The susceptibility of parental and hybrid willows to plant enemies under contrasting soil nutrient conditions.

We conducted an experimental study of the effects of nutrient addition on the susceptibility of two species of willows (Salix eriocephala and S. sericea) and their hybrid to a pathogen and several herbivores. We hypothesized that the relative susceptibility of parental and hybrid willows would depend upon soil nutrient availability and vary among plant enemies. Using potted plants in a common garden, we found that S. eriocephala was significantly more susceptible to attack by a fungal rust (Melampsora sp.), a leaf-chewing beetle (Popillia japonica), and a leaf-folding sawfly (Phyllocolpa nigrita) than was S. sericea. Conversely, S. sericea was significantly more susceptible to attack by a spider mite (Tetranychus sp.) and a leaf-mining caterpillar (Phyllocnistis sp.) than was S. eriocephala. Hybrid susceptibility to Melampsora sp. and to Phyllocnistis sp. resembled S. eriocephala's, while hybrid susceptibility to Phyllocolpa nigrita, Tetranychus sp., and Popillia japonica resembled S. sericea's. Susceptibility to a sixth enemy, another leaf-mining caterpillar, Phyllonorycter salicifoliella, did not differ among the parents and hybrid. Susceptibility to herbivores and pathogens increased along a gradient of increasing fertilizer application, and this effect was independent of plant taxon or enemy. The results of our study point to the contrasting influences of a taxonomically and functionally diverse enemy community, reinforce the hybrid dominance model of enemy susceptibility, and demonstrate that physiological stress and enemy susceptibility can be inversely related.

Interspecific hybridization of plants and resistance to herbivores: hypotheses, genetics, and variable responses in a diverse herbivore community.

We studied the morphology, molecular genetics, and hebivory of two species of willows (Salix sericea and S. eriocephala) and their interspecific hybrids to test four alternative hypotheses concerning the effects of hybridization on plant resistance. Individually marked plants were identified using morphological traits in the field and measurements of stipule and leaf pubescence were made and compared using Canonical Discriminant Function Analysis. DNA was extracted from the leaves of a sample of the marked plants and RAPD-PCR analysis was performed to establish the genetic status of parental and hybrid plants. RAPD band analysis generally verified the genetic status of parental plants. Hybrid plants were usually correctly identified in the field with a few exceptions. However, the hybrid plants were a heterogeneous group of plants made up of most plants that appear to be F1s and a few plants that appear to be backcrosses to S. sericea. Morphological variables were useful for distinguishing S. sericea from S. eriocephala and hybrids, but were not as dependable in distinguishing between S. eriocephala and hybrids. We compared the densities of 11 herbivore species and the infection by a leaf rust pathogen (Melampsora sp.) on the leaves and stems of two parents and the hybrids in the field. We found support for the Additive hypothesis (3 species), the Dominance hypothesis (2 species) and the Hybrid Susceptibility hypothesis (7 species, 6 herbivores and the Melampsora rust). We found no evidence for the Hybrid Resistance hypothesis. Guild membership was not a good predictor of similar responses of species to hybrid versus parental plants. A Canonical Discriminant Function Analysis showed discrete separation of the taxa based on herbivore densities, illustrating different community structures on hybrid and parental plants. This study demonstrates the diversity of responses of phytophages in response to interspecific hybridization.

The effects of hybridization in plants on secondary chemistry: implications for the ecology and evolution of plant-herbivore interactions.

Natural hybridization is a frequent phenomenon in plants. It can lead to the formation of new species, facilitate introgression of plant traits, and affect the interactions between plants and their biotic and abiotic environments. An important consequence of hybridization is the generation of qualitative and quantitative variation in secondary chemistry. Using the literature and my own results, I review the effects of hybridization on plant secondary chemistry, the mechanisms that generate patterns of chemical variation, and the possible consequences of this variation for plants and herbivores. Hybrids are immensely variable. Qualitatively, hybrids may express all of the secondary chemicals of the parental taxa, may fail to express certain parental chemicals, or may express novel chemicals that are absent in each parent. Quantitatively, concentrations of parental chemicals may vary markedly among hybrids. There are five primary factors that contribute to variation: parental taxa, hybrid class (F(1), F(2), etc.), ploidy level, chemical class, and the genetics of expression (dominance, recessive vs. additive inheritance). This variation is likely to affect the process of chemical diversification, the potential for introgression, the likelihood that hybrids will facilitate host shifts by herbivores, and the conditions that might lead to enhanced hybrid susceptibility and lower fitness.

Preserving leaves for tannin and phenolic glycoside analyses: A comparison of methods using three willow taxa.

Members of the Salicaceae often produce phenolic glycosides and condensed tannins. There is much debate on the best method for the preservation of leaf material prior to chemical analysis. Published results indicate freeze-drying, a method commonly used for tannin analysis, may be inappropriate for phenolic glycosides, unless done in a manner to prevent thawing during the drying process. Another commonly employed method, air-drying, is appropriate for phenolic glycosides but inappropriate for condensed tannins. I present evidence using willow leaves that demonstrates that: (1) leaves freeze-dried in external flasks without temperature control contain lower concentrations of phenolic glycosides (salicortin and 2'-cinnamoylsalicortin), (2) air-dried leaves have reduced concentrations of condensed tannins, while (3) vacuum-dried fresh leaves have high concentrations of both phenolic glycosides and condensed tannins. Freeze-drying caused salicortin and 2'-cinnamoyl salicortin concentrations to drop by 20 mg/g and 4 mg/g, respectively. Salicin, a product of salicortin and 2'-cinnamoyl salicortin degradation, is absent in vacuum-dried leaves, present in air-dried leaves and very high in freeze-dried leaves. Thus, the presence of salicin in this system is an artifact of the preservation technique. Condensed tannin concentrations dropped nearly 20 mg/g when leaves were air-dried. Thus, vacuum-drying fresh leaves allows researchers to quantify phenolic glycosides and condensed tannins from the same leaf material.

Secondary chemistry of hybrid and parental willows: Phenolic glycosides and condensed tannins inSalix sericea, S. eriocephala, and their hybrids.

Salix sericea andS. eriocephala differ markedly in secondary chemistry.S. sericea produces phenolic glycosides, salicortin and 2'-cinnamoylsalicortin, and low concentrations of condensed tannin. In contrast,S. eriocephala produces no phenolic glycosides, but high concentrations of condensed tannins. Hybrid chemistry is intermediate for both types of chemicals, suggesting predominantly additive inheritance of these two defensive chemical systems from the parental species. However, there is extensive variation among hybrids. This variation may be due to genetic variation among parental genotypes, which genes were passed on, or to subsequent back-crossing. The differences in chemistry are likely to exert a strong effect on the relative susceptibility of hybrid and parental willows to herbivores.

Morphological and molecular evidence for hybridization and introgression in a willow (Salix) hybrid zone.

Hybrid zones provide biologists with the opportunity to examine genetic and ecological interactions between differentiated populations. Accurate identification of hybrid genealogies is considered a necessary prerequisite to understanding observed patterns of hybridization-related phenomena. We analysed molecular and morphological data from individuals in a hybrid zone between two species of willows (Salix sericea Marshall and S. eriocephala Michaux) and report the use of randomly amplified polymorphic DNA (RAPD), chloroplast DNA (cpDNA), and ribosomal DNA (rDNA) markers, as well as vegetative morphology and foliar chemistry data to identify individuals in terms of hybrid genealogy and to infer the direction and extent of backcrossing and introgression within the hybrid zone. A novel version of a maximum likelihood estimate approach (developed for this study) was used to calculate hybrid index scores from RAPD marker data; this method produced results similar to those obtained using traditional arithmetic methods. Distribution of rDNA, cpDNA, and chemistry data were examined within the graphical context of RAPD-based hybrid index score histograms and principal component analyses (PCA) on RAPD and morphology data. Seven of the 21 plants classified as S. eriocephala in the field were possible introgressants. Another plant presented an unequivocal example of backcrossed S. sericea chemistry and RAPD markers. Inter- and intraspecific chloroplast diversity found within the hybrid zone suggests both historic introgression (perhaps in a glacial refugium), and contemporary hybridization. Patterns of inheritance and expression within the hybrid zone suggest that morphological characters are often not expressed in a simple additive fashion, and problems associated with both morphological and molecular data are considered.