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Ecological consequences of hybridization between a wild species (Echinacea purpurea) and related cultivar (E. purpurea `White Swan')
Scientia Horticulturae 76 (1998) 73±88
Ecological consequences of hybridization between a wild species (Echinacea purpurea) and related cultivar (E. purpurea `White Swan') Tami M. Van Gaal1, Susan M. Galatowitsch*, Mark S. Strefeler Department of Horticultural Science, University of Minnesota, 305 Alderman Hall, 1970 Folwell Avenue, St. Paul, MN 55108, USA Accepted 20 February 1998
Abstract In the first such study of its kind, we examined gene flow potential between a valued native plant and a popular cultivar. We studied outcomes of controlled hybridization between Echinacea purpurea and a conspecific cultivar, E. purpurea `White Swan', by comparing levels of competitive ability and reproductive potential. Differences in plant performance of wild-types and F10 s were studied at three densities (20.3, 45.7, and 182.9 plants/m2) using a field competition experiment. Wild-types were slightly larger but F1s had higher reproductive output. These differences can be attributed to floricultural breeding of the cultivar parent. Based on the results of this research, if the initial hybridization between the wild-type and cultivar were to occur, it is anticipated that the resulting F1 generation would survive and reproduce, creating the potential for continued gene flow. This methodology has potential for use in risk assessment of plant introductions, including transgenic crops. # 1998 Elsevier Science B.V. All rights reserved Keywords: Echinacea purpurea; Purple cone¯ower; Competition; Fitness; Gene ¯ow; Hybridization; Minnesota; Prairie; Risk assessment
1. Introduction While most introduced plants cannot compete with indigenous flora and can only be maintained in cultivation (Harper, 1965), the exceptional introductions * Corresponding author. Fax: +1 612 624 4941; e-mail: [email protected] 1 Present address: Wagner Greenhouses, 6024 Penn Avenue South, Minneapolis, MN 55419, USA. 0304-4238/98/$19.00 # 1998 Elsevier Science B.V. All rights reserved PII S 0 3 0 4 - 4 2 3 8 ( 9 8 ) 0 0 1 1 9 - 8
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that do become invasive have severe impacts on natural and/or cultural ecosystems (Ruesink et al., 1995). Within the context of cultural landscapes, the evolution of new or more aggressive weedy species via the introduction and/ or interbreeding with exotic, transgenic, or cultivated plants is a common concern (National Research Council, 1989; Ellstrand and Hoffman, 1990; Klinger et al., 1992; Till-Bottraud et al., 1992; Klinger and Ellstrand, 1994; Raybould and Gray, 1994). One perceived risk is that gene flow can affect existing populations (Abbott, 1992; Harper, 1965), resulting in altered relative fitness levels within the wild or crop populations. Although certain traits of cultivated plants are not expected to confer a fitness advantage in the wild (dwarfism, non-shattering seed heads, absence of dormancy), other traits (resistance to diseases, pests, and stresses) are advantageous in the wild and are expected to persist once integrated into weedy populations (Ellstrand and Hoffman, 1990; National Research Council, 1989; Klinger et al., 1992; Raybould and Gray, 1994; Till-Bottraud et al., 1992). Predicting the ecological and genetic consequences of such plant interactions to reduce the likelihood of future harmful invasions (Ruesink et al., 1995) is gaining interest. Very little work has been performed, however, to determine the effects of gene flow on hybrid fitness ± see reviews by Arnold and Hodges (1995) and Rieseberg (1995). Arnold and Hodges (1995) conclude that (1) most hybrids studied do not exhibit the lowest fitness when compared to both parents and (2) hybridization may lead to production of relatively fit genotypes. Three studies illustrate these points. Levin and Schmidt (1985) determined that although the hybrids of Phlox drummondii did not possess a significant increase in fitness over the parents, they did not suffer a fitness disadvantage, and would likely persist. Klinger and Ellstrand (1994) showed that the fitness of Raphanus sativus hybrids was greater than the fitness of the weedy parents, evidenced by the increased fruit and seed production. Hybrids of Oryza sativa possessed intermediate fitness compared with the parental types with respect to vegetative traits (Langevin et al., 1990). The resulting convergence in morphology could result in a dramatic increase in fitness in an agricultural setting for the F1 (Langevin et al., 1990). Together these studies provide evidence that introduced plants, through the action of gene flow, have the potential to impact wild populations. Thus far, no studies have examined the potential for gene flow between valued native plants and commonly used cultivars. Many important horticultural cultivars are derived from wild species, and these cultivars are often planted near populations of wild relatives, or even amongst them. To address the potential impact of gene flow from cultivars to native forbs, we examined the consequences of gene flow between Echinacea purpurea and a conspecific cultivar, E. purpurea `White Swan'. Echinacea purpurea, an herbaceous perennial and obligate outcrosser, is a common indigenous member
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of the tall grass prairie community in the central United States valued as a medicinal herb. By obtaining hybrids through controlled greenhouse crosses between wild-type and cultivar, we studied the consequences of gene flow instead of the process itself. We used a field competition experiment as a tool to compare the relative fitness of the wild-type and the F1 hybrids to investigate the hypothesis that the F1 generation will have different levels of fitness than the wild-type parent. To test the hypothesis, the study determined if the wild-type or F1 plants had a competitive advantage and/or differing reproductive potential. 2. Methods 2.1. Experimental design The design of the competition study was based on that proposed by Goldberg and Werner (1983) and implemented by Goldberg (1987). This design takes advantage of the benefits of a neighborhood design (Mack and Harper, 1977), allowing the response of a target species to be measured over a range of densities of a neighbor species. Competition can be measured by regressing the performance of the target individuals against the `amount' of neighbor species (e.g. density or biomass). This research used the basic experimental design proposed by Goldberg and Werner (1983) with two modifications: (1) all plants were planted in an agricultural field in a predetermined design instead of planting the target plants into natural populations of neighbor plants and (2) the neighbors were arranged in hexagonal annuli ensuring that each plant was equidistant from each immediate neighbor. Our variation of Goldberg and Werner's design was a 223 factorial design comparing target type, neighbor type, and density. A single plant of either the wild-type parent or the F1 served as the target plant in each plot. The neighbor group consisted of 18 plants of either the wild-type or F1 in two hexagonal annuli (Fig. 1). The effects of three density levels of neighbors were examined by relaxing the annuli of neighbor plants around the target individual. In the high density plots, the plants were separated by 0.1 m to create a density of 183 plants/m2. The plants in the medium density treatment were separated by 0.2 m with a resulting density of 46 plants/m2. The low density treatment (20 plants/m2) was achieved by separating the plants by 0.3 m. A total of 12 treatments were applied; each treatment was represented once in each of four blocks across the field. 2.2. Plant material Seeds of wild-type E. purpurea were obtained from Prairie Moon Nursery (Winona, Minnesota). These seeds were progeny of plants grown from seeds
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Fig. 1. Hexagonal plot design used for neighborhood competition experiment. The target plant is located in the center of the plot (T), surrounded by two rings of neighbor s (N).
collected from the restored Curtis Prairie at the Arboretum of the University of Wisconsin, Madison (43.18N, 89.48W; Fig. 2) ca. 1970. E. purpurea was established within the prairie using mostly transplants and some seeds collected from wild stands in Illinois (V. Kline, Univeristy of Wisconsin-Madison, personal communication). Echinacea purpurea had a pre-settlement distribution ranging from Oklahoma to Louisiana and Georgia north to Iowa, Illinois, Michigan, and Ohio (McGregor, 1968; Fig. 2). The species has since naturalized in a larger range that includes Wisconsin and Ontario (Cody and Boivin, 1973; Fig. 2). Seeds of the cultivar parent, E. purpurea 'White Swan', were obtained from G.S. Grimes Seed Company (Concord, Ohio). E. purpurea `White Swan' differs from the wild-type in flower color, flower number, and plant stature. This dwarf cultivar produces more flowers, and the flowers are white as compared to the purplish-pink flowers of the wild-type. After purchase all seeds were stored at 78C, 35% relative humidity. The F1 progeny were obtained by performing crosses (wild typecultivar) in the greenhouse in late spring, 1994. Plants used in crosses were in their first year of growth in the greenhouse. Resulting F1 seeds were harvested in late summer of 1994 and stored at 78C, 35% relative humidity. Seeds used in the competition study were germinated in petri dishes in a growth chamber (258C, 16 h light). Germination followed a 2 week stratification (4±18 October 1994) in 1 mM ethephon at 48C under 24 h light (Feghahati and Reese, 1994). Germinated seeds were moved into the greenhouse (DT278C, NT158C, 14 h photoperiod) in 98-cell plug trays for 3 weeks, then transferred to randomly arranged 10 cm standard pots. On 21 December 1994 plants were
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Fig. 2. Distribution of Echinacea purpurea (shaded area; adapted from McGregor, 1968). The location of the St. Paul, Minn. field plot, site of the neighborhood competition experiment, is indicated by the upright triangle. The inverted triangle indicates the location of Curtis Prairie at the University of Wisconsin-Madison Arboretum, the source population of the wild-type Echinacea purpurea.
transferred to a cooler to begin vernalization. The cooler environment (initially DT228C, NT198C, 12 h photoperiod) was altered weekly by dropping the night temperature 38C and decreasing the photoperiod by 1.5-h per week (ending environment DT228C, NT108C, 8 h photoperiod). On 20 January 1995, the cooler environment was adjusted to 48C without a photoperiod. After nine weeks of vernalization the plants were repotted to 13 cm pots and placed into a heated coldframe. 3. Experimental site The experiment was conducted in an agricultural field on the St. Paul campus of the University of Minnesota in east central Minnesota, USA (458N, 938W).
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The field had been used for forage production (corn) two years before the experiment and was planted with buckwheat (Fagopyrum sagittatum) during the previous year. On 8 May 1995, 56 plot centers were established in a grid at 15 m intervals across the field. A circular area with a diameter of 1 m was cleared around each plot center. The plots were divided into 4 blocks, and treatments were randomly assigned to plots within blocks. On 9 May 1995, plants were transplanted to the field. The plots were irrigated at the time of planting and for two weeks after planting to ensure establishment. Plants that died within the first two weeks of the study were replaced. Fertilizer was not applied during the study. Throughout the study, plots were kept clear of weeds and the buckwheat immediately surrounding the plots was pruned to reduce shading. 3.1. Plant response Three measures of plant performance ± biomass production, reproductive output, and plant structure were examined to determine the competitive effects of target type, neighbor type, and density on target plant performance. Observations were collected from target and first annulus plants only. Initial measurements for each plant, including number of leaves and plant height, were made at the time of planting. Plants were harvested for determination of aboveground biomass on 21 and 22 August 1995 (peak biomass). Plants were harvested to ground level, separated into vegetative and floral components, dried at 658C for 48 h, and weighed. The number and dry weight of inflorescences represented reproductive output. Because plants were harvested at peak biomass (before full seed maturation), mature seed weight could not be used as a reliable measure of fitness. Plant structure was measured by recording number of shoots, number of leaves, and plant height at four-week intervals throughout the study. 3.2. Analysis Through preliminary analyses, we found that a multiplicative model best fit the data, therefore, further analyses were performed on log transformed data (Mendenhall and Sincich, 1989). General main effects of target type, neighbor type, and density on target plant performance (total biomass) were determined using ANOVA (PROC GLM; SAS Institute, 1994). Two components of competitive ability, competitive response and competitive effect, were examined using regression analysis (Goldberg and Werner, 1983; Goldberg, 1987; PROC REG, SAS Institute, 1994). Competitive response was defined as the effect of target type on target plant performance. Competitive effect was defined as the effect of neighbor type on target plant performance. Competitive response and effect were further explained by regressing total target biomass on leaf number,
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plant height, shoot number, inflorescence number and average inflorescence biomass (PROC REG, SAS Institute, 1994). 4. Results Overall survival rates of wild-type and F1 plants were similar (99%). Only three plants (all neighbors) failed to establish; two F1 and one wild-type individuals were replaced in six days after planting. All remaining plants survived the duration of the experiment. The effect of neighbor density on the growth of target plants was strong in all experimental combinations (ANOVA, F36.29, df2, p0.0001). Plants in low density plots were taller, possessed more leaves, and had more numerous stems compared to medium and high density plants (Table 1). For both wild-type and F1 plants, total biomass and floral biomass under high density were 25±31% of the values at low density and Table 1. Under high density compared to low density, plant height was reduced by 16±17% and shoot, leaf, and inflorescence number were reduced by 42±68% (Table 1). With few exceptions, plant growth and morphology of wild-type and F1 plants were similar within densities. Differences between wild-type and F1 plants were not easily distinguished in the field; plants appeared similar in height, stem number, leaf number, and inflorescence number. Only four measures of plant growth and morphology revealed statistically significant differences between wild-type and F1 target plants in five specific comparisons: plant height at low density, number of shoots at medium density, floral biomass at medium density, and average inflorescence biomass at medium and high densities (Table 1). Wild-type plants were taller at low densities and possessed more stems at medium densities (p0.03 and p0.09, respectively). Although differences between wild-type and F1 plants for the remaining comparisons were not statistically different (p>0.1), several trends were apparent. First, measures of plant size (total biomass, plant height, stem number and leaf number) were generally greater in wild-type plants compared to F1 plants. For a given density, combined total biomass of neighbors was always higher when the neighbors were wild-type (Table 2). Although inflorescence number was generally higher in wild-type plants, floral biomass and average inflorescence biomass were greater in F1 plants. With respect to target type and neighbor type, differences in biomass production in target plants could be attributed to differences in plant structure and reproductive output. Plants with greater biomass tended to have larger and more numerous leaves, increased branching, and increased inflorescence number. Although final measures of plant structure affected total biomass, initial height and leaf number of plants were not significantly correlated with total biomass
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Plant type
Density
Total biomass
Floral biomass
Height (cm)
Shoot number
Leaf number
Inflorescence number
Average inflorescence biomass (g)
Wild-type
20.3 45.7 182.0
75.3 (33.8) 39.9 (22.5) 20.7 (22.5)
13.0 (8.3) 6.6 (6.9)* 4.0 (3.5)
79.7 (15.3)** 71.6 (21.1) 66.3 (194.4)
6.4 (2.2) 5.2 (2.1)** 3.7 (1.5)
78.1 (32.2) 49.7 (24.8) 31.9 (16.1)
8.9 (6.3) 4.9 (5.3) 2.8 (2.4)
1.6 (1.0) 1.0 (0.9)*** 1.1 (0.8)**
F1
20.3 45.7 182.9
70.2 (29.5) 40.4 (21.0) 17.3 (11.1)
14.3 (8.3) 9.4 (10.0) 4.4 (3.3)
73.9 (13.3)** 72.4 (15.7) 61.9 (14.0)
6.4 (2.5) 4.5 (1.6)** 3.5 (1.2)
77.9 (31.6) 48.6 (198.8) 30.0 (13.7)
8.1 (4.3) 4.7 (3.1) 2.8 (1.9)
1.8 (0.8) 1.7 (1.3)*** 1.4 (0.7)**
*P<0.1;**P<0.05;***P<0.01.
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Table 1 Mean (SD) effect of density on growth and morphology of wild-type and F1 Echinacca purpurea plants used as target and neighbor plants in neighborhood competition, separated by the density at which they were grown. Asterisks indicate significant differences between wild-type and F1 plants at the same density
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Table 2 The effect of density (mean, SD) on target total biomass and combined neighbor total biomass of wild-type and F1 Echinacea purpurea plants used in neighborhood competition experiment Target type
Neighbor type
Density (plants/m2)
Target total biomass (g)
Combined neighbor total biomass (g)
Mean
sd
Mean
sd
Wild-type
Wild-type
20.3 45.7 182.9
73.9 36.1 14.0
36.2 17.5 5.1
469.6 287.9 142.4
70.0 98.7 31.7
Wild-type
F1
20.3 45.7 182.9
67.6 23.2 11.5
35.2 13.1 3.5
437.9 257.2 113.8
98.3 66.9 40.1
F1
F1
20.3 45.7 182.9
46.4 28.0 17.2
17.8 8.3 4.2
427.7 252.2 102.0
94.8 72.1 48.9
F1
Wild-type
20.3 45.7 182.9
70.7 28.4 8.6
35.2 16.3 5.3
451.6 210.7 120.1
99.0 48.1 32.7
(p0.16 and p0.21 respectively). Of the three measures of plant structure (leaf number, height, and stem number) and two measures of reproductive output (inflorescence number and average inflorescence biomass) used in regression analysis, three contributed to differences in total biomass of target plants (Table 3). Leaf number was the only measure of plant structure to affect target total biomass for all target type and neighbor type combinations (Table 3). Average inflorescence biomass contributed to differences in target total biomass only if the neighbors were of the same type as the target, while inflorescence number affected total biomass of wild-type targets with wild-type neighbors only (Table 3). Total plant biomass was not affected by plant height or stem number under any treatment (Table 3). The only significant relationship between plant structure or reproductive output and target total biomass observed was inflorescence number on wild-type targets with wild-type neighbors. The competitive effect of F1 neighbors on F1 target plants generated the most significant competitive difference observed in the study; F1 target plants performed differently with wild-type or F1 neighbors. The significant difference in slope of the regression of total biomass of F1 targets on combined total biomass of F1 neighbors is explained by a negative relationship between target plant performance and combined neighbor biomass at high densities (Fig. 3(a)±(d)). Within the high density plots, the competitive effect of wild-type neighbors (open circles; ûÿ2.285, p0.038) was stronger than the effect of F1 neighbors (solid circles; ûÿ0.526, p0.0076; t3.80 p<0.005), indicating the wild-type plants
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Target type
Wild-type Wild-type F1 F1
Neighbor type
Wild-type F1 F1 Wild-type
Leaf number
Height
Coefficient
p
3.09 0.90 0.90 1.44
0.001 0.051 0.098 0.118
Coefficient 0.54 ÿ0.76 ÿ0.17 1.40
Shoot number
Inflorescence number
p
Coefficient
p
0.597 0.301 0.696 0.354
ÿ0.44 ÿ0.18 0.33 ÿ0.81
0.445 0.550 0.466 0.132
Coefficient ÿ1.30 0.44 ÿ0.17 ÿ0.21
Average inflorescence biomass p
Coefficient
p
0.002 0.265 0.725 0.774
0.73 ÿ0.33 1.12 ÿ0.56
0.060 0.202 0.002 0.437
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Table 3 The effects of plant structure and reproductive effort on total target biomass of wild-type and F1 Echinacea purpurea used as target plants in a neighborhood competition experiment. Coefficients and p-values were determined using multiple regression on log/log transformed data
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Fig. 3. Comparisons of competitive abilities of wild-type and F1 Echinacea purpurea plants used in a neighborhood competition experiment, including the effect of wild-type neighbors on competitive response of wild-type and F1 target plants (Fig. 3(a)), effect of F1 neighbors on competitive response of wild-type and F1 target plants (Fig. 3(b)), competitive effect of wild-type and F1 neighbors on wild-type target plants (Fig. 3(c)), and competitive effect of wild-type and F1 neighbors on F1 target plants (Fig. 3(d)). Ovals enclose data points from different planting densities represented by symbols: circles for high, diamonds for medium, and squares for low density plots (filledF1, openwild type). The solid line shows the regression function for F1 plants; the dashed line is for wild type plants.
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were slightly stronger competitors. Unlike F1 target plants, wild-type targets performed similarly when surrounded by wild-type or F1 neighbors (t0.669; p>0.1). Differences in competitive response of target plants were not strong (ANOVA: F0.07, df1, p0.7974); target performance (biomass) did not vary within a density for either neighbor. 5. Discussion Hybridization between the wild-type, Echinacea purpurea, and cultivar, E. purpurea `White Swan', can produce an F1 generation that would survive and reproduce, creating the potential for continued gene flow. Survival and reproduction of the F1 generation can be anticipated based on the similar fitness levels of the wild-type parent and F1. Measures of plant structure and competitive ability indicated the wild-type population had a slight fitness advantage but measures of reproductive output indicated the F1 had an advantage, although these differences did not evoke a strong biological response. Overall, the two plant classes were more similar than they were different. Wild-type and F1 plants did, however, exhibit differences in resource allocation, which may affect overall plant performance over time. Gene flow will likely have a long term effect on Echinacea populations for two reasons: (1) the high survival rates of the F1s and (2) the greater reproductive output of the F1s. The high survival rates of the F1, which were similar to the wild-type parent, indicate the hybrid product of gene flow between the cultivar and wild-type populations could be expected to survive once established. These initial results are similar to the findings of Levin and Schmidt (1985), who determined that hybrids of two subspecies of Phlox did not exhibit dramatic decreased or increased fitness relative to either parent population, and are consistent with the views of Arnold and Hodges (1995), who have concluded that most hybrids studied are of similar or intermediate fitness compared to the parents. In every case, the F1 plants had a greater floral biomass and greater average inflorescence biomass compared to the wild-type plants. Based on increased reproductive output, the F1s possess marginally greater fitness than the wild-type plants. Differences in resource allocation are likely a direct result of commercial breeding of the cultivar parent. Floricultural breeders favor a pattern of resource allocation that leads to increased reproductive output (more flowers). The cultivar parent, E. purpurea `White Swan' has been subject to stringent selection through breeding for compactness and high floral output, including high flower number, large flower size, and extended bloom time. In addition to these ornamental traits, E. purpurea `White Swan' was also bred to be seed propagated, with selection for high seed count per inflorescence and strong seed performance (high seed viability and germination rates). The smaller size and greater reproductive output
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of the F1 plants are likely traits inherited from the cultivar parent. These morphological traits are known to be highly heritable in other genera (Hallauer and Miranda, 1982), justifying this conclusion. The results of this research suggest that traits acquired from a cultivar parent can be expressed in hybrid generations. For many plant species, larger plants have greater competitive ability and therefore greater fitness (Goldberg, 1987). In a neighborhood competition experiment, Goldberg (1987) found that larger sized neighbors had a greater competitive effect on target Solidago plants. A positive correlation between plant size and competitive ability was also described within the genus Echinacea (Snyder et al., 1994). With respect to plant structure, a general trend in the data from our experiment indicates that wild-type plants are slightly larger than F1 plants, suggesting the wild-type plants could possess marginally greater fitness than F1 plants. The slightly higher fitness of the wild-type plants, conferred through larger plant size, could result in increased long-term performance of wild-types relative to the F1s in a prairie community. Had the cultivar parent been selected for increased biomass, as are some forage grasses, the F1 would likely exhibit increased competitive ability relative to the wild-type plant. The contrast between the F1 plants, which exhibit greater reproductive output, and the wild-type plants, which exhibit greater vegetative growth, suggests differences in resource allocation. Compared to annuals, herbaceous perennials allocate more resources to vegetative growth, especially below-ground biomass, at the expense of reproductive output. This pattern of resource allocation, especially for nitrogen, has been shown to result in superior competitive ability for resources (Tilman, 1990). In this experiment, the F1 plants appeared to allocate more resources to reproductive output than the wild-type plants. Increased reproductive output would result in greater seed production only if this were not outweighed by decreased long term performance and survival due to decreased acquisition of light, nutrients, and water (Harper, 1965). Decreased performance of target plants with increasing density of neighbor plants is a typical plant response that has been attributed to the effects of competition (Goldberg, 1987; Antonovics and Fowler, 1985; Shaw and Platenkamp, 1993). In our study, the decreased performance of target plants at high densities indicated that competition for resources did occur; therefore, comparisons of competitive ability were possible. Patterns in the variance of total biomass (higher variance around measures at low densities compared to high densities) suggested that competition was an important factor in plant growth and survival, i.e. fitness (Goldberg, 1987). The differences in competitive effect of wild-type and F1 neighbors can be attributed to the cumulative effects of competition. These cumulative effects, called diffuse competition, have been detected among species in natural communities (MacArthur, 1972; Dyer and Rice, 1997). The observed differences
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in competitive effect resulted from the cumulative effects of slightly stronger neighbors. Although the wild-type plants were slightly stronger competitors, the biological effects were small and apparent only under very high densities. The slightly weaker competitive ability of the F1 plants observed at high densities reflects the influence of breeding on the cultivar parent. Ornamental plants, including E. purpurea `White Swan', are bred for garden performance in an environment where moisture, light, and nutrients are not limiting (i.e. reduced competition). E. purpurea is a successful competitor in natural settings, and that competitive ability under field conditions was reflected in this study. Since F1 plants exhibited similar levels of survival and performance to wild-type plants in high density plots, we anticipate F1 plants would also survive under natural conditions. The survival of F1 plants would represent the successful initiation of gene flow between the wild-type E. purpurea and E. purpurea `White Swan', as has been suggested for Sorghum weed X crop hybrids (Arriola and Ellstrand, 1997). This study can serve as a model for risk assessment of other plant introductions, including transgenic plants. The main strength of our study is the use of a competition experiment to evoke differences in plant performance as indicators of fitness. Several points should be considered when using competition experiments for risk assessment. First, our study showed that although a cultivar can possess a trait that many would predict to be maladaptive under natural conditions, i.e. dwarfism (Ellstrand and Hoffman, 1990; National Research Council, 1989; Klinger et al., 1992; Raybould and Gray, 1994; Till-Bottraud et al., 1992), the F1 may not be dramatically less fit than the wild-type parent. At the same time, adaptive traits, i.e. reproductive output of the cultivar may not increase fitness of the F1 with respect to the wild-type. No risk assessment should be considered complete if the hybrid itself has not been studied. Second, researchers should note that significant differences in competitive ability were observed at only one density in this experiment, demonstrating the need to design experiments to encompass a wide range of densities to insure inclusion of the proper, but likely unknown, density at which differences can be observed. Third, although this work examined only intraspecific competition, interspecific competition would provide additional valuable information about the relative fitness of hybrids with other members of the community. Finally, subsequent generations should be examined, as in ongoing work from this experiment studying the potential for introgression between the cultivar and wild-type in a field setting. Acknowledgements We thank N. Anderson, J. Luby, and R. Mack for their review and suggestions that improved early versions of the manuscript. This research was supported in
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part by a Grant-in-Aid of Research from Sigma Xi, The Scientific Research Society and by a graduate fellowship from the University of Minnesota Center for Community Genetics. This is Minnesota Agricultural Experiment Station Publication, 981210012. References Abbott, R.J., 1992. Plant invasions, interspecific hybridization and the evolution of new plant taxa. Trends Ecol. Evol. 7, 401±405. Antonovics, J., Fowler, N.L., 1985. Analysis of frequency and density effects on growth in mixtures of Salvia splendens and Linum grandiflorum using hexagonal fan designs. J. Ecology 73, 219±234. Arnold, M.L., Hodges, S.A., 1995. Are natural hybrids fit or unfit relative to their parents?. Trends Ecol. Evol. 10, 67±71. Arriola, P.E., Ellstrand, N.C., 1997. Fitness of interspecific hybrids in the genus Sorghum: persistence of crop genes in wild populations. Ecological Applications 7, 512±518. Cody, W.J., Boivin, B., 1973. Purple coneflower, Echinacea purpurea, in Ontario. The Canadian Field Naturalist 87, 70. Dyer, A.R., Rice, K.J., 1997. Intraspecific and diffuse competition: the response of Nassella pulchra in a California grassland. Ecological Applications 7, 484±492. Ellstrand, N.C., Hoffman, C.A., 1990. Hybridization as an avenue of escape for engineered genes. BioScience 40, 438±442. Feghahati, S.M.J., Reese, R.N., 1994. Ethylene-, light-, and prechill-enhanced germination of Echinacea angustifolia seeds. J. Am. Soc. Hortic. Sci. 119, 853±858. Goldberg, D.E., 1987. Neighborhood competition in an old-field plant community. Ecology 68, 1211±1223. Goldberg, D.E., Werner, P.A., 1983. Equivalence of competitors in plant communities: a null hypothesis and a field experimental approach. Am. J. Botany 70, 1098±1104. Hallauer, A.R., Miranda, J.B., 1982. Quantitative Genetics in Maize Breeding. Iowa State University Press, Ames, Iowa, USA. Harper, J.L., 1965. Establishment, aggression and cohabitation in weedy species. In: Baker, H.G., Stebbins, G.L. (Eds.), The Genetics of Colonizing Species. Academic Press, New York, USA, pp. 243±265. Klinger, T., Arriola, P.E., Ellstrand, N.C., 1992. Crop-weed hybridization in radish (Raphanus sativus): effects of distance and population size. Am. J. Botany 79, 1431±1435. Klinger, T., Ellstrand, N.C., 1994. Engineered genes in wild populations: fitness of weed-crop hybrids of Raphanus sativus. Ecological Applications 4, 117±120. Langevin, S.A., Clay, K., Grace, J.B., 1990. The incidence and effects of hybridization between cultivated rice and its related weed red rice (Oryza sativa L.). Evolution 44, 1000±1008. Levin, D.A., Schmidt, K.P., 1985. Dynamics of a hybrid zone in Phlox: an experimental demographic investigation. Am. J. Botany 72, 1404±1409. MacArthur, R.H., 1972. Geographical Ecology: Patterns in the Distribution of Species. Harper and Row, Publishers, New York, USA. Mack, R.N., Harper, J.L., 1977. Interference in dune annuals: spatial pattern and neighbourhood effects. J. Ecology 65, 345±363. McGregor, R.L., 1968. The taxonomy of the genus Echinacea (Compositae). The University of Kansas Science Bulletin 48, 113±142.
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Mendenhall, W., Sincich, T., 1989. A Second Course in Business Statistics: Regression Analysis. Dellen Publishing Company, San Francisco, CA, USA. National Research Council, 1989. Field Testing Genetically Modified Organisms: Framework for Decisions. National Academy Press, Washington, District of Columbia. Raybould, A.F., Gray, A.J., 1994. Will hybrids of genetically modified crops invade natural communities?. Trends Eco. Evol. 9, 85±88. Rieseberg, L.H., 1995. The role of hybridization in evolution: old wine in new skins. Am. J. Botany 82, 944±953. Ruesink, J.L., Parker, I.M., Groom, M.J., Kareiva, P.M., 1995. Reducing the risks of nonindigenous species introductions. BioScience 45, 465±477. SAS Institute, Inc., 1994. SAS system for windows, release 6.1. SAS Institute, Inc. Cary, North Carolina, USA. Shaw, R.G., Platenkamp, G.A.J., 1993. Quantitative genetics of response to competitors in Nemophila menziesii: a greenhouse study. Evolution 47, 801±812. Snyder, K.M., Baskin, J.M., Baskin, C.C., 1994. Comparative ecology of the narrow endemic Echinacea tennesseensis and two geographically widespread congeners: relative competitive ability and growth characteristics. Int. J. Plant Sci. 155, 57±65. Till-Bottraud, I., Reboud, X., Brabant, P., Lefranc, M., Rherissi, B., Vedel, F., Darmency, H., 1992. Outcrossing and hybridization in wild and cultivated foxtail millets: consequences for the release of transgenic crops. Theor. Appl. Genet. 83, 940±946. Tilman, D., 1990. Contraints and tradeoffs: toward a predictive theory of competition and succession. Oikos 58, 3±15.
Ecological consequences of hybridization between a wild species (Echinacea purpurea) and related cultivar (E. purpurea `White Swan') Tami M. Van Gaal1, Susan M. Galatowitsch*, Mark S. Strefeler Department of Horticultural Science, University of Minnesota, 305 Alderman Hall, 1970 Folwell Avenue, St. Paul, MN 55108, USA Accepted 20 February 1998
Abstract In the first such study of its kind, we examined gene flow potential between a valued native plant and a popular cultivar. We studied outcomes of controlled hybridization between Echinacea purpurea and a conspecific cultivar, E. purpurea `White Swan', by comparing levels of competitive ability and reproductive potential. Differences in plant performance of wild-types and F10 s were studied at three densities (20.3, 45.7, and 182.9 plants/m2) using a field competition experiment. Wild-types were slightly larger but F1s had higher reproductive output. These differences can be attributed to floricultural breeding of the cultivar parent. Based on the results of this research, if the initial hybridization between the wild-type and cultivar were to occur, it is anticipated that the resulting F1 generation would survive and reproduce, creating the potential for continued gene flow. This methodology has potential for use in risk assessment of plant introductions, including transgenic crops. # 1998 Elsevier Science B.V. All rights reserved Keywords: Echinacea purpurea; Purple cone¯ower; Competition; Fitness; Gene ¯ow; Hybridization; Minnesota; Prairie; Risk assessment
1. Introduction While most introduced plants cannot compete with indigenous flora and can only be maintained in cultivation (Harper, 1965), the exceptional introductions * Corresponding author. Fax: +1 612 624 4941; e-mail: [email protected] 1 Present address: Wagner Greenhouses, 6024 Penn Avenue South, Minneapolis, MN 55419, USA. 0304-4238/98/$19.00 # 1998 Elsevier Science B.V. All rights reserved PII S 0 3 0 4 - 4 2 3 8 ( 9 8 ) 0 0 1 1 9 - 8
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that do become invasive have severe impacts on natural and/or cultural ecosystems (Ruesink et al., 1995). Within the context of cultural landscapes, the evolution of new or more aggressive weedy species via the introduction and/ or interbreeding with exotic, transgenic, or cultivated plants is a common concern (National Research Council, 1989; Ellstrand and Hoffman, 1990; Klinger et al., 1992; Till-Bottraud et al., 1992; Klinger and Ellstrand, 1994; Raybould and Gray, 1994). One perceived risk is that gene flow can affect existing populations (Abbott, 1992; Harper, 1965), resulting in altered relative fitness levels within the wild or crop populations. Although certain traits of cultivated plants are not expected to confer a fitness advantage in the wild (dwarfism, non-shattering seed heads, absence of dormancy), other traits (resistance to diseases, pests, and stresses) are advantageous in the wild and are expected to persist once integrated into weedy populations (Ellstrand and Hoffman, 1990; National Research Council, 1989; Klinger et al., 1992; Raybould and Gray, 1994; Till-Bottraud et al., 1992). Predicting the ecological and genetic consequences of such plant interactions to reduce the likelihood of future harmful invasions (Ruesink et al., 1995) is gaining interest. Very little work has been performed, however, to determine the effects of gene flow on hybrid fitness ± see reviews by Arnold and Hodges (1995) and Rieseberg (1995). Arnold and Hodges (1995) conclude that (1) most hybrids studied do not exhibit the lowest fitness when compared to both parents and (2) hybridization may lead to production of relatively fit genotypes. Three studies illustrate these points. Levin and Schmidt (1985) determined that although the hybrids of Phlox drummondii did not possess a significant increase in fitness over the parents, they did not suffer a fitness disadvantage, and would likely persist. Klinger and Ellstrand (1994) showed that the fitness of Raphanus sativus hybrids was greater than the fitness of the weedy parents, evidenced by the increased fruit and seed production. Hybrids of Oryza sativa possessed intermediate fitness compared with the parental types with respect to vegetative traits (Langevin et al., 1990). The resulting convergence in morphology could result in a dramatic increase in fitness in an agricultural setting for the F1 (Langevin et al., 1990). Together these studies provide evidence that introduced plants, through the action of gene flow, have the potential to impact wild populations. Thus far, no studies have examined the potential for gene flow between valued native plants and commonly used cultivars. Many important horticultural cultivars are derived from wild species, and these cultivars are often planted near populations of wild relatives, or even amongst them. To address the potential impact of gene flow from cultivars to native forbs, we examined the consequences of gene flow between Echinacea purpurea and a conspecific cultivar, E. purpurea `White Swan'. Echinacea purpurea, an herbaceous perennial and obligate outcrosser, is a common indigenous member
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of the tall grass prairie community in the central United States valued as a medicinal herb. By obtaining hybrids through controlled greenhouse crosses between wild-type and cultivar, we studied the consequences of gene flow instead of the process itself. We used a field competition experiment as a tool to compare the relative fitness of the wild-type and the F1 hybrids to investigate the hypothesis that the F1 generation will have different levels of fitness than the wild-type parent. To test the hypothesis, the study determined if the wild-type or F1 plants had a competitive advantage and/or differing reproductive potential. 2. Methods 2.1. Experimental design The design of the competition study was based on that proposed by Goldberg and Werner (1983) and implemented by Goldberg (1987). This design takes advantage of the benefits of a neighborhood design (Mack and Harper, 1977), allowing the response of a target species to be measured over a range of densities of a neighbor species. Competition can be measured by regressing the performance of the target individuals against the `amount' of neighbor species (e.g. density or biomass). This research used the basic experimental design proposed by Goldberg and Werner (1983) with two modifications: (1) all plants were planted in an agricultural field in a predetermined design instead of planting the target plants into natural populations of neighbor plants and (2) the neighbors were arranged in hexagonal annuli ensuring that each plant was equidistant from each immediate neighbor. Our variation of Goldberg and Werner's design was a 223 factorial design comparing target type, neighbor type, and density. A single plant of either the wild-type parent or the F1 served as the target plant in each plot. The neighbor group consisted of 18 plants of either the wild-type or F1 in two hexagonal annuli (Fig. 1). The effects of three density levels of neighbors were examined by relaxing the annuli of neighbor plants around the target individual. In the high density plots, the plants were separated by 0.1 m to create a density of 183 plants/m2. The plants in the medium density treatment were separated by 0.2 m with a resulting density of 46 plants/m2. The low density treatment (20 plants/m2) was achieved by separating the plants by 0.3 m. A total of 12 treatments were applied; each treatment was represented once in each of four blocks across the field. 2.2. Plant material Seeds of wild-type E. purpurea were obtained from Prairie Moon Nursery (Winona, Minnesota). These seeds were progeny of plants grown from seeds
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Fig. 1. Hexagonal plot design used for neighborhood competition experiment. The target plant is located in the center of the plot (T), surrounded by two rings of neighbor s (N).
collected from the restored Curtis Prairie at the Arboretum of the University of Wisconsin, Madison (43.18N, 89.48W; Fig. 2) ca. 1970. E. purpurea was established within the prairie using mostly transplants and some seeds collected from wild stands in Illinois (V. Kline, Univeristy of Wisconsin-Madison, personal communication). Echinacea purpurea had a pre-settlement distribution ranging from Oklahoma to Louisiana and Georgia north to Iowa, Illinois, Michigan, and Ohio (McGregor, 1968; Fig. 2). The species has since naturalized in a larger range that includes Wisconsin and Ontario (Cody and Boivin, 1973; Fig. 2). Seeds of the cultivar parent, E. purpurea 'White Swan', were obtained from G.S. Grimes Seed Company (Concord, Ohio). E. purpurea `White Swan' differs from the wild-type in flower color, flower number, and plant stature. This dwarf cultivar produces more flowers, and the flowers are white as compared to the purplish-pink flowers of the wild-type. After purchase all seeds were stored at 78C, 35% relative humidity. The F1 progeny were obtained by performing crosses (wild typecultivar) in the greenhouse in late spring, 1994. Plants used in crosses were in their first year of growth in the greenhouse. Resulting F1 seeds were harvested in late summer of 1994 and stored at 78C, 35% relative humidity. Seeds used in the competition study were germinated in petri dishes in a growth chamber (258C, 16 h light). Germination followed a 2 week stratification (4±18 October 1994) in 1 mM ethephon at 48C under 24 h light (Feghahati and Reese, 1994). Germinated seeds were moved into the greenhouse (DT278C, NT158C, 14 h photoperiod) in 98-cell plug trays for 3 weeks, then transferred to randomly arranged 10 cm standard pots. On 21 December 1994 plants were
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Fig. 2. Distribution of Echinacea purpurea (shaded area; adapted from McGregor, 1968). The location of the St. Paul, Minn. field plot, site of the neighborhood competition experiment, is indicated by the upright triangle. The inverted triangle indicates the location of Curtis Prairie at the University of Wisconsin-Madison Arboretum, the source population of the wild-type Echinacea purpurea.
transferred to a cooler to begin vernalization. The cooler environment (initially DT228C, NT198C, 12 h photoperiod) was altered weekly by dropping the night temperature 38C and decreasing the photoperiod by 1.5-h per week (ending environment DT228C, NT108C, 8 h photoperiod). On 20 January 1995, the cooler environment was adjusted to 48C without a photoperiod. After nine weeks of vernalization the plants were repotted to 13 cm pots and placed into a heated coldframe. 3. Experimental site The experiment was conducted in an agricultural field on the St. Paul campus of the University of Minnesota in east central Minnesota, USA (458N, 938W).
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The field had been used for forage production (corn) two years before the experiment and was planted with buckwheat (Fagopyrum sagittatum) during the previous year. On 8 May 1995, 56 plot centers were established in a grid at 15 m intervals across the field. A circular area with a diameter of 1 m was cleared around each plot center. The plots were divided into 4 blocks, and treatments were randomly assigned to plots within blocks. On 9 May 1995, plants were transplanted to the field. The plots were irrigated at the time of planting and for two weeks after planting to ensure establishment. Plants that died within the first two weeks of the study were replaced. Fertilizer was not applied during the study. Throughout the study, plots were kept clear of weeds and the buckwheat immediately surrounding the plots was pruned to reduce shading. 3.1. Plant response Three measures of plant performance ± biomass production, reproductive output, and plant structure were examined to determine the competitive effects of target type, neighbor type, and density on target plant performance. Observations were collected from target and first annulus plants only. Initial measurements for each plant, including number of leaves and plant height, were made at the time of planting. Plants were harvested for determination of aboveground biomass on 21 and 22 August 1995 (peak biomass). Plants were harvested to ground level, separated into vegetative and floral components, dried at 658C for 48 h, and weighed. The number and dry weight of inflorescences represented reproductive output. Because plants were harvested at peak biomass (before full seed maturation), mature seed weight could not be used as a reliable measure of fitness. Plant structure was measured by recording number of shoots, number of leaves, and plant height at four-week intervals throughout the study. 3.2. Analysis Through preliminary analyses, we found that a multiplicative model best fit the data, therefore, further analyses were performed on log transformed data (Mendenhall and Sincich, 1989). General main effects of target type, neighbor type, and density on target plant performance (total biomass) were determined using ANOVA (PROC GLM; SAS Institute, 1994). Two components of competitive ability, competitive response and competitive effect, were examined using regression analysis (Goldberg and Werner, 1983; Goldberg, 1987; PROC REG, SAS Institute, 1994). Competitive response was defined as the effect of target type on target plant performance. Competitive effect was defined as the effect of neighbor type on target plant performance. Competitive response and effect were further explained by regressing total target biomass on leaf number,
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plant height, shoot number, inflorescence number and average inflorescence biomass (PROC REG, SAS Institute, 1994). 4. Results Overall survival rates of wild-type and F1 plants were similar (99%). Only three plants (all neighbors) failed to establish; two F1 and one wild-type individuals were replaced in six days after planting. All remaining plants survived the duration of the experiment. The effect of neighbor density on the growth of target plants was strong in all experimental combinations (ANOVA, F36.29, df2, p0.0001). Plants in low density plots were taller, possessed more leaves, and had more numerous stems compared to medium and high density plants (Table 1). For both wild-type and F1 plants, total biomass and floral biomass under high density were 25±31% of the values at low density and Table 1. Under high density compared to low density, plant height was reduced by 16±17% and shoot, leaf, and inflorescence number were reduced by 42±68% (Table 1). With few exceptions, plant growth and morphology of wild-type and F1 plants were similar within densities. Differences between wild-type and F1 plants were not easily distinguished in the field; plants appeared similar in height, stem number, leaf number, and inflorescence number. Only four measures of plant growth and morphology revealed statistically significant differences between wild-type and F1 target plants in five specific comparisons: plant height at low density, number of shoots at medium density, floral biomass at medium density, and average inflorescence biomass at medium and high densities (Table 1). Wild-type plants were taller at low densities and possessed more stems at medium densities (p0.03 and p0.09, respectively). Although differences between wild-type and F1 plants for the remaining comparisons were not statistically different (p>0.1), several trends were apparent. First, measures of plant size (total biomass, plant height, stem number and leaf number) were generally greater in wild-type plants compared to F1 plants. For a given density, combined total biomass of neighbors was always higher when the neighbors were wild-type (Table 2). Although inflorescence number was generally higher in wild-type plants, floral biomass and average inflorescence biomass were greater in F1 plants. With respect to target type and neighbor type, differences in biomass production in target plants could be attributed to differences in plant structure and reproductive output. Plants with greater biomass tended to have larger and more numerous leaves, increased branching, and increased inflorescence number. Although final measures of plant structure affected total biomass, initial height and leaf number of plants were not significantly correlated with total biomass
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Plant type
Density
Total biomass
Floral biomass
Height (cm)
Shoot number
Leaf number
Inflorescence number
Average inflorescence biomass (g)
Wild-type
20.3 45.7 182.0
75.3 (33.8) 39.9 (22.5) 20.7 (22.5)
13.0 (8.3) 6.6 (6.9)* 4.0 (3.5)
79.7 (15.3)** 71.6 (21.1) 66.3 (194.4)
6.4 (2.2) 5.2 (2.1)** 3.7 (1.5)
78.1 (32.2) 49.7 (24.8) 31.9 (16.1)
8.9 (6.3) 4.9 (5.3) 2.8 (2.4)
1.6 (1.0) 1.0 (0.9)*** 1.1 (0.8)**
F1
20.3 45.7 182.9
70.2 (29.5) 40.4 (21.0) 17.3 (11.1)
14.3 (8.3) 9.4 (10.0) 4.4 (3.3)
73.9 (13.3)** 72.4 (15.7) 61.9 (14.0)
6.4 (2.5) 4.5 (1.6)** 3.5 (1.2)
77.9 (31.6) 48.6 (198.8) 30.0 (13.7)
8.1 (4.3) 4.7 (3.1) 2.8 (1.9)
1.8 (0.8) 1.7 (1.3)*** 1.4 (0.7)**
*P<0.1;**P<0.05;***P<0.01.
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Table 1 Mean (SD) effect of density on growth and morphology of wild-type and F1 Echinacca purpurea plants used as target and neighbor plants in neighborhood competition, separated by the density at which they were grown. Asterisks indicate significant differences between wild-type and F1 plants at the same density
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Table 2 The effect of density (mean, SD) on target total biomass and combined neighbor total biomass of wild-type and F1 Echinacea purpurea plants used in neighborhood competition experiment Target type
Neighbor type
Density (plants/m2)
Target total biomass (g)
Combined neighbor total biomass (g)
Mean
sd
Mean
sd
Wild-type
Wild-type
20.3 45.7 182.9
73.9 36.1 14.0
36.2 17.5 5.1
469.6 287.9 142.4
70.0 98.7 31.7
Wild-type
F1
20.3 45.7 182.9
67.6 23.2 11.5
35.2 13.1 3.5
437.9 257.2 113.8
98.3 66.9 40.1
F1
F1
20.3 45.7 182.9
46.4 28.0 17.2
17.8 8.3 4.2
427.7 252.2 102.0
94.8 72.1 48.9
F1
Wild-type
20.3 45.7 182.9
70.7 28.4 8.6
35.2 16.3 5.3
451.6 210.7 120.1
99.0 48.1 32.7
(p0.16 and p0.21 respectively). Of the three measures of plant structure (leaf number, height, and stem number) and two measures of reproductive output (inflorescence number and average inflorescence biomass) used in regression analysis, three contributed to differences in total biomass of target plants (Table 3). Leaf number was the only measure of plant structure to affect target total biomass for all target type and neighbor type combinations (Table 3). Average inflorescence biomass contributed to differences in target total biomass only if the neighbors were of the same type as the target, while inflorescence number affected total biomass of wild-type targets with wild-type neighbors only (Table 3). Total plant biomass was not affected by plant height or stem number under any treatment (Table 3). The only significant relationship between plant structure or reproductive output and target total biomass observed was inflorescence number on wild-type targets with wild-type neighbors. The competitive effect of F1 neighbors on F1 target plants generated the most significant competitive difference observed in the study; F1 target plants performed differently with wild-type or F1 neighbors. The significant difference in slope of the regression of total biomass of F1 targets on combined total biomass of F1 neighbors is explained by a negative relationship between target plant performance and combined neighbor biomass at high densities (Fig. 3(a)±(d)). Within the high density plots, the competitive effect of wild-type neighbors (open circles; ûÿ2.285, p0.038) was stronger than the effect of F1 neighbors (solid circles; ûÿ0.526, p0.0076; t3.80 p<0.005), indicating the wild-type plants
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Target type
Wild-type Wild-type F1 F1
Neighbor type
Wild-type F1 F1 Wild-type
Leaf number
Height
Coefficient
p
3.09 0.90 0.90 1.44
0.001 0.051 0.098 0.118
Coefficient 0.54 ÿ0.76 ÿ0.17 1.40
Shoot number
Inflorescence number
p
Coefficient
p
0.597 0.301 0.696 0.354
ÿ0.44 ÿ0.18 0.33 ÿ0.81
0.445 0.550 0.466 0.132
Coefficient ÿ1.30 0.44 ÿ0.17 ÿ0.21
Average inflorescence biomass p
Coefficient
p
0.002 0.265 0.725 0.774
0.73 ÿ0.33 1.12 ÿ0.56
0.060 0.202 0.002 0.437
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Table 3 The effects of plant structure and reproductive effort on total target biomass of wild-type and F1 Echinacea purpurea used as target plants in a neighborhood competition experiment. Coefficients and p-values were determined using multiple regression on log/log transformed data
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Fig. 3. Comparisons of competitive abilities of wild-type and F1 Echinacea purpurea plants used in a neighborhood competition experiment, including the effect of wild-type neighbors on competitive response of wild-type and F1 target plants (Fig. 3(a)), effect of F1 neighbors on competitive response of wild-type and F1 target plants (Fig. 3(b)), competitive effect of wild-type and F1 neighbors on wild-type target plants (Fig. 3(c)), and competitive effect of wild-type and F1 neighbors on F1 target plants (Fig. 3(d)). Ovals enclose data points from different planting densities represented by symbols: circles for high, diamonds for medium, and squares for low density plots (filledF1, openwild type). The solid line shows the regression function for F1 plants; the dashed line is for wild type plants.
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were slightly stronger competitors. Unlike F1 target plants, wild-type targets performed similarly when surrounded by wild-type or F1 neighbors (t0.669; p>0.1). Differences in competitive response of target plants were not strong (ANOVA: F0.07, df1, p0.7974); target performance (biomass) did not vary within a density for either neighbor. 5. Discussion Hybridization between the wild-type, Echinacea purpurea, and cultivar, E. purpurea `White Swan', can produce an F1 generation that would survive and reproduce, creating the potential for continued gene flow. Survival and reproduction of the F1 generation can be anticipated based on the similar fitness levels of the wild-type parent and F1. Measures of plant structure and competitive ability indicated the wild-type population had a slight fitness advantage but measures of reproductive output indicated the F1 had an advantage, although these differences did not evoke a strong biological response. Overall, the two plant classes were more similar than they were different. Wild-type and F1 plants did, however, exhibit differences in resource allocation, which may affect overall plant performance over time. Gene flow will likely have a long term effect on Echinacea populations for two reasons: (1) the high survival rates of the F1s and (2) the greater reproductive output of the F1s. The high survival rates of the F1, which were similar to the wild-type parent, indicate the hybrid product of gene flow between the cultivar and wild-type populations could be expected to survive once established. These initial results are similar to the findings of Levin and Schmidt (1985), who determined that hybrids of two subspecies of Phlox did not exhibit dramatic decreased or increased fitness relative to either parent population, and are consistent with the views of Arnold and Hodges (1995), who have concluded that most hybrids studied are of similar or intermediate fitness compared to the parents. In every case, the F1 plants had a greater floral biomass and greater average inflorescence biomass compared to the wild-type plants. Based on increased reproductive output, the F1s possess marginally greater fitness than the wild-type plants. Differences in resource allocation are likely a direct result of commercial breeding of the cultivar parent. Floricultural breeders favor a pattern of resource allocation that leads to increased reproductive output (more flowers). The cultivar parent, E. purpurea `White Swan' has been subject to stringent selection through breeding for compactness and high floral output, including high flower number, large flower size, and extended bloom time. In addition to these ornamental traits, E. purpurea `White Swan' was also bred to be seed propagated, with selection for high seed count per inflorescence and strong seed performance (high seed viability and germination rates). The smaller size and greater reproductive output
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of the F1 plants are likely traits inherited from the cultivar parent. These morphological traits are known to be highly heritable in other genera (Hallauer and Miranda, 1982), justifying this conclusion. The results of this research suggest that traits acquired from a cultivar parent can be expressed in hybrid generations. For many plant species, larger plants have greater competitive ability and therefore greater fitness (Goldberg, 1987). In a neighborhood competition experiment, Goldberg (1987) found that larger sized neighbors had a greater competitive effect on target Solidago plants. A positive correlation between plant size and competitive ability was also described within the genus Echinacea (Snyder et al., 1994). With respect to plant structure, a general trend in the data from our experiment indicates that wild-type plants are slightly larger than F1 plants, suggesting the wild-type plants could possess marginally greater fitness than F1 plants. The slightly higher fitness of the wild-type plants, conferred through larger plant size, could result in increased long-term performance of wild-types relative to the F1s in a prairie community. Had the cultivar parent been selected for increased biomass, as are some forage grasses, the F1 would likely exhibit increased competitive ability relative to the wild-type plant. The contrast between the F1 plants, which exhibit greater reproductive output, and the wild-type plants, which exhibit greater vegetative growth, suggests differences in resource allocation. Compared to annuals, herbaceous perennials allocate more resources to vegetative growth, especially below-ground biomass, at the expense of reproductive output. This pattern of resource allocation, especially for nitrogen, has been shown to result in superior competitive ability for resources (Tilman, 1990). In this experiment, the F1 plants appeared to allocate more resources to reproductive output than the wild-type plants. Increased reproductive output would result in greater seed production only if this were not outweighed by decreased long term performance and survival due to decreased acquisition of light, nutrients, and water (Harper, 1965). Decreased performance of target plants with increasing density of neighbor plants is a typical plant response that has been attributed to the effects of competition (Goldberg, 1987; Antonovics and Fowler, 1985; Shaw and Platenkamp, 1993). In our study, the decreased performance of target plants at high densities indicated that competition for resources did occur; therefore, comparisons of competitive ability were possible. Patterns in the variance of total biomass (higher variance around measures at low densities compared to high densities) suggested that competition was an important factor in plant growth and survival, i.e. fitness (Goldberg, 1987). The differences in competitive effect of wild-type and F1 neighbors can be attributed to the cumulative effects of competition. These cumulative effects, called diffuse competition, have been detected among species in natural communities (MacArthur, 1972; Dyer and Rice, 1997). The observed differences
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in competitive effect resulted from the cumulative effects of slightly stronger neighbors. Although the wild-type plants were slightly stronger competitors, the biological effects were small and apparent only under very high densities. The slightly weaker competitive ability of the F1 plants observed at high densities reflects the influence of breeding on the cultivar parent. Ornamental plants, including E. purpurea `White Swan', are bred for garden performance in an environment where moisture, light, and nutrients are not limiting (i.e. reduced competition). E. purpurea is a successful competitor in natural settings, and that competitive ability under field conditions was reflected in this study. Since F1 plants exhibited similar levels of survival and performance to wild-type plants in high density plots, we anticipate F1 plants would also survive under natural conditions. The survival of F1 plants would represent the successful initiation of gene flow between the wild-type E. purpurea and E. purpurea `White Swan', as has been suggested for Sorghum weed X crop hybrids (Arriola and Ellstrand, 1997). This study can serve as a model for risk assessment of other plant introductions, including transgenic plants. The main strength of our study is the use of a competition experiment to evoke differences in plant performance as indicators of fitness. Several points should be considered when using competition experiments for risk assessment. First, our study showed that although a cultivar can possess a trait that many would predict to be maladaptive under natural conditions, i.e. dwarfism (Ellstrand and Hoffman, 1990; National Research Council, 1989; Klinger et al., 1992; Raybould and Gray, 1994; Till-Bottraud et al., 1992), the F1 may not be dramatically less fit than the wild-type parent. At the same time, adaptive traits, i.e. reproductive output of the cultivar may not increase fitness of the F1 with respect to the wild-type. No risk assessment should be considered complete if the hybrid itself has not been studied. Second, researchers should note that significant differences in competitive ability were observed at only one density in this experiment, demonstrating the need to design experiments to encompass a wide range of densities to insure inclusion of the proper, but likely unknown, density at which differences can be observed. Third, although this work examined only intraspecific competition, interspecific competition would provide additional valuable information about the relative fitness of hybrids with other members of the community. Finally, subsequent generations should be examined, as in ongoing work from this experiment studying the potential for introgression between the cultivar and wild-type in a field setting. Acknowledgements We thank N. Anderson, J. Luby, and R. Mack for their review and suggestions that improved early versions of the manuscript. This research was supported in
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