while other traits (trichomes and nitrogen content) just vary at the continental scale. These results, combined with our previous evidence for plant population differentiation based on larval performance on fresh fruits, suggest that U. ornatrix is subjected to divergent selection even at the regional scale. Finally, with a microsatellite study we investigated population structure of U. ornatrix. We found that population structure is not stable over time: we found population differentiation at the regional scale in the first year of sampling, but not in the second year. Unstable population structure of the herbivore is the most likely cause of the lack of regional adaptation.

Evolutionary constraint and ecological consequences.

One of the most important shifts in evolutionary biology in the past 50 years is an increased recognition of sluggish evolution and failures to adapt, which seem paradoxical in view of abundant genetic variation and many instances of rapid local adaptation. I review hypotheses of evolutionary constraint (or restraint), and suggest that although constraints on individual characters or character complexes may often reside in the structure or paucity of genetic variation, organism-wide stasis, as described by paleontologists, might better be explained by a hypothesis of ephemeral divergence, according to which the spatial or temporal divergence of populations is often short-lived because of interbreeding with nondivergent populations. Among the many consequences of acknowledging evolutionary constraints, community ecology is being transformed as it takes into account phylogenetic niche conservatism and the strong imprint of deep history.

Dietary specialization influences the efficacy of larval tortoise beetle shield defenses.

Plant chemical defenses and escape from natural enemies have been postulated to select for dietary specialization in herbivorous insects. In field and laboratory bioassays, we evaluated the effectiveness of intact and chemically modified larval shield defenses of the generalist Chelymorpha alternans and the specialists Acromis sparsa and Stolas plagiata (Chrysomelidae: Cassidinae) against three natural predators, using larvae reared on two morning glory (Convolvulaceae) species. We assessed whether: (1) specialists were better defended than generalists when both were fed and assayed on the same plant; (2) larval shield defenses were chemical, physical, or both; and (3) specialists exploit chemistry better than generalists. Live specialist larvae survived at higher rates than did generalists in predator bioassays with the bug Montina nigripes (Reduviidae), but there were no differences among groups against two species of Azteca ants (Hymenoptera: Dolichoderinae). Solvent leaching by H(2)O or MeOH significantly reduced shield efficacy for all species compared to larvae with intact shields. In contrast, freshly killed specialist larvae exhibited significantly lower capture rates and frequencies than the generalists. Although solvent leaching significantly reduced overall shield efficacy for freshly killed larvae of all species, the pattern of leaching effects differed between specialists and generalists, with H(2)O-leaching having a greater impact on the specialists. The overall vulnerability of the generalists appears due to lower chemical protection, which is ameliorated by increased escape behaviors, suggesting a selective trade-off between these defensive components. These experiments indicate that shield defenses are essential for larval survival and that specialists are superior at exploiting plant compounds residing in the aqueous fraction. Our results support the hypothesis that diet-specialized herbivorous insects have more effective defenses than generalists when both feed on the same plant due to the differential ability to exploit defensive precursors obtained from the host. The evolution of dietary specialization may therefore confer the advantage of enhanced enemy-free space.

THE ORIGIN AND DISTRIBUTION OF CLONAL DIVERSITY IN ALSOPHILA POMETARIA (LEPIDOPTERA: GEOMETRIDAE).

A survey of spatial and temporal variation in the frequency of electrophoretically defined genotypes in the geometrid moth Alsophila pometaria revealed a high diversity of uncommon or rare asexual genotypes and clinal distributions of two of the more common clones. There was substantial year-to-year variation in genotype frequencies in seven of eleven sites. Progeny tests have revealed that sexual reproduction is uncommon in two populations and that new asexual genotypes arise from the sexual population. The recurrent origin of asexual genotypes is likely to account for the high genetic and ecological diversity of the asexual contingent of this species' populations, in contrast to the lower genetic diversity in some obligately asexual species in which such recruitment does not occur.

PHYLOGENY AND THE EVOLUTION OF HOST PLANT ASSOCIATIONS IN THE LEAF BEETLE GENUS OPHRAELLA (COLEOPTERA, CHRYSOMELIDAE).

Species of Ophraella, a North American genus of leaf beetles (Chrysomelidae), feed variously on eight genera in four tribes of Asteraceae. A phylogenetic analysis, based on morphological features and allozymes, was undertaken to deduce the history of host affiliation within the genus. The two data sets are combined to arrive at a provisional phylogeny of the species, onto which host associations are parsimoniously mapped. Among and within the 12 species studied, at least two shifts are postulated to have occurred among congeneric plant species, five between genera in the same tribe, and four between different tribes of Asteraceae. The phylogeny of Ophraella appears not to be congruent with that of its hosts. This and other evidence indicates that many host shifts in Ophraella postdate the divergence of the host plants, a conclusion that may apply commonly to phytophagous insects. A phenetic analysis of the plants' secondary compounds provides modest support for the hypothesis that host shifts are facilitated by commonalities in plant chemistry. A possible trend in host shifts is evident, from chemically simpler to chemically more forbidding plants. The chemical barriers to host shifts in Ophraella appear to require adaptation in both behavior and in physiological attributes. There is no evidence that the host associations of these insects or the divergence in secondary chemistry of their hosts can be attributed to coevolution.

GENETIC VARIATION AND COVARIATION IN RESPONSES TO HOST PLANTS BY ALSOPHILA POMETARIA (LEPIDOPTERA: GEOMETRIDAE).

Progeny of uncommon parthenogenetic genotypes of the polyphagous geometric Alsophila pometaria were reared on four host plant species representing the same genus, different confamilial genera, and different families. On the supposition that uncommon asexual genotypes have arisen recently from the sympatric sexual population, they may be viewed as a representation of variation in the sexual population that has been captured by parthenogenesis. In both the laboratory and the field, significant effects of genotype, host, and their interaction were found for survival. Live weight displayed significant effects of genotype, host, and genotype x host interaction in the laboratory, and significant main effects in the field. The broad heritability of live weight within hosts was significantly greater than zero in all cases, ranging from 0.18 to 0.33. Heritability values in the field were similar to those in the laboratory. Genetic correlations between weight attained on pairs of host plants, calculated from uncorrected family means, were all positive and many were statistically significant. When the data were corrected for differences in generalized vigor by taking the deviations from genotype means on a standard host plant, one of nine genetic correlations was significant and positive, and one was significant and negative. Within their limits of precision, these data suggest that genetic factors enhancing performance on one host do not generally have strong antagonistic pleiotropic effects on performance on the other hosts in this population. The relevance of these observations to the evolution of resource specialization in general and host specialization by phytophagous insects in particular is discussed. The common assumption that trade-offs in efficiency of utilization cause the evolution of specialized resource use requires more empirical evidence than seems to exist.

GENETIC CONSTRAINTS AND THE PHYLOGENY OF INSECT-PLANT ASSOCIATIONS: RESPONSES OF OPHRAELLA COMMUNA (COLEOPTERA: CHRYSOMELIDAE) TO HOST PLANTS OF ITS CONGENERS.

We ask whether patterns of genetic variation in a phytophagous insect's responses to potential host plants shed light on the phylogenetic history of host association. Ophraella communa feeds chiefly, and in eastern North America exclusively, on Ambrosia (Asteraceae: Ambrosiinae). Using mostly half-sib breeding designs, we screened for genetic variation in feeding responses to and larval survival on its own host and on seven other plants that are hosts (or, on one case, closely related to the host) of other species of Ophraella. We found evidence for genetic variation in feeding responses to five of the seven test plants, other than the natural host. We found no evidence of genetic variation in feeding responses to two plant species, nor in capacity for larval survival on six. These results imply constraints on the availability of genetic variation; however, little evidence for constraints in the form of negative genetic correlations was found. These results are interpreted in the context of a provisional phylogeny of, and a history of host shifts within, the genus. Ophraella communa does not present evidence of genetic variation in its ability to feed and/or survive on Solidago, even though it is probably descended from a lineage that fed on Solidago or related plants, possibly as recently as 1.9 million years ago. Genetic variation in performance on this plant may have been lost. Based on evidence for genetic variation and on mean performance, by far the greatest potentiality for adaptation to a congener's host was evinced in responses to Iva frutescens, which not only is related and chemically similar to Ambrosia, but also is the host of a closely related species of Ophraella that may have been derived from an Ambrosia-associated ancestor. Genetic variation in O. communa's capacity to feed and/or survive on its congeners' hosts is less evident for plants that do not represent historically realized host shifts (with one exception) than for those that may (but see Note Added in Proof). The results offer some support for the hypothesis that the evolution of host shifts has been guided in part by constrained genetic variation.

GENETIC CONSTRAINTS ON MACROEVOLUTION: THE EVOLUTION OF HOST AFFILIATION IN THE LEAF BEETLE GENUS OPHRAELLA.

We hypothesize that the evolution of an ecologically important character, the host associations of specialized phytophagous insects, has been influenced by limitations on genetic variation. Using as a historical framework a phylogenetic reconstruction of the history of host associations in the beetle genus Ophraella (Chrysomelidae), we have employed quantitative-genetic methods to screen four species for genetic variation in larval survival, oviposition (in one species only), and feeding responses to their congeners' host plants, in the Asteraceae. We here report results of studies of one species and evaluate the results from all four. Analysis of half-sib/full-sib families and of progenies of wild females of O. notulata, a specialist on Iva (Ambrosiinae), provided evidence of genetic variation in larval consumption of five of six test plants and in adult consumption of four of six. Larval mortality was complete on five plants; only on Ambrosia, a close relative of the natural host, was there appreciable, and genetically variable, survival. Oviposition on Ambrosia showed marginally significant evidence of genetic variation; a more distantly related plant elicited no oviposition at all. In compiling results from four Ophraella species, reported in this and two other papers, we found no evidence of genetic variation in 18 of 39 tests of feeding responses and 14 of 16 tests of larval survival on congeners' hosts. This result is consistent with the hypothesis that absence or paucity of genetic variation may constrain or at least bias the evolution of host associations. The lower incidence of genetic variation in survival than in feeding behavior may imply, according to recent models, that avoidance is a more common evolutionary response to novel plants than adaptation. The usually great disparity between mean performance on congeners' hosts and the species' natural hosts, and an almost complete lack of evidence for negative genetic correlations, argue against the likelihood that speciation has occurred by sympatric host shift. The presence versus apparent absence of genetic variation in consumption was correlated with the propinquity of relationship between the beetle species tested and the species that normally feeds on the test plant, suggesting that the history of host shifts in Ophraella has been guided in part by restrictions on genetic variation. It was also correlated with the propinquity of relationship between a test plant and the beetle's natural host. The contributions of plant relationships and insect relationships, themselves correlated in part, to the pattern of genetic variation, are not readily distinguishable, but together accord with phylogenetic evidence that these and other phytophagous insects adapt most readily to related plants. In this instance, therefore, the macroevolution of an ecologically important character appears to have been influenced by genetic constraints. We hypothesize that absence of the structural prerequisites for genetic variation in complex characters may affect genetic variation and the trajectory of evolution.

TESTS OF INBREEDING EFFECTS ON HOST-SHIFT POTENTIAL IN THE PHYTOPHAGOUS BEETLE OPHRAELLA COMMUNA.

Although inbreeding, on average, decreases additive genetic variance, some inbred populations may show an increase in phenotypic variance for some characters. In those populations with increased phenotypic variance, character changes by peak shifts may occur because of the effects of the higher variance on the adaptive landscape. A population's increased phenotypic variance may place it in the domain of attraction of a new adaptive peak or increase the likelihood of a selection-driven peak shift as the landscape of mean fitness flattens. The focus of this study was to test for increased variance, in inbred populations, in a behavioral character involved in adaptive diversification and probably speciation. We examined the effect of inbreeding on feeding responses of the leaf beetle Ophraella communa in a series of inbred lineages across a range of levels of inbreeding (f = 0.25, 0.375, 0.5). We measured the feeding response of inbred lineages of O. communa on its normal host, Ambrosia artemisiifolia, and on two novel plants, Chrysopsis villosa and Iva frutescens, that are the hosts of other Ophraella species. The results show that feeding responses on the different plants are not correlated, indicating that the feeding responses to the different plants are to some degree genetically independent. Despite apparent genetic variation in lineage feeding responses, we could not statistically demonstrate increases in phenotypic variance within the lineages. Thus, the experimental results do not support the idea that host shifts in this beetle evolved by peak shifts in bottlenecked populations.

INSIGHT INTO SPECIATION FROM HISTORICAL DEMOGRAPHY IN THE PHYTOPHAGOUS BEETLE GENUS OPHRAELLA.

Speciation in phytophagous insects is commonly associated with shifts in host use. Using a phylogenetic framework to identify recently diverged taxa that have undergone a radical host shift, this study focuses on how reconstruction of the historical demography of a species, in conjunction with branching patterns between species, provides insight into mode of speciation. Analyses of mitochondrial cytochrome oxidase I sequences indicate that the leaf beetle Ophraella communa exhibits significant population structure, as shown by patterns of genealogical relationships among mitochondrial haplotypes and high FST -values. However, the absence of regional localization of old clades of haplotypes, negative Tajima's D, and unimodal rather than bimodal frequency distribution of the number of pairwise differences between sequences suggests an absence of long-term barriers to gene flow. Furthermore, we found no evidence of isolation by distance. This pattern of genetic variation is consistent with episodes of gene flow on a large geographic scale, perhaps owing to Pleistocene changes in climate. Ophraella communa and its sister species O. bilineata diverged during the early Pleistocene. The evidence of dynamic population structure in O. communa, potentially including episodic but massive gene flow, suggests that reproductive isolation evolved quite rapidly on a localized geographic scale, because speciation would probably have been reversed by gene flow if the evolution of reproductive isolation had been prolonged. That is, gene flow occasioned by range shifts during the Pleistocene would likely have interrupted speciation unless it occurred very rapidly. Sequence diversity implies a large effective population size (> 106 ) in both O. communa and O. bilineata. However, a model based on a drastic bottleneck did not have a lower likelihood than a model with no bottleneck, simply because the time since speciation has been great enough for coalescence to a single ancestor that existed after the speciation event. Sequence diversity in itself, without reference to the time since speciation, cannot provide evidence on the demography of speciation.