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Widrlechner, M.P., and K.A. McKeown. 2002. Assembling and characterizing a comprehensive Echinacea germplasm collection. p. 506508. In: J. Janick and A. Whipkey (eds.), Trends in new crops and new uses. ASHS Press, Alexandria, VA.
Mark P. Widrlechner and Kathleen A. McKeown
During the 1990s, the popularity of the genus Echinacea Moench (Asteraceae) as a dietary supplement in the United States increased markedly, as the general public learned of its possible efficacy in fighting colds and other illnesses (Bauer and Wagner 1991; Li 1998). Plant and medical scientists responded to this phenomenon by increasing their efforts to understand the biology, cultivation, and pharmacology of these plants. Unfortunately, very few well-documented living collections of Echinacea were readily available to support that research. Well-documented germplasm collections could also be used to broaden the genetic base of ornamental Echinacea cultivars, which are widely cultivated as attractive landscape perennials.
In response, during the mid-1990s, the North Central Regional Plant Introduction Station (NCRPIS) began assembling germplasm collections from wild Echinacea populations from throughout its native range in the United States and Canada. These efforts were given a great boost in 1997, when the United States Department of Agriculture, Agricultural Research Service sponsored the location and collection of seed samples representing the diversity of all known Echinacea taxa (McKeown 1999a). By the end of the decade, the NCRPIS had acquired samples of more than 130 different wild populations, including all recognized taxa (Table 1, Fig. 1). In 1999, the NCRPIS began a project to produce sufficient quantities of seeds from these populations, both to conserve the populations (which are often threatened in nature by commercial exploitation) and to make seeds freely available to the research community. The regeneration project cultivates individual populations within screened cages (Fig. 2) with honeybees as pollinators to produce control-pollinated seeds (Widrlechner et al. 1997).
Fig. 1. Map of the natural range of Echinacea in the United States and collection sites (McKeown 1999b).
Table 1. Echinacea germplasm conserved at the North Central Regional Plant Introduction Station.
|E. angustifolia DC.||17|
|E. angustifolia var. angustifolia||22|
|E. angustifolia var. strigosa McGregor||2|
|E. atrorubens Nutt.||5|
|E. laevigata (F.E. Boynton & Beadle) S.F. Blake||10|
|E. pallida (Nutt.) Nutt.||39|
|E. paradoxa (Norton) Britton var. neglecta McGregor||4|
|E. paradoxa var. paradoxa||5|
|E. purpurea (L.) Moench||13|
|E. sanguinea Nutt.||9|
|E. simulata McGregor||8|
|E. tennesseensis (Beadle) Small||4|
Fig. 2. Control-pollinated seed multiplication of E. pallida in a screen cage (image by A.P. Ovrom).
In 2000, sufficient numbers of control-pollinated seeds were produced from more than 80 populations to allow their distribution, with additional harvests made during the late summer and autumn of 2001. During the course of the regeneration project, notes and measurements were collected on a wide range of morphological descriptors, and taxonomic identities were verified. These characterization data are being prepared for inclusion in the Germplasm Resources Information Network (GRIN) database, which is accessible on the Internet at <www.ars-grin.gov/npgs>. In late September, 2001, at the end of three years in the field and a final seed harvest, roots were dug, dried, and shipped for chemical analysis of bioactive compounds (caffeic acid derivatives and alkamides) by James Simons laboratory at Rutgers University.
Beyond conducting morphological and biochemical characterization, there is a need to understand the genetic basis of such variation. Which genes are responsible for the unusual biochemistry of this genus? How are they shared among the taxa? Can they be transferred to existing crops, such as sunflower, to confer natural insect or fungal resistance? How can Echinacea best be domesticated, both as an ornamental and as a dietary supplement to replace wild harvesting?
To begin to answer these questions a preliminary evaluation of genetic diversity within both the genus and selected accessions is underway. Phylogeographic analysis, a novel approach to plant germplasm evaluation of genetic diversity, will be the primary methodology utilized. The power of phylogeographic analysis lies in consideration of the spatial distribution of genealogical lineages inferred from nucleotide sequence variation, which is an absolute measure of genetic diversity. Echinacea taxa are morphologically and chemically similar and exhibit broad phenotypic plasticity, particularly under varying photoperiod regimens (K. McKeown, unpubl. obs.) A sequence-based analysis of diversity among Echinacea taxa is an essential counterpart to phenotypic-based evaluations.
Analyses of nucleotide sequence variation are conducted on the same region in a given locus for all taxa considered. In particular, a gene region that is selectively neutral in molecular evolutionary terms is required. In this context, selectively neutral indicates a region in the genome where mutations (leading to nucleotide substitutions, insertions, or deletions) have no effect on the fitness of the organism. For example, introns are generally neutral, and a number of statistical tests on the sequence data are utilized to demonstrate this neutrality (Li 1997). Nonrandom geographic patterns of genetic variation would suggest significant genetic divergence. The potential utility of phylogeography to plant germplasm conservation is apparent when the pattern of sequence variation is considered in association with the geographic spatial distribution of the taxa in question (Avise 2000).
Glyceraldehyde 3-phosphate dehydrogenase (G3PDH) is an enzyme in the glycolytic pathway, encoded by a nuclear gene in plants. Sequence variation at the G3pdh locus has recently been used to unravel the geographic origin of domesticated cassava (Olsen and Schaal 1999). A sample of 40 Echinacea populations was chosen for analysis by selecting populations collected nearest to the intersection of whole-number longitudes and latitudes between 78°99° West and 30°40° North. Preliminary nucleotide sequence data of the noncoding region of the G3pdh locus across these populations reveal useful variation, and geographic mapping of haplotypes is underway. This will be the first phylogeographic analysis of a medicinal plant based on the nuclear genome, the genome which characteristically has the greatest degree of nucleotide polymorphism in plants.
An Amplified Fragment Length Polymorphism (AFLP) analysis (Vos et al. 1995) of Echinacea populations will also be conducted. This will generate a large set of markers, the frequency and identity of which will be used for calculating genetic distances, quantifying overall levels of diversity and fine-scale genetic mapping. These genome-wide markers will also provide an important comparison for findings based on the analysis of G3pdh.