Trisetum Spicatum Complex (Poaceae)
.Biosystematic Studies in the North American Trisetum spicatum Complex (Poaceae)
by
John L. Randall Jr.
thesis submitted to the Faculty of the
Virginia Polytechnic Institute and State University
in partial fulfillment of the requirements for the degree of
MASTER OF SCIENCE
in
botany
APPROVED:
Khidir W. Hilu
Duncan M. Por~er W. dattterJohnson
September, 1984 Blacksburg, Virginia
ACKNOWLEDGEMENTS
My sincerest gratitude extends out to the many people who have influenced and encouraged my interests in biology. In particular, wish to thank Dr. Khidir Hilu who has been the guiding light throughout my graduate carrer. His moral, logistic, and financial support throughout this project has been unending. He was never too busy to lend an ear when I needed help with a problem.
I owe my first academic glimpse into biology to Dr. Herbert
Hechenbleikner whose teachings on natural history will always be with me. Dr. Larry Mellichamp deepened my interests in plant science with his knowledge of plant and field biology in addition to encouraging me into graduate studies.
I also wish to thank Dr. Ed Clebsch, Dr. Dwight Billings, and Ms.
Shirley Billings for providing invaluable information and seed material of
Trisetum splcatum. Without these collections my project would have been incomplete. Thanks is also, extended to the USDA and Native
Plants, Inc. for seed material. For the loan of herbarium specimens, wish to thank the curators of DUKE, US, and TENN. Funding for this project was granted through Sigma Xi, the Virginia Academy of Science, and the VPl&SU Biology Department.
The assistant curator and students in the Virginia Tech herbarium always provided advice, encouragement, and most importantly
i i friendship. This friendship added reality to graduate student life.
These special people include Tom Wieboldt, Conley McMullen, Garrie
Rouse, Alan Koller, and Hazel Beeler. Special thanks goes to Rytas
Vilgalys for his instrumental advice in my electrophoretic studies.
I also wish to thank Dr. Duncan Porter and Dr. Carter Johnson for serving as committee members. Their support, encouragement, and expertise in taxonomy and ecology, respectively, made my investigation more complete.
My parents, John and Helen Randall, deserve greatest thanks for their desire for me to choose my own direction in life and their faith in my decisions. To them this project is dedicated.
Lastly, much more than thanks goes to my wife and partner in life,
Debbie Tolman. She has assisted in every aspect of this project. Her energy, dynamism, and love never cease to amaze me.
iii TABLE OF CONTENTS
ACKNOWLEDGEMENTS ii
Chapter page
INTRODUCTION 1
TAXONOMIC HISTORY I CONCEPTS OF THE SPECIES 3
DISTRIBUTION ...... 11
Worldwide Distribution . . . . .• 11 North American Distribution .. 13 Habitat· ...... 18
MATERIALS AND METHODS ... 20
·Collection of specimens 20 Cytology ...... 21 Numerical analysis . . . . 22 Stomate size ...... 25 Enzyme electrophoresis .. 26
RESULTS ...... 29
Numerical I Morphological Analyses 32 Electrophoresis · 61
DISCUSSION 68
LITERATURE CITED 73
VITAE ...... 78
iv LIST OF FIGURES
Figure page
1 . Worldwide distribution of the species complex. 12
2. North American distribution of the species complex. 14
3. Geographic distribution of the species complex in North America (spicatum , molle ----- , and pilosiglume ( ..... )·...... -.-...... 16
4. Populations surveyed for electrophoretic study. 28
5. Range in mean stomate length between cytotypes. 30
6. Expansion of cluster analysis demonstrating geographic distribution...... 31
7. Cluster analysis phenogram. 33
8. Inconsistency of inflorescence interruption within the same individual...... 47
9. Correlation between characters used in the final analysis. See Table 5 for description of character acronyms. 48
10. Plot of principal coordinate. 50
11. Graph of morphology versus subgroup. 54
12. Graph of morphology versus geographic distribution. 56
13. Inconsistency of culm height within the same population. 59
14. Variation in inflorescence length, width, and shape within the same population. 60
15. Enzyme system zymograms. 62
16. Esterase: interpopulation similarities and differences. 66
v LIST OF TABLES
Table page
1 . Synonymy of the North American species complex from Nash (1939), and Dore & McNeil (1980). . ... 5
2. Taxonomic treatments of the North American species complex. 6
3. Synopsis of the three generally recognized subgroups .. 17
4. Morphological characters used in the numerical analyses. 23
5. Populations surveyed for electrophoretic study. Population numbers can be correlated with the map in Fig. 4. 27
6. Summary of stepwise selection of variables. 52
7. Summary of basic statistics. 53
vi INTRODUCTION
The species complex Trisetum spicatum (L.) Richt. (Poaceae) is circumpolar as well as bipolar (Du Rietz 1940; Hulten 1959). Its populations are essentially discontinuous and often times widely disjunct. Morphological diversity is high, leading to its separation into several races, varieties, subspecies, and even species (Fernald 1916;
Hulten 1959; Bacher 1959; Morrison 1959; Love and Love 1962). Both tetraploid and hexaploid cytotypes occur. The morphological diversity coupled with wide distribution and polyploidy has resulted in a very confusing taxonomy. Previous taxonomic treatments (Fernald 1916;
1950; Louis-Marie 1928; Hulten 1927, 1937, 1959; Bacher 1959, Love and
Love 1962) were inconsistent and largely inconclusive. These treatments were often based on the examination of few specimens with emphasis on highly variable characters of the inflorescence and stem pubescence. In order to assess the trends of variation in Trisetum spicatum, a large number of specimens representing the the entire
North American range of distribution has to be examined, and the consistency of the characters should be evaluated.
The objective of the present study is to analyze the morphological and genetic diversity of the species complex in North America, elucidate the trends of variation, and compare these results to previous studies.
To approach these goals the techniques of numerical analysis and electrophoresis were utilized. 2
Numerical techniques were used to analyze the morphological data.
Since grasses possess such reduced reproductive structures, analysis of many different variables is useful and has proven successful in elucidating trends of variation in this group (Hilu and Wright 1982).
Electrophoretic techniques are a relatively recent innovation in plant systematics. Although developed in the sixties ( Fai rbrothers
1962), these techniques did not become popular until the seventies
(Avise 1974; Hamrick and Allard 1975; Gottleib 1975, 1977; Jensen et al. 1979), and are presently a common approach (Soltis 1981; Doebly et al. 1984; Crawford and Smith 1984). One of the most useful and successful electrophoretic techniques is the staining of specific enzymes. These isozymes represent different molecular forms of the same enzyme or locus. Both quantitative and qualitative information can be resolved from the particular number and mobility (position) of the stained bands. These allelic frequency data give estimates of the genetic variation within and between populations. Estimates of this type are especially useful at the intrageneric and infra specific levels.
The use of these biosystematic techniques in the light of traditional taxonomy is expected to help clarify the taxonomic difficulties in the
North American Trisetum spicatum group and elucidate the degree of genotypic and phenotypic variability within and between populations of this species complex. TAXONOMIC HISTORY I CONCEPTS OF THE SPECIES
The taxon recognized today as Trisetum spicatum was first known as
"Gramen avenaceum panicufatum alpinum humile". It was originally described by Scheuchzer (1719, p. 221), whose translated description follows (after Clebsch 1960):
The plant (17-30 cm. high) is described as having glabrous, striate leaves; ligule (about 1 mm. long) obtuse; culm densely tomentose; panicle spikelike (about 1.5 cm. long, 0.07-1 cm. wide), purplish, shining; spikelets (about 4 mm. long) 2-flowered; glumes unequal (I shorter and narrower, 11 about 2 mm. wide and 4 mm. long), glabrous; lemma (4 mm. long) awned dorsally on the upper one forth; awn (about 4 mm. long) reflexed; palea equal or subequal to the lemma; rachilla densely villous; anthers 0.6-1 mm. long.
Since Scheuchzer, T. spicatum has been assigned by various taxonomists to the genera Aira, Avena, Deyeuxia, Koleria, Melica,
Rupestrina, Trisetaria, and Trisetarium.
The long history of nomenclatural confusion within the species began with Linnaeus (1753). He gave "Gramen avenaceum paniculatum alpinum humile" the binomial Aira spicata in his first edition of Species
Plantarum. Linnaeus, however, named two plants Aira spicata, on pages 63 and 64. This mistake was corrected in the errata, with A. spicata (p. 64, habitat in Lapland) maintained and A. spicata (p. 63, habitat in India) transferred to A. indica. Richter (1890) transferred
Aira spicata to the genus Trisetum. Linnaeus' nomenclatu ral snafu resulted in published arguements between Mackenzie (1929) and
Hitchcock (1929) regarding the correct specific epithet for Trisetum
3 4
spicatum. The problem was eventually rectified by Farwell (1930), who settled the matter by pointing out that both Aira spicata I and Aira spicata 11 are valid according to Articles 48 and 53 of the International
Rules of Botanical Nomenclature. These in essence state that when transfers are made by later botanists to different genera, the specific epithets must be maintained unless the genus to which it is transferred already has a species by that name. Trisetum spicatum (L.) Richt. is thus the correct name for the species.
The confusion associated with the infraspecific classification is reflected in the 56 synonyms for the species in North America alone
(Table 1).
This confusion 1s also exhibited by the inconsistent classification of different authors, and even the same author in separate publications
(Table 2). There are, nevertheless, three infraspecific taxa that have been most often recognized by taxonomists in the North
American Trisetum spicatum complex. These are T. spicatum var. maidenii (Gand.) Fern. (T. spicatum var. spicatum of Hulten), T. spicatum var. mo/le (Ku nth) Beal, and T. spicatum var. pilosiglume
Fern.
Fernald (1916) studied the components of the species in eastern
North America. He indicated that "true" European T. spicatum
(referring to plants similar to the type specimen from Lapland) did not occur in North America except for solljle arctic and northwestern populations. Specimens from Greenland and other, arctic American and TABLE 1. Synonymy of the North American Trisetum spicatum complex, from Nash (1939), Hulten (1959), and Dore & McNei I (1980). Aira spicata L. Trisetum congdoni i Scribn. & Merr. Aira subspicata L. Trisetum americanum Gand. Avena airoides Koller Tri set um !fil!.i!l§_ Ryd b. Avena iiioTTTsMichx. Avena spicata Fedtsch. Avena subs~atB_ Gia i rv. Trisetum spicatum var. pi losiglume Fern. Trisetum subspicatum Beauv. Trisetum spicatum var. laxius Holmberg Aven~ elongata H.B.K. Trisetum disjunctum Louis-Marie Avena tolucensis H.B.K. Trisetum humi le Louis-Marie MeTTca triflora Bigel. Trisetum sprcatum var. alaskanum Malte. Trisetaria airoide.§ Baumg. Trisetum spicatum var. brittoni i Louis-Marie Trisetum airoides Beauv. Trisetum spicatum congdoni i Hitchc. TrTSetarium elongatum Pair. Trisetum spicatum var. nivosum Louis-Marie Trisetarium tolucense Pair. Trisetum spicatum var. vi I losissimum Louis-Marie AvQna squa rrosa Schrank Trisetum albidum Sodiro Trisetum mol le Kunth Trisetum gracile Sodiro Jrisetuni elongatum Kunth Trisetum spicatum subsp. phleoides (D'Urv.) Hult. Tri setum-tol ucen-se- Ku nth i::r i set um sp Lea tum subsp. sp i ca tum Hu It.- Koeleria sutiSPTCata Reichenb. T r i setum spicatum var. laxius Lange Trisetum labradoricum Steud. Trisetum spicatum subsp. congdoni i Hult. frisetum groenTandTcum steud. Trisetum spicatum var. spicatiforme Hult. Trisetum subs~_1:!tum var. molle A. Gray' Trisetum spicatum subsp. molle Hult. Rupestrina pubescens Prov. Trisetum spicatum subsp. pi losiglume Hult. Trisetum subspicatum var. laxius Lange Trisetum _§_pjcatum subsp. ma jus Hult. Trisetum subspicatum var. vi I loissimum Lange Trisetum spicatum subsp. alaskanum Hult. Oeyeuxia graci I is Fourn. Trisetum triflorum subsp. triflorum (Bigelow) A. & D. Love Trisetum nivosum Fourn. Trisetum triflorum subsp. mol le (Hult.) A. & D. Love Trisetuiii interruptum Fourn. Trisetum spicatum var. mol le Beal Trisetum brittoni i Nash Trisetum alaskanum Nash TABLE 2. Taxonomic treatments of the North American l. spicatum complex.
Ferna Id ( 1916, 1950) Louis-Marie ( 1928) Hu I ten ( 1927) Hulten (1937) Hulten (1959) l. spicatum l. spicatum l. spicatum l. spicatum l. spicatum var. Mai den i. i var. Mai den i i var. mo I le var. mo 11 e ssp. spicatum
var. mol le var. mo I le var. Mai den i i var. laxius
var. pi losiglume var. alaskanum va r. v i I I o s i s s i um
var. alaskanum var. phleoides ssp. congdon i i
var. spicatiforme ssp. mo I le
ssp. pi losiglume
ssp. ma,ius
ssp. alaskanum
Bacher (1959) Morrison (1959) Love & Love (1962) Anderson ( 1952) l. spicatum _§_p. str. l. sp icatum l. spicatum l· spicatum l. mol·le ssp. spicatum ssp. spicatum ( po I ymo rph i c)
var. mo I I e l· mol le l· triflorum var. pi I os i g I ume var. mo I le ssp. triflorum
var. pi losiglume (formerly pi losigl_ume) 7
Eurasian populations were recognized as T. spicatum var. maidenii, which tends to be ta.lier than the other North American subtaxa.
Fernald cites the temperate areas of North America as containing T. spicatum var. mo/le. He recognized this variety as an extreme variation within the spec.ies with long, slender culms and silvery-green and much
interrupted inflorescences. Fernald goes so far as to suggest specific
status for it (regardless of its obvious affinities with the other varieties). Fernald (1916) also described a new taxon, T. spicatum var. pi/osiglume, as "unique in the maze of plants which go
undistinguished as T. spicatum" (p. 196). He further stated that its
spikelets were "somewhat intermediate between the more arctic-alpine T. spicatum var. maidenii and T. spicatum var. mo/le." Although he
considered T. spicatum var. mo/le to be quite .distinct from European T.
spicaturri, he indicated that the series "seems to merge through var.
maidenii and again through var. pilosiglume" (p. 197).
Louis-Marie (1928), a student of Fernald, named fourteen varieties
of T. spicatum in the Americas. These include five in the Andes, one
in Mexico, two in Greenland, and six in the United States and Canada.
Hulten (1927, 1937, 1959, 1968) has made the most extensive
investigation of this species complex. In his 1927 Flora of Kamtchatka
and the adjacent islands, he noted that many specimens were similar to
those familiar to him in Scandinavia (T. spicatum s. str.). He found
many specimens to contain various character differences, especially with
respect to pubescence. Hulten stated, however, that these characters 8
"do not seem to possess any considerable systematic value." In a later treatment (Hulten 1937), he found that most specimens of T. spicatum in the Aleutian Islands do differ from Scandinavian plants in size and glume characteristics. Hulten placed these taxa in T. spicatum var. maidenii. He also determined that the taxon T. a/askanum Nash could not be successfully distinguished from T. spicatum var. maidenii. He noted that all intergrading forms seem to occur between these two taxa.
Hulten reported that several different forms could be distinguished, but it would be impracticable to draw lines of demarcation between these.
Therefore, he concluded that there is "little use to try to group the materials as varieties."
In 1959 H ulten revised the species complex worldwide. Twenty,-one subtaxa were described through new combinations based on examination of approximately one.:.hundred herbarium specimens. Nine of these subtaxa (Table 2) represented the North American component of the species. They include T. spicatum subsp. alaskanum (Nash) Hult., T. spicatum subsp. pi/osiglume (Fern.) Hult., T. spicatum subsp. mo/le
(Michx.) Hult., T. spicatum subsp. majus (Vasey) Hult., and T. spicatum subsp. congdonii (Scrib. & Merr.) Hult. Hulten recognized these taxa as subspecies but refers to them as "races" in the text.
This entire revision Is extremely difficult to understand. In his conclusions he stated that, "in the Himalayas,· in Kamtchatka, in southern Alaska, and in the Rocky Mountains, where the range of subsp. spicatum overlaps that of other races, the differentiation 9
between the different taxa becomes somewhat arbitrary." In addition,
his final statement was "that the construction of the spikes is very
alike in the different races and that the descriptions given in diagnoses
usually say nothing about the characteristic differences'' (p. 227). If
Hulten found characteristic differences in the construction of the
spikes, they should be critical in delineation of his infraspecific
categories.
Trisetum spicatum was also treated in the Flora of Alaska and
adjacent parts of Canada (Anderson 1959). Anderson, well familiar with
T. spicatum and arctic and alpine ecology, states the following:
This species has been segregated into a great many intricately critical subspecies (e.g. Hulten, E. 1959, Svensk. Bot. Tidskr. 53:203-28), of which five subspecies are reported from Alaska and Yukon. Mostly the ·diagnostic features are involved with variation in pubescence, relative length of spikelet parts, and differences in spike characteristics. None of these, taken either singly or in combination, appears to be definitive. Therefore it se.ems best to recognize our materials as portions of a polymorphic species.
The above authors concentrated primarily on morphology. Others
believed chromosome number to be an important criterion for taxonomic
subdivisions.
Becher (1959) concentrated on the northern European and
Greenland components of the species. Becher based his revision of the
species on cytology, floral and vegetative morphology, ecology, and
geographic distribution. The name Trisetum spicatum (L.) Richt. was
maintained for the more arctic tetraploids, whereas the low~arctic and · 10
temperate hexaploids were elevated to Trisetum mo/le (Michx.) Ku nth with the former T. spicatum var. pilosiglume included as its subspecies.
Morrison (1959) reviewed the differences between the North
American tetraploid and hexaploid Trisetum spicatum. He recognized the heterogeneous nature of the taxon and agreed with Bocher's
(origonally Ku nth 's) suggestion to create a new species, T. mo/le, for the hexaploids. Morrison concluded that the taxonomy of the species will remain unsatisfactory until it is examined with biometric techniques and cultural experiments.
Love and Love (1962), in the most recent revision of the the North
American group,· maintained the tetraploids in Trisetum spicatum but relegated the hexaploids T. spicatum var, pilosiglume and T. spicatum var. mo/le to T. triflorum Bigelow subsp. triflorum Love & Love and
T. triflorum subsp. mo/le (Hulten) Love & Love, respectively. They cite chromosome number, panicle shape and interruption, glume length and color, and geographic distribution as diagnostic characteristics.
However, they list a number of exceptions to these rules of segregation. DISTRIBUTION
WORLDWIDE DISTRIBUTION
Trisetum spicatum is subcosmopolitan (no plant species is truly
cosmopolitan) its distribution being basically circumboreal and bipolar
(Fig. 1). ·The species occurs also in Mexico, Haiti, tropica·I South
America, New Guinea, Borneo, Australia, and Tasmania (Hulte"n 1959).
Few plants are as widespread as T. spicatum. DuRietz (1940) indicated
a number of taxa with bipolar distributions, with none as ubiquitous as
T. spicatum. He included T. spicatum as a species with "one large
circumpolar boreal population and two widely separated austral ones
with connecting tropical mountain populations". (page 219). Only the
moss Sphagnum mage/lanicum and the lichens Thamnolia vermicu/aris and
Cetraria islandica show similar distributions. These three taxa differ in
their distribution from T. spicatum only in their absence in Borneo
(Du Rietz 1940) .
Subcosmopolitan distributions may be divided into 1)
Eurasian-North American, 2) South American-Australian, and 3) North
American-South American (Thorne 1972). The Eurasian-North American
distribution describes the bulk of the taxa whose ranges are
circumboreal, arctic, boreal, and temperate, including wide
intercontinental disjuncts. In lower temperate latitudes, T. spicatum is
found only in the high elevations of the Rocky Mountains, Cascades,
Sierra Nevada, and Appalachians in North America and the Pyrenees,
11 N
IJ
Figure 1: Worldwide distribution of the Trisetum spicatum complex. 13
Alps, Caucasus and Himalayas in Eurasia. Many populations of the
South American-Australian distribution are disjunct, with occurrences in the Andes, Tierra del Fuego, Falkland Islands, Borneo, Australia, New
Zealand, and Tasmania. The North American-South American (bipolar) distribution links the above two types.
This amazing distribution has been denoted by Hulte'n (1959) as the world's most extensive in terms of continuous connecting localities.
Hulten speculates on the factors responsible for this distribution as follows (p. 228):
It seems evident that T. spicatum in an earlier period with a colder and wetter climate, probably not later than during the maximal glaciation, has been able to spread from the main area in the Northern hemisphere along the mountain chains that border the Pacific, on one hand over Formosa, the Himalayas and Borneo to Australia and New Zealand and on the other hand over Mexico and the Andes to Patagonia and Tierra del Fuego and to bridge the lowland gaps occurring there. The long-scaled Pacific subsp. alaskanum type has wandered both these ways, while the broad-scaled subsp. spicatum type which is mainly circumpolar has wandered to the Southern hemisphere only on the American side. When the climate in the tropical parts of the ·range got warmer and drier in the intergalcials T. spicatum was isolated on the high mountains in its former pathway.
NORTH AMERICAN DISTRIBUTION
Specimens from North America seen by this author have the following distribution (Fig. 2). These populations essentially follow the high elevations of the temperate United States. In Canada and Alaska, exposed rock or tundra provide suitable habitats. Clebsch (1960) describes the North American distribution as arctic-boreal montane
("montane" used in the sense of mountains, not life zones). 14
' I
• • •• •• •
Figure 2: North American distribution of the species complex. Dots represent populations surveyed through herbarium specimens. Squares indicate populations seen by the author. 15
The three most often recognized subtaxa of T. spicatum (T. spicatum var. spicatum, T. spicatum var. mo/le, T. spicatum var. pilosig/ume) in North America have also been separated on the basis of geographic distribution (Fig. 3), in addition to morphology and ploidy level (Table 3).
The spicatum subgroup has been regarded as more or less the model for the entire species complex. Its distribution is considered as circiJmarctic, circumboreal, and bipolar (Hulten 1959; Bocher 1959;
Morrison 1959; Love and Love 1962). The North American range of the spicatum subgrovp includes Alaska, Canada, and the high elevations of the Rockies, Cascades, and Sierra Nevada (Hulte'n 1959), in addition to its occu rren.ce around the north shore of Lake Superior and the Mingan
Islands, Quebec .(Dore and. McNeil 1980). This subgroup has also been characterized by its tetraploid chromosome number (Becher 1959; Hultfn
1959; Morrison 1959; Love and Love 1962).
The mo/le subgroup has been reported to reside in the more temperate areas of Canada and in the transition regions between boreal and temperate environments, including New England (Fernald 1916,
1950), as well as Pennsylvania (Pretz 1919), the high mountains of
North Carolina (Hitchcock and Chase 1950), and Haiti, (Hulten 1959).
Hulten also reports the mo/le subgroup from northeastern Asia.
The pi/osiglume subgroup is only reported from northeastern North
America, including Greenland (Fernald 1916, 1950; Hulten 1959;
Morrison 1959; Becher 1959; Dore and McNeil 1980). These last two 16
-...... ' ' '\ ' '\
A ...... , ...... >.~~ \· .. ·-·~·,: S T A T E ,_,~' NATIONAL SCl-IOOL OF AERONAUTICS, INC. NORTH AMERICA COPYRIGHT G.r.1..1..UP MAP a STATIONERY COMPANY 10,00 SCALl Of MILES Figure 3: Geographic distribution of the species complex in North America (spicatum , molle -----, and pilosiglume ( ..... ) . 17 TABLE 3 Synopsis of the three generally recognized subgroups. After Dore & McNeil (1980). spicatum subgroup Plants 10-30 cm high; panicle short, compact, not interrupted, 3-4 times as long as thick, rounded at the top; glumes extending to the top ofthe florets; lemmas brightly colored with purple and brown; arctic species; tetraploid. · mo/le subgroup Plants 20-50 cm high; pan icle slender, looser, and somewhat interrupted especially toward thebase, about 10 times as long as thick, tapering to tip; glumes extending almost to top of the florets; lemmas pale green, drying to a light straw color; boreal species; hexaploid. pi/osiglume subgroup Same description as mo/le with the addition of pi lose glumes. 18 subgroups represent the hexaploid component of the species in North America. In summary, the spicatum subgroup is tetraploid and arctic-boreal whereas the mo/le and pilosiglume subgroups are temperate-boreal and hexaploid .. It should be noted that in view of the many exceptions to be found in distribution and morphological variation, these statements are generalizations. Exceptions will be discussed later in the text. HABITAT Trisetum spicatum occupies open, relatively exposed habitats that include spits, tundra, heathlands, rock outcrops, bars, snow flushes, river banks, roadsides, landing strips, ridge tops, moraines, open woods (Anderson 1959), moist subalpine and alpine meadows (Fernald 1916; Cronquist et al. 1977), boreal forest and in crevices of rocks on exposed shores of lakes (Dore and McNeil 1980), gravelly habitats (Baldwin 1958), and boulder fields (Massey et al. 1983). The exposure generally depends on the latitude and elevation. For example, in arctic and boreal habitats a southern exposure or level plain is characteristic for the species, in contrast to its restriction to east- and north-facing exposures in most temperate environments. Trisetum spicatum also grows on a variety of bedrock types, including granite, shale, limestone, sandstone, chert, precambrian shield, peaty loam, and sand (Massey et al. 1983; personal observation). These varied soil and rock types imply that the species is relatively substrate neutral. 19 In terms of its temporal (successional) position, T. spicatum has been characterized as both pioneer or early successional to late successional or climax (Massey et al. 1983). Massey et al., however, fail to qualify what is meant by late successional. The "late successional" habitats they describe are those associated with alpine and arctic tundra. However, it is generally accepted that succession does not occur in extreme environments such as tundra. My investigation substantiates the above habitat and substrate descriptions, but makes no attempt to define the successional status of T. spicatum, although the taxon appears to be a pioneer species. In eastern North American, the species is primarily found on rock ledges and in crevices, often associated with waterfalls. The association with waterfalls is due perhaps to the requirement for sunny, exposed sites. Association with water may also effectively buffer the extremes in high temperatures for this boreal taxon. In addition, streams and rivers may facilitate seed dispersal to other suitable sites downstream. Nearly all sites visited by the author were established with no evidence of recent disturbance. However, two populations were found in "disturbed" sites. These included an abandoned gravel parking area near a river in Magpie, Quebec, and on a sandy road cut in Manicougan, Quebec. In summary, T. spicatum is a plant of open habitats and various soils of many rock types. Clebsch (1960) classified the species as a "tundra weed which can compete successfully in closed herbaceous vegetations or can invade bare areas of harsh environment." MATERIALS AND METHODS COLLECTION OF SPECIMENS Seed collections were made available from the USDA, Native Plants, Inc., and from Prof. Dwight Billings of Duke University. These last collections were especially important since they were from the American west, including Alaska. They also included detailed locality and habitat descriptions. Furthermore, voucher specimens were made by Billings and Clebsch from many of these populations (deposited at DUKE and TENN). Most of the Duke collections were at least twenty years old with unknown viability, but they had been stored below zero degrees C in glass vials. Since there were no seed collections from eastern North America, a trip was made in the summer of 1983 to the New England states, Quebec, and Ontario. Herbarium specimens from known populations were mapped to expedite the finding of collecting sites. In addition, all major herbaria in the northeastern U.S. were visited for more information concerning exact location of populations. Figure 2 maps the populations visited by the investigator (squares). Collections from these populations were described as to habitat, including exposure, aspect, and substrate. Inflorescences were collected for seed material, along with living plants and voucher specimens (deposited at VPI). Collections were made from thirty-one populations. Seven of these were from historical sites, with the remainder in new locations. 20 21 CYTOLOGY For chromosome counts, ca ryopses were germinated on four thicknesses of moistened filter paper, and were kept evenly moist throughout the germination tests. Caryopses received approximately a nine hour photoperiod under fluorescent lights at 25 ·degrees C. Seed germination was rapid and successfu I, with 85 % to 100 % of seeds germinating in five to twelve days. For a more detailed experiment on seed germination in Trisetum spicatum see Clebsch and Billings (1976). Seedlings with ten millimeter roots were pretreated with 0.05 96 colchicine for three hours and fixed in absolute ethanol-glacial acetic acid (3: 1 by volume) for at least ten minutes. Root tips were squashed directly or transferred to 70 % ethanol and refrigerated at four degrees C for later microscopy. Squashes were made using the Fuleguen technique (Darlington and LaCour 1963). Two cytotypes of Trisetum spicatum were confirmed, tetraploid and hexaploid. These data support the findings of Becher (1959), Morrison (1959), and Love (1962). All tetraploids came from high elevation or upper latitude populations, whereas most hexaploids came from the more temperate regions (32 to 65 degrees) of eastern North America. 22 NUMERICAL ANALYSIS Three-hundred-fifty herbarium specimens (from US, DUKE, and TENN), representing the entire North American range of the species, were measured for thirty-three quantitative. and qualitative morphological characters (Table 4). Fifteen of these were vegetative, while the remainder were reproductive. Basic statistics for means, standard deviation, range, standard error of mean, coefficient of variation, etc., were calculated for the different variables by the SAS means procedure (SAS, 1982). The multivariate numerical techniques of cluster analysis, principal coordinate, and discriminant function were used to analyze the data. Cluster analysis and principal coordinate were calculated with the NT-SYS package of Rolf et al. (1977). Discriminant function was computed with the SAS (1982) program. Matrices were generated for both Q-correlation and distance measurements. Clustering was performed with the unweighted pair group method using arithmetic averages (UPGMA). In addition, analysis of variation between OTU's (operational taxonomic units) was determined th rough factor analysis. Factor analysis also calculated the correlation between characters. Five different combinations of characters were analyzed in the cluster analysis. These combinations include: 1) all of the original 33 characters; 2) those characters selected through factor analysis (i.e. Table 4. Morphological characters used in this study, including their acronyms, description, and scoring method (continuous (c), bistate (bs), and multistate (ms). Characters dropped from the final analysis are also listed with the reason for elimination (E =environmental basis, I= invariant, M = missing values, * = inconsistant, and ! = kept for final analysis). MORPHOLOGICAL CHARACTERS Vegetative Characters Reproductive Characters Acronym Character Seo r i ng E I i m. Acronym Character Seo r i ng E I i m. N CULMHEIT .... culm height ( c) E INFLINTR •. ~. inflorescence base interruption (bs) w SHEATHLG .... sheath length (c) E INFLLGTH .. ~. inflorescence length (c) * SHEATHCL .... sheath color (bs) E INFLWDTH .••• inflorescence width (c) SHEATHSF .... sheath surface (bs) I INFLSHAP •••. inflorescence shape (ms) SHEATHPB .... sheath pubescence (bs) E SPKLLGTH .... spikelet length (c) BLADELGT .... blade length (c) E SPKLWDTH .•.. Spikelet width (c) BLADEWDT .... blade width ( c) E SPKLCOLR ••.. spikelet color (bs) BLA~ECLR .. ~.blade color (bs) E GLUMERTO ..•. glume ratio (c) BLADESUR .... blade surface (bs) I GLUMESUR .... glume surface (bs) BLADEPUB .... blade pubescence (bs) E GLUMEKEL .••. glume keel surface (bs) LIGULEHR .... i igule hairs (bs) I LEMMALGT .•.. le~ma length (c) STEMSFAB .... stem surface above(ms) LEMAWNLG .... lemma awn length (c) STEMSFBL .... stem surface below(ms) ! LEMAWNSP .... lemma awn shape (bs) STEMCOLR .•.. stem color (bs) E PALEALGT .... palea length ( c) RACEJNFL. .•. racemes per inflorescence (c) FLTSSPKL .... florets per spike let · (c) ! ANTHERLG .... anther 1ength (c) M 24 variate vs. in variate); 3) floral characters alone; 4) characters used in previous taxonomic treatments; and 5). weighting the characters in number two above. Character weighting was performed after Adams (1975), who suggested weighting characters by one minus their F-values (1-F). This study deviates from the ones suggested by Adams in that characters were weighted by the inverse of their respective F-values. This 1/F calculation should then weight all characters proportionally, according to their respective variances. By using reciprocal F-values the characters with less variance are emphasized, while demphasizing those characters with a high degree of variance. This a posteriori design attempted to eliminate some of the "noise" created by the more variable characters and to allow more subtle differences to surface. Principal coordinate analysis (Gower 1966), using standardized data, outlines the trends of variation as well as reduces the dimensionality of the matrix. Matrix dimensionality is reduced by factors being created from the variables. These variables are then selected and included in the analysis according to their variance (high variances are included first). Principal coordinate was used to analyze: 1) all of the original 33 characters; 2) the characters selected through factor analysis; and 3) the weighted characters chosen through factor analysis. Five factors were plotted (two at a time) to produce two-dimensional scatter diagrams. 25 Discriminant function utilized both stepwise Wilks and test list sub-programs to analyze the data. The stepwise procedure selects characters on the basis of F ratios. Characters with the highest F ratios are selected one by one for entry into the analysis. Test list predicts group membership from the matrix and compares this against the a posteriori assignment made th rough cluster analysis. OTU's may then be reclassified according to the group with the highest affinity. STOMATE SIZE Differences in ploidy level (within the same species) have been associated with changes in plant morphology (Stebbins 1950) and in particular, stomate (guard cell) size (De Wet 1954). To correlate stomate size and ploidy level, plants with known chromosome number were· grown from seed and surveyed for differences in stomate size. All fresh plants were pressed and dried in order to provide fair comparison with herbarium specimens. Nail-polish peels for grass leaf epidermis study were made following the technique of Hilu and Randall (1984). Peels were always made on the abaxial surface of the leaf midpoint. 26 ENZYME ELECTROPHORESIS Polyacrylamide gel electrophoresis was employed to produce isozyme profiles of 600 individual seedlings representing eight populations (Table 5; Fig. 4). Eight enzyme systems were assayed for allele frequencies among and between populations. Ultimately only 5 enzyme systems were assayed because of poor staining in the other three. Caryopses were germinated by the same method used for the cytological study. Two-week-old seedlings of uniform height were ground individually in buffer (appendix B), placed in microcentrifuge tubes, and frozen at -20 degrees C until used. . Seven or ten percent polyacrylamide gels were used depending on the enzyme system. Enzymes were run on an LKB PROTEAN electrophoresis unit with a Buchler 3-1500 power supply. All polyacrylamide gels and buffers were prepared according to methods described by Davis (1964) and Ornstein (1964). Gels were stained according to the recipes of Shaw and Prasad (1970), Gabriel (1971), or Harris and Hopkinson (1976). This study assumes that all electrophoretic data represent enzyme loci, and no attempt was made to discover the genetic bases for the alleles surveyed. All electrophoretic data were analyzed qualitatively and quantitatively, according to the mobility and presence or absence of enzyme loci, to group tax a in terms of their "genetic" (electrophoretic) differences (and similarities). 27 TABLE 5 Populations surveyed for electrophoretic study. Population numbers can be correlated with the map in Fig. 4. LOCATION ELEVATION COLLECTOR and DATE Alaska, Thompson Pass 793 m P. Godfrey 1963 2 Alberta, Highwood Pass 2,205 m D. Billings 1958 3 California, Carson Pass 2, 743 m H. Mooney 1960 4 Wyoming, Medicine Bow Mountains 3,292 m D. Hillier 1962 5 Ontario, High Falls 60 m J. Randall 1983 6 Vermont, Smugglers Notch 914 m J. Randall 1983 7 Quebec, Chibougamau 305 m J. Randall 1983 8 Quebec, Magpie sea level J. Randall 1983 28 A N © T E D S NATIONAL SCHOOL m: AERONAUTICS, INC. NORTH AMERICA COPYRIGHT G,ALL.UP MAP&: STATIO~.ERY _COMPANY StAU 'Of MILU Figure 4: Populations surveyed for electrophoretic study. . RESULTS Stomate Size The extremes in stomate size reflect the respective ploidy levels. However, correlating stomate size with chromosome number was not always successful since there was considerable overlap between stomate size (Figure 5). If the stomates are less than 31 microns the plants are tetraploid and if greater than 43 microns they are hexaploid. These criteria were used to identify the ploidy levels of the herbarium specimens included in the numerical analyses. Although the results of the numerical analyses have not yet been discussed, the results of stomate size are included first to facilitate a better understanding of the numerical results. OTU's from the two cytotypes clustered randomly within and between "groups" (Fig. 6). Random groupings of OTU's occurred regardless of character combination or th rough character weighting. Perhaps there are more differences in stomate size between diploids and higher ploidy levels than between taxa of high ploidy levels. 29 30 hexap/oid tetraploid 25.3 31.7 42.3 55.5 Microns Figure 5: Range in mean stomate length between cytotypes. 31 NHALKD86 Alaska (4X) I I NHCNLL33 Canada (4X) I I NHCONA36 Colorado (4X) NHMEML26 Maine (GX) I I JR RAG ENE Quebec (GX) I I TNMNNA74 Minnosota (GX) I I NHLANA07 Labrador I I l_I JRCROWPT Ontario (GX) I I NHALML39 Alaska I I NHGLGF48 Greenland I I I I NHBCGM79 British Columbia (4X) 11 11 NHALPC92 Alaska II I II I NHMTFC57 Montana (4X) 111 I I NHVTML28 Vermont (GX) II II NHRIML47 Rhode Island (GX) II II NHWYNA05 Wyoming (4X) Figure 6: Expansion of cluster analysis demonstrating geographic distribution. 32 NUMERICAL I MORPHOLOGICAL ANALYSES Cluster analysis reveals the mode of groupings of the different OTU's. OTU's are grouped (clustered) according to their overall similarities or differences. Phenograms were generated for: 1) all of the original thirty-three characters; 2) those characters selected through factor analysis; 3) floral characters alone; 4) characters used in previous taxonomic treatments; and 5) weighting the characters in number 2). The distance equation always gave the highest degree of clustering (Fig. 7), whereas Q-correlation considered all OTU's identical. The distance technique is more effective than similarity when closely related taxa are analyzed, and because of the use of both quantitative and qualitative characters (Sneath and Sokal 1973). The number of characters used in the analyses above was trimmed by dropping characters known to vary with the environment. This choice (Table 4) was based on the variation seen on individual herbarium sheets, from plants grown in the greenhouse, and from field observations. These characters include culm height, sheath length, sheath color, sheath pubescence, blade length, blade width, blade color, blade pubescence, stem color, and spi kelet color. Factor analysis was used next to test the variance between OTU's and the characters contributing to that variance. This approach allowed the deletion of variables with invariant characteristics. These included sheath surface, blade surface, ligule hairs, lemma awn shape, and 33 PHENOGRAM FROM MATRIX Q 13.500 11.000 8.500 6.000 3.500 1.000 -1.500 1~~~~1~~~~1~~~~1~~~~1~~~~1~~~~1 !DENT NHALKD86 4X I I NHCNLL33 4X I _I NHCONA36 4X I I NHMEML26 6X I I JRRAGENE 6X I I I I TNMNNA74 6X II II NHLANA07 11 I I I JRCROWPT 6X 11 11 NHALML39 II 11 NHNLGF48 11 11 I NHBCGM79 4X 111 II I NHALPC92 II I I I II I NHMTFC57 4X I II I 11 I NHVTML28 6X Figure 7: Cluster analysis phenogram. 34 111 11 NHRIML47 6X I II I II NHWYNA05 4X I I NHALGR85 I I I I NHWANA38 I I I I NHCNB103 4X I 11 I I NHALMG26 I I I I NHORCL46 I I I I NHALKD62 6X I I I I I NHLAFR17 4X I I I I NHCOGP81 I I I I I I NHALOB58 I 11 _II NHALNA32 4X I _I TNCA1033 4X I I NHMEML31 I I I I NHQBIL05 6X I I I NHORNA22 6X I I I I NHMENA48 I I I _I JRSMUGLR 6X I I I JRDENMRK 6X I I I JRMCRIFL 6X I I I _l_l_I JRBRUNSW 6X I I NHWAML10 NHALML60 4X I I NHALML55 6X 11 I NHORNA21 4X I NHCOGU61 NHALCR83 Fig. 7. continued. 35 NHCAYT54 NHNVNA09 I _J NHALML33 4X I I NHALNA89 I I I I NHALAN41 6X 11 I NHWYBP98 6X I I DKNVRM98 I I I NHCONA60 I I DKCA1145 I I NHCOGBOO I I I I NHCOML59 I I I l_l_I NHC01245 NHOR5T29 4X NHALMC91 JRRAP I DE 6X I JRKAPUSK 6X I I JRBURLIN 6X 11 11 JRVALJAM 6X I I JRRDROCK 6X 11 11 JRSILVIS 6X 111 11 JRRIVAUT 6X I I JRSTJBCH 6X I I NHALMC91 I I NHAR1230 I I TNALNA57 I I NHWANA40 I I NHALNA34 4X Fig. 7. continued. 36 I I JRNEYPRP 6X I I NHMNNA53 II II NHALNA42 I II II NHCNNT25 I NHWYNT91 6X NHALNA19 NH390671 I I NHWAMR13 4X I NHCOGU58 4X I I NHNHPG64 I DK149920 I I Nll07571 II II NHOTML72 I II II DKWY1116 I II I I NHCOGU60 II _II NHNCML36 NHNVML62 4X I I NHALNA76 II II NHCONA12 4X I I N2523837 II II DK144720 II I _, ,_, DKMT9T20 NHMINA56 I I DKMNNA 11 I II I _II DKMNNA 11 II II JRMTMCRA II II JRISLCAR Fig. 7. continued. 37 11 11 I 11 11 I JRALGASH 11 111 II 11 JRTERBAY II II I II II I JRMTWASH I II I I II _1_11_1111 JRCHIBML I I NHALAM65 I I I _I NHALKY40 I I _I I NHALNA37 I I NHCNL071 NHCANA33 I NHMNNA44 I I NHGLNA13 11 I NHGLNA18 I I NHALML77 I I DKAL4T30 11 I NHLANA74 11 I DKCNBC31 I I DKWY1145 I I l_I DKWY1284 I I NHALNA38 I I I I NHQNPG48 I I I NHGLPG94 I I NHALMLOO II II NHQBPD89 I I NHALAB96 I I I I NHALAl23 I I I I DKCN8T34 I II I II NHORAT58 Fig. 7. continued. 38 I I r NHBINABO I I NHALSP13 I I DKALSR22 I I I I NHBINA46 I I I NHCNYK32 I I I I NHLANA49 I I I I l_I JRTADOUS NHALNA71 DKNV1181 I I JRRBFALL I I NHQBHB35 I NHBINA31 I I DKALNA21 I I I I JRMATAMC II II NHCAYT23 II I le-I I I JRMAGPI E I I NHALSP03 NHALNA43 I I NHGLNA67 I TN EUROPE I NHALJR70 I -' I NHALNA70 I I ,_, DKC01232 I I NHAL3T51 I I I DKWY1236 I I I _I DKNZ5T82 II II NHBINA44 Fig. 7. continued. 39 I II I _l_ll_I DKWY1288 I I TNMNNA74 I I I _I NHVTNA61 I I I I N2436514 I I NHIDNT07 I I l __ I NHALNA69 I I NHCOML34 I I DKALNA30 I NHALNA31 I I -' NHALNA 11 I I NHCNPH51 I I I NHWANA15 I II _I_! I NHBINA84 I I NHCNHU79 II 11 JROLDWMB II II JRP I NBAY I I NHUTML53 I I I I NHWYML34 I I I I l __ l_I __ , NHALMA01 NHCNAB18 I NHWY8T69 I I NHCNML64 I I NHMTNA77 I I NHALNA73 I I NHWYML18 II II NHNYNA30 II I I I NHMTNA48 Fig. 7. continued. 40 NHAL2M04 DKCANA15 NHC01477 I I NHORNA23 11 I NHMT8T42 11 11 NHMEML20 111 11 NHMT6T97 I I NHORMH24 NHMEML25 I I NHALNA98 I I NHBCNA74 11 11 NHCONA34 I I NHMENA30 I I I I NHOR1623 I 11 I 11 NHUT7Tl 3 I I I I NHWY7T60 I I NHALMA54 I I NHORML25 I I _l_I NHCONA32 NHALML81 I I NHALMLB5 11 11 NHALMLOO I I NHNMPB79 I NHOR7T28 NHALKD84 NHALNA40 NHCNNT20 Fig. 7. continued. 41 I I NHNYPG58 I I I I NHVTNA36 I I I I NHNYNA51 I I -' I I DKCA7521 II II NHUT1037 II II NHWYNA77 II II NHID8T03 II II NHVTML27 I I I TNVTNA97 II I NHMENA13 I I NHMENA49 I I I DKID9205 I I _I NHNHNA76 I I JRMANICO NHTD1206 NHCABM70 I I NHCABM70 I I NHWYMLOO I I NHUTML54 I I I I NHNYNA39 II I NHBCTC21 II I NHCNNT98 I I NHMENA34 11 II NHC01241 I I NHUTNA05 11 I NHALNA39 II II NHMINA07 Fig. 7. continued. 42 II I II I ___ DKMAML59 111 11 DKMEML94 111 _Ill ___ NHCN7T38 NHCOGU20 I I ____ NHALNA45 I I NHALNA10 I I I I ___ NHALAL83 I I I NHC08T23 I I I I NHOR7068 111 Ill l_I ___ NHIDML24 1111 1111 ____ NHC01252 I I I JRHIGHFL I I I _l_l_I _____ JRPTLAND I I NH507315 I I NHALKl11 I I I NHALSF23 I I I I l_l_l_I ____ NHALSB96 I I N1646290 I I N2436568 I I _1 __ 1______1 NHNHMLOW I NHALNA16 I I ___ ----~--- NHALNA46 NHCOML23 DKNVB806 I l_I _____ NHWYYS89 I NHQBNA32 I I I I DKWA6265 I I I I I ___ NHNCML31 I 11 I I NH868122 I 11 I II ___ NHARNA25 I I _l_l ___--'- DKWY1018 1 I NHUTML55 I I I _I ____ NHWY1066 I I I I N2040267 I I I I _I ____ JRDICKEY Fig. 7. continued. I 11 43 I I 11 NHBCPR91 I I' 11 I I 11 DKSDNA42 I I 11 I I I 11 _I NHMENA24 I I 11 I I I 11 I DKSDNA44 I I 11 I I I I _I l_l_I NHMTNA94 I 11 I 11 DKC01199 I 11 I I 11 I NHVTML19 I 11 I I 11 I NHALAM96 I 11 I I 11 NHAE6T50 I 11 I I 11 I DKAL 1807 I 11 11 I 11 11 NHWAMR03 I 111 I 111 NHIDNA39 I I N2436570 I I N1768865 I I l_I NHALMD17 I _I NHALNA58 I I NHC04T01 I I NHCOPH86 I I I _I NHC01076 I I I I NHCNRP32 I I I I NHMTNA47 I I I I I I NHCOML94 I I I I I I NHC07T34 I I I I I I __ l_I NHC01043 I I I I I I N1768864 I I I I I I DKWYNA16 I I I I I I I I NHWYSS96 I I I I I I I I TNMENA16 I I I I I I I I NHORNA26 I I I I I I I I I NHVTML29 I I I I I I l_l __ l_I NHPNML47 I I NHMTNA94 _I !DENT 13.500 11.000 8.500 6.000 3.500 1. 000 -1.500 Fig. 7. continued. 44 ""'_uu-- i Le== IL== 11 I! r~=c::= IJ IE-· !ir:"-== t :tl r- ' I I I! i I I I I 3 I I I ,-i-'. __: 4 :/;i! .:=== = ,_i_u,~--· ,_,_.:..---:.....--•-·-·-·-· - ·- ..... ·_ ·-· ·- ·-. ·- ··- Fig. 7, m. Condensed phenogram. Numbers indicate groups used in discriminant analysis. Dots represent individuals from the same population. 45 First two spaces US = US National Herbarium DK = Duke University TN = University of Tennessee JR = J. Randall collection Second two spaces AL = Alaska OR = Oregon CA = Ca·lifornia UT = Utah NV = Nevada BC = British Columbia NM = New Mexico AB - Alberta CO = Colorado QB = Quebec NC = North Carolina NT = Northwest Territory SD = South Dakota FC = Fort Churchill WY= Wyoming FR = Frazier River ID = Idaho GF = Grand Falls MT = Montana Kl = Kodiak Island LO = Lover MH = Mt. Hood LA = Labrador GL = Greenland ·. OT = Ontario Third two spaces KO = Kodiak, A.laska AL = Aleutians GR = Grahm, Alaska GM = Green Mountain ML = mo/le YT = Yosemite PG = pilosiglume IL= Is .. Lemoine SP = spicatum PR = Pasaytin Ridg.e MD = maidenii SS = Sulphur Spring CL = Crater Lake G 13 = Glacier Bay AT = above timberline TC = Telegraph Creek AB = Aurilix Bay YS = Yellowstone SB = saltwater beach BM = Black Mountain BP = Breccia Park Bl = Bihriny Island JR = Jago River PC = Port Clarance YK = Yukon PH = Point Henderson BH = Berthound Pass CR = Corville River De.ita RM = Ruby Mountains AM = Alalogoshuk Mt. PD = Pointe Dutilly · GP = Gray's Peak GU = Gunnison Basin . PB = Pecos Baldy Lake OB = Olga Bay LL = Lake Louise . AT = Attu Is. MR = Mount Ranier SR = Sagavanirhtok River HB = Hood Bay HU = Hudson Bay MC = Mt. McKinley RP = Rocky Mt. Park NA = nothing available Last two spaces represent the last two numbers on the respective herbarium sheet. Fig. 7. continued. Key to OTU labels 46 racemes per inflorescence. The inflorest:erice interruption variable was dropped sin.ce it varied between populations, within populations, and even within individuals (Fig. 8), and was thus of little use as a taxonomic character. Anther length was dropped because many harbarium specimens were either immature or past anthesis. To delimit the correlation of the characters used in the final analysis, a phenogram was generated (fig 9). Representation of the matrix was quite good, with a cophenetic value of 0.82. The only highly correlated characters were lemma length (LEMMALGT) and palea length (PALEALGT). Otherwise, no other characters are highly correlated. Many characters of the same structure do "cluster" (at very low levels), for example, characters representing pubescence (STEMSFAB, STEMSFBL, GLUMESUR), relative size of the inflorescence (INFLLGTH, INGLWDTH), and floret characteristics (LEMMALGT, PALEALGT, LEMAWNLG). These characters were weakly correlated, indicating their potential taxonomic value. The expanded top of the phenogram (Fig. 6) of Figure 7 demonstrates the· random clustering of OTU's from different geographic regions. It was possible to find taxa clustered in the same group from Alaska, Newfoundland, Colorado, Maine, Quebec, Minnesota, Ontario, British Colum!;:>ia, Montana, and Vermont (Fig. 6). Often, several taxa from within the same population would cluster widely from one another (Fig. 7,m). Therefore, no meaningful relationships can be found in the 47 Figure 8: Inconsistency of inflorescence interruption within the same individual. 48 0. 120 0.270 0.420 0.570 0. 720 0.870 _I ___ ------I Character 1 I . STEMSFAB STEMSFBL GLUMESUR INFLLGTH I __ ! ______INFLWDTH I I SPKLLGTH I I I I SPKLWDTH I I I I LEMMALGT I I I I PALEALGT I I I I __ l ____ l_l_I __-'---_ LEMAWNLG I I INFLSHAP I I I '-~~~~~~- SPKLCOLR ! I - _I ! ______FLTSSPKL I I GLUMERTO I I __ ------GLUMEKEL Figure 9: Correlation between characters used in the final analysis. See Table 5 for description of character acronyms. 49 groups generated by cluster analysis in terms of geographic distribution. Similar results were obtained through principal coordinate analysis. Five factors were created, but the first two factor loadings accounted for 99 % of the total variance. This degree of extraction is essentially complete and the plot of the first two factors should, therefore, be the best example of the relationship between OTU's. Principal coordinate analysis utilizes axis rotation to achieve the best separation of points in the matrix. Two factors at a time are plotted against each other with the arrangement of points displayed on a two-dimensional graph. The two-dimensional graph (Fig, 10) displayed no segregation of tax a where all OTU' s appeared at the bottom of the Y-axis (factor 2), with all variation along the X-axis (factor 1). Since 95 % of the variation was contained in factor 1, OTU separation should occur along this axis. Principal coordinate, unfortunately, does not list the variables comprising the respective factor loadings. It is assumed though, that the inflorescence length (I NFLLGTH), inflorescence width (IN FLWDTH), and inflorescence shape (INFLSHAP) characters are included in factor 1 since they are corelated and possess a high variance. As in the cluster analysis, the OTU's fell out in a seemingly random fashion. No correlation could be found in terms of geographic distribution or chromosome number.· If the effect of factor 2 is eliminated and factor 1 is projected on one axis, no separation of groups can be seen whatsoever. Therefore classification would be arbitrary. . I 42.463 - I I I I -,-I I it I I I F 12.337 I 31. A I c it it T I 0 it I it 01 R I it it it 0 it it it I 2 2 it it it it it it it it it I it it it it it it it I it 2 2itititit it 2.295 2 it it2 it 2 I it 22 it it it 41 it 2 it itit I 2 22 it it 2 it lit__ ? __ititititititit 2it it itit it it it I it 2 it2it 4ititit2"it it"°"itit- itit 3it it it it I it it 2it it itit it 21 22 ititit 2 it .it .it it it it it I it it it itit it I it it- 22 2 it it it I it 2it2 2itit it 2 --it it I 2 2 I it2 ititi+it 2 it itl it it it I it it it it it I it it I it -:-7.748 * 51 -:-1 _____ 1 l _____ I____ _ I I I I -37.715 -24.384 -11.053 2.278 15.609 28.939 42.270 55.601--~'-- 68.932 FACTOR 1 Figure 10. Two-dimensional plot of principal coordinate. 51 Discriminant function acts to test the a posteriori assignment of groups. This method was utilized to "test" the "groups" generated by the cluster analysis (as indicated by the lines drawn through the phenogram in Fig. 7) ..·Only a ten percent misclassification was detected. · These results support the "groups" produced by the cluster analysis even though the clusters were not meaningful Un terms of chromosome number or geographic distribution). Discriminant function also lists the characters most important in the discrimination (Table 6). These characters were of the inflorescence and stem pubescence. The res1Jlts of the SAS· means procedure are listed in Table 7. lhose characters scored as continuous variables (INFLLGTH, INFLWDTH, LEMAWNLG, PALEALGT, and LEMMALGT) are graphed in Figure 11. OTU's were assembled into into subgroups according to their herbarium sheet annotation labels. Subgroups were then compared morphologically. Subgroups were not distinguishable using the morphological . differences among the characters analyzed (Fig. 11). Overlap was considerable betwen the range and standard deviations of all subgroups. Thus, none of these characters resolved a particular subgroup as defined by herbarium sheet annotations. This investigator is aware of the problem with herbarium sheet. annotations, but it was the only means of selecting subgroups since this study fails to recognize subspecific separation. 52 TABLE 6 Summary of stepwise selection of variables. Variable F prob > F INFLLGTH 57.809 0.0001 STEMSFAB 12.378 0.0001 INFLWDTH 8.635 0.0003 LEMAWNLG 7.812 0.0006 INFLSHAP 5.067 0.0075 PALEALGT 4.168 0.0174 LEMMALGT 3.489 0.0331 STEMSFBL 3.368 0.0372 SPKLWDTH l.956 0. 1451 FLTSSPKL 0.962 0.3847 GLUMESUR . 0.876 0.4188 GLUMERTO 0.708 0.4944 SPKLCOLR 0.670 0.5134 GLUMEKEL 0.255 0.7751 SPKLLGTH 0.175 0.3971 TAB{E 7. Basic statistics on 15 morpllologicai characters maintained for the final analysis of the ll.ill..l.l.f:, spicatum, and pi losiglume subgroups .. ya r i ab I e N Mean Stand a rd Minimum Maximum Range Variance Std Error Devi at ion Value Va I ue of Mean ---~··~------·--·------JH()J_J_f) .§ilQ9J'_Q!lQ ··~·---~-~-··-----~-- STLMSFAB IJ2 0.2380952 0. 53231;52 0.0000000 2.000000 2.0000000 0.28339 o.oe21l1265 STEMSFBL 42 0.42857111 0, '.?4711044 0.0000000 2.000000 2.0000000 0. 29965 o. 084116633 INFLLGTH l,t2 605. !1761 'JO~> 21!,1317053 ;..200. 0000000 1110.000000 910.0000000 1111576. 59698 32. 57B329111 INFLWDTH lt,2 108.8095238 3'.c'. 17311917 .50. 0000000 180.000000 130.0000000 1035.13357 11. 96111n7511 I NFLSllAP 42 1 . 88095211 0. 83231155 l. 0000000 3.000000 2.0000000 0.69280 0.128113370 SPKLLGTH 42 56.9761905 6.2161185 Lj.8. 0000000 70.000000 22.0000000 39.38966 0. 968112611 SPKLWDTH 42 19.8333333 3.8121543 13.0000000 32.000000 19.0000000 14.53252 0.58822818 SPKLCOLR 112 0.8095238 0.5516315 0.0000000 2.000000 2.0000000 .0. 301130 0.08511859 GLUMLRTO 112 11. 92857111 0.9.211037 10.0000000 15.000000 5.0000000 0. 8118113 0.111212939 GLUM(SUF< 42 0.01176190 0.21551103 0.0000000 1. 000000 1.0000000 0. Ol.16/.16 fJ.03325859 U1 GLUM EK EL 42 0.7857143 O. l.fl52997 0.0000000 1.000000 1.0000000 0.172117 0.06408214 w LEMMALGT 112 l;[j. 5952381 4. 1218203 41. 0000000 60.000000 19.0000000 22.29559 0. 72859268 LEMAHNLG 112 ii]; 9047619 8. 18031110 30.0000000 6B.OOOOOO 38.0000000 66.91754 1.262249B6 PALEALGT 112 39.9.5238i0 4.3837415 32.0000000 50.000000 18.0000000 19.21719 0.67642599 FLTSSPKL 42 l.1904762 0.3974366 1.0000000 2.000000 1.0000000 0.15796 0.06132580 Vil r i ab I e Co. Var. Skewness l Variable N Mean St fl nda rd Min i rnurn Milximum Range Variance Std Error Deviation Value Value. of Mean STEMSFAB 33 0.4545455 0.7111131 0.0000000 2.0000000 2.0000000 0.50568 0.12378890 STEMSFBL 33 0.9696970 0.6839613 0.0000000 2.0000000 2.0000000 0.46180 0.11906238 INFLLGTI~ ~3 531.2121212 170.5644595 200.0000000 830.0000DOO 630.0000000 29092.23485 29.69146130 I NFLVDTH :fl 99. 6969697 32. M246 I 7 50. 0000000 200.0000000 150. 0000000 1065.53030 5.68232321 INFLSHAP 33 2.18181B2 0.6351449 1.0000000 3.0000000 2.00GOOOO 0.403~1 0.11056454 SPKLLGTH 33 54.5115115115 12.8211030 5.0000000 17.0000000 72.0000000 164.38068 2.23186755 SPKLHDTH 33 1.8. 7272727 3 .. 98611B28 13~0.008000 30 .OO(+OfHlfi--- -1-·1-. O.!J-i}(}00-0 .l'i.cl'l92tJ5 Cc_.i); 69395759 ·sP1 Variable Co. Var Sl{ewness Kurtosis STEMSFAB 156.445 1. 2798L~ 195 0.290B5933 STEMSFBL 70.53lf 0.03761630 -0.72569050 INFLLGTH 32. 109 -o. 08734921 -i.07978320 INFLWDTH 32. 7Ll2 J .00:425060 1 . 73 7011809 I NFLSHAP 29. 111 -0.160911950 -o .115252610 SPKLLGTH 23.505 -1.495548111 6.01058061 SPKLWDTH 21.287 1.18131911 1.35998958 SPKLCOLR 56.827 0. 000301~39 0.20639805 GLUMERTO 10. 553 -0. 089114646 -o. 3211.39650 GLUMESUR 321.131 2.98340886 7.34279570 GLUMEl [! i Io s i g I ume subg rou12 STEMSFAB 8 0.6250000 0. 517511917 0.0000000 1.0000000 1.0000000 0.267851 0. 18298126 STEMSFBL 8 0.3750000 0.517511917 0.0000000 1.0000000 1.0000000 0.267851 0. Hl298126 INFLLGTH 8 430.0000000 81.1. 68'~28763 320.0000000 590.0000000 270.0000000 7171.428571 29. 9t101.11 702 I NFLWDTH 8 73.7500000 28. 75388173 40.0000000 120.0000000 80.0000000 826.785714 10.16603238 INFLSHAP 8 2.1250000 0. 611086994 1 . 0000000 3.0000UOO 2.0000000 0.11HH14 0.226581711 SPKLLGTH 8 67.3750000 13.86606855 48.0000000 81.0000000 33.0000000 192.267857 l.j. 90239555 SPKLWDTH 8 18.2500000 2.05287255 15.0000000 22.0000000 7.0000000 4.2142136 0.72580005 SPKLCOLR 8 0.5000000 0. 531152248 0.0000000 1.0000000 1.0000000 0.285714 0. 188982211 GLUMERTO 8 12.5000000 0.75592895 12.0000000 ·111. 0000000 2.0000000 0.571429 0.26726124 GLUMESUR 8 0.3750000 0.51754917 0.0000000 1.0000000 1.0000000 0.2671351 0. 18298.126 GLUMEKEL 8 1. 1250000 0.99103121 0.0000000 2.0000000 2.0000000 0.9821!13 0. 350382411 LEMMALGT 8 111 • 0000000 4. 40778532 37.0000000 51.0000000 111. 0000000 19. 1128571 1. 5583871.1/1 LEMAWH.G 8 37.5000000 7.03054560 26.0000000 116. 0000000 20.0000000 49. 1:28571 2 .1rn557323 PALEALGT 8 33.6250000 l. 890998119 16.0000000 42.0000000 26.0000000 62.267857 2. 7898892'1 FLTSSPKL 8 1.0000000 0.00000000 1.0000000 1.0000000 0.0000000 0.000000 0.00000000 Variable Co. Var. SKEWNESS f =f-4 molle 0 ~ spic a tum ~ 'l!C ~ pilosiglume ~ ::i:: f-4 molle Q ~ ~ spic a tum t-J I ~ z pilosiglume ~ + I I I I I I 0 20 40 60 80 100 mm Figure 11: Graph of morphology versus subgroup. 55 c:> io-::1 Z molle ~ < spicatum ~ ~ pilosiglume ~ ~ c:> molle io-::1 ~ spicatum ~ < pilosiglume ~ ~ c:> molle ~ ~ spicatum ~ pilosiglume ~ io-::1 0 2 4 6 8 10 mm Fig. 11: continued. 56 :c: ..... (!) _J _J LL z Key to Geographic Location ::c: ...... Alaska 0 -- Northern Canada 3: _J Western Canada LL Rocky Mountain Canada z Rocky Mountain U.S. Eastern Canada Eastern U.S. 0 20 40 60 80 100 120 mm Figure 12: Graph of morphology versus geographic distribution. 57 :c I- C!) ...J ...J ~ a.. en (.!) ...J z 3: <( ::E w ...J I- C!) ...J <( w ...J <( a.. I- C!) ...J <( ::E ::E w ...J 0 2.0 4.0 6.0 a.o 1 o.o mm Fig. 12: continued. 58 In addition to subgroup versus morphology, geographic distribution versus morphology was tested. Eight geographic regions were assigned and compared graphically (Fig. 12). No distinctive differences between morphology and geographic distribution were elucidated. Specimens from the respective geographies" overlap in their ranges and standard deviations. Again, morphological differences fail to separate the taxon into distinct units. The past taxonomic treatments rely on the characters of culm height, inflorescence length, inflorescence width,· inflorescence shape, inflorescence interruption, lemma color, and glume length (Table 3). Culm height is a highly variable character with a strong environmental basis for this variation (Fig. 13). Inflorescence interruption is also highly inconsistent. Lemma color and glume length were characters eliminated by factor analysis because of their invariate nature. Statistics indicated that some of the above characters were non-significant (p > 0.05). However, inflorescence length, inflorescence width, and inflorescence shape were found to be significantly different between OTU's (p < 0.05). Inflorescence characters are variable even within the same population (Fig. 14) The other characters found to be statistically significant are stem surface above (STEMSFAB), stem surface below (STEMSFBL), lemma length (LEMMALGT), lemma awn length (LEMAWNLG), and palea length ( PALEALGT), which often vary at the intrapopulational level. 59 Figure 13: Inconsistency of culm height within the same population. 60 f'IMola "' doe C....,._ Baain ~ s.p.cioe, and Hu..ci.le c.-tioe. Calorello TRIS , UM ~PICATUM IL . ) Rlcble Figure 14: Variation in inflorescence length, width, and shape within the same population ;· 61 The numerical analyses in summary: 1) cluster analysis gave the best separation of groups using the distance coefficient, although none of the clusters demonstrated any meaningful relationships, 2) character weighting through inverse F-values to proportionally weight all characters relative to their respective variance failed to help emphasize any "hidden" or potential groups, 3) principal coordinate failed to elucidate any trends of variation or group segregation; and 4) discriminant function supported the selection of groups by the cluster analysis regardless of its failure to produce meaningful groups. Finally, the characters used in previous taxonomic studies are extremely variable and essentially unreliable taken either singly or together. Thus, there are more exceptions to the past "rules" of subtaxa delineation than consistancies. ELECTROPHORESIS Of the eight enzyme systems surveyed, only five gave useful results. These included acid phosphatase (ACP), alcohol dehydrogenase (ADH), esterase (EST), ma late dehydrogenase (MDH), and peroxidase (PER). Of these, all but PER exhibited good resolution. All enzymes migrated a nodally. Zymog rams (Fig. 15) depict the relative mobilities of the different enzyme systems. (only the dark-staining bands are included). Individual populations showed complete homogeneity. There is also a remarkable degree of homogeneity between populations, as evidenced ACID PHOSPHATASE High Fa I ls (6X) Chibougamau (6X) .80 .79 .75 .75 Magpie (6X) Smugglers Notch (6X) .BO .79 .74 .74 Ca I i fo rn i a ( 4X) A I be rta ( 4X) .74 .74 ------CJ') .72 N Wyoming (4X) Alaska ( 4X) .75 .74 .74 .72 .73 FIGURE 15. Zymograms of the enzyme systems surveyed of 15 individuals in eight populations. Al I enzymes migrated anodal ly with their origins at the bottom of each zymogram. Rf values are included to th~ left of each I ine. Dashes (---) represent bands with faint resolution. Alcohol Dehydrogenase !!.i.9..!:! Fa I I s ( 6X ) Chibougamau (6X) .72 .72 .63 .63 .55 .55 Magpie (6X) Smugglers Notch (6X) .72 .72 ------.63 .63 .55 .55 Ca I i fo rn i a ( 4X) Alberta (4X) .57 .56 .57 ------.55 .55 ------ Wyoming (4X) Al a ska ( 4X) .77 ------.76 .68 ------.67 .58 ------~- -- -- .57 .57 .56 FIGURE 15. continued. EST ERASE .!:!l.91:! Fa I I s ( 6X) Chibougamau (6X) .40 .40 .39 .39 .38 .36 Magpie ( 6 X) Smugglers Notch (6X) .39 .39 .38 .38 .36 .36 Ca I i fo rn i a ( 4X) A I be rta ( 6X) . 39 .39 .38 .38 .36 .36 Wyoming (4X) A I a ska ( 4X) • L10 ? ? ? ? ? .40 .39 .39 .38 FIGURE 15. continued. MALATE DEHYDROGENASE High Fal Is (6X) Chibougamau (6X) .71 ? ? ? ? ? .71 .64 .64 ------.61 .61 Magpie (6X) Smugglers Notch (6X) .71 .68 ------.64 .66 ------.61 .63 Ca I i fo rn i a ( 4X) A I be rta ( 4X) ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? .66 Wyoming (4X) Alaska (4X) .66 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? FIGURE 15. continued. 66 pop. 3, / pop. 6, 6X --- pop. 5, 6X pop. 7, 6X - pop. 8, 6X ~l, 4X Figure 16: Esterase: interpopulation similarities and differences. A - difference between tetraploid and hexaploid. B - difference between hexaploid and hexaploid. C - simi l arity between hexaploid and tetraploid. 67 by their similar Rf values (distance band traveled I total length of gel). Although all populations appear to demonstrate monomorphism, heterozygosity is extremely high. EST was the most uniform enzyme regardless of geographic distribution or ploidy level (Fig. 15, c, 16). It also has the most enzyme bands. This result is significant since EST is a highly polymorphic enzyme system (Harris and Hopkinson 1976). The only exception to this EST homogeneity was between the High Falls and Chibougamau populations, both hexaploid, where Chibougamau demonstrated only two dark-staining bands. The most variable populations were Wyoming and Alaska, both tetraploid. The Wyoming population had six dark-staining bands for AOH yet only one for MOH. Both AOH and MOH are dimeric. Therefore, the AOH gene is heterozygous while the MOH gene is homozygous for these populations. A zymogram was not included for PER because of its poor resolution, although the enzyme appeared to be entirely monomorphic. The enzyme systems for each population appear to be fixed. Most populations are fixed for the heterozygote. Only the California and Wyoming populations are fixed for the homozygote of MOH, and the Alberta population for ACP. Genetic variability is absent at the i nfrapopulation level for the enzyme systems surveyed and relatively low at the interpopulation level. Hexaploid populations are more similar to one another than are tetraploid populations. DISCUSSION Trisetum spicatum has had a long history of taxonomic controversy. This is exemplified by the different taxonomic treatments (Table 1) and the survey of its voluminous synonomy through nine different genera (Table 2). Regardless of the infraspecific problems associated with morphological variation I T. spicatum possesses distinct specific boundaries. In the morphological assessment of T. spicatum, cluster analysis and principal coordinate failed to elucidate meaningful subspecific groups in the North American species complex. This situation prevailed after the elimination of characters. under environmental influence and of those determined to be invariant through factor analysis. Even when the more variable characters w.ere demphasized or the less variable characters emphasized through weighting, a meaningful subspecific classification· did not emerge. Variable characters are, of course, required for the subdivision of groups. In this case the vari.able charact.ers overlap to the extent that they are useless for taxonomic purposes. Perhaps most importantly, the characters traditionC311y employed by past investigators as diagnostic demonstrate an exceeding amount of variability. These traditional diagnostic characters with p < 0.05 include: INFLLGTH (p = 0.0001), STEMSFAB (p = 0.0001), INFLWOTH (p = 0;0003}, INFLSHAP (p = 0.0075], and STEMSFBL (p - 0.0372). INFLLGTH also has an F""value of 57.809, indicc:ltirig its tremendous variability. The above characters are quite variable at both 68 69 interpopulation and infrapopulation levels. Lastly, discriminant function tested the OTU classification generated by the cluster analysis and listed the characters most useful in the discrimination. The discrimination of group membership, when applied, essentially paralleled the cluster analyses of the different character combinations. The characters selected in order of their ability to maximize differences between OTU's are primarily the ones formerly considered to be taxonomically useful. This investigation finds these characters, taken either. singly or together, inadequate for subspecific assignment. Correlation between morphology and cytotype apparently does not exist. These results were consistant throughout. This situation is not unique._ Chinnappa and Morton (1984) also discovered no correlation between cytotype and morphology in the boreal Ste/laria /ongipes Goldie (Ca ryophyl laceae) polyploid series. Fernald (1950), notOrious for the assignment of varieties, commented on the variability in T. spicatum, and only assigned to it three subspecific tax a. He did not even recognize the additional varieties created by his own student, Louis-Marie (1928). Fernald based his classification on plant height, panicle length, color, and base interruption, and glume pilosity. Numerical analysis recognized panicle length to be the only one of these with statistical merit. .However, this character is the most variable and fails to diagnose group membership. Louis-Marie (1928) never published the descriptions of his T. spicatum subtaxa. Hutten (1927, 1937, 1959, 1968) was initially aware of the 70 polymorphic nature of the species, yet eventually he recognized nine subspecific taxa in North America. His taxonomy was based on. variation in pubescence, relative lengths of spikelet parts, and differences in spike characteristics. None of these characters taken either singly or together appears to be difinitive. Other workers held the philosophy that different infraspecific cytotypes deserve specific reinstatement. Becher (1959) and Love and Love (1962) considered the tetraploids to be morphologically distinct from the hexaploids. Their diagnostic characters were primarily of the inflorescence shape, color, and base interruption, glume. length, color, plant size, and geographic distribution. Variation was found in all of these characters, and they fail to separate the majority of plants easily recognized as T. spicatum s. I. Electrophoresis revealed that in some cases hexaploids expressed more loci coding for the enzyme system th rough what appeared to be gene duplication (Fig. 16). In other cases the tetraploids expressed more enzyme loci, perhaps because of heterozygosity. Finally, some tetraploid and hexaploid populations were identical in their staining patterns. This may be explained by the decrease or masking of infraspecies variation in isozymes with the increase in ploidy level in autogamous species ( Bahatia et al. 1968). The results of enzyme electrophoresis show the species to be highly heterozygous. lnfrapopulation variation is nearly absent for the enzyme systems surveyed and relatively low for the interpopulation 71 variation. This last situation is particularly interesting since these populations are separated by great distances and are essentially reproductively isolated from one another. Tlius, morphological diversity is great, while genetic variability is small. Preliminary studies show the species to .be self compatible. Therefore, genetic uniformity may be explained through the founder principle, where a single introduction is capable of creating a genetically homogeneous population. Populations are relatively isolated from one another, although reintroductions are Ii kely when they occur along rivers and streams. The result of polyploidy makes the taxonomy of a species very difficult (Cronquist 1968). This phenomenon has no doubt played a role in the taxonomic confusion in T. spicatum. Polyploidy complicates .the taxonomy through genome duplication, which increases the potential for variability. This also increases the potential for phenotypic plasticity (Ennes 1983). Plasticity may be morphological as well as physiological (Bradshaw 1965). Clebsch (1960) demonstrated the physiological plasticity in T. spicatum by observing dramatic changes in rates of respiration and photosynthesis in the same .individual under uniform environmental conditions. Morphological plasticity is strongly suggested in T. spicatum because of its wide range of phenotypes in different habitats and low genetic diversity as shown by electrophoresis. In addition, many .different genotypes no doubt exist · from polyploid segregation. They respond individually to the environment, resulting in different phenotypes. 72 The phenotypic plasticity and polyploid seg reg at ion in T. spicatum probably account for the polymorphic status of the species. Individuals of the two different cytotypes cannot be separated, because of overlapping morphologies. Taxa of different cytotypes should not be recognized as distinct sp~cies, but as chromosomal races. The assessment of variability and assignment of names to those variations associated with polymorphism has burdened the past taxonomy. This study concludes that T. spicatum (L.) Richt. is a polymorphic species complex in North America without distinct subspecific boundaries. This investigator wishes in no way to discredit any of the former students of Trisetum spicatum. It is an extremely difficult task to assign names to the different forms of a polymorphic species. In other words, some species resist pigeonholes more than others and remind us that plant systematics is a dynamic science, ·not one with cut and dried answers. Furthermore, we al I occasionally forget that we exist and perceive the world around us at only one point in time. Thus, we are permitted only a fleeting glance at the dynamics of plant evolution and systematics. LITERATURE CITED Adams, R. P. 1975. Statistical character weighting and similarity stability. Brittonia 27(4) :305-316. Anderson, J.P. 1959. Flora of Alaska and the adjacent parts of Canada. Iowa State Univ. Press. 543 p. Avise, J. C. 1974. Systematic value of electrophoretic data. Syst. Z ool. 23: 465-481. Baldwin, W. K. W. 1958. Plants of the clay belt of northern Ontario and Quebec. Nat. Mus. Canada Bull. 156. Ottawa, Queen's Printer. 324 p. Bahatia, C. R., et al. 1968. Polyploidy and isoenzymes. Genetics 60: 162 (abstract). BMDP. 1983. Biomedical Computer Programs, University of California Press. Becher, T. 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New York. Thorne, R. F. 1972. Major disjunctions in the geographic ranges of seed plants. The quarterly review of biology 47(4) :365-411. The vita has been removed from the scanned document Biosystematic studies in the North American Trisetum spicatum complex ( Poaceae) by John Randall (Abstract) Trisetum spicatum (Poaceae) is circumboreal as well as bipolar in distribution. The species is comprised of both tetraploid and hexaploid cytotypes that display high morphological diversity. Previous taxonomic treatments have resulted in the separation of the species into a series of species, subspecies, varieties, races, and types. Many of these treatments were inconclusive, based on highly variable characters, and few specimens were examined. For instance, the characters of the inflorescence and relative stem pubescence are the characters most relied on for subspecific separation. These characters have been found to be extremely inconsistent, even within the same population. This study is concerned with the North American component of the species complex. Three-hundred and fifty herbarium specimens and field collections were measured for thirty-three characters. These data were analyzed numerically with basic statistics, cluster analysis, principal coordinate, and discriminant function techniques. The results indicate that extreme variation exists within the species complex with no distinct subgroups. The species was also analyzed genetically with isozyme electrophoresis. Eight populations were surveyed for. five enzyme systems. These results indicate the species to possess complete allelic homogeniety within populations with few interpopulation differences. Previous recognition of the hexaploid as a distinct species is not supported morphologically or genetically.