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Systematics and evolution of the Carex pachystachya c o m p le x (Cyperaceae)
Whitkus, Richard, Ph.D.
The Ohio State University, 1988
UMI 300 N. Zeeb Rd. Ann Arbor, MI 48106
SYSTEMATICS AND EVOLUTION OF THE CAREX PACHYSTACHYA
COMPLEX (CYPERACEAE)
DISSERTATION
Presented in Partial Fulfillment of the Requirements
for the Degree Doctor of Philosophy in the Graduate
School of The Ohio State Univerity
By
Richard Whitkus, B.A., M.Sc.
*****
The Ohio State University
1988
Dissertation Committee: Approved by:
Dr. Daniel J. Crawford
Dr. Gary L. Floyd
Dr. Ronald L. Stuckey Adviser ^
Department of Botany ACKNOWLEDGMENTS
I wish to express my deep gratitude to Dr. Daniel J. Crawford for his scholarly advice, extreme patience, and ever-ready humor. Sincere thanks is extended to the members of my various committees, Drs. D. J. Crawford, G. L.
Floyd, K. L. Gross, T. N. Taylor and R. L. Stuckey who have devoted much of their time to molding me into some semblance of a scientist, as well as the drudgery of helping edit this and other of my written works. Pleasure is remembered in the interactions I have experienced with many of my fellow graduate students with whom many thoughtful discussions about research has help me along in many ways. Finally, to Marilyn, whose constant support and encouragement has helped ease the burden of graduate study, even though it brought hardship upon herself, I cannot express my heartfelt appreciation enough.
Financial support for the research presented herein was provided by an NSF doctoral dissertation improvement grant BSR-8311123. VITA
August 15, 1956...... Born - Passaic, New Jersey
197 8 ...... B.A., Rutgers U niversity, Newark, New Jersey
197 9 ...... Grant-In-Aid, Boreal Institute for Northern Studies, The University of Alberta, Edmonton, Alberta
1981 ...... MSc., The University of Alberta, Edmonton, Alberta
1983 ...... Doctoral Dissertation Research Improvement Grant, NSF.
1986 ...... The College of Biological Sciences Dean’s Graduate Research Fellowship, The Ohio State University, Columbus, Ohio
PUBLICATIONS
1981. Whitkus, R. Chromosome numbers of some northern New Jersey carices. Rhodora 83:461-464.
1982. Packer, J. G. and R. Whitkus. In: A. Love (ed). lOPB chromosome number reports LXXV. Taxon 51:363-364.
1984. Whitkus, R. and J. G. Packer. A contribution to the taxonomy of the Carex macloviana D’Urv. aggregate (Cyperaceae) in western Canada and Alaska. Canad. J. Bot. 62:1592-1607.
1985. Whitkus, R. A FORTRAN program for computing genetic statistics from allelic frequency data. J. Heredity 76:142.
1987. Crawford, D. J., R. Whitkus, and T. F. Stuessy. Plant evolution and spéciation on oceanic islands. Pp. 183-199. In: K. M Urbanska (ed). Differentiation Patterns in Higher Plants. Academic Press, London.
Ill 1987. Wolf, S. J. and R. Whitkus. A numerical analysis of flavonoid variation in Arnica subgenus Austromontana (Asteraceae). Amer. J. Bot. 74:1577- 1584.
1987. Whitkus, R., F. A. Bryan, D. H. Les, and L. E. Tyrrell. Genetic structure in a heterocyanic population ofTrillium sessile (Liliaceae). PI. Sp. Biol. 2:67-73.
1988. Whitkus, R. Modified version of GENESTAT: a program for computing genetic statistics from allelic frequency data. PI. Genet. Newslet. 4:10.
1988. Whitkus, R. Experimental hybridizations among chromosome races of Carex pachystachya and the related species C. macloviana and C. preslii (Cyperaceae) Syst. Bot. 13:146-153.
1988. Crawford, D. J. and R. Whitkus. Allozyme divergence and the mode of spéciation for Coreopsis gigantea and C. maritima (Compositae). Syst. Bot. 13:256-264.
1988. Les, D. H., R. Whitkus, P. A. Bryan, and L. E. Tyrrell. The biochemical basis of floral color polymorphism in a heterocyanic population of Trillium sessile L. (Liliaceae). Amer. J. Bot. 75: in press.
1988. Craw ford, D. J., B. J. Post, and R. Whitkus. Allozyme variation within and between populations ofCoreopsis latifolia (Asteraceae). PI. Sp. Biol. 3: in press.
FIELDS OF STUDY
Major Field: Botany
Studies in plant systematics and evolution. Dr, Daniel J. Crawford, Adviser.
IV TABLE OF CONTENTS
ACKNOWLEDGEMENTS...... ii
VITA ...... iii
LIST OF TABLES...... vii
LIST OF FIGURES...... ix
INTRODUCTION...... 1
CHAPTER PAGE
I. MEIOTIC PAIRING PATTERNS IN HYBRIDS AMONG CHROMOSOME RACES OF CAREX PACHYSTACHYA, THE RELATED SPECIES C. MACLOVIANA AND C. PRESLII (CYPERACEAE), AND THE RELEVANCE TO AGMATOPLOID EVOLUTION...... 4
Introduction...... 4 Materials and Methods...... 6 Results...... 7 Discussion...... 9
II. ALLOZYME VARIATION WITHIN THE CAREX PACHYSTACHYA (CYPERACEAE) COMPLEX...... 27
Introduction...... 27 Materials and Methods...... 28 Results...... 30 Discussion...... 32 CHAPTER PAGE
III. PHENETIC ANALYSIS OF THE CAREX PACHYSTACHYA COMPLEX (CYPERACEAE) AND ITS BEARING ON THE EVOLUTION AND SYSTEMATICS OF THE COMPLEX ...... 44
Introduction...... 44 Materials and Methods...... 46 Choice of Characters...... 46 Choice of Specimens and Initial Grouping...... 46 Phenetic Methods...... 47 A nalyses...... 48 R esu lts...... 48 Analysis of Groups...... 48 Analysis of Individuals...... 50 Analysis of Species Pairs...... 52 Intraspecific Variation...... 54 D iscussion...... 55 T axonom y ...... 57 Key to Members of C. pachystachya C om plex...... 63 Species Descriptions and Distributions...... 65
SUMMARY...... 140
APPENDICES...... 141
A. Experimental Hybridizations Among Chromosome Races o f Carex pachystachya and the Related Species C. macloviana and C. preslii (Cyperaceae)...... 141
B. Determination of Maximum Observed Affinity Pairing Formula ...... 149
C. Table of Allelic Frequencies
LITERATURE CITED ...... 164
VI LIST OF TABLES
TA BLE PAGE
1. Observed pairing patterns from hybrids of crosses am ong races o f C. pachystachya (n = 37, 38, 39, 41), C. macloviana (n = 43) and C. preslii (n = 40), MOAP formulae and estimated number of translocations...... 14
2. Percent pollen stainabilities of hybrids among crosses of races of C. pachystachya {n = 37, 38, 39, 41), C. macloviana (n = 43) and C. preslii (« = 40)...... 16
3. Crosses among races of C. pachystachya (n = 37, 38, 39, 41), C. macloviana (n = 43) and C. preslii {n = 40) with unresolved trivalents...... 18
4. Population designation (by collection number) and locality for populations of C.pachystachya complex, C. preslii an d C. macloviana...... 37
5. Measures of genetic variability in 31 polymorphic populations of the C.pachystachya complex and in one population of C.preslii ...... 39
6. Diversity statistics for C. pachystachya com plex, C. preslii and C. macloviana...... 40
7. Matrix of identity (I) values between species of C. pachystachya complex, C. preslii and C. m acloviana...... 41
8. Range of identity values for interspecific population comparisons...... 42
9. Mean and range of identity values for intraspecific p o p u la tio...... n 43 comparisons
10. Synopsis of literature treatment of members of C. pachystachya com plex ...... 86 vii TABLE PAGE
11. Characters used in phenetic analysis and mode of assessment...... 87
12. Taxa and morphological groups used in phenetic analyses...... 88
13. Characterization of data sets for phenetic analyses...... 89
14. Factor loadings for first three axes of group PCA...... 90
15. Classification matrix in DFA of individual OTU’s using ’typical’ groups. Total «©rrs-st classification for ’typical’ groups is 75.7%...... 91
16. Factor loadings for the first two axes of the C. subbracteata-C. gracilior PC A ...... 92
17. Classification matrix for C. subbracteata- C. gracilior DFA. Total correct classification is 90.6%...... 93
18. Factor loadings for first two axes of C. pachystachya-C. subfusca PCA...... 94
19. Classification matrix for C. subfusca-C. pachystachya DFA. Total correct classification is 89.8% ...... 95
20. Classification m atrix for C. subfusca. Total correct classification is 57.2% ...... 96
21. Classification m atrix for C. pachystachya DFA. Total correct classification is 87.3% ...... 97
Vlll LIST OF FIGURES
FIGURE PAGE
1. Number of translocations necessary to account for observed pairing patterns at metaphase I. Numerator is the largest number, denominator the smallest. Single number indicates only value...... 20
2. Average pollen stainabilities within and among races and species...... 22
3. Diagramatic representation of resolution of interlocks between non-homologous chromosomes during prophase. A: Events in plant with localized centromeres resulting in reciprocal translocation. B: Events in plant with unlocalized centromeres resulting in a translocation and two fragments...... 24
4. Metaphase I configuration of plant with unlocalized centromeres from resolution of interlocked non-homologous chromosomes resulting in a translocation and two fragments, subsequent viable gametes, and chromosome complement in siblings ...... 26
5. Groups analysis phenogram based on correlation coefficient pnd average linkage clustering. Group acronyms follow Table 12...... 99
6. Groups analysis phenogram based on average taxonomic distance and average linkage clustering ...... 101
7. Projection of groups onto the first two principal component axes. Group numbers follow Table 12, ellipses enclose species of Table 12...... 103
8. Projection of groups onto principal components axes I and III ...... 105
9. Projection of group centers onto the first two canonical axes of the DFA of individuals ...... 107
IX FIGURE PAGE
10. Illustration of F-statistics from DFA of individuals. Within each species, groups not significantly different at p = 0.05 level are joined ...... 109
11. P rojection of ellipses of C. montereyensis and C. harfordii onto first two principal component axes. H = type of C. harfordii', M = type of C. montereyensis...... 111
12. Projection of group ellipses for C.subbracteata an d C. gracilior onto first two principal component axes. G = type of C. gracilior, S = type of C. subbracteata ...... 113
13. Projection of ellipses of ’typical’ groups of C. subfusca an d C. pachystachya onto first two principal component axes...... 115
14. P rojection of group ellipses of C.subfusca onto first two principal component axes. S = type specimens of C.subfusca', T = type specimens o f C. teneraeformis...... 117
15. Projection of ’typical’ group ellipses of C. pachystachya onto first two principal component axes...... 119
16. P rojection of group ellipses of C.pachystachya onto first two principal component axes. OTU’s of known chromosome number are shown as #: n = 41; A : n = 39; ■: w = 38; O: « = 37 ...... 121
17. D istrib u tio n o f C. integra...... 123
18. D istrib u tio n o f C. abrupta...... 125
19. D istrib u tio n o f C. subbracteata ssp. subbracteata ...... 127
20. Distribution of C. subbracteata ssp. gracilior...... 129
21. Distribution of C. harfordii...... 131
22. Distribution of C. m ariposana...... 133
23. Distribution of C. subfusca...... 135
24. Distribution of C. pachystachya ssp. pachystachya ...... 137
25. Distribution of C. pachystachya ssp. com pacta...... 139
X INTRODUCTION
Carex is considered a taxonomically difficult genus, partly as a result of its size (1100 to 2000 species, depending on author), and of the slight morphological distinctions among species. Although recent taxonomic studies have generally led to a reduction in the number of species, there is no denying the great diversity present in the genus. Large phylads have been interpreted as indicating ancient lineages, or, rapid rates of evolution. The fossil record for
Carex or Carex-like plants is sparse, with definite records dating from the late
Miocene for North America. This evidence, taken with the fact that many extant species are morphologically very similar, argues the situation inCarex appears to be one of recent and/or rapid evolutionary divergence.
One feature of the genus suggested as a factor in its evolution is agmatoploidy. Agmatoploidy is a cytological condition present in the Cyperaceae,
Juncaceae and certain insect groups and is characterized by chromosomes lacking localized centromeres. Centromeric activity appears to be located on many parts of the chromosome (polycentry) confering the ability of chromosome fragments to migrate towards the cellular poles during mitosis and meiosis. Since chromosomal fission and fusion do not seriously hamper cell division, cells still possess a complete genome after division. This cytological condition has probably resulted in the aneuploid series present inCarex, with haploid numbers ranging from 6 to
56 with every number from 12 to 43 represented. Workers in the genus have
1 hypothesized that agirîJfoploidy has played an important part in the evolution of genome in Carex by imparting a structural flexibility on the karyotype in which linkage groups may easily be rearranged.
With an appreciation that the intergrating morphologies of species in the genus may represent recent divergence, and agmatoploidy may play a role in this divergence, we are better prepared to understand the pattern of relationships among extant taxa as well as the process involved in producing the pattern. One of the largest sections of Carex in North America, the Ovales, appears to be a prime example of recent evolutionary diversification. The Ovales is distinguished by a cespitose growth form, gynaecandrous spikes (pistillate flowers distal), evidently winged perigynia, and lenticular achenes. Within the section are a number of species groups (aggregates, complexes), one of which is the C. pachystachya Cham, ex Steud. complex. This group of species in the Festivae assemblage, is readily distinguished by characteristically plump or plano-convex perigynia. Eleven species, one variety and one seggregate comprise the complex, with a number of these having been noted for apparant intergradation and affinities to C. pachystachya.
As indicated for the genus, problems in the C. pachystachya complex center
on morphological and cytological variation which appears to be a result of recent
evolution. Three sets of investigations were initiated to study the morphological
variation within the complex and see if correlates existed with chromosome and
genetic divergence. The first step was an investigation into the degree of
relatedness of the chromosome races of C. pachystachya utilizing meiotic pairing patterns and pollen stainabilities in F, hybrids (Chapter 1). The second step was an investigation of genetic variation within and among populations and species of the complex using enzyme electrophoresis (Chapter 2). The data obtained from these studies was finally combined with a multivariate statistical analysis of
morphological variation in the complex to provide insights into the evolutionary history and contribute to a revision of the complex (Chapter 3). CHAPTER I
MEIOTIC PAIRING PATTERNS IN HYBRIDS AMONG CHROMOSOME RACES
O F CAREX PACHYSTACHYA, THE RELATED SPECIES C. MACLOVIANA AND
C. PRESLII, AND THE RELEVANCE TO AGMATOPLOID EVOLUTION
T he genus C arex is noted for its long aneuploid chromosome series (Davies
1956; Heilborn 1924, 1932; Tanaka 1949) with haploid numbers ranging from 6 to
56 (Davies 1956). A number of explanations have been proposed to account for
the aneuploid series, including quantitative aneuploidy (Heilborn 1924, 1932,
1939; Schmid 1982; Tanaka I940a,b, 1941,1949), secondarily balanced polyploids
(Tanaka 1940, 1949; Wahl 1940), structural rearrangements (Faulkner 1972;
Schmid 1982), and hybridization with segregation of aneuploid numbers (Heilborn
1924, 1939). The most recurrent hypothesis is qualitative aneuploidy, that is,
change in chromosome number without duplication or loss of chromatin. The
presence of polycentric chromosomes is thought to be most conducive to this
hypothesis by allowing fragments to maintain centromeric activity and not
become lost during cell division (Lima-de-Faria 1949). This process of chromsome
fragmentation and fusion is specifically labeled agmatoploidy (Malherios-Garde
and Garde 1951). Distinguishing which of these alternative hypotheses plays a
major role in agmatoploid evolution in the genus is difficult. In reality, no one
process may be of primary importance, and several probably interact in the genus (Cayouett and Morisset 1986a; Schmid 1982), yet confirmation for any of the processes is equivocal. Direct evidence for the natural occurrence of agmatoploidy in C arex can be derived from artificial hybridizations among different aneuploid numbers within a species. As was shown in experimental studies in Eleocharis (Hakansson 1954) and Luzula in the Juncaceae (Nordenskiold
1963), a strict agmatoploid change results in one large chromosome pairing with
two small chromosomes in a hybrid between plants with two different numbers.
Pairing relationships have been documented in the genus, but from either natural
or artificial interspecific hybrids (Cayouett and Morisset 1985, 1986a,b; Davies
1955; Faulkner 1972; Schmid 1982). In these instances, the effect of agmatoploidy
on meiotic pairing can be complicated by differences that may result from
divergence through spéciation.
A single study has examined intraspecific hybrids in the species C.
oxyandra, C. gibba and C. parciflora (Tanaka 1949). The complex pairing patterns
observed in the hybrids led Tanaka to conclude that a combination of
quantitative aneuploidy and structural rearrangements were responsible for the
aneuploid numbers in C. oxyandra and C. gibba, and that secondarily balanced
allopolyploidy for C.parciflora. Agmatoploidy was not considered a feasible
pathway because of the observed complex pairing relationships.
This study continues with the approach of Tanaka (1949) using another
species with an aneuploid number series. Meiotic associations were studied in
crosses among four chromosome races of C. pachystachya Cham , ex Steud. (« = 37,
38, 39 and 41). These chromosome races are morphologically indistinguishable,
(except for the « = 41 race; Whitkus and Packer 1984), genetically similar as
measured by isozyme analysis (Whitkus 1985, unpublished), and cross-compatible
(Whitkus 1988). If agmatoploidy alone were responsible for the aneuploid numbers in this species, then the expected pairing at metaphase I should be a single large chromosome paired with two smaller chromosomes in crosses between plants with consecutive numbers. Alternatively, if more complex pairing patterns are found such as those seen in interspecificC arex hybrids, then this would argue that chromosome number evolution is not coupled with spéciation in the genus as suggested by Heilborn (1924), Faulkner (1972) and Schmid (1982). To see if additional effects on chromosome pairing may be due to species differences, crosses among C. pachystachya and two closely related species, C. macloviana
D’Urv. (n = 43) and the « = 40 race of C. preslii Steud. were included.
MATERIALS AND METHODS
Seeds of Fj hybrids among the various cytological races and species were produced in crossing experiments described in Whitkus (1988). Seeds (in reality, the achenes and enveloping perigynia) were stored for two month's at 0° C, sown on moist, standard potting soil in 17.0 X 12.5 cm flats, and germinated under ambient light and temperature in the greenhouses at Ohio State University. After establishment, the F | seedlings were transplanted individually into 10 cm diameter plastic pots. After a winter cold period, inflorescences were produced in April. The best time to collect material for meiotic activity is in the early morning and inflorescences just emergent from the sheath of the fertile culm show greatest meiotic activity. These were fixed in a 6:3:2 methanol: chloroform: propionic acid mixture and stored at -20 °C. Anthers were dissected from the inflorescences and squashes made with 2% lactic acetic orcein (Cooperrider and
Morrison 1967).
Since several pairing patterns were commonly observed in each cross, a single pattern needed to be derived for a cross to compare meiotic pairing patterns among crosses. The method of inferring a single pattern is based on the fact that multivalents can break into simpler combinations. Thus a quadrivalent can appear as a trivalent and univalent, two bivalents, a bivalent and two univalents, or four univalents (Sybenga 1975). Cayouett and Morisset (1985,
1986a,b) have used such an approach called maximum affinity pairing (MAP) formulae. Their method of producing MAP formulae can result in a number of complications. Several equally possible formulae can be derived from a series of pairing patterns, and MAP formulae may require inferring the presence of multivalents not actually observed in any of the cells. For the present study the
MAP approach has been modified to account for only observed pairing patterns and to accommodate the more complex patterns observed. The result is a maximum observed affinity pairing (MOAP) formula an example of which is given in Appendix B.
To determine pollen stainability, five or more unopened anthers per plant were collected in the morning and placed in 1.5 ml microcentrifuge tubes. Five drops of 2% lactophenol analine blue were added to each tube, and the stain allowed to be absorbed for at least one week. Anthers were removed, broken on glass slides and four hundred pollen grains counted. All meiotic observations, pollen counts and photographs were made with a Zeiss Standard microscope using phase contrast optics at 1000 X magnification.
RESULTS
Meiotic pairing was observed best at diakinesis or metaphase I. The small
(2.1-3.7 wm for bivalents) and numerous chromosomes prevented observation at pachytene, the most informative stage for determining pairing relationships
(Jackson 1984). 8
A range of configurations was observed. Complete sets of bivalents were seen only in crosses within the races of C. pachystachya, and within C. macloviana and C. preslii (Table 1). More complex patterns were observed in interracial and interspecific crosses with univalents, trivalents, quadrivalents, pentivalents and sextivalents present. Bivalents and multivalents were both homomorphic and heteromorphic, with heteromorphic pairing seen in every cross. In all interracial and interspecific crosses, and in several intraracial crosses, inferred MOAP formulae require the presence of multivalents (Table 1). As no evidence of bridges were observed during anaphase, all quadrivalents, pentivalents and sextivalents in the MOAP formulae were assumed to be a result of reciprocal translocations with a quadrivalent counted as a single translocation event, a pentivalent as a single translocation in conjunction with a fragmentation, and a sextivalent as resulting from a minimum of two translocations among three chromosomes. The minimum number of translocations needed to account for each
MOAP formula are shown in Table 1. These range from a single translocation in the intraracial cr 110 to ten in cr 24. Figure 1 presents the maximum and minimum number of translocations necessary to account for the MOAP formulae.
W ithin C. pachystachya, the n = 37, 38, and 39 races share fewer structural differences than those observed between these races and the n = 41 race. The n =
41 race exhibits three translocation differences with C. macloviana, th e sam e number by which C. macloviana and C. preslii differ. No interspecific differences in the number of translocations were detected since interspecific crosses result in fewer differences than crosses among some of the races of C. pachystachya.
A range of pollen stainabilities was also found (Table 2). Stainabilities of selfed plants did not differ from intraracial crosses in C. pachystachya or intraspecific crosses in C. malcoviana an d C. preslii (P > 0.1, Wiîcoxon two sample test). Selfing data were then combined with intraracial and intraspecific crosses and averages within and among the races and species calculated (Fig. 2). Overall, highest stainability was found in the crosses within races of C. pachystachya and w ithin C. macloviana and C. preslii, although the n = 38 race has an average below
50%. The Wilcoxon two sample test indicates, however, that intraracial and intraspecific crosses result in higher average stainabilities than interracial and interspecific crosses (P < 0.05). Interspecific crosses generally result in stainability below 50%. Two exceptions are the 37 X 43 and 40 X 41 crosses.
The 40 X 43 cross is also relatively high.
The pollen stainabilities do not appear to reflect the number of inferred translocation differences. However, the data show concordance when tested by
Spearman rank correlation. Pollen stainability is highly correlated (P < 0.01, rho
= -0.700 for high numbers, rho = -0.702 for low numbers, n = 16) with both the high and low number of inferred translocation differences. Although the theoretical concordance between the two data sets in not obtained, the significant correlation satisifies the assumption that increasing the number of structural rearrangements results in decreasing pollen stainabilities.
DISCUSSION
The pairing patterns and pollen stainabilities obtained from this study
indicate that the aneuploid races of C.pachystachya have differentiated in both
chromosome number and structural arrangement. The least amount of
differentiation is seen between the n = 38 and 39 races, and the greatest between
the « = 41 race and all other races. There is no increase in the degree of
chromosome differentiation when the races of C. pachystachya are compared to C.
macloviana or C. preslii. The low amount of morphological differentiation 10
(Whitkus and Packer 1984), high genetic identity as measured by isozymes
(Whitkus 1985, Chapter 2), and the cross compatibilities among the races and species (Whitkus 1988) indicate the races, as well as the species, have undergone
recent evolutionary divergence. The data from this study demonstrate that this divergence has been accompanied by numerous chromosomal structural rearrangements. A similar pattern of rapid chromosomal evolution with little to
no morphological or isozyme divergence has been observed in annual species of
Clarkia (Lewis 1953), Caura (Gottlieb and Pilz 1976) and Stephanomeria (Gottlieb
1973). The expected pattern in herbaceous perennials (excluding island groups) is
gradual divergence of all features such as in the generaLayia (Warwick and
G ottlieb 1985) or Coreopsis (Crawford and Bayer 1981; Craw ford and Whitkus
1988). The pattern of evolution seen in C. pachystachya and its related species
raises the possibility that evolution in perennials can also follow the pathway of
rapid chromosomal evolution as seen in annuals with divergence in other
characters following reproductive isolation via chromosomal repatterning.
There are no data from this or previous studies that other proposed modes
of chromosome evolution for the genus have occurred in this group. Polyploidy,
including auto- and allopolyploidy and their various consequences, is not
considered a factor in the section Ovales as haploid numbers range from 26 to 45
(Fedorov 1969). Hybridization with segregation of aneuploid numbers in also
not a likely explanation for this pattern because the plants are known to be self
compatible and likely not cross-breeding in nature. In over 3000 sheets of
specimens of the complex observed by the author, none appear morphologically of
hybrid nature, nor show reduced seed set that was found in artificial,
interspecific hybridizations (Whitkus 1988). 11
A question arises as to whether the aneuploid numbers have resulted from quantitative or qualitative aneuploidy. Quantitative aneuploidy is assumed when homomorphic pairing patterns are found, especially among trivalents, or when unpaired univalents are present (Faulkner 1972; Schmid 1982). With the abundant evidence of structural rearrangements, the chance that the homomorphic trivalvents in the present data represent decomposed quadrivalents or other multivalents is quite possible. Trivalents are removed from many MOAP formulae if univalents are available and the two construed to be decomposed quadrivalents. This would be the case if MAP formulae were used instead of
MOAP formulae. This still leaves eleven crosses with unresolved trivalents (Table
3). Decomposed multivalents are suspect for a number of these crosses since other crosses involving these races lack trivalents. Unresolved trivalents are then left in the crosses involving the 41 and 39 races of C. pachystachya, and the cross of
C. preslii with the 37 race of C. pachystachya. Since these crosses contain
heteromorphic trivalents, they may well be the expected trivalent association of
agmatoploid evolution.
Heteromorphic pairing patterns seen in every cross are strong evidence for
agmatoploid evolution. However, simple, heteromorphic trivalent associations are
not as prévalant as heteromorphic quadrivalents. When the structural
rearrangements via translocations are considered, this pairing pattern suggests a
model of chromosomal evolution for this group. For both centric and polycentric
chromosomes, structural rearrangements necessitate a break. This fragmentation
is also a requirement for agmatoploid evolution. A parsimonious solution should
explain how both types of fragmentation can occur with a single mechanism.
Current evidence suggests that during zygotene end pachytene, synapsis
and chromosome condensation can result in non-homologous chromosomes 12
becoming interlocked (Wettstein et al. 1984; see Fig. 3a). These interlocks are
resolved by breaks in the chromosomes (Fig 3a), and errors in their resolution are
thought to be the most important factor in inducing structural rearrangements
(Imai et al. 1986). There are a number of ways in which resolution can occur.
Homologous fragments can rejoin, resulting in no change in chromosome structure. Alternatively, non-homologous fragments can combine, resulting in a
reciprocal translocation (Fig. 3a). These are the only viable alternatives for
plants with centric chromosomes. For plants with polycentric chromosomes,
additional options are available for the resolution of interlocked chromosomes.
Fragments may fail to rejoin, leading to one chromosome breaking into two fragments and the other reforming a single chromosome. Alternatively, both chromosomes may persist as four fragments; or a translocation and two fragments
(Fig. 3b). With a translocation and two fragments, the result is the same as a reciprocal translocation, a classic translocation heterozygote is seen at metaphase I
(Fig. 4). A translocation of this nature reduces fertility. Half of the remaining
viable gametes will be of the original type and the other half will carry the new
structural type, which includes an increase in haploid chromosome number (Fig.
4). Low initial numbers of individuals within a population containing the
rearrangement, and reduced fertility of heterozygotes, makes it difficult for the
rearangement to become fixed in homozygous form. However, as selfing provides
one means for the new structural type to become fixed (Hedrick 1981; Lande
1979, 1984), the self compatible nature of C. pachystachya opens the way for the
fixation of new structural types.
This hypothesis may be extended to other species of Carex. There is ample
evidence for the occurrence of structural rearrangements from natural and
artificial hybrids in Carex (see: Cayouette and Morisset 1985, 1986a; Davies 1955; 13
Faulkner 1972: Schmid 1982: Tanaka 1949). Chromosomal rearrangements are also known within members of a species (see: Cayouette and Morisset 1986a,b;
Faulkner 1972; Hoshino 1981; Schmid 1982; Standley 1985; Tanaka 1941; Wahl
1940). In addition, self compatibility has been demonstrated in the majority of the studies that have examined breeding systems in the genus (see: Faulkner 1973;
Handel 1976, 1978; Pojar 1974; Schmid 1982; Standley 1985; Vonk 1979). It thus appears that instances exist where conditions permit agmatoploid change to be accomplished by the mechanism presented here. This does not rule out other modes of chromosome number change in Carex, it merely demonstrates that the characteristic long aneuploid series of the genus could arise by a variety of mechanisms.
Further work is needed to test the present hypothesis. Low chromosome number species in Carex with intraspecific aneuploidy must be explored so that unambiguous pairing affinities can be assigned in artifically produced hybids. If the hypothesis is found to hold in other species, then it would be demonstrated that agmatoploid evolution is not be a unique event, but simply an alternate pathway open to plants with polycentric chromsomes. The mechanism by which it occurs being the same as in any plant group undergoing structural chromosomal evolution. 14
Table 1. Observed pairing patterns from hybrids of crosses among races of C. pachystachya {n = 37, 38, 39, 41), C. macloviana (n = 43) and C. preslii (n = 40), MOAP formulae and estimated number of translocations (ENT).
Cross Confieuration Number MOAPENT cr 59 37 X 37 37II 10 37II cr 110 37 X 37 35II IIV 8 35II IIV 1 cr 111 37 X 37 37 II 7 37II cr 93 37 X 38 31II 2IV IV 3 31II 2IV IV 3 cr 19 37 X 39 31II 2IV IVI 1 29II 31V IVI 5 11 29II 31V IV 1 29II 31V IVI 1 cr 76 37 X 39 21 35II IIV 10 36II IIV 1 cr 37 37 X 41 21 22II 3III 3IV IV IVI 2 11 20II 2III 5IV IV IVI 8 31 26II 4III IV IVI 1 41 19II 2III 5IV 2V 1 31 24II 3III 3IV IVI 1 cr 41 38 X 38 38II 12 38II cr 48 38 X 39 35II IIII IIV 13 33II IIII 2IV 2 33II IIII 2IV 1 cr 51 38 X 39 11 34II 21V 15 11 34II 2IV 2 37II IIII 3 cr 8 38 X 40 151 17II 2III 2IV 3V 1 151 17II 2III 21V 3V 5 cr 89 39 X 39 39II 11 39II cr 109 39 X 37 21 35II IIV 6 34II 2IV 2 21 33II 2IV 12 cr 34 39 X 38 35II IIII IIV 15 35II IIII IIV 1 cr 47 39 X 38 35II IIII IIV 8 35II IIII IIV 1 37II IIII 1 cr 95 39 X 38 11 36II IIV 1 11 34II 2IV 2 11 34II 2IV 19 cr 102 39 X 38 11 36II IIV 11 3511 IIII IIV 1 35II IIII IIV 3 15
Table 1 (continued) cr 66 39 X 41 34II IIII IIV IV 5 34II nil IIV IV cr 29 41 X 41 39II IIV 7 39II IIV cr 49 41 X 41 39II IIV 19 39II IIV 41II 3 II 39II IIII cr 70 41 X 37 51 24II IIII 4IV IVI 21II IIII 3IV 3V IVI 61 21II IIII 4IV IV IVI 41 19II IIII 3IV 3V IVI 61 22II 2III 4IV IVI cr 79 41 X 37 11 35II IIII IIV 35II 2IV 2 35II 2IV cr 24 41 X 38 91 15II IIII 5IV IV 2VI 21II 5IV IV 2VI 10 71 20II IIII 3IV IV 2VI 61 20II 2III 4IV IV IVI 61 20II 3III 2IV 2V IVI 51 21II 3III 3IV IV IVI cr 74 41 X 39 81 25II IIII 2IV IV IVI II 25II 2III 3IV IV IVI 51 26II 2III 3IV IV 91 25II 3III 3IV 41 2511 2III 5IV cr 80 41 X 39 11 34II 2III IV II 34II 2III IV 21 35II nil IV
cr 5 41 X 43 21 37II 2IV 3511 2I1Ï 2IV 51 34II nil 2IV 21 34II 2III 2IV
cr 26 41 X 43 51 34II nil 2IV 21 34II 2III 2!V 61 34II 2III IIV 21 34II 2III 2ÎV'
cr 32 43 X 41 61 33II 3IV 36II 3IV 38II 2IV 41 36II 2IV
cr 9 40 X 37 41 18II 9III IIV IVI 20II 9III IIV IVI 71 20II 7III IIV IV 61 20II 9III IIV
cr 11 40 X 38 131 191! 3III 2ÏV 2V 91 20II 2IIÏ 3IV IV IVI 91 20II 2III 3IV IV IVI
cr 2 40 X 43 281 16II 4III IV IVI 141 23II 4III IV IVI 181 23II 5III IIV 16
Table 2. Percent pollen stainabilities of hybrids among crosses of races of C. pachystachya (n = 37, 38, 39, 41), C. malcoviana (« = 43) and C, preslii (« = 40).
Cross/ID % Cross/ID %, 37 X 37 41 X 41 cr 59 96.0 cr 27 99.8 cr 110 98.0 cr 28 1.6 cr 111 88.4 cr 29 89.8 cr 30 93.0 37 X 38 cr 49 93.8 cr 93 12.4 cr 57 92.2
37 X 39 41 X 37 cr 19 21.4 cr 70 13.9 cr 52 27.4 cr 79 69.2 cr 76 18.2 cr 99 22.4 41 X 38 cr 104 15.4 cr 24 7.2
37 X 41 41 X 39 cr 37 98.0 cr 21 5.8 cr 78 89.6 cr 67 54.8 cr 74 6.6 38 X 38 cr 80 16.2 cr 41 17.6 cr 42 82.0 41 X 40 cr 14 87.4 38 X 37 cr 94 21.8 41 X 43 cr 103 42.2 cr 5 6.4 cr 26 32.0 38 X 39 cr 48 87.2 40 X 40 cr 51 98.4 cr 18 97.1 cr 96 60.4 40 X 37 38 X 41 cr 9 2.2 cr 23 2.8 40 X 38 38 X 40 cr 7 0.0 cr 8 3.6 cr 11 3.0 cr 10 49.5 40 X 41 38 X 43 cr 4 89.4 cr 12 3.4 17
Table 2 (continued)
40 X 43 39 X 39 cr 2 0.2 cr 63 92.2 43 X 37 cr 65 87.8 cr 44 97.2 cr 72 76.6 cr 88 94.6 43 X 41 cr 89 94.2 cr 32 21.8 cr 90 87.4 cr 91 93.4 43 X 40 cr 98 90.6 cr 3 96.9
39 X 37 37 self 45.4 cr 20 8.4 cr 45 94.6 39 self 90.3 cr 53 25.4 cr 100 16.4 cr 109 90.4 41 self 97.4
39 X 38 cr 34 93.2 40 self 88.1 cr 47 89.0 cr 95 78.8 cr 102 89.2 43 self 88.6
39 X 41 cr 22 5.8 cr 55 84.4 cr 66 8.0 cr 75 3.4 18
Table 3. Crosses among races of C. pachystachya (n = 37, 38, 39, 41) C. macloviana {n = 43) and C. preslii (n = 40) with unresolved trivalents.
Cross Without Cross/ID Trivalents Trivalents 37 X 41 cr 37 1 41 X 37 cr 70 1 cr 79
38 X 39 cr 48 1 cr 51 39 X 38 cr 34 1 cr 47 1 cr 95 cr 102 1
39 X 41 cr 66 1 41 X 39 cr 74 1 cr 80 1
41 X 43 cr 5 2 cr 26
40 X 37 cr 9 9 Figure 1. Number of translocations necessary to account for observed pairing patterns at metaphase I. Numerator is the largest number, denominator the smallest. Single number indicates only value
19 PRESLII MACLOVIANA
4 0 43
6/6 3/2
8/2 37
1/0 5/1
6/1
38 39 2/1
PACHYSTACHYA
NI Figure 1 O Figure 2. Average pollen stainabilities within and among races and species
21 PRESLII MACLOVIANA
40 64.7 43 92.6 88.6
88.4
2.2 20.1
97.2
37 56.9
82.0 4.0 83.1
25.5 21.1 23.1 '3.4 5.0 38 39 85.2 49.8 89.7 PACHYSTACHYA N) Figure 2 K) Figure 3. Diagramatic representation of resolution of interlocks between non-homologous chromosomes during prophase. A: Events in plant with localized centromeres resulting in reciprocal translocation. B: Events in plant with unlocalized centromeres resulting in a translocation and two fragments
23 B
Figure 3 Interlock break resolution Figure 4. Metaphase I configuration of plant with unlocalized centromeres from resolution of interlocked non-homologous chromosomes resulting in a translocation and two fragments, subsequent viable gametes, and chromosome complement in siblings
25 metaphase I
I I n:2
/
I viable gametes I c II ) selfing or Inbreeding
n;2 n:3 W Figure» 4 O n CHAPTER II
ALLOZYME VARIATION WITHIN THE CAREX PACHYSTACHYA COM PLEX
(CYPERACEAE)
T he Carex pachystachya Cham, ex Steud. complex consists of eleven species that have been noted for complex morphological variation within species and apparant intergradation among the species. As in most other members of section
Ovales to which the C. pachystachya complex belongs, the morphological variation lies in quantitative features of the inflorescence and perigynium, making the species difficult to distinguish. The complex is one part of the C.macloviana aggregate (Whitkus and Packer 1984) which is thought to have arisen recently, thus obstensibly making C. pachystachya and its derivatives fairly recent species.
The perplexing morphological variation among species may thus be attributed to this recent origin. Since modern taxonomic decisions on limits of species attempt to reflect actual genetic differences, it would be advantageous to know if the morphological differences among the species reflect a greater amount of genetic differentiation than found within species. Information on levels of genetic variation within species would also be favorable in discussing evolutionary processes that may have contributed to the present taxonomic situation.
Since many quantitative characters have a complex genetic basis, allozyme analysis can provide an alternative estimate of genetic variation. This is possible
27 28 because of the known simple genetic basis of isozyme variants (Crawford 1983,
1985; Gottlieb 1977, 1981b) and relative ease of obtaining the inform ation.
Reviews by Gottlieb (1981) and Crawford (1983) have shown that the level of genetic differentiation increases when moving from infraspecific populations to congeneric species. Additionally, Gottlieb (1977, 1981b) and Crawford (1985) have shown how data from allozyme studies can be useful in inferring ideas about evolutionary processes such as primary spéciation and hybridization, while others (Hamrick et al. 1979; Loveless and Hamrick 1984) have related isozyme data to various populations paramaters in plants.
The purpose of the present study was to conduct an electrophoretic survey of allozyme variation in the C. pachystachya complex to 1) determine levels of genetic variation within and among populations of several species; 2) determine levels of genetic divergence among the species, and; 3) use the data to infer some aspects of the evolutionary history of the complex.
MATERIALS AND METHODS
A total of 1194 progeny from 67 populations were assayed (Table 1). From each population, seed was collected from one to thirty individuals and the progeny grown in the greenhouses at Ohio State University. Six species from the complex are represented in the study, as well as one populations each ofC. preslii and a bulked collection of C. macloviana. The latter two species are closely related but not members of the C. pachystachya complex and were included to provide an estimate of the amount of genetic variation between the C. pachystachya complex and other related species. Additionally, populations of the four known chromosome races of C. pachystachya were included (Whitkus and
Packer 1984). A population of C. pachystachya was also included which has a 29
chromosome number of m = 40 (unpublished). As it is presently known from only
a single population, it is not considered a chromosome race.
Enzymes were extracted from leaves, culms and roots of young progeny
that had grown until they had at least three expanded leaves, in Gottlieb’s (1981a) extracting buffer. Samples were centrifuged for 2 minutes and the supernatant
blotted onto paper wicks for application to starch gels. Samples were run on 12%
starch gels using two buffer systems. The lithium borate system (pH 8.3; Gottlieb
1981a) was used to resolve the enzymes leucine aminopeptidase (LAP), aspartate
aminotransferase (AAT), alcohol dehydrogenase (ADH), glutamate dehydrogenase
(GDH), esterase (EST), phosphoglucose isomerase (PGI), glucose-6 phosphate
dehydrogenase (G-6PGD), and aldolase (ALD). A histidine citrate system (pH 6.5,
Cardy et al. 1981) was used to resolve the enzymes malate dehydrogenase (MDH),
shikimate dehydrogenase (SKDH), isocitrate dehydrogenase (IDH), 6-
phosphogluconate dehydrogenase (6-PGD), phosphoglucomutase (PGM) and
superoxide dismutase (SOD).
Gels using the lithium borate system were run at 75 millamps and those
using the histidine citrate system at 250 volts. Gels were run until a bromphenol
blue marker reached the anodal end of the gel. Staining protocols followed
Cardy et al. (1981) for tray stained systems (LAP, AAT, ADH, GDH, EST, G-
6PGD, MDH) and Soltis et al. (1983) for those using the agarose overlay procedure
(PGI, ALD, SKDH, IDH, 6-PGD, PGM). The system SOD was scored on any
dehydrogenase system on which it was best developed.
Genetic interpretation of the banding patterns were inferred from the
known isozyme number for diploid plants and the active subunit structure for
these enzymes (Crawford 1983; Gottlieb 1981b, 1982), and from obtaining the
expected additive banding patterns in F | hybrids between different allelic types. 30
Allelic frequencies were determined for each population. These were used to calculate mean number of alleles per locus (k), mean number of alleles per polymorphic locus (kp) where a locus was considered polymorphic if the most common allele occurred at a frequency of 0.99 or less, and expected heterozygosities (HET). Observed heterozygosities were obtained by direct count of individuals. Additional statistics were the unbiased genetic identity for all pair-wise population comparisions (Nei 1978) and unbiased gene diversity statistics (Nei and Chesser 1983), calculated using the GENESTAT2 program
(Whitkus 1988a).
RESULTS
Fourteen enzyme systems, presumably encoded by 21 loci, were scored.
The number of genes obtained for each enzyme were: LAP, three; IDH, two;
MDH, three; ADH, two; AAT, two; EST, one; 6-PGD, one; PGM, one; PGI, one;
GDH, one; SOD, one; G-6PD, one; SKDH, one; ALD, one. At least two additional genes were expressed in EST, but the highly variable nature of the enzyme, both in activity and probable number of loci, precluded interpretation of the banding patterns. Confidence in the EST locus used was gained by obtaining the expected two banded pattern in the Fj hybrid between two different electromorphs of a presumably monomeric gene.
Eighteen of the 21 loci examined were polymorphic. Allelic frequencies for all populations and the number of progeny examined per locus in each population are presented in Appendix C. Nearly half (31 of 65) of the populations in this complex are polymorphic at one or more loci, the rest are fixed for an allele at all loci, even though the locus may be polymorphic in the complex. A total of 13 alleles are unique to species in the complex, with an 31 additional unique allele found in C.preslii. Ten of the unique alleles are found in C. pachystachya, with six present at frequencies of less than 0.01. Each of the remaining three unique alleles is found in C.integra, C. abrupta an d C. subbracteata.
The mean number of alleles per locus is 1.5 or less, and 2.8 or less for polymorphic loci in the polymorphic populations (Table 2). The percent polymorphic loci indicate relatively moderate values of generally less than 33%, although values of 40% (ABRU 3971) and 45% (PACH 2440) are obtained.
Observed heterozygosity is zero in most populations and in all cases save one, are lower than expected. The exception is INTE 4097 which results from a single polymorphic locus (MDH-1) being at or near equilibrium.
Total genetic diversity (H j) ranges from 0.019 to 0.170 for taxa with two or more populations (Table 3). In all cases where two or more populations were surveyed, the average within populaion genetic diversity (Hg) is much less than the total diversity. This results in average gene diversity among populations
(Dgj) representing a large amount of the total diversity, and thus gives high values of the coefficient of genetic differentiation (Ggj). Only C. integra and C. abrupta have a G gj value of 0.5 or less (Table 3).
Genetic indentity (I) values among the taxa (Table 4) appear evenly distributed, ranging from 0.745 (SUBF X SUBB) to 0.946 (ABRU X HARF). The range of pair-wise population comparisons (Table 5) shows values as low as 0.354 to as high as 1.00. Within each taxon, mean identity values are generally above
0.84, with C. subfusca being the exception at 0.730 (Table 6). Like the interspecific pair-wise population comparisons, intraspecific pair-wise population comparisons can be quite low (0.632), values usually associated with differences among species (Crawford 1983, Gottlieb 1981b). 32
DISCUSSION
The low levels of genetic variation obtained in this study, along with the high amount of genetic differentiation among populations agree with the values reported in the literature for predominately self-pollinating species. Gottlieb
(1981b) reports a value of 18.3% polymorphic loci for self pollinating species versus 51% for outcrossers. Similar values are given by Hamrick et al. (1979).
Total genetic diversity (Hj) obtained for members of the C. pachystachya complex, however, is much lower than the value of 0.291 reported by Loveless and Hamrick (1984) for autogamous species, and is even lower than the value of
0.251 given for outcrossing species. In contrast, the coefficients of genetic differentiation obtained for the C. pachystachya complex are much higher than the value of 0.523 given for autogamous species by Loveless and Hamrick (1984).
Thus the diversity statistics indicate that members of the C. pachystachya complex are likely to be selfing in nature, but as a whole, carry much less genetic variation than has been reported previously for autogamous species.
The identity values obtained in this study are also generally lower than reported for selfing species. Gottlieb (1981b) and Crawford (1983) report values of 0.90 or more for pair-wise population comparisons within species. The average identity values in Table 6 are all lower than 0.931 for the species. The range of identities within each species is quite large, with low values similar to what
Gottlieb (1981b) and Crawford (1983) report for intraspecific comparisons. The lowered identity values between populations within the species are not unprecedented. Roberts (1983) reported intraspecific identity values as low as
0.688 for the selfing Bidens discoidea. Hamrick and Allard (1972) reported identity values as low as 0.451 between populations ofAvena barbata. A separate 33 analysis of the data given by Crawford and Wilson (1979) on several diploid species of Chenopodium results in intraspecific identities of 0.401 for C. leptophyllum, 0.486 for C. atrovirens, 0.500 for C. hians, and 0.429 for C.incognitum, while other species have values above 0.750 (Whitkus and Crawford, unpublished).
Thus the low identity values obtained for the C, pachystachya complex are not unknown for other selfing species, and comparisons with mean values in survey papers can be misleading.
Two factors can account for the pattern of genetic variation found this study. The first and probably overriding factor is self pollination in populations.
Aside from the genetic statistics from this study agreeing with other studies on selfing species, data from controlled crosses also point to the autogamous nature of the plants. Whitkus (1988b) has shown that self pollinated plants in all races of C. pachystachya result in higher seed set and germination, on average, than outcrossed plants. Yet the crossing data and the isozyme data are at variance with the fact that all members of Carex are wind pollinated, a mechanism generally thought to promote outcrossing. In addition, species ofCarex, including members of the C. pachystachya complex, have some form of dichogamy (Schmid
1982; Standley 1985; Whitkus 1988b). How then can these species be selfing in nature? First, all species of Carex thus far investigated are self-compatible
(Davies 1955; Faulkner 1973; Handel 1976, 1978; Pojar 1974; Schmid 1982;
Standley 1985; Vonk 1979; Whitkus 1988b). Second, although the plants are dichogamous, there is a large degree of overlap in maturation times of flowers within an inflorescence and among inflorescences on a plant (Vonk 1979; pers. obs.). Third, with the cespitose habit, a single culm will more than likely have as a nearest neighbor, another culm from the same plant, thus promoting geitonogamy. Only in rare instances will plants be close enough for culms to 34 intermingle and allow outcrossing. When the large difference in observed and expected heterozygosities from polymorphic populations are considered, the evidence strongly suggests that members of the C. pachystachya complex are autogamous in nature.
A second factor accounting for the results is the age of the complex.
Whitkus (1981) suggested that C. pachystachya differentiated from its parental complex (the C. macloviana aggregate) during the end of the Pleistocene at about
20,000 y.b.p. Since it is widely assumed that isozymes are generally neutral to selection (Hartl et al. 1985, 1986; Kimura 1983), their origin and maintance in populations is largely determined by drift (Kimura 1983). With the assumption that other factors remain essentially stable (mutation rate, population sizes, mating system) then the number of isozymic variants that accumulate between populations can roughly be attributed to divergence time. Indeed, Nei (1975) has suggested genetic distance (D = -In I) can be used to estimate this time. Yet in this study, low values of I among conspecific populations would argue for long periods of isolation. The paradox is resolved when overall measures of variability are used, i.e.; HET and H p The low values for these indicate little accumulated genetic variation, both within populations and species. The reason populations are highly differentiated from one another likely results from a lack of gene flow, not only among members of a population, but among populations as well. This would allow drift to predominate, fixing various populations for different alleles, as is found in most populations in this study. Lastly, the range of identities among the species are all above the mean of 0.67 reported by
Gottlieb (1981b) for conspecific species, indicating lower levels of divergence among the species, on average, than found in most other isozyme studies. 35
Additional evidence for a short divergence time is the number of unique alleles within the species. One unique allele is found in each of three species, and the remainder in C. pachystachya. The ten unique alleles in C.pachystachya likely come from the large number of populations sampled, thus increasing the chance of finding such alleles. Also, six of the ten alleles in C. pachystachya are in frequencies of less than 1%, indicating rare alleles that may have recently arisen, but may just as likely disappear via drift. Kimura (1983) points out that many populations carry such alleles at all times, only a few of which become higher in frequency or fixed. Thus seven unique alleles are distributed among four species, with the presumed ancestor of the ccmplc?,C. pachystachya, containing the majority of unique alleles. This is likely a reflection of its greater genetic variation probably due to being in existence for a longer period of time than any of its presumed derivative species.
Finally, a word about divergence among the chromosome races of C. pachystachya. Examination of Table 4 shows identity values among the ti = 37, 38, and 39 races are above 0.913. When the « = 41 race is added, the values drop to below 0.873, similar to what the races show when compared to other species in the complex, as well as the related species C. preslii an d C. macloviana. T hus the n =
41 race is as differentiated from the other races of C. pachystachya as are different species. Data from meiotic pairing patterns in Fj hybrids among the various races also shows the « = 41 race to be as different from the remaining races as C. macloviana and C. preslii. These data show that the « = 41 race of C. pachystachya is at least a biological species, although Whitkus and Packer (1984) could find no morphological evidence for this conclusion.
It is interesting to compare this study with the only other available electrophoretic study in Carex. Bruederle and Fairbrothers (1988) found in the C. 36 crinita complex total genetic diversity (H j) of 0.256, a coefficient of genetic differentiation of 0.658, and a range of identity values within taxa of 0.866 to
1.00, and between taxa of 0.66 to 1.00. In almost all of this statistics, this study differs with the results of the C. crinita complex. For the C. pachystachya complex, there is a higher amount of expected heterozygosity and thus a greater difference between the observed and expected. Total diversity is lower and genetic differentiation much higher. Finally, identity values both between populations within a species and between population from different species are lower. Thus much less genetic variation as measured by isozyme loci, is found in th e C. pachystachya complex in comparison to the C. crinita complex, but it is apportioned more between the populations rather than among individuals within populations as in C.crinita. Although members of both complexes are wind pollinated and predominately self-pollinated, there is a major difference. The C. crinita complex occurs in wetlands of eastern North America. These are generally more stable habitats than the serai habitats of western North America where m em bers o f th e C. pachystachya complex occur. Loveless and Hamrick (1984) indicate that plants of early successional stages have lower genetic diversity (H j) within populations and higher levels of genetic differentiation (Ggy) than do plants of later successional stages. The contrasting patterns of diversity between these two complexes, therefore agrees with the data from Loveless and Hamrick
(1984). 37
Table 4. Population designation (by collection number) and locality for populations of C.pachystachya complex, C. preslii and C. macloviana.
Population Locality C. preslii 2029 Rogers Pass, British Columbia C. macloviana Bulked seed from localities in Alaska and British Columbia C. pachystachya M = 40 1818 Stewart, British Columbia n = 37 1871 Vanderhoof, British Columbia 1879 58 km E of Vanderhoof, British Columbia « = 38 1911 28 km SB of Dawson Creek, British Columbia « = 39 1517 Moose Pass, Alaska 1540 Mile 38, Seward-Anchorage Hwy., Alaska 1787 200 km S of Dease Lake, British Columbia 1799 250 km S of Dease Lake, British Columbia 1826 73 km S of Meziadin Jt., British Columbia 1836 3 km E of South Hazelton, British Columbia 1884 75 km N of Prince George, British Columbia 1899 27 km N of McLeod Lake, British Columbia « = 41 1462 76 km S of Cantwell, Alaska 1491 16 km S of Portage, Alaska 1511 1 km N of Hope, Alaska 1682 Chilkat Pass, British Columbia 1847 Smithers area, British Columbia 1860 7 km E of Houston, British Columbia 1903 87 km NE of McLeod Lake, British Columbia 2023 Rogers Pass, British Columbia 2043 Rogers Pass, British Columbia « = ? 2152 Cameron Lake, Alberta 2159 Kootenay Summit, British Columbia 2166 Vermilion Range, Kootenay Natl. Park, British Columbia 2172 Mt. Robson Prov. Park, British Columbia 2192 24 km S of Princeton, British Columbia 2197 Agassiz, British Columbia 2259 Columbia Co., Oregon 2268 Tillamook Co., Oregon 2276 Benton Co., Oregon 2284 Benton Co., Oregon 2304 Curry Co., Oregon 2314 Curray Co., Oregon 2360 Klamath Co., Oregon 2379 Klamath Co., Oregon 2440 Linn Co., Oregon 2469 Clackamas Co., Oregon 2480 Skamanai Co., Washington 2500 White Pass, Lewis and Yakima Cos., Washington 2502 Lewis Co., Washington 2521 Yakima Co., Washington 2528 Yakima Co., Washington 2537 King Co., Washington 38
Table 4 (continued)
2557 Steven’s Pass, King and Chelan Cos., Washington 2616 Fend Oreille Co., Washington 3076 Mendocino Co., California C. subfusca 2343 Jackson Co., Oregon 2344 Jackson Co., Oregon 3905 Alpine Co., C alifornia 4418 T rinity Co., California C. integra 3191 Tulare Co., C alifornia 3805 Calaveras Co., California 3988 Eldorado Co., California 4097 Sierra Co., California C. abrupta 3784 Tuolumne Co., California 3891 Alpine Co., California 3971 Eldorado Co., California 4230 Sierra Co., California C. harfordii 2769 San Luis Obispo Co., California 2781 San Luis Obispo Co., California 2911 San Mateo Co., California 2918 M arin Co., California 2950 Sonoma Co., California 3074 Mendocino Co., California C. subbracteata 3035 Humboldt Co., California 39
Table 5. Measures of genetic variability in 31 polymorphic populations of the C. pachystachya complex and in one populations ofC. preslii.
Population k HET H P PRES 2028 1.1 2.0 0.10 0.000 0.040 40 1818 1.4 2.0 0.35 0.000 0.146 39 1787 1.1 2.0 0.05 0.000 0.010 41 1847 1.1 2.0 0.10 0.000 0.004 PAC 2152 1.2 2.0 0.18 0.000 0.003 PAC 2616 1.1 2.0 0.06 0.009 0.025 PAC 2304 1.2 2.0 0.17 0.003 0.056 PAC 2557 1.4 2.8 0.21 0.009 0.078 PAC 2379 1.3 2.2 0.28 0.021 0.135 PAC 2521 1.1 2.0 0.21 0.000 0.025 PAC 2502 1.2 2.0 0.21 0.000 0.063 PAC 2314 1.1 2.0 0.10 0.007 0.039 PAC 2197 1.3 2.0 0.31 0.000 0.137 PAC 2528 1.3 2.0 0.25 0.000 0.029 PAC 2360 1.2 2.5 0.12 0.002 0.074 PAC 2440 1.5 2.2 0.45 0.000 0.088 PAC 2469 1.1 2.0 0.13 0.003 0.038 PAC 2284 1.3 2.0 0.26 0.000 0.092 PAC 2276 1.3 2.0 0.32 0.000 0.081 SUBF 3905 1.1 2.0 0.11 0.000 0.011 SUBF 4418 1.1 2.0 0.11 0.000 0.011 INTE 3805 1.4 2.2 0.33 0.018 0.048 INTE 3191 1.1 2.0 0.11 0.000 0.033 INTE 4097 1.1 2.0 0.07 0.017 0.015 INTE 3988 1.3 2.3 0.27 0.039 0.093 ABRU 3971 1.4 2.0 0.40 0.000 0.081 ABRU 4230 1.4 2.0 0.36 0.000 0.098 ABRU 3891 1.3 2.0 0.27 0.000 0.076 HARF 2781 1.1 2.0 0.06 0.000 0.012 HARF 2950 1.2 2.0 0.17 0.000 0.056 HARF 2769 1.1 2.0 0.06 0.000 0.025 SUBB 3035 1.1 2.2 0.06 0.000 0.006
X 1.2 2.0 0.20 0.004 0.054 40
Table 6. Diversity statistics for C. pachystachya complex, C. preslii and C. macloviana. # Pod % % -SI ^ S I PRES 1 0.041 0.041 0.000 0.000 MAC 1 0.000 0.000 0.000 0.000 PACK n = 40 1 0.184 0.184 0.000 0.000 n = 31 2 0.000 0.024 0.024 1.000 n = 38 I 0.000 0.000 0.000 0.000 n = 39 7 0.001 0.019 0.018 0.947 n = 41 9 0.001 0.062 0.061 0.984 All PACK 45 0.025 0.127 0.102 0.803 SUBF 4 0.005 0.150 0.145 0.967 INTE 4 0.097 0.169 0.072 0.426 ABRU 4 0.085 0.170 0.085 0.500 HARF 6 0.015 0.052 0.037 0.712 SUBB 1 0.009 0.009 0.000 0.000 41
Table 7. Matrix of identity (I) values between species of C. pachystachya complex, C. preslii and C. macloviana.
PRES MAC 40 37 38 39 41 PRES -— 0.741 0.815 0.717 0.663 0.736 0.749 MAC ----- 0.914 0.884 0.810 0.900 0.873 n = 40 ----- 0.926 0.840 0.945 0.938 n = 37 0.915 0.993 0.857 n = 38 0.913 0.782 n = 39 ---- 0.873
PACH SUBF INTE ABRUHARFSUBB PRES 0.758 0.685 0.746 0.797 0.818 0.742 MAC 0.902 0.753 0.712 0.858 0.936 0.823 n = 40 ——' 0.828 0.826 0.902 0.932 0.842 n = 37 0.816 0.801 0.904 0.920 0.933 n = 38 -— 0.800 0.791 0.872 0.894 0.882 n = 39 -— 0.820 0.800 0.921 0.934 0.909 n = 41 -— 0.822 0.777 0.924 0.904 0.881 PACH -— 0.857 0.823 0.926 0.943 0.919 SUBF 0.789 0.807 0.813 0.745 INTE 0.774 0.769 0.756 ABRU ----- 0.946 0.938 HARF -— 0.908 42
Table 8. Range of identity values for interspecific population comparisons.
Comparison Range
PACH X SUBF 0.517 - 0.888 PACH X INTE 0.574 - 0.874 PACH X ABRU 0.404 - 0.997 PACH X HARF 0.703 - 1.000 SUBF X INTE 0.354 - 0.788 SUBF X ABRU 0.586 - 0.889 SUBF X HARF 0.604 - 0.781 INTE X ABRU 0.583 - 0.809 INTE X HARF 0.650 - 0.867 ABRU X HARF 0.556 - 0.996 43
Table 9. Mean and range of identity values for intraspecific population comparisons.
Taxon I Range PACH « = 37 0.952 0.952 - 0.952 « = 39 0.979 0.949 - 1.000 n = 41 0.937 0.750 - 1.000 All PACH 0.889 0.632 - 1.000 SUBF 0.730 0.665 - 0.797 INTE 0.926 0.883 - 0.999 ABRU 0.842 0.661 - 1.000 HARF 0.931 0.846 - 1.000 CHAPTER III
PHENETIC ANALYSIS OF THE CAREX PACHYSTACHYA COMPLEX
(CYPERACEAE) AND ITS BEARING ON THE EVOLUTION AND
SYSTEMATICS OF THE COMPLEX
Phenetic methods can aid in the resolution of taxa in morphologically difficult groups, that is, where variational patterns in morpholgical features overlap to such a degree that delimitation of discreet taxa becomes problematic.
This situation can especially arise in groups where quantitative characters provide the majority of the discriminating features.
Many members of the genus Carex provide prime examples of the above mentioned situation. This problem has been recognized particularly in section
Ovales in which the Carex pachystachya complex occurs:
All students of Carex admit that the species must often be recognized by small technical differences, especially in the details of the structure of the perigynium. In some groups, notably the section Ovales, precise measurements of the mature perigynium and achene may be necessary for accurate identification. In some sections the species, though narrowly limited, are sharply defined. In others, again notably the Ovales, the boundaries are apt to be vague and there appears to be considerable intergradation between taxa which it seems necessary to hold at the specific level (Cronquist 1969: 221).
The C. pachystachya complex is distinguished in section Ovales as those members of Mackenzie’s (1931) Festivae group having plump or plano-convex perigynia. Although 17 Mackenzien species fit this description, six are readily
44 45 eliminated. Carex macloviana D’Urv. characteristically possesses flattened perigynia, but varies towards plano-convex forms (Whitkus and Packer 1984).
Carex bonplandii K unth and C. purdeii Boott are Mexican species with elongated rootstocks (Mackenzie 1931), the presence of which can be used to remove them from the Ovales which has cespitose members. Carex preslii Steud. has been traditionally placed in the Festivae group, but its large achenes and generally flattened perigynium beaks indicate it belongs in the Festucaceae group (Whitkus and Packer 1984). Carex illota L. H. Bailey is a distinct and pivotal species which is most often placed in the section Ovales but may also be placed in the section
Stellulatae (Mackenzie 1922a,b; 1923) because of the absence of a winged perigynium. Carex amplectans Mack, possesses elongate, amplectate lower bracts like C. alhrostachya Olney of the Athrostachyae group. This leaves eleven species comprising the complex, with members having been noted for their apparant intergradation and affitnites to C. pachystachya (Table 1).
In addition to vague boundaries among species, a great deal of variation occurs within species. The best example of this is C. pachystachya which is widespread geographically, contains four chromosome races (Whitkus and Packer
1984), a taxonomic variety (gracilis fida Mackenzie 1931), and a morphological seggregate (stubby) without taxonomic rank (Whitkus and Packer 1984). Thus the problem presented by the morphological variation within the complex is two- tiered: the question of relationships among species, and the question of seggregates within species. To provide a better understanding of the morphological relationships within the complex, a phenetic analysis was conducted, and the results used to explore the taxonomic and evolutionary relationships within the complex. 46
MATERIALS AND METHODS
Choice of Characters
Keys and descriptions of members of the complex in the works of
Cronquist (1969, 1977), Hermann (1970), Howell (1959), Mackenzie (1916, 1922b,
1923, 1931), and Whitkus and Packer (1984) provided the characters used in the
phenetic analyses. Characters dealing with shape were reduced to measurements
of continuous characters. This avoided multistate, unordered characters (which
are not suitable for discriminant analysis) and provided characters with normal
distributions, or that could be transformed to a normal distribution to allow
hypothesis testing with discriminant anaylsis (Neff and Marcus 1980). The shape
characters used are; plants phyllopodic vs. aphyllopodic; degree of aggregation of
spikes in the inflorescence; inflorescence shape; scale length to perigynium length;
beak length to perigynium length; proportion of perigynium filled by achene;
perigynium shape; proportion of perigynium beak margined; degree of
constriction of beak from perigynium body. A number of shape characters could
not be reduced, but are considered important and were maintained: perigynium
cross-sectional shape; perigynium texture; perigynium base. A total of 26
characters was chosen for the analyses (Table 2).
Choice of Specimens and Initial Grouping
Herbarium specimens and field collections were examined initally to see if
all characters were present. After removing immature or incomplete specimens,
examination of all remaining specimens was conducted to place each in a species
based on the available keys, species descriptions, and type material. Dissection
and examination of perigynia with a 20X dissecting microscope was necessary in
these determinations. A great deal of morphological variation was evident in the
initial taxon determinations. To sample better the variation within each taxon. 47 specimens were microscopically examined a second time and placed into morphlogically similar groups in each taxon. The final groups used in the analyses and a short description of each are given in Table 3.
After placing specimens into groups, a third and fourth round of examinations by eye sorting were conducted to see if further separation could be accomplished. This placed specimens into piles of subgroups for sampling in the phenetic analyses.
The range of morphological variation within each taxon was sampled by random drawing of specimens from the subgroups. An exception to random sampling was done if the chromosome number of the specimen was known (in C. pachystachya). If the total number of subgroups per group was 10 or more, a maximum of five specimens (if available) were selected. If there were less than
10 subgroups, a maximum of 10 specimens (if available) were selected. This sampling scheme roughly kept the number of specimens equal among groups within a taxon. A total of 618 specimens was scored for each of the 26 characters.
Phenetic Methods
Clustering of OTU’s was accomplished by standardizing the data (mean of zero, standard deviation of one), calculating pair-wise OTU similarities or distances, and clustering the resulting matrices. The similarity coefficient used was the Pearson product-moment correlation coefficient (Sneath and Sokal 1973), and the distance coefficient was the average taxonomic distance of Sokal (1961).
Phenograms were produeed by elustering with the unweighted pair group method using arithmetic averages (Sneath and Sokal 1973). All calculations were carried out with the NT-SYS program package (Rohlf et al. 1972). 48
Principal components analysis (PCA) was performed using the correlation matrix of the standardized data, and extracting the first five components.
Stepwise discriminant function analysis (DFA) was performed using only characters which had a univariate normal distribution. This was checked by calculating the skewness, gj, and kurtosis, g2 > of each character in each data set, and testing for normality by means of a t-test. Values of t were obtained by dividing each statistic by its standard error. A test of significance was done by comparing the t-value with the value of tggg and infinite degrees of freedom
(Sokal and Rohlf 1981) with a significant value indicating a non-normal distribution. Transformations were performed on characters that had significant t-values for gj and g2 . The new distributions were tested again, and new transformations carried out until non-significant t values were obtained. If a character could not be sucessfully transformed to a normal distribution, it was excluded from the DFA. Tests of normality, transformations, PCA and DFA were conducted with the BMDP program package (Dixon 1981).
Analyses
Analyses were conducted to address specific questions. These questions, along with program limitations and the unsuitability of characters or OTU’s, required different data sets. The general composition of each data set in terms of number of OTU’s and characters used is presented in Table 4. The questions addressed for each analysis are given in the beginning of the appropriate results section.
RESULTS
Analysis of Groups 49
The first analysis was performed to ascertain if clusters of related OTU’s corresponded to any previously recognized taxa. Because of the numerous OTU’s and large amount of variation, mean variable values for each group were calculated and a cluster analysis performed on the groups. The phenogram using the correlation coefficient (Fig. 1) placed the groups into their respective species better than the phenogram employing the distance coefficient (Fig. 2). Both phenograms indicate a number of consistent clusters of groups. 40MONT and
42HARF cluster, and then form a larger cluster with 23PACH and 29ABRU using correlation (Fig. 1, cluster 2), or 14PACH using distance (Fig. 2, cluster 2). All groups ofC. gracilior form a cluster that joins with the groups of C.subbracteata
(Fig. 1, cluster 7 and Fig. 2, cluster 6). The groups of C.mariposana are found in two clusters; the 35MARI, 36MARI and 37MARI groups form one cluster in the distance phenogram (Fig. 2, cluster 8) but Join with 34MARI and 24PACH using correlation (Fig 1, cluster 4). The 38MARI and 39MARI groups are part of the
51INTE, 52PAUC and 12SUBF cluster in the correlation phenogram (Fig. 1, cluster 1) but join with the 34MARI group as a cluster using distance (Fig. 2, cluster 4). The C. abrupta groups also occur as two clusters: 25ABRU, 26ABUR,
27ABUR, 30ABUR, 31ABUR and 33ABRU as one (Fig 1, cluster 5), and 29ABRU in the 23PACH, 40HARF, 42MONT cluster in the correlation phenogram, while the distance phenogram places 27ABRU and 29ABRU in the large C. pachystachya and C. subfusca cluster (Fig. 2, cluster 3). The C.subfusca groups occur as four clusters with 12SUBF and 13SUBF in separate clusters. In the correlation phenogram, 13SUBF is part of a C.subfusca cluster (Fig. 1, cluster 3). The
ISUBF, 3SUBF, 4SUBF, 6SUBF, 9SUBF, and 11 SUBF groups are found in the large cluster of C. pachystachya and C. subfusca groups in the distance phenogram
(Fig. 2, cluster 3). The C. pachystachya groups, excluding 14PACH, form part of 50 the large cluster of C. pachystachya and C. subfusca groups with the distance coeffecient (Fig. 2, cluster 3). In the correlation phenogram the 23PACH and
24PACH groups occur in different clusters than that containing the remaining C. pachystachya groups.
The relationships seen in the phenograms are reflected in the plot of the groups onto the first two principal components (Fig. 3). The plot of Axes I and
III (Fig. 4) changes the relative positions of the C. mariposana groups which become more distinct, and C. abrupta groups which become further incorporated into the center of the plot. These plots also indicate relationships among the taxa, and levels of variation within them. The C. pachystachya groups are centrally located, as expected, but exhibit as a whole, less variation than some of the other species. For example, the C. mariposana groups range from -0.2 to 2.3 on
Axis I and -1.8 to 0.8 on Axis II. Carex abrupta and C. subfusca also show a great degreee of variation. The eight characters highly correlated (> 70%) with Axis I
(Table 5) deal with size of the achene, perigynium, scale and spike, and are consistent with characters used to delimit the species. Axis II is mostly comprised of the perigynium margin width and distance from the top of the achene to the beak base, with the C. abrupta groups showing high values for these characters
(Table 5, Fig. 3). The degree of compactness of the inflorescence makes up Axis
III, showing the open nature of 12SUBF and 13SUBF, and the C.mariposana groups. Also note the position of the C.abrupta groups close to the C.mariposana groups.
Analysis of louiviuuals
The analysis of groups suggests that boundaries among the species are not clear, even when based on group means. To see if individual OTU’s could
objectively be placed into their a priori determined species, a discriminant 51 analysis was performed. A maximum of 10 OTU’s per group (when available) representing every group and subgroup, was subjected to a stepwise discriminant analysis by using the ’typical’ groups to derive the discriminant function.
Plotting individual OTU’s onto the first two canonical axes produces too complicated a plot and so only the group centers are shown (Fig. 5). The pattern of relationships among groups on the canonical axes is similar to that seen in the group PCA. As in the group PCA, the 27ABRU and 29ABRU group centers occur near C. pachystachya and C. subfusca. Carex harfordii and C. montereyensis occur within the range of C. pachystachya. The 12SUBF group is far removed from the remaining C. subfusca groups and C.paucifructus is close to the centers of
39MARI and 44SUBB.
Since each character used in the DFA originally had, or was transformed to a normal distribution, the F-statistics produced by the DFA can be used to test
whether groups are significantly different from one another, based on variation
in the characters. The results show that within each species groups can be linked
through a series of non-significant (P > 0.05) F-statistics (illustrated in Fig. 6). In
C. subfusca the 12SUBF and 13SUBF groups are significantly different from the
remaining C. subfusca groups as well as each other. In C.pachystachya all groups
can be linked except the 24PACH group. In C. abrupta the 27ABRU and 29ABRU
groups are significantly different from the remaining groups. In C.mariposana
the 39MARI group is separate from the rest. The 40HARF and 42MONT groups
are not significantly different, as are the C. subbracteata and C. gracilior groups.
Carex intégra is significantly differernt from all other groups, while C.
paucifructus is not significantly different from 12SUBF, 34MARI, 37MARI,
38MARI, and 39MARI (not shown). Most other inter-group F-statistics are
significantly different, but a few are not. These are: 3SUBF to 17PACH, 52
19PACH, 21 PA CH and 27ABRU; 4SUBF to 17PACH; 9SUBF to 17PACH,
21 PACH, 23PACH and 29ABRU; 17PACH to 29ABRU, 40HARF, 42MONT and
44SUBB; and 24PACH to 29ABRU and 31 ABRU.
The classification function used seven characters that could significantly differentiate groups. In total, 75.7% of the a priori classifications were considered correct by the function for the ’typical’ groups. OTU’s for each group were placed both within the species for the group as well as within other species
(Table 6). Interestingly, all I2SUBF OTU’s and individuals from other taxa are placed in 52PAUC. OTU’s of other C. subfusca groups were placed into C. pachystachya, C. montereyensis, C. harfordii, and C. integra. O T U ’s o f C. pachystachya groups occur among each other, as well as in C.abrupta, C. montereyensis, C. harfordii, an d C. paucifructus. OTU’s of C. abrupta are placed in
C. pachystachya and C. montereyensis. Carex mariposana O T U ’s w ere placed in C. abrupta and C. subbracteata. OTU’s of C. gracilior an d C. subbracteata are shared with each other, and are also found in C. montereyensis, C. harfordii, and C. paucifructus. A number of C. integra OTU’s occur in C. subbracteata.
Analysis of Species Pairs
Analyses up to this point have shown that three pairs of species cluster together: C. montereyensis-C. harfordii, C. subbracteata-C. gracilior, and C. pachystachya-C. subfusca. Additional analyses were performed to see if these pairs contained different variation patterns (through PCA) and if the members of each pair could be separated using the characters available (through DFA).
C. montereyensis-C. harfordii
The plot of the first two axes of the PCA shows complete overlap in the
OTU’s of the two species (Fig. 7). Separation is not accomplished when the third axis is employed (not shown). The three axes account for 52.5% of the total 53 variation. DFA of the two taxa results in a significant F-value (P < 0.001) indicating the two can be separated. Overall, 71.1% of the a priori determinations were correctly classified.
C. subbracteata - C. gracilior
The two taxa separate well on the principal components plot (Fig. 8), even though these two components account for only 33% of the total variation. As the two taxa separate on the second axis, characters dealing with the inflorescence are important in this separation (Table 7), characters that have also been used to separate the two taxa in previous works. In the DFA, all OTU’s of C. gracilior were correctly classified. 89.5% of the OTU’s of ’typical’ C. subbracteata were classified correctly, while 44SUBB had 75% of its OTU’s placed into C. subbracteata and 25% in C. gracilior (Table 8). Overall, 90.6 % of the a priori determinations were correct (Table 8). The ’typical’ groups are significantly different at the 0.001 level.
C. pachystachya - C. subfusca
Due to the large size of both data sets, only OTU’s from the ’typical’ groups were used in this analysis (ISUBF, 22PACH, I9PACH, 14PACH). The
ISUBF and 14PACH groups show unique variation patterns in principal components space (Fig. 9). Inclusion of the 22PACH and 19PACH groups fills in the gap, however. Separation along Axis I is due to characters dealing with the size of the scale, perigynium and achene (Table 9) with 14PACH having the larger values, as has been used in keys. Separation along the second axis is mainly due to inflorescence length, with both ISUBF and 14PACH having larger values than 22PACH and 19PACH. However, notice that ISUBF also possesses individuals with these lower values. 54
In the DFA, a better separation of ISUBF from the remaining groups is obtained (not shown). Percent correct classification is 94.8% for ISUBF, with
5.2% of its OTU’s classified in C. pachystachya (Table 10). No OTU’s of C. pachystachya are misclassified into ISUBF, only into other groups within the species.
Intraspecific Variation
Analysis of the large and variable species C. subfusca and C. pachystachya was conducted to find whether intraspecific variation existed, using PCA and
DFA.
C. subfusca
Ellipses of the various groups of OTU’s on the first two principal components shows considerable overlap (Fig. 10). Only the 3SUBF and 9SUBF groups are separated from the others. In the DFA, all groups except 9SUBF and
3SUBF are significantly different at the 0.05 level. However, overall correct classification of the a priori determ inations is 57.2% (Table 11) w ith only the
12SUBF and 13SUBF groups having a value of 100%.
C. pachystachya
Ellipses of the three ’typical’ groups of this species on the principal components shows overlap of all three (Fig. 11). Although 22PACH is more distinct from 14PACH, 19PACH overlaps both. The plot is complicated when the remaining group OTU’s are added (Fig. 12). Most of these occur towards the middle of the plot, for the most part separate from the 22PACH OTU’s. The
21P.4.CH OTU’s fall between 22PACH and 14PACH. Looking at chromosome numbers, all n = 41 specimens fall within the 22PACH ellipse, while the remaining numbers are found throughout the rest of the plot. 55
In the DFA, the three ’typical’ groups are significantly different from each other at the 0.001 level. The classification matrix (Table 12) shows none of the ’typical’ groups has a 100% correct a priori classification. OTU’s of 22PACH may still be placed into 14PACH or 19PACH, as well as OTU’s of 14PACH and
19PACH being placed into 22PACH. As might be expected, the 21 PACH OTU’s are divided among the 14PACH and 22PACH groups.
DISCUSSION
The analyses show that members of the complex do not represent distinct morphological entities. The degreee of overlap varies however, from distinct species such as C. integra to near completely merged taxa such as C. harfordii and
C. montereyensis. As mentioned in the Introduction, this pattern is expected when dealing with members of section Ovales. The critical question is what causes the intergradation. Two likely answers are recent evolutionary divergence with the taxa representing budding seggregates of a progenitor species; and/or reticulate evolution where distinct species are merging into a single unit through hybridization. Review of the evidence suggest that recent, and even ongoing divergence, appears to be the cause.
The strongest evidence for divergence comes from crossing studies among the chromosome races of C. pachystachya. All the races are cross compatible, but compatibilities among the races are much lower than compatibilities within races
(Whitkus 1988). Meiotic pairing in F| hybrids indicates that numerous structural rearrangements in the chromosomes have occurred so that pollen stainabilities of
F 2 hybrids are significantly reduced (chapter 2). The evidence suggests that races have arisen as a result of chromosomal repatterning and that significant reproductive isolation has developed. The phenetic analyisis of the OTU’s in C. 56 pachystachya shows that except for the n = 41 race, all other races are morphologically Indistinguishable, and there are numerous intermediates between the « = 41 race and the remaining members of the species. Thus reproductive isolation has arisen among the races without a concomitant amount of morphological divergence.
The above observations can be extended to other members of the complex that are more divergent morphologically. Two further lines of evidence can support the conclusion that the species represent morphologically similar but reproductively isolated products of recent evolution. The first is that isozyme analysis shows high levels of similarity among populations and no complimentary hybrid patterns (chapter 2). The second argument deals with geographic distribution in the complex. Carex pachystachya is a northern boreal species, occurring from coastal Alaska to northern California, eastward to the Rocky
Mountains (Whitkus 1981; Whitkus and Packer 1984). Carex integra and C. subfusca are Cascade-Sierrean species (Cronquist 1969; Howell 1959), and the remaining species endemic or centered in California (Howell 1959). C arex pachystachya occurs in moist habitats such as river, stream or lake shores, damp roadside depressions or mountain meadows. These are habitats that remain moist throughout most of the summer (Whitkus 1981). Carex integra, C. subfusca, C. abrupta, and C. mariposana occur in seasonally moist habitats that quickly dry out in late spring (pers. obs.). Carex gracilior, C. subbracteata, C. harfordii and C. montereyensis are coastal endemics, generally found in very sandy, moist soil along the Californian coast and Coastal Ranges (Howell 1959; pers. obs.). If C. pachystachya is considered the progenitor of the complex, then the southern taxa have diverged to a drier habitat. This pattern of a northern temperate element giving rise to southern, more dry adapted taxa is well known for many 57
C alifornian plants (Raven and Axelrod 1978; Stebbins 1982) and is thought to be a response to the increasing aridity afforded by the orogeny of the Cascade,
Sierra Nevada and Coast Ranges. Since the origin of the aggregate tc which C. pachystachya belongs is thought to have been since the end of the Pleistocene glaciation (Whitkus 1981), then the origin of the species within the complex must have either arisen during or subsequent to this time. This leaves about 20,000 years for the taxa to have originated and argues for recent evolutionary divergence. The general separation of habitats, especially of C. pachystachya from
C. subfusca, C. abrupta, and C. mariposana argues that the species do not have the opportunity to occur together to produce hybrids. With chromosomal repatterning likely to have accompanied evolution in this complex (chapter 4), the combination of habitat and chromosomal differences maintains reproductive isolation among the species. The only species that commonly can be found in the same habitat are
C. subfusca, C. integra, and C. abrupta in mountain meadows and forest openings.
However, few intermediates are found among these taxa. The two species pairs,
C. harfordii'C. montereyensis and C. gracilior-C. subbracteata are also found in the same habitat, but again, no intermediates occur between these species pairs, although many are present between the species within the pairs.
Taxonomy
Taxonomic decisions in this complex must achieve a balance between the variation that tends to blur taxonomic boundaries (a result of the evolutionary process) and the need to recognize the modes in the variation that likely represent independently evolving species. The analyses indicate that morphologically recognizable groups do exist. As Cronquist (1969) has stated, the variation within and among these groups forces the recognition of taxa which are not delimited by a discontinuity in morphological variation. The analyses have shown that some 58 of the previously recognized species cannot continue to be recognized at the specific level without further data. Alternatively, morphology and biosystematic data have shown that biological entities exist within the polymorphic C. pachystachya. The following taxonomic suggestions are based mainly on the analysis of this study and incorporates, where possible, all pertinent data on the group.
Carex integra
No realignment is needed for C. integra. The species is eonsistently distinct in the analyses. Affinities with C. subfusca are evident from the group phenograms and PCA, and the DFA of individuals. In addition to the morphological differences, the two also differ in their chromosome number.
Carex harfordii
Consistent clustering and overlap of phenetic variation patterns of C. montereyensis and C. harfordii argue for not recognizing two species. Although
Mackenzie delim ited C. harfordii on the basis of larger heads and perigynia, the analyses effeetively show that the two represent ends of a morphological continuum. The significant separation in the DFA of the two species suggest the two taxa may indeed represent separate evolving populations systems. However, the lack of any consistent phenetic as well as habitat and distribution differences, argues that until further data become avialable, the recognition of a single morphological species is necessary.
Carex subbracteata
Carex subbracteata and C. gracilior mirror the situation seen in C. harfordii and C. montereyensis. Similarity of the two taxa is seen in the group PCA and phenograms. Unlike the previous pair, the two taxa are readily distinguished from each other in their separate analyses, mainly on the basis of inflorescence 59 characters. Mackenzie (1916) separated the two on the basis of elongate inflorescences in C. gracilior with spikes separate, while C. subbracteata has a more dense inflorescences. Some intergradation is still evident from the analyses.
The sharing of a common distribution and habitats argue that these two taxa are member of the same species. The clear morphological separation in the analysis of the species pair however, indicates they can be recognized as subspecies. This contrasts with the lack of evident differences in C. montereyensis and C. harfordii.
Carex mariposana
The case of C. mariposana is puzzling. The type shows a large inflorescence with light colored perigynia contrasting with the chestnut brown scales. This is reminescent of C. straminiformis in the Festuceae group, but the perigynia are unmistakably those of the C. pachystachya complex. The problem arises because many specimens identified as C. mariposana are really other species.
This is a result of Mackenzie’s key to the species and the fact that many workers are not familar with Ovales. Essentially, any member of the Festiva group that has elongate inflorescences and perigynia over 3.5 mm is placed into C. mariposana by many workers; this includes some C. subfusca and many C. cbrupta.
The character of evidently nerved perigynia on the ventral side is another feature used to place specimens in this species, but is often misunderstood.Carex mariposana, C. abrupta, C. harfordii a n d C. montereyensis have several prominent nerves extending the entire length of the perigynium body; that is, from the base to the top of the achene or base of the beak. Other members of the complex may have nerves on the ventral surface of the perigynium, but these may not be prominent, or they may extend partially up the perigynium body. In a few instances, a single nerve may extend the length of the perigynium body. Once this difference is understood, C. mariposana can recognized be more clearly. 60
After removing obviously misidentified specimens, C. mariposana still contains a wide range of variation. The group PCA shows the 34MARI, 38MARI and 39MARI groups are separated from the 35MARI, 36MARI and 37MARI groups. It thus appears that differentiation is taking place within this species, and that it is similar to the level seen in C. harfordii and C. montereyensis. The same decision is made here for C. mariposana as was made for C. harfordii, i.e., not to recognize infraspecific categories.
Because very few collections (only two mature specimens were available in the study) of C. paucifructus are known it is currently recognized as an endangered species in California. However, because the type is not a fully matured specimen, the validity of this species is questionable. The taxon occurs in proxim ity to C. mariposana in the group PCA, DFA of all OTU’s, and correlation group phenogram. OTU’s of C.paucifructus are not significantly different from many of the groups of C.mariposana in the DFA of individuals.
In light of the many morphological variants present in each species of the complex, especially C. mariposana, it is best to assume C. paucifructus represents one of those variants.
Carex abrupta
As mentioned in the discussion of C. mariposana, many specimens of C. abrupta are misidentified as C. mariposana because of the presence of more elongate inflorescences. However, the distinguishing characters for C. abrupta are perigynia with a relatively small achene, resulting in a longer distance from top of the achene to beak base than in other members of the complex. This difference is the same character used to differentiate the C. microptera complex from the C. pachystachya group (Whitkus and Packer 1984). Indeed, many specimens of C. abrupta seem to represent members of the C. microptera group. 61
This species may better be placed in that complex, or at least represents an intermediate between the two complexes. Without further evidence, the position of C. abrupta cannot be decided unambigously, so it is maintained as a member of the C. pachystachya complex.
Carex abrupta shows a wide range of morphological variation. The
27ABRU and 29ABRU groups show a distinct separation from the remaining groups. Since both of these groups represent intermediate forms, their placement is not surprising. It is likely they actually contain specimens of C.pachystachya and C. subfusca, resulting in their proximity to these species in the analyses.
These two groups were not included, therefore, in the species circumscription.
Carex subfusca
Carex subfusca represents a large and polymorphic species. The nature of
13SUBF and 12SUBF and the status of C. teneraeforinis must be addressed.
Depending on the analysis, 13SUBF either is found with other C.subfusca groups, or like 12SUBF is quite distinct. All OTU’s of 13SUBF are classified by the discriminant function in the DFA of individuals as members of C. subfusca.
Thus 13SUBF (and C. teneraeformis which it represents) is an extreme form of C. subfusca. Whether or not to grant it taxonomic standing depends on how other extreme forms are treated in this variable species (see below). Without evidence indicating any reproductive or habitat differences and no clear morphological distinction, the view of Cronquist (1969) and others that C. teneraeformis is not worthy of taxonomic standing is followed. The 12SUBF group does not help to clear the situation since the small stature of the plants indicate this group represents a depauperate version of the 13SUBF group. At this point, there are no indications of habitat or distributional differences. 62
Other divergent forms also occur in C. subfusca. The PCA of the species highlights the differences of the 3SUBF and 9SUBF groups from the remaining groups. The 9SUBF group is a restricted form, as it is found in the Charleston
Mts. of Clarke Co., Nevada and the White Mts. of Mono Co., California. The
3SUBF group is more widely distributed, occurring in Tehama, Madera and San
Bernardino Cos., California. Both forms appear to be adapted drier conditions than the remainder of the species. However, the limited number of specimens and their clustering with the remainder of the species indicate these groups should not be taxonomically recognized.
Carex pachystachya
Carex pachystachya shows considerable geographic, chromosomal and morphological diversity, probably the result of it being the most intensely studied member of the complex. In an earlier work (Whitkus and Packer 1984) no justification could be found to recognize infraspecific taxa in the species. The analysis presented here, together with other information, now suggest the need for taxonomic recognition for one of these variants. The stubby group consists of the « = 41 chromosome race and is distinct morphologically, although morphological intermediates do occur. Crossing studies, pairing in FI hybrids, and isozyme analysis indicate that it is as distinct from other elements in C. pachystachya as more distantly related species (i.e.: C. macloviana and C. preslii), and on the basis of these data, stubby should be recognized as a new species. The lack of clear morphological distinction, with the numerous intermediate forms, however, indicates this would be problematic taxonomically. The most desirable solution is to recognize that stubby is a species in the sense that it has completed divergence from C. pachystachya in terms of reproductive isolation, but has not 63 diverged sufficiently in morphology. For these reasons, stubby is accorded the status of a subspecies.
The arguments presented above for stubby do not apply to C.olympica because this morphological form contains the chromosome races 37, 38, and 39, and is closer morphologically to C. pachystachya. Thus C. olympica is like forms in the other species, that is, diverging from the typical form, but not to a point where it can be granted taxonomic status.
Key to Members of C. pachystachya Complex
1. Spikes closely aggregated, length of first two internodes of inflorescences less
than 1/3 of overall inflorescence length
2. Mature achenes filling 1/2 or more of body of perigynia; beaks less than
2/5 of overall perigynia length; plants of coastal regions C. harfordii
2. Mature achenes filling less than 1/2 of body of perigynia; beaks more than
2/5 of overall perigynia length; plants of inland, montane regions C.
abrupta
1. Spikes in moniliform inflorescence to loosely aggregated, length of first two
internodes of inflorescences greater than 1/3 of overall inflorescence length
3. Perigynia wings very narrow (<0.1 mm), mainly absent from upper body
and beaks ...... C. integra
3. Perigynia wings wider or prominent on upper body and beaks, often
serrulate
4. Perigynia nerved on ventral side with thin, prominent nerves
extending into beaks ...... C. mariposana
4. Perigynia smooth on ventral side or with thick nerves, often extending
no further than middle of body of perigynia, rarely one or two
extending into beaks 64
5. Mature perigynia thick-walled, membranous-coriaceous to
coriaceous; scales averaging 3.5 mm long or more
6. Length of first two internodes of inflorescences 2.3-9.4 mm,
comprising up to 2/5 of inflorescence length; leaves 1.5-3.9
mm wide ...... C. subbracteata ssp. subbracteata
6. Length of first two internodes of inflorescences 3.9-13.8 mm,
comprising up to 1/2 of inflorescence length; leaves 1.2-3.0
mm wide ...... C. subbracteata ssp. gracilior
5. Mature perigynia thin-walled, membranous; scales usually less than
3.5 mm long
7. Perigynia stramineous, sometimes light lustrous brown; upper
margins becoming stramineous...... C. subfusca
7. Perigynia lustrous, reddish to coppery brown or nearly black;
upper margins becoming concolorous with body of perigynia,
often dark-edged
8. Dorsal surface of beaks dark lustrous brown to black,
extending to upper margins and upper body of
perigynia, contrasting with lighter color of remainder of
body of perigynia; spikes tightly aggregated in
inflorescences C. pachystachya ssp. compacta
8. Dorsal surface of beaks concolorous with body of
perigynia, or upper third of beaks dark brown; spikes
aggregated to loosley aggregated in inflorescences, at
least lowest spikes separate and distinct ....C. pachystachya
ssp. pachystachya 65
SPECIES DESCRIPTIONS AND DISTRIBUTIONS
Carex integra Mackenzie, Bull. Torrey Bot. Club 43: 608. 1917.
-Type: California: Placer Co., Summit, 7,000 ft., July 16, 1909,Heller 9841
(holotype: NY!).
Culms stiff, (11) 13-46 cm tall, exceeding the leaves; leaves straight, blades
1.1-2.9 (3.1) mm wide, flat; inflorescences 11.6-22.9 (24.0) mm long, 6.3-13.1 (14.1) mm wide, first two internodes 2.S-7.9 mm, comprising up to 2/5 length of the inflorescence, averaging 1/3; spikes 5-9, aggregate, 4.5-9.1 (10.2) mm long, (2.1)
2.1-5.1 mm wide; bracts scale-like, lower bracts awned, awn not extending beyond subtended spike; scales lustrous, light to dark chestnut brown to dark brown, (1.9)
2.1-3.5 mm long, 3/4 to slightly longer than perigynia, 0.8-1.4 mm wide, centers green to light green, margins narrow white hyaline; perigynia lustrous, stramineous to light brown, upper margins green to light green, membranous, plano-convex, lance-ovate to ovate, generally constricted towards base when dry,
(2.1) 2.3-3.6 mm long, 0.8-1.4 mm wide, thin winged or margined, wings less than
0.1 mm wide, mostly absent from beak and upper body, ventral side nerveless or with up to 4 thick nerves extending into the body with one or two extending into the beak, beaks gradually to abruptly tapering from body of perigynia, brown to dark brown, 0.7-1.6 mm long, up to 1/2 length of perigynia, averaging 1/3, distance from top of achene to beak base less than 0.4 mm; achenes elliptic to quadrate, 1.1-1.4 mm long, 0.7-1.0 mm wide, filling up to 2/3 of the perigynium body, averaging 1/2, and distending body to the margins; n = 41.
From southern Washington to southern California in the Cascade, Klamath,
Warner, Sierra Nevada, San Bernandino and Santa Rosa Mountains (Fig. 13).
Meadows and roadsides, mountain to subalpine conditions from 900 to 3350 meters. 6 6
As noted by previous authors (eg; Cronquist 1969; Mackenzie 1931), C. integra resembles C. subfusca. The most striking difference is the lack of a wing
on the beaks and upper body of the perigynia in C. integra. This feature makes
C. integra sim ilar to C. illota which lacks a wing margin along the entire length of
the perigynium. The two species are easily distinguished by the small, dark, tightly congested inflorescences of C. illota versus the more open, lighter colored inflorescences of C. integra. The difference in color of the inflorescences is due to the much darker colored scales and perigynia of C. illota. The two species grow in different habitats; C. illota is found in bogs and wet meadows in
subalpine and alpine situations, while C.integra occurs in montane to subalpine
meadows that may be seasonally moist but tend to be quite dry through most of
the summer.
Representative Specimens. Washington: Yakima Co., Mt. Adams, Suksdorf
6863 (CAS, WS). Oregon: Deschutes Co., near summit of McKenzie Pass,Detling
3244 (CAS). Douglas Co., Diamond Lk., Peck 19278 (CAS). Lake Co., west side of
McKenzie Pass, Whitkus 2405 (NY). Klamath Co., Cresent Lk., Peck 27250
(WILLU). California: Alpine Co., Luther Pass,Eastwood & Howell 8374 (CAS).
Butte Co., Jonesville, July 4, 1938, Copeland s. n. (CAS). Eldorado Co., Echo
Summit ski area, Whitkus 4051 (NY). Fresno Co., Huntington Lk., Whitkus 3447
(NY). Glenn Co., Plaskett Meadow, Howell 19013 (CAS). Kern Co., Mt.
Breckenridge, Twisselmann 5386 (CAS). Klam ath Co., 3.2 km N of Ft. Klamath,
Whitkus 2379 (NY). Madera Co., Devils Postpile, Howell 14456 (CAS). Modoc Co.,
Warner Mts., Alexander 18310 (CAS). Placer Co., near Emigrant Gap, Rose 39216 (CAS, UC). Plumas Co., 12.8 km SW of Johnsville, Rose 51030 (UC). Riverside Co., Santa Rosa Pk., Munz 15348 (CAS). San Bernardino Co., 7.2 km W of Bear Valley, Whitkus 3828 (NY). 67 Shasta Co., 33.5 km S of McCloud, Hermann 24778 (CAS). Sierra Co., Yuba Pass, Whitkus 4262 (NY). Siskiyou Co., Spirit Lk., Howell 15002 (CAS, DS). Trinity Co., N of Carrville, Howell 12761 (CAS). Tulare Co., Little Whitney Meadow, Howell 25893 (CAS). Tuolumne Co., Yosemite Valley,Abrams 4399 (DS). Carex abrupta Mackenzie, Bull. Torrey Bot. Club 43: 618. 1917. -Type: California: Butte Co., West Branch of North Fork of Feather River, near Sterling, 3,000 ft., June 7, 1913, Heller 10820 (holotype: NY!). Culms stiff to +lax, 18-61 (66) cm, exceeding leaves; leaves straight, blades 1.5-3.7 (4.9) mm wide, flat; inflorescences bicolorous, the scales contrasting with lighter perigynia margins, 11.9-19.5 (21.8) mm long, (6.0) 8.9-17.7 (17.9) mm wide, first two internodes 1.7-5.5 (6.3) mm long, comprising up to 1/3 length of the inflorescence, averaging 2/10; spikes 4-9, loosely to densely aggregate, the lowest spike sometimes separate and distinct, (4.4) 5.4-10.0 (10.8) mm long, (2.7) 3.5-8.0 (8.4) mm wide; bracts scale-like, lowest with short awn not extending beyond subtended spike; scales lustrous, light to dark chestnut or coppery brown, (2.4) 2.6- 3.9 mm long, 1/2 to nearly as long as perigynia, 0.9-1.4 mm wide, centers green or lighter, margins concolorous with scales to white hyaline; perigynia lustrous, light to dark chestnut or coppery brown, upper margins light green becoming stramineous, membranous, plano-convex or sometimes flat and distended by achenes, elliptic to lance-ovate or ovate, (2.9) 3.6-5.4 mm long, 1.0-2.1 mm wide, wings less than 0.3 mm wide, ventral side with 2-8 distinct (fine) nerves, most extending into beak, beaks gradually tapering from body of perigynia, dark chestnut or coppery brown, (0.8) 1.3-2.6 mm long, up to 1/2 length of the perigynia, averaging 2/5, distance from top of achene to beak base 0.2-1.2 mm; 6 8 achenes narrow-oblong to ovate-quadrate, 1.2-1.8 mm long, 0.7-1.1 mm wide, filling up to 1/2 of perigynium body, averaging 2/5. Growing in the San Bernandino, Sierra Nevada, Klamath and North Coast Ranges of California; also in the southern Cascade, Steens and Wallowa Mountains of Oregon, Washoe Co., Nevada, and the Wind River Range of Wyoming (Fig. 14). In moist meadov/s and slopes in montane to subalpine habitats, 1400 to 3300 meters. In th e C. pachystachya complex, C. abrupta is closest to C. mariposana. The key differences are technical and overlap to quite an extent. Generally, C. abrupta has more congested spikes while the spikes in C. mariposana are m ore loosely arranged. The contrasting color of the upper body of the perigynia with the scales given by Mackenzie (1916) for C. mariposana is variable in that species and can be present in C. abrupta. Another notable difference is their relative abundance. Carex abrupta is quite common and is expected in all highland meadows throughout California while C. mariposana is less abundant and occurs from the central Sierra Nevada through the Klamath Ranges. Representative Specimens. Oregon: Harney Co., 32 km ESE of Frenchglen, Hansen 580 (CSC). Klamath Co., Munson Valley, Crater Lk., Applegate 11270 (CAS). Wallowa Co., Mirror Lk., Mason 6649 (ORE). California: Alpine Co., 1.1 km W of Sonora Pass, Wiggins 14229 (DS). Eldorado Co., Mt. Tallac, Alexander & Kellogg 3470 (UC). Fresno Co., Kaiser Pass, Whitkus 3514 (NY). Glenn Co., Plaskett Meadows, Howell 19225A (CAS). Inyo Co., near Lk. Sabrina, Raven & Stebbins 15 (CAS, UC). Lassen Co., Juniper Lk., Cillett 867 (JEPS). Madera Co., Garnet Lk., Howell 16638 (CAS). Mono Co., Tioga Pass, Whitkus 3670 (NY). Nevada Co., Donner Summitt, Gierisch & Esplin 3591 (RM). Plumas Co., 16 km S of Quincy, Cantelow 3637 (CAS). Riverside Co., Mt. San Jacinto, Reed 2499 (DS). 69 San Bernardino Co., San Gorgonia Mts., Grant 6403 (DS). Shasta Co., 2.4 km N of Summit Lk., Whitkus 4387 (NY). Sierra Co., Yuba Pass, Whitkus 4263 (NY). Siskiyou Co., Marble Mts., Howell 15190 (CAS). Tehama Co., near Sulphur Works, Mt. Lassen Park, Aug. 30, 1955, Leschke s. n. (CAS). T rinity Co., N orth Yolla Holla Pk., Munz 16678 (DS). Tulare Co., Mineral King, Howell 17244 (CAS). Tuolumne Co., White Wolf Meadow, Yosemite Park, Bracelin 876 (CAS). Nevada: Clark Co., Charleston Mts., Clokey 7471 (CAS, DS, ORE). Washoe Co., Tahoe Meadows, Lewis 451 (RM). Idaho: Custard Co., below Castle Pk., Hitchcock & Muhlick 10824 (CAS). Wyoming: Subulette Co., SE of Pinedale, Porter & Miller 6154 (WTU). Carex subbracteata Mackenzie, Bull. Torrey Bot. Club 43: 612. 1917. -Type: California: Oakland, Bolander (holotype: NY!). Culms stiff to +lax, 27-89 (103) cm tall, exceeding the leaves; leaves straight, blades 1.3-3.7 (4.6) mm wide, flat; inflorescences 12.9-31.0 (34.9) mm long, 6.9-19.3 (23.3) mm wide, first two internodes 2.3-13.8 (14.3) mm, comprising up to 1/2 length of inflorescence, averaging 1/3; spikes 4-11, majority separate and distinct, more commonly loosly aggregate to aggregate with lowest 1-3 spikes separate and distinct, 6.0-11.4 (12.3) mm long, 3.8-8.0 (9.1) mm wide; bracts scale like, lowest with awn extending up to length of inflorescence; scales lustrous, light to dark brown, red-brown, or coppery brown, 3.1-4.5 (4.7) mm long, 3/4 to as long as perigynia, 1.2-2.1 mm wide, centers green, turning concolorous with scales or lighter, margins white hyaline; perigynia lustrous, stramineous to coppery brown, upper margins green, turning stramineous to concolorous with body, membranous-coriaceous to coriaceous, plano-convex to biconvex, ovate to wide ovate, (2.9) 3.6-5.1 (5.7) mm long, 1.3-2.2 mm wide, wings mostly less than 0.2 mm l(j wide, ventral side with 0-7 indistinct nerves occasionally extending past middle of perigynium body, beaks gradually to +abruptly tapering from body of perigynia, concolorous with body to darker brown, 1.1-2.2 (2.6) mm long, up to 1/2 length of the perigynia, averaging 2/5, distance from top of achene to beak base less than 0.7 mm; achenes wide ovate to quadrate, 1.5-2.1 mm long, 1.0-1.5 (1.7) mm wide, filling up to 3/4 of perigynium body, averaging 1/2, usually distending body to the margins. C. subbracteata ssp. subbracteata Inflorescences mostly comprised of aggregate spikes, first two internodes 2.3-9.4 (11.2) mm long, distance from base to widest part 3.6-14.4 (15.3) mm; perigynia ventral sides with up to 7 nerves extending up to top of achene, distance from top of achene to beak base 0.1-0.7 mm. Ranging from Humboldt Co. south to Santa Barbara Co. (Fig. 15). Along the coast and into Coastal Ranges, in meadows, swales and along roadsides from sea level to 900 meters. The two subspecies differ in the given key characters. The subspecies subbracteata has its spikes more closely aggregated in the inflorescence and has slightly wider leaf blades. Both subspecies contain evident nerves on the ventral side of the perigynia, in contrast to Mackenzie’s (1916) description of both taxa being essentially nerveless ventrally. Representative Specimens. California: Alameda Co., hills near Berkeley, Tracy 1415 (UC). Contra Costa Co., Richmond, Robbins 3984 (JEPS). Humboldt Co., Eel Riv. near Scotia, Tracy 4695 (UC). Marin Co., Tomales, June 10, 1945, Leschke s. n. (CAS). Mendocino Co., Mendocino City, Eastwood 11444 (CAS). Monterey Co., Ft. Sur, Howell 42038 (CAS). Napa Co., E of St. Helena, Tracy 18287 (UC, WTU). San Francisco Co., San Francisco, Bolander 1568 (UC). San 71 Luis Obispo Co., Santa Lucia Mts.,Hardham 2228 (CAS). San Mateo Co., near South San Francisco, Howell 13779 (CAS). Santa Barbara Co., Highway 150, May 27, 1938, Pollard s. n. (CAS). Santa Clara Co., road to Madrone Springs, Eastwood & Howell 4530 (CAS, UC). Santa Cruz Co., 3.2 km NE of Santa Cruz, Hesse 2515 (CAS). Sonoma Co., Petaluma, May 7, 1933, Stacey s. n. (CAS). C. subbracteata ssp. gracilior (Mackenzie) Whitkus, comb. nov. C. gracilior Mackenzie, Bull. Torrey Bot. Club 43: 614. 1917. -Type: California: Sonoma Co., Cloverdale, April 1864, Bolander 3822 (holotype: DS!; isotype: DS!). Inflorescences mostly moniliform, varying to loosely aggregate spikes, first two internodes 5.0-13.8 (14.3) mm long, distance from base to widest part 7.3-19.5 mm; perigynia ventral sides smooth or with up to 4 nerves to top of achene, distance from top of achene to beak base G.G-0.5 mm. Along the California coast and in Coast Ranges from Mendocino Co. to Santa Barbara Co., inland to the Sierra Nevada foothills (Fig. 16), in meadows, sloughs and along roadsides, from sea level to 6G0 meters. Subspecies gracilior has spikes more loosely arranged in the inflorescence, narrower leaf blades and generally has fewer nerves on the ventral side of the perigynia than the other subspecies. Subspeciesgracilior also has a wider range, occurring in scattered localities in the foothills of the Sierra Nevada Mts. Representative Specimens. California: Alameda Co., North Berkeley, Howell 13937 (CAS), Butte Co., Chico-Hamilton City road, Heller 14853 (CAS). Calaveras Co., Douglas Flat, Eastwood & Howell 8734 (CAS). Contra Costa Co., 0.8 km E of Ft. San Pablo, Robbins 3984 (RM). Lake Co., Nicasio, Rose 39079 (UC). Mendocino Co., near Navarro, Eastwood & Howell 2593 (CAS). Napa Co., Sarco 72 Creek, E of Napa, Raven 3047 (CAS, UC). Sacramento Co., 6.4 km NE of Elk Grove, True 6478 (CAS). San Francisco Co., Presidio, Howell 32684 (CAS). San Luis Obispo Co., Santa Lucia Mts., Green’s Lk., Hardham 1963 (CAS). San Mateo Co., Stanford Univ. pasture land, Benson 2065 (WTU). Santa Barbara Co., Mountain Drive near Sheffield Res., April 19, 1952,Pollard s. n. (CAS). Santa Clara Co., Mt. Hamilton, April 23, 1939, Thorsteinsm s. n. (CAS). Sonoma Co., 4.8 km E of Valley Ford, Howell 5241 (CAS). Carex harfordii Mackenzie, Bull. Torrey Bot. Club 43: 615. 1917. -Type: California, 1868-9, Kellogg & Harford 1073 (holotype: NY!). Carex montereyensis Mackenzie, Erythea 8: 92. 1922. -Type: Californai: Monterey Co., Pacific Grove, C. P. Smith 1055 (holotype: GHI). Culms stiff to +lax, 29-112 (134) cm tall, exceeding the leaves; leaves straight, blades 1.8-5.0 (9.9) mm wide, flat; inflorescences 10.8-28.5 (38.1) mm long, 8.6-18.7 (20.0) mm wide, first two internodes 1.7-8.2 (12.8) mm long, comprising up to 1/3 length of the inflorescence, averaging 2/10; spikes 4-11, aggregate, 4.7-10.8 (11.8) mm long, (3.7) 4.0-7.6 mm wide; bracts scale-like or lower bracts enlarged and awned, awn longer than the inflorescence; scales lustrous, pale green, stramineous or light through dark red or coppery brown, 2.2-4.1 (4.7) mm long, 3/4 as long to slightly longer than perigynia, 1.0-1.6 (1.8) mm wide, centers green becoming stramineous to concolorous with scales, margins concolorous with scales to white hyaline; perigynia lustrous, stramineous to light coppery brown, upper margins green turning stramineous, membranous, plano-convex or slightly biconvex, ovate to round-ovate, 2.6-4.4 (4.6) mm long, 1.1-2.0 mm wide, wings less than 0.2 mm wide, ventral side smooth or more commonly with up to 7 thick 73 nerves extending to top of achene, beaks gradually tapering from body of perigynia, concolorous with body or darker, 0.8-1.7 mm long, up to 2/5 length of the perigynia, averaging 1/3, distance from top of achene to beak base <0.1-0.9 mm; achenes elliptic to quadrate, 1.3-1.9 mm long, 0.9-1.4 mm wide, filling up to 2/3 of perigynium body, averaging 1/2. In California from Humboldt Co. south to Santa Barbara Co., and in the Santa Cruz Islands (Fig. 17). Along the coast in moist swales and marshes, in moist to boggy open areas into the Coastal Ranges, up to 600 meters. Representative Specimens. California: Humboldt Co., Eureka, Tracy 13811 (CAS, UC). Marin Co., near Mill Valley, Howell 19523 (CAS). Mendocino Co., Westpoint Union Landing St. Park, Whitkus 3011 (NY). Monterey Co., near Monterey, Brewer 697 (UC). San Francisco Co., Presidio, Howell 13006 (CAS, DS, UC, WTU). San Luis Obisco Co., Cambria, Hardham 6183 (CAS). San Mateo Co., near South San Francisco, Howell 13782 (CAS). Santa Barbara Co., Santa Barbara, May 1, 1951, Pollard s. n. (CAS). Santa Clara Co., 14.4 km W of Gilroy, Whitkus 2884 (NY). Santa Cruz Co., San Vicente Road, 11.2 km from Empire Grade, Hesse 1637 (CAS). Sonoma Co., S of Plantation, Whitkus 2989 (NY). Carex mariposana L. H. Bailey ex Mackenzie, Bull. Torrey Bot. Club 43: 619. 1917. C. albomarginata Mackenzie ex Parish, PI. World 20: 176. 1917. nom. nud. -Type: California: Tuolumne Meadows, Sierra Nevada, 8,800 ft., July 20, 1911, Jepson 4476 (holotype: NY!). Carex paucifructus Mackenzie, Bull. Torrey Bot. Club 43: 615. 1917. -Type: California, Eldorado Co., Devil’s Basin, 8,300 ft., July 19, 1897, Brainerd 200 (holotype: ). 74 Culms stiff to ±lax, 20-81 (91) cm tall, exceeding the leaves; leaves straight, blades 1.6-3.7 (3.9) mm wide, flat; inflorescences variable, large and coarse to slender and moniliform to small and aggregated, 16.6-40.5 (48.1) mm long, 7.2-16.9 (22.6) mm wide, first two internodes 2.8-17.5 (27.6) mm long, comprising up to 1/2 length of the inflorescence, averaging 1/3; spikes 3-12, majority separate and distinct, or loosely aggregate to aggregate with the lower spikes usually separate and distinct, 6.0-12.6 (14.4) mm long, 3.7-7.8 (9.2) mm wide; bracts scale-like or lower bracts expanded and awned, awn up to half the length of the inflorescence; scales lustrous, light chestnut to red-chestnut to brown or hyaline, (2.5) 2.7-4.4 (4.7) mm long, 2/3 to as long as perigynia, 1.0-1.7 (2.0) mm wide, centers green, turning lighter or hyaline, margins concolorous with scales to wide hyaline; perigynia lustrous, stramineous to light coppery, upper margins green turning stramineous, membranous, plano-convex, lance-ovate to ovate, 3.4- 5,3 mm long, 1.1-2.2 mm wide, wings mostly less than 0.2 mm wide, ventral side with up to 8 distinct nerves to the top of achene, often extending into beak, beaks gradually tapering from body of perigynia, red-coppery to brown, 1.1-2.3 mm long, up to 2/5 length of the perigynia, averaging 1/3, distance from top of achene to beak base 0.1-1.1 mm; achenes elliptic to ovate, 1.5-2.0 mm long, 0.9-1.3 mm wide, filling up to 2/3 of perigynium body, averaging 1/2. In the Klamath, North Coast and Sierra Nevada Mountains of California, and Washoe Co., Nevada (Fig. 18). Moist meadows and slopes in montane to subalpine habitats from 1200 to 3200 meters. See discussion under C.abrupta. Representative Specimens. California: Alpine Co., Ebbetts Pass, Whitkus 3885 (NY). Butte Co., Butte Mt.,Hall 9792 (UC). Fresno Co., G ranite Basin, Kings Canyon Natl. Park, Aug. 8, 1946, Leschke s. n. (CAS). Lake Co., Lakeport, 75 Eastwood & Howell 5723 (CAS). Mariposa Co., Porcupine Flats, Yosemite Natl. Park, Howell 20697 (CAS). Placer Co., N of Bowman, Smith 1904 (JEPS). Shasta Co., Mt. Lassen, Sep. 15, 1945, Leschke s. n. (CAS). Siskiyou Co., Marble Mt. Wilderness, Oettinger 733 (UC). T rinity Co., T rinity Mts., Alexander & Kellogg 5469 (DS, RM). Tulare Co., Hamilton Lks., Howell 17655 (CAS). Tuolumne Co., Tuolumne Meadows,Howell 20071 (CAS). Yuba Co., Browns Valley, Rose 40380 (CAS, UC). Carex subfusca W. Boott in S. Wats., Bot. Calif. 2: 234. 1880. C. macloviana var subfusca (W. Boott in S. Wats.) Kukenth., Pflanzenr. IV. 20 (Heft 38): 197. 1909. -Type: Lake Tahoe, Kellogg. Nevada: Virginia City, Bloomer. Carex festiva var. stricta L. H. Bailey, Mem. Torrey Bot. Club 1: 51. 1889. C. festiva var. horneri Piper, Contr. U. S. Natl. Herb. 11: 164. 1906. C. macloviana var. stricta (L. H. Bailey) Kukenth., Pflanzenr. IV. 20 (H eft 38): 196. 1909. -Type: California, Palmer 389. Buck Creek, Oregon, Howell 938\. Carex teneraeformis Mackenzie, Bull. Torrey Bot. Club 43: 609. 1917. -Type: California: Butte Co., in shade of thicket, Jonesville, 5,100 ft., July 25, 1914, Hall 9781 (holotype: NY!; isotype: UC!). Carex stenoptera Mackenzie, Erythea 8: 22. 1922. -Type: California: Ice House Canon, San Antonio Mts.,Johnston 1505 (holotype: NY!; isotype UC!). Culms stiff to lax, 20-81 (104) cm tall, exceeding the leaves; leaves straight, blades 1.2-3.3 (3.7) mm wide, flat; inflorescences 11.0-27.1 (29.5) mm long, (5.4) 6.3-14.4 (16.3) mm wide, first two internodes 1.9-10.7 (16.3) mm long, 76 comprising up to 1/2 length of the inflorescence, averaging 1/3; spikes 4-11 (14), majority separate and distinct, or loosely aggregate to aggregate with lowest spikes usually separate and distinct, 5.0-9.0 (10.5) mm long, (2.7) 3.0-6.5 (7.1) mm wide; bracts scale-like or the lower bracts slightly expanded and awned, awn occasionally as long as inflorescence; scales lustrous, mostly chestnut, varying from pale chestnut to chestnut-red or chestnut-brown to dark brown, 2.1-3.3 (3.9) mm long, 2/3 to as long as perigynia, 0.8-1.6 mm wide, centers green turning white, margins hyaline; perigynia lustrous, pale to shiny stramineous, sometimes light brown, upper margins green turning stramineous, membranous, plano convex, lance-ovate to ovate, (2.4) 2.6-4.0 (4.3) mm long, 0.9-1.7 (1.9) mm wide, wings mostly less than 0.2 mm wide, ventral side smooth or nerved towards base or with up to 6 distinct (thick) nerves to top of achene, beaks gradually to +abruptly tapering from body of perigynia, pale to light chestnut, or dark lustrous brown, 0.8-1.7 (2.1) mm long, up to 1/2 length of perigynia, averaging 2/5, distance from top of achene to beak base <0.1-0.8 (1.0) mm; achenes elliptic to +quadrate, 1.0-1.6 mm long, 0.7-1.2 mm wide, filling up to 3/4 of perigynium body, averaging 1/2; « = 42. Found throughout the mountains of Washington, Oregon, western Idaho, California, Nevada, Utah and Arizona; also in Grant Co., New Mexico (Fig. 19). In seasonally wet habitats that turn dry during the season, in meadows, along watercourses and open fields, from 100 to 3500 meters. Several specimens have been located with Kellogg’s and Bloomer’s names, but without the exact localities given by Boott. Until such time as they can be located, no lectotype can be chosen. In the case of C. festiva var. stricta, three specimens arc listed by Bailey, one of which, Kellogg & Harford 1073, was chosen as the holotype for C. harfordii. 77 From the two remaining specimens, Mackenzie (1931) indicates Palmer 389 is the type because it is the first cited specimen. As there is no indication at this time that Mackenzie actually examined the specimen, a lectotype for this name is still lacking. This should be resolved when the specimen is located. Representative Specimens. Washington; Clallam Co., Olympic Mts.,Elmer 2702 (WS). K itsap Co., K ingston, Oils 1450 (WS). S kagit Co., R ain y Pass, Whitkus 2588 (NY). Skamania Co., Butterfly Lk., Suksdorf 3130 (WS). Oregon; D ouglas Co., Anchor, Peck 7778 (WILLU). Jackson Co., 27.2 km E of Ashland, Peck 15022 (DS, WTU). Josephine Co., 17.6 km NW of Grants Pass, Gould 1061 (CAS, UC). Klamath Co., Lake-of-the-Woods, Peck 16954 (WILLU). Lake Co., Warner Mts., M unz 18444 (CAS). Linn Co., jt. of Rts. 22 and 126, Whitkus 2446 (NY). Umatilla Co., Blue Mts., Hitchcock & Muhlick 13795 (CAS, DS, RM, UC). Wasco Co., near M aupin, Peck 17347 (WILLU). California; Alpine Co., N side of Pigeon Flat, Hoover 5352 (CAS). Butte Co., Chico Meadows, Heller 12018 (CAS, DS, UC). Del Norte Co., 11.2 km E of Smith Riv., Parks & Tracy 11485 (UC). Eldorado Co., Luther Pass, July 26, 1936,Cantelow s. n. (CAS). Fresno Co., Tollhouse, Howell & Barneby 29347 (CAS). Humboldt Co., Dinsmore, Eastwood & Howell 4792 (CAS). Kern Co., Tehachapi-Kernville region, Twisselmann 2492 (CAS). Lake Co., L akeport, Bradshaw 116 (UC). Lassen Co., Butte Lk., Gillett 913 (CAS, JEPS). Los Angeles Co., Big Pines, Thorne & Thorne 38559 (CAS). Madera Co., Pumice Flats Rec. Area, Raven 3165 (CAS). Mariposa Co., Yosemite Valley, Abrams 4401 (CAS, DS). Modoc Co., Cedar Pass, Whitkus 4487 (NY). Mono Co., NE of Bridgeport, Alexander & Kellogg 3848 (JEPS, UC). Monterey Co., Santa Lucia Mts., Howitt 1326 (CAS). Napa Co., 10.2 km SW of Middletown, Johnson 646A (CAS). Nevada Co., Donner Pass, Howell 18311 (CAS). Placer Co., Lake Tahoe, Howell 18871 (CAS). Plumas Co., Johnsville, Howell 27651 (CAS). Riverside Co., Pipe Ck., San 78 Jacinto Mts., Munz 5801 (UC). San Bernardino Co., Lk. Arrowhead, Munz 13248 (UC). San Diego Co., Cuyamaca St. Park, Fiker 3563 (UC). Santa Cruz Co., 4.8 km from Boulder Ck., Hesse 1119 (CAS). Shasta Co., Burney Springs, Peirson 10865 (CAS). Sierra Co., Sierra Buttes, Hall & Babcock 4487 (DS). Siskiyou Co., Mt. Shasta, Stacey 13732 (OCS). Tehama Co., M ineral, Cooke & Cooke 52774 (CAS). T rinity Co., above Trinity Alps Resort, Aug. 22, 1953, Pollard s. n. (CAS). Tulare Co., 27.2 km N of Johnsondale, Whitkus 3137 (NY). Tuolumne Co., Pinecrest, Quick 52-202 (CAS). Ventura Co., Mt. Pinos, Hall 6572 (UC). Nevada: Clark Co., Lee Canyon, Clokey 8662 (CAS, DS, ORE, UC, WTU). Douglas Co., Glenbrook, July 25, 1935, Rose s. n. (CAS). Storey Co., Adams 189 (RM). Washoe Co., 3.8 km from Incline, Howell 14016 (CAS). Arizona: Cochise Co., Chiricahua Mts., Blumer 1401 (DS). Coconino Co., Shulz Pass, near Flagstaff, Hanson 250 (CSC). Gila Co., W side of Sierra Ancha Mts., Gould 3574 (UC). Pima Co., Santa Catalina Mts., Thornber 7676 (CAS). New Mexico: G rant Co., mountains near Pinos Altos, June 26, 1936, Stewart s. n. (CAS). Utah: San Juan Co., N slope of Aba jo Mts., Howell & True 44946 (CAS). Wayne Co., Thousand Lake Mt., Lewis 216 (CAS). Idaho: Washington Co., Weiser Arm, Lomasson 89 (RM). Carex pachystachya Cham, ex Steud., Syn PI, Glum. 2: 197. 1855. C. festiva var. pachystachya (Cham, ex Steud.) L. H. Bailey, Mem. Torrey Bot. Club 1: 51. 1889. C. macloviana var. pachystachya (Cham, ex Steud.) Kukenth., Pflanzenr. IV. 20 (Heft 38): 197. 1909. C. macloviana ssp. pachystachya (Cham, ex Steud.) Hulten, FI. Alaska and Yukon 2: 138. 1942. -Type: Alaska: Unalaska, Aleutian Islands, Chamisso s. n. (isotype: GH!). Culms stiff to ±lax, 15-85 (120) cm tall, exceeding the leaves; leaves straight or curved, blades 1.2-4.2 (6.5) mm wide, fiat; inflorescences (9.0) i 0.0-21.9 79 (23.8) mm long, (6.7) 7.1-15.8 (16.3) mm wide, first two internodes 1.8-7.3 (9.0) mm long, comprising up to 2/5 length of the inflorescence, averaging 1/3; spikes 3-9 (13), loosely aggregate to aggregate, 4.4-8.8 (9.9) mm long, 3.2-7.8 (8.7) mm wide; bracts scale-like, lowest occasionally awned, awn shorter than inflorescence; scales lustrous, reddish to dark brown or coppery, 2.2-3.8 (4.2) mm long, 2/3 to nearly as long as perigynia, 0.9-1.7 (1.9) mm wide, centers mostly concolorous with scales, margins concolorous with scales to white hyaline; perigynia lustrous, reddish to coppery brown or nearly black, upper margins green, turning concolorous with perigynia, membranous, plano-convex, ovate to elliptic-ovate, 2.8-4.7 (5.1) mm long, 1.1-2.1 (2.3) mm wide, wings less than 0.2 mm wide, ventral side smooth, nerved at base, or with up to 6 nerves to top of achene, 1 or 2 extending into beak, beaks gradually to ±abruptly tapering from body of perigynia, concolorous to darker than perigynia, 0.8-2.0 (2.2) mm long, up to 1/2 length of the perigynia, averaging 1/3, distance from top of achene to beak base <0.1-0.8 (1.0) mm; achenes elliptic to ovate, 1.2-1.9 mm long, 0.7-1.3 mm wide, filling up to 2/3 of perigynium body, averaging 1/2; n = 37, 38, 39, 41. C. pachystachya ssp. pachystachya Carex festiva var. gracilis Olney ex W. Boott in S. Wats., Bot Calif. 2: 234. 1880. C. festiva var. gracilis Olney in Gray, Proc. Am. Acad. Arts, 8: 407. 1872. nom. nud. C. multimoda L. H. Bailey, Bot. Gaz. (Crawfordsville) 21: 5. 1896. C. macloviana var. gracilis (Olney ex W. Boott) Kukenth., Pflanzenr. IV. 20 (Heft 38): 197. 1909. C. pachystachya var. gracilis (Olney ex W. Boott) Mackenzie, N. Am. Flora, 18: 136. 1931. -Type: Oregon, 1871, Hall 586 (isotype: BUF!) Carex pyrophila Gandoger, Bull. Soc. Bot. Fr. 60: 420. 1913. 80 -Type: Kamtschatka, Peninsula, Siberia, Komarov 3286 (holotype: LE, photo: ALTA!; isotype: LYl). Carex olympica Mackenzie, Bull Torrey Bot. Club, 43: 610. 1916. -Type: Sequin, Washington, 1915,J. M. Grant 703 (lectotype: NY!). Carex pachystachya var. monds-coulteri Kelso, Biol. Leafl. 64: 2. 1953. C. pachystachya f. monds-coulteri (Kelso) F. J. Hermann, Leafl. W. Bot. 9: 16. 1959. -Type: Aspen, Pitkin Co., Col. Kelso 6662 (not located). Culms 17-89 (120) cm tall, inflorescences variable, (9.9) 10.3-21.9 (23.8) mm long, first two internodes 2.1-7.3 (9.0) mm long; spikes loosely aggregate to aggregate, the lowest often separate and distinct; perigynia 3.0-4.7 (5.1) mm long, variable in color, from light reddish or brown coppery to nearly black, ventral side smooth or nerved at base or with up to 6 nerves to top of achene, beak margins extending 0.5-1.6 mm; achenes filling up to 4/5 of body of perigynia, averaging 2/3; n = 37, 38, 39. From southern coastal Alaska, although extending inland to the Alaska Range, south to California in Coast Ranges to Marin Co., in the Sierra Nevada to Tuolumne and Mono Cos., in the Rocky Mtns. of Colorado (Glenn Co.) and Unita Mts. of Utah (Summit Co.; Fig. 20). In moist locations from coastal to subalpine habitats, sea level to 3000 meters. The lectotype for C. olympica was chosen from among the syntypes collected in Washington State. This agrees with the author’s intentions since Mackenzie (1931) indicates the type is from the state of Washington. The collection from the Olympic Mountains,Elmer 2700, is actually a specimen of C. preslii. Of the remaining collections. Grant 703 .is the most complete, fits the 81 original description, and was examined by Mackenzie, and is thus chosen as the lectotype. The type for C. pachystachya var. monds-coulteri could not be located. This has been a problem for other workers in Carex who have tried to find Kelso types (Standley 1985; per. comm.). Since the taxon is based on the variable character of nerves on the ventral side of the perigynium, and the original description fits the subspecies pachystachya, it most likely belongs as a synonym under this subspecies. Representative Specimens. Canada. British Columbia: 200 km S of Dease Lk. on Rt. 37, Whitkus 1787 (ALTA). 73 km S of Meziadin Jt. on Rt. 37, Whitkus 1826 (ALTA). Nelson, Eastham 10037 (RM, WS). US-CAN b o rd e r betw een K ettle and Columbia Rivs., Macoun 63308 (WS). V ancouver, Eastham 11383 (WS). Y ukon Territory: Carcross, Henderson 14781 (ORE). U. S. A. Alaska: Expedition Isl., U nalaska, Jepson 202 (JEPS). 0.8 km fro m H aines, Viereck 8512 (RM). Moose Pass, Kenai Pen., Whitkus 1514 (ALTA). Ketchikan, B aker 355 (ORE). Mendenhall near Juneau, Anderson 6198 (RM, WTU). Washington: Chelan Co., 4 km NW of Ardenvoir, Hermann 19014 (WTU). Clallam Co., Deer Lk., Eyerdam 6328 (WS). C lark Co., 4.8 km W o f O rchards, Whitkus 2496 (NY). Columbia Co., Rucannon Riv., Blue Mts., Darlington 232 (RM, WS). G rays H a rb o r Co., S end of Humptulips,Whitkus 2244 (NY). King Co., Seattle, June 9, 1884, Piper s. n. (WS). Kitsap Co., Port Orchard, Eyerdam 1588 (WS). K ittita s Co., Johnson’s Ck. near m outh, Thompson 9526 (WTU). Klickitat Co., near Glenwood, Suksdorf 6199 (WS). Lewis Co., Chehalis, Whitkus 2247 (NY). Linn Co., 4 km E of Corvallis, Lawrence 1575 (OSC). Mason Co., Shelton, Becking 270 (WTU). Okanogan Co., Disautel, Fiker 1197 (WTU). Pend Oreille Co., 3.2 km S of Tiger Pass, Whitkus 2626 (NY). San Juan Co., Cattle Pt., Peck 12676A (WS). S kagit Co., H id d en L ake M eadows, 82 D e N e ff 673 (WS). Skam ania Co., 1.6 km E o f B onneville D am , Hitchock 13752 (WTU). Spokane Co., Newman Lk., Suksdorf 8785 (WS). Thurston Co., Mud Bay, Otis I9 I4 (WS, WTU). Whatcom Co., E end of Newhalem, Whitkus 2577 (NY). Whitman Co., head of Rock Lk., Beattie & Lawrence 2381 (WS). Y akim a Co., Mt. Adam s, Suksdorf 5800 (WS). Oregon: Baker Co., 9.6 km NW of Haines, Hermann 18784 (WTU). Benton Co., Mary’s Pk., Whitkus 2300 (NY). Clackamas Co., Timberline Lodge, Whitkus 2469 (NY). Clatsop Co., Saddle Mt., June 10, 1928, Paterson s. n. (ORE). Columbia Co., 1.6 km SW of Berkenfield, Whitkus 2267 (NY). Coos Co., S Fork Coquille Riv. near Powers, Henderson 10050 (ORE). Curry Co., 4.8 km N of Carpenterville, Whitkus 2309 (NY). Deschutes Co., 1.6 km N of Three Creeks Lk., Halpern 585 (OSC). Douglas Co., 32 km W of Crater Lk., Peck 3379 (WILLU). Hood River Co., Mt. Hood, July 24, 1884, Henderson s. n. (ORE). Jackson Co., 14.4 km NE of Ashland, Whitkus 2352 (NY). Jefferson Co., Camp Sherm an, Peck 2586 (WILLU). Josephine Co., 3.2 km S of Takilma, Peck 8030 (WILLU). Klamath Co., Ft. Klamath, Peck 19699 (WILLU). Lane Co., 43.2 km SB of Oakridge, Peck 22063 (WILLU). Linn Co., Clear Lk., Sheldon 12680 (ORE). Marion Co., Salem, Peck 1237 (WILLU). Multnomah Co., Savvies Isl., Thompson 4237 (WTU). Tillamook Co., 1.6 km E of Sunset Summit, Whitkus 2273 (NY). Umatilla Co., Meacham, Peck 3388 (WILLU). Wallowa Co., Ice Lk., S of Enterprise, Cole 219D (ORE). California: Del Norte Co., Shelly Ck., Eastwood & Howell 3693 (CAS). Eldorado Co., Meeks Bay, Whitkus 4081 (NY). Glenn Co., Plaskett Meadows, Howell 19236 (CAS). Humboldt Co., Eureka, Tracy 13938 (UC). Marin Co., Olema, Henderson 15321 (ORE). Mendocino Co., Philo, Eastham, Howell & Stacey 4397 (CAS). Plumas Co., Drakesbad, Howell 35958B (CAS, OSC). Shasta Co., Lower Twin Lk., Lassen Natl. Park, Leschke 1708 (CAS). Siskiyou Co., Mt. Shasta, Cooke 13732 (DS). Sonoma Co., near Forestville, Rubtzoff 112 (CAS). 83 Tehama Co., 8 km S of Childs Meadow, Whitkus 4319 (NY). Trinity Co., between Eagle and Bear Cks., Eastwood & Howell 4980 (CAS). Tuolumne Co., near Pinecrest, Hesse 2275 (CAS). Nevada; Washoe Co., near Mt. Rose, Howell 14141 (WTU). Idaho: Boise Co., headwaters of S Fork Payette Riv., Hitchcock & Muhlick 9856 (WS). Bonner Co., Indian Ck., Daubenmire 44345 (WS). C learw ater Co., 1.6 km W of Bovill-Elk Rivs. summit, Cronquist 5881 (WTU). Idaho Co., 4.8 km W of Low ell, Jones 59 (WTU). Valley Co., Payette Lk., Ertter 4073 (WTU). Montana: Flathead Co., 28 km WNW of Whitefish, Stickney 2222 (RM). Madison Co., W Fork Madison Riv., Kovalchik 314 (RM). Mineral Co., 20.8 km E of Wallacea, Cronquist 6722 (ORE, WTU). Missoula Co., Lolo Pass, Hitchcock & Muhlick 14652 (WTU). Wyoming: Teton Co., Treasure Mt., South Camp,Anderson 397 (RM). Utah: Summit Co., 11.2 km E of Kamas, Whitkus 4530 (NY). Colorado: Grand Co., Buffalo Park Rd., 5.4 km N of Gore Pass Rd., Colson 78-54 (RM ). C. pachystachya ssp. compacta Whitkus subsp. nov. -Type: Alaska: Coeur d’Alene Campground, 12km from Hope,Whitkus 1511 (holotype: ALTA; isotype NY). Culms 16-56 cm tall; inflorescences mostly compact, 9.0-17.6 mm long, first two internodes 1.8-5.5 (5.8) mm long; spikes aggregate, rarely lowest spike separate; perigynia 2.8-4.1 mm long, dark brown coppery or coppery to nearly black, upper margins green, turning concolorous with body of perigynia, ventral side smooth or with up to 3 nerves extending to top of achene, at least 1 extending into beak, beaks dark coppery brown to black, beak margins extending 0.3-1.2 (1.4) mm; achenes filling up to 3/4 of body of perigynia, averaging 1/2; n = 41. 84 Coastal southern Alaska south to northern California (Mt. Lassen) and in the Rocky Mtns. to Colorado (Ouray Co.; Fig. 21). In moist locations near sea level in northern part of range, more commonly in montane to subalpine habitats from 550 to 3500 meters. This subspecies comprises the taxon ’stubby’ in this and previous works (Whitkus and Packer 1984; Whitkus 1988). The subspecies compacta differs from the typical subspecies in the key characters of a more congested inflorescence and darker scales and perigynia. It occurs in generally higher elevations and is typically found in high montane to subalpine habitats. Representative Specimens. Canada. British Columbia: Chilkat Pass on Haines Rd., Whitkus 1691 (ALTA). Mountains above Kennedy Riv., Vancouver Isl., Colder & Mackay 32016 (RM). 17.6 km W of Revelstoke, Hitchcock & Martin 7552 (WTU). Rogers Pass, Whitkus 2042 (ALTA). U. S. A. Alaska: Hope, Anderson 6674 (RM). Moose Pass, Anderson 6841 (WTU). Road from Hyder to Stewart, Whited 1157 (WS). Washington: Chelan Co., Mt. Ingalls, July 4, 1937, Eyerdam s. n. (WS). Clallam Co., Hurrican Ridge, Whitkus 2209 (NY). Columbia Co., Tucannon Riv., Barkworth 456 (WS). King Co., Stevens Pass, Whitkus 2564 (NY). Lewis Co., 3.2 km W of White Pass, Whitkus 2508 (NY). Pend Oreille Co., Abercrombie Mt., Layser 1231 (WS). Pierce Co., Reflection Lk, July 18, 1919, Flett s. n. (WS). Skagit Co., Rainy Pass, Whitkus 2586 (NY). Skamania Co., Peter’s Prairie, Suksdorf 3140 (WS). Thurston Co., Capitol Pk., Meyer 1632 (WS). Whatcom Co., Nooksack Riv., Suksdorf 3119 (WS). Yakima Co., 3.2 km E of Chinook Pass, Whitkus 2527 (NY). Oregon: Benton Co., Mary’s Pk., Whitkus 2278 (NY). Clackamas Co., Mt. Hood, Nelson 4768 (OSC). Clatsop Co., Saddle Mt., Howell 28426 (OSC, WTU). Deschutes Co., Elk Lk., Whitkus 2390 (NY). Grant Co., John Day River Valley, Henderson 5308 (ORE). Hood R iver Co., Mt. Hood, Henderson 972 (ORE). Jackson Co., N of 85 Prospect, Quick 54-60 (CAS). Klamath Co., 3.2 km N of Ft. Klamath, Whitkus 2383 (NY). Lane Co., Horse Pasture Mt., Peck 3287 (WILLU). Linn Co., 4.8 km N of Rt. 126 on Rt. 22, Whitkus 2458 (NY). Marion Co., Salem, Peck 592 (WILLU). Umatilla Co., Langdon Lk., Peck 22238 (WILLU). Wallowa Co., Lostine Canyon, 30.4 km above Lostine, Peck 17759 (DS, WILLU). California: Shasta Co., Hat Lk., Leschke 1602 (CAS). Siskiyou Co., Medicine Lk., Heller 13708 (CAS, DS, RM, WTU). Idaho: Idaho Co., NE of Buffalo Hump,Baker 12468 (CAS). Montana: Gallatin Co., Rat Lk., Hitchcock & Muhlick 15243 (WS). L ake Co., road from R t. 93 to Kicking Horse Res., Lackshewitz 6502 (WTU). Wyoming: Bighorn Co., head of Granite Ck., M ay 11 (RM). Teton Co., Jackson Hole, Williams 310 (RM). Colorado: Routt Co., Meadows Campground W of Rabbit Ears Pass, Whitkus 4614 (NY). 86 Table 10. Synopsisof literature treatment of members of C.pachystachya complex. Reference Taxon Nature of problem or treatment Whitkus and C. pachystachya Morphological and cytological Packer (1984) variation in a widespread species. Possible seggregate called ’stubby’. Mackenzie (1931) C. pachystachya Connected to C. pachystachya by var. gracilis numerous intermediates. Cronquist (1977) C. mariposana Forms from Sierra Nevada C. abrupta which are possibly no more than varietally distinct forms of C. pachystachya Cronquist (1969, C. subfusca In forms, transitional to 1977) C. pachystachya. Cronquist (1969) C. teneraeformis Not distinct from C. subfusca, a more slender form. Cronquist (1969) C. integra Allied to C. illota but diverges to C. subfusca. Hoover (1970) C. montereyensis A form of C. subfusca. C. subbracteata Forms of C. harfordii. C. gracilior 87 Table 11. Characters used in phenetic analyses and mode of assessment. Character (mode of assessment^ 1. Height of divergence of leaf greater than 3 cm long (cm) 2. Culm height (cm) 3. Leaf blade width (mm) 4. Inflorescence length (mm) 5. Inflorescence width (mm) 6. Length of first two internodes of inflorescence (mm) 7. Distance from base to widest part of inflorescence (mm) 8. Number of spikes in inflorescence (count) 9. Spike length (mm) 10. Spike width (mm) 11. Scale length (mm) 12. Scale w idth (mm) 13. Perigynium cross-sectional shape (flat, distended by achene/plano- convcx/biconvex) 14. Perigynium texture (thick, coriaceous/thin, membraneous) 15. Perigynium base (constricted/not constricted) 16. Perigynium length (mm) 17. Perigynium width (mm) 18. Perigynium margin width (mm) 19. Distance from base of perigynium to widest part (mm) 20. Number of ventral nerves extending to base of beak (count) 21. Beak length (mm) 22. Beak base width (mm) 23. Distance from beak base to wingless portion (mm) 24. Distance from top of achene to beak base (mm) 25. Achene length (mm) 26. Achene width (mm) Table 12. Taxa and morphological groups used in phenetic analyses. C. subfusca (SUBF) ISUBF ’Typical’ C. subfusca 3SUBF Wide-ovate perigynia 4SUBF Very small (<3 mm), thin membraneous perigynia 6SUBF Very small (<3 mm), thin winged perigynia 9SUBF Shiny stramineous-brown perigynia; compact inflorescences 11 SUBF Perigynia with red beaks and smooth, shiny ventral surfaces 12SUBF Plants slender and small; inflorescence slender and loose 13SUBF Plants more robust than 12SUBF; agrees with C. teneraeformis C. pachystachya (PACH) 14PACH ’Typical’ C. pachystachya 17PACH Perigynia green, long tapering into beak 19PACH ’Typical’ gracilis 21 PACH Intermediates between C. pachystachya and stubby 22PACH ’Typical’ stubby 23PACH Perigynia lanceolate, thin margined 24PACH Intermediates between C. pachystachya and C. abrupta C. abrupto (ABRU) 25ABRU ’Typical’ C. abrupta 26ABRU Perigynia light colored, contrasting with dark beaks 27ABRU Intermediates between C. abrupta and C. subfusca 29ABRU Intermediates between C. abrupta and stubby 30ABRU Large, flattened perigynia 31ABRU Perigynia wide ovate, dark glossy brown 33ABRU Intermediates between C. abrupta and C. microptera C. mariposana (MARI) 34MARI Intermediates between C. mariposana and C. abrupta 35MARI ’Typical’ C. mariposana 36MARI Intermediates between C. mariposana and C. festivella 37MARI Lanceolate perigynia 38MARI Perigynia narrow ovate, very strongly nerved ventrally 39MARI Perigynia small (ca. 3.5 mm), thick textured. C. harfordü (HARF) 40HARF ’Typical’ C. harfordii C. montereyensis (MONT) 42MONT ’Typical’ C. montereyensis C. subbracteata (SUBB) 43SUBB ’Typical’ C. subbracteata 44SUBB Perigynia smaller than 43SUBB 45SUBB Perigynia membraneous, not coriaceous 46SUBB Perigynia with thick beak bases C. gracilior (GRAC) 47GRAC ’Typical’ C. gracilior 48GRAC Perigynia wide ovate 49GRAC Intermediates between C. gracilior and C. subbracteata C. integra (INTE) 51INTE ’Typical’ C. integra C. paucifructus (PAUC) 52PAUC ’Typical’ C. paucifmetis 89 Table 13. Characterization of data sets for the phenetic analyses. Analysis Characters OTU’s Groups Cluster analysis 1-26 39 PCA 1-26 39 OTU’s of all taxa DFA 1-12, 16, 17, 19-26 272 Species Pairs MONT-HARF PCA 1-12, 14-26 84 DFA 1-12, 16-26 84 SUBB-GRAC PCA 1-12, 14-26 67 DFA 1-12, 16-18, 20-26 67 SUBF-PACH PCA 1-26 196 DFA 1-12, 16-19-21-26 196 Infraspecific Analyses SUBF PCA 1-12, 16-26 150 DFA 1-12, 16-26 150 PACH PCA 1-12, 16-26 148 DFA 1-12, 16-19,21-26 148 90 Table 14. Factor loadings for first three axes of group PCA. icter Axis I Axis II Axis III 1 0.049 -0.041 -0.391 2 0.464 -0.219 -0.318 3 0.652 0.282 -0.303 4 0.513 -0.383 0.652 5 0.655 0.462 -0.327 6 0.206 -0.536 0.712 7 0.252 -0.527 0.748 8 0.114 0.206 0.060 9 0.829 -0.052 0.292 10 0.661 0.437 -0.313 11 0.947 -0.133 -0.037 12 0.642 -0.504 -0.385 13 -0.137 0.556 0.097 14 0.541 -0.540 -0.319 15 -0.189 -0.553 0.453 16 0.889 0.267 0.234 17 0.752 0.224 -0.458 18 -0.046 0.835 0.110 19 0.883 -0.098 0.337 20 0.287 0.525 0.608 21 0.767 0.386 0.246 22 0.833 0.170 -0.108 23 0.647 0.372 0.482 24 0.150 0.762 0.362 25 0.827 -0.431 -0.092 26 0.687 -0.533 -0.435 91 Table 15. Classification matrix for taxa in DFA of individual OTU’s using ’typical’ groups. Total correct classification for ’typical’ groups is 75.7%. 1 14 19 22 25 35 40 42 43 47 51 52 ISUBF 70.0 0.0 10.0 0.0 0.0 0.0 10.0 0.0 0.0 0.0 10.0 0.0 3SUBF 0.0 0.0 50.0 0.0 0.0 0.0 25.0 25.0 0.0 0.0 0.0 0.0 4SUBF 20.0 0.0 0.0 30.0 0.0 0.0 0.0 20.0 0.0 0.0 30.0 0.0 6SUBF 40.0 0.0 0.0 10.0 0.0 0.0 10.0 0.0 0.0 0.0 40.0 0.0 9SUBF 25.0 25.0 12.5 0.0 12.5 0.0 25.0 0.0 0.0 0.0 0.0 0.0 1 ISUBF 40.0 0.0 20.0 0.0 0.0 0.0 20.0 0.0 0.0 0.0 10.0 10.0 12SUBF 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 100.0 13SUBF 100.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 14PACH 0.0 60.0 0.0 0.0 10.0 10.0 10.0 0.0 10.0 0.0 0.0 0.0 17PACH 0.0 0.0 100.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 19PACH 0.0 0.0 90.0 0.0 0.0 0.0 10.0 0.0 0.0 0.0 0.0 0.0 21 PACH 0.0 20.0 10.0 40.0 0.0 0.0 10.0 20.0 0.0 0.0 0.0 0.0 22PACH 0.0 10.0 10.0 60.0 0.0 0.0 0.0 10.0 0.0 0.0 0.0 10.0 23PACH 16.7 0.0 0.0 16.7 16.7 0.0 50.0 0.0 0.0 0.0 0.0 0.0 24PACH 0.0 11.1 33.3 0.0 33.3 0.0 0.0 0.0 0.0 0.0 11.1 11.1 25ABRU 0.0 10.0 0.0 0.0 80.0 0.0 10.0 0.0 0.0 0.0 0.0 0.0 26ABRU 0.0 10.0 0.0 0.0 80.0 0.0 0.0 10.0 0.0 0.0 0.0 0.0 27ABRU 0.0 0.0 22.2 11.1 44.4 0.0 0.0 11.1 0.0 0.0 0.0 11.1 29ABRU 0.0 0.0 25.0 37.5 25.0 0.0 0.0 12.5 0.0 0.0 0.0 0.0 30ABRU 0.0 20.0 0.0 0.0 80.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 31 ABRU 0.0 0.0 0.0 50.0 50.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 33ABRU 0.0 0.0 0.0 0.0 100.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 34MARI 10.0 10.0 20.0 0.0 10.0 50.0 0.0 0.0 0.0 0.0 0.0 0.0 35MARI 0.0 0.0 0.0 0.0 0.0 88.9 0.0 0.0 0.0 0.0 0.0 11.1 36MARI 0.0 0.0 0.0 0.0 0.0 50.0 0.0 0.0 50.0 0.0 0.0 0.0 37MARI 0.0 0.0 0.0 0.0 0.0 100.0 0.0 0.0 0.0 0.0 0.0 0.0 38MARI 0.0 0.0 0.0 0.0 25.0 75.0 0.0 0.0 0.0 0.0 0.0 0.0 39MARI 0.0 0.0 0.0 0.0 0.0 33.3 0.0 0.0 33.3 33.3 0.0 0.0 40HARF 0.0 0.0 0.0 0.0 0.0 0.0 80.0 20.0 0.0 0.0 0.0 0.0 42MONT 0.0 10.0 0.0 10.0 0.0 0.0 10.0 70.0 0.0 0.0 0.0 0.0 43SUBB 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 80.0 20.0 0.0 0.0 44SUBB 0.0 0.0 12.5 12.5 0.0 0.0 0.0 0.0 62.5 0.0 12.5 0.0 45SUBB 0.0 0.0 0.0 0.0 0.0 0.0 33.3 0.0 66.6 0.0 0.0 0.0 46SUBB 0.0 0.0 0.0 0.0 0.0 0.0 0.0 16.7 66.7 0.0 0.0 16.7 47GRAC 0.0 0.0 0.0 0.0 0.0 10.0 0.0 0.0 10.0 80.0 0.0 0.0 48GRAC 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 100.0 0.0 0.0 0.0 49GRAC 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 100.0 0.0 0.0 51INTE 20.0 0.0 10.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 70.0 0.0 52PAUC 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 100.0 92 Table 16. Factor loadings for the first two axes of thesubbracteata-C. C. gracilior PCA. ;ter Axis I Axiç II 1 0.022 -0.055 2 0.289 0.105 3 0.509 -0.441 4 0.261 0.688 5 0.705 -0.471 6 -0.087 0.908 7 0.174 0.789 8 0.509 -0.331 9 0.688 -0.031 10 0.580 -0.023 11 0.564 -0.043 12 0.368 0.245 14 -0.422 0.056 15 -0.159 -0.122 16 0.734 0.173 17 0.628 0.021 18 0.444 -0.030 19 0.424 -0.050 20 0.035 -0.049 21 0.574 0.427 22 0.631 0.140 23 0.504 0.196 24 0.511 -0.096 25 0.240 -0.049 26 0.401 -0.109 93 Table 17. Classification matrix for C. subbracteata-C. gracilior DFA. Total correct classification is 90.6%. 43 47 43SUBB 89.5 10.5 44SUBB 75.0 25.0 47GRAC 0.0 100.0 49GRAC 0.0 100.0 94 Table 18. Factor loadings for first two axes ofpachystachya-C. C. subfusca PCA. icter Axis I Axis 11 1 0.130 0.499 2 0.193 0.655 3 0.495 0.522 4 -0.239 0.808 5 0.634 0.488 6 -0.432 0.344 7 -0.450 0.625 8 -0.209 0.746 9 0.296 0.710 10 0.654 0.121 11 0.761 0.030 12 0.611 -0.159 16 0.895 -0.013 17 0.763 -0.028 18 0.001 0.481 19 0.659 -0.034 20 -0.134 0.216 21 0.739 0.087 22 0.795 -0.065 23 0.535 0.178 24 0.012 0.162 25 0.791 -0.191 26 0.747 -0.094 95 Table 19. Classification matrix for C. subfusca-C. pachystachya DFA. Total correct classification is 89.8%. 1 14 19 22 ISUBF 94.8 0.0 3.1 2.1 14PACH 0.0 80.6 8.3 11.1 19PACH 0.0 4.8 88.1 7.1 22PACH 0.0 4.5 9.1 86.4 96 Table 20. Classification matrix for C. subfusca DFA. Total correct classification is 57.2%. 1 3 4 6 9 11 12 13 ISUBF 47.3 1.1 7.7 13.2 1.1 19.8 5.5 4.4 3SUBF 0.0 75.0 25.0 0.0 0.0 0.0 0.0 0.0 4SUBF 0.0 0.0 66.6 8.3 16.7 8.3 0.0 0.0 6SUBF 8.3 0.0 0.0 83.3 0.0 0.0 8.3 0.0 9SUBF 0.0 25.0 0.0 0.0 62.5 12.5 0.0 0.0 11 SUBF 0.0 0.0 10.0 0.0 20.0 60.0 0.0 10.0 12SUBF 0.0 0.0 0.0 0.0 0.0 0.0 100.0 0.0 13SUBF 0.0 0.0 0.0 0.0 0.0 0.0 0.0 100.0 97 Table 21. Classification matrix for C. pachystachya DFA. Total correct classification is 87.3%. 14 19 22 14PACH 89.2 5.4 5.4 17PACH 0.0 100.0 0.0 19PACH 4.7 90.7 4.7 21PACH 41.4 6.9 51.7 22PACH 13.6 9.1 77.3 23PACH 83.3 0.0 16.7 24PACH 11.1 55.6 33.3 Figure 5. Groups analysis phenogram based on correlation coefficient and average linkage clustering. Group acronyms follow Table 12 98 99 51INTE 12SVBF 38MARI 39MARI 52PAUC 40HARF 42MONT 23PACH 29ABRU OISUBF 06SUBF 04SUBF nSUBF 13SUBF 24PACH 34MARI 3SMARI 37MARI 36MARI 25ABRU 26ABRU 30 ABRU 33ABRU 27ABRU 31ABRU 14PACH 21PACH 22PACH 03SUBF 09SUBF 17PACH 19PACH 43SUBB 45SUBB 46SUBB 47GRAC 48GRAC 49GRAC 44SUBB Figure 5 Figure 6. Groups analysis phenogram based on average taxonomic distance and average linkage clustering 100 101 5HNTE 12SUBF 52PAUC 40HARF 42MONT 14PACH 17PACH 19PACH H = i 22PACH 21PACH 09SUBF 03SUBF 24PACH 27ABRU 29ABRU 23PACH OISUBF IISUBF 04SUBF 06SUBF 34MARI 38MARI 39MARI 25ABRU 4= 26ABRU 30ABRU -C 33ABRU 31ABRU 43SUBB 45SUBB 6 46SUBB 44SUBB 47GRAC 49GRAC 48GRAC 13SUBF 35MARI 37MARI 36MARI Figure 6 Figure 7. Projection of groups onto the first two principal component axes. Group numbers follow Table 12, ellipses enclose species of Table 12 102 33 25 27 36 '23 21 3 L 22 % 38 45, 43, 52 l4 7 39] II O Figure 7 w Figure 8. Projection of groups onto principal components axes I and III 104 37 35 Q8 34 52 33 30 3 8 '24i '49 4 8 22 42 4 6 ? O Figure 8 Ui Figure 9. Projection of group centers onto the first two canonical axes of the DFA of individuals 1 0 6 30 26 14 2 ^ 35 3y 34, 22 46 43 52 39 i49 47 o Figure 0 - j Figure 10. Illustration of F-statistics from DFA of individuals. Within each species, groups not significantly different at p = 0.05 level are joined 108 109 6 11 SUBF 1 3 9 f \ PACH 14-17-21 24 19 25 \ 27 ABRU 3 0 -3 1 / I 26 29 MAR I 34-36-35- 37 39 HARF-MONT 4 0 -4 2 SUBB-GRAC 44— 43— 48— 49—47 4 ' s ^ Figure 10 Figure 11. Projection of ellipses of C.montereyensis and C. harfordii onto first two principal component axes. H = type of C.harfordii', M = type of C. montereyensis 110 42 40 Figure 11 Figure 12. Projection of group ellipses for C.subbracleata and C. gracilior onto first two principal component axes. G = type ofC. gracilior, S = type of C. subbracleata 112 47 49 43 44 45 Figure 12 Figure 13. Projection of ellipses of ’typical’ groups ofsubfusca C. and C. pachystachya onto first two principal component axes 114 22 19 Figure 13 Figure 14. Projection of group ellipses of C.subfusca onto first two principal component axes. S = type specimens of C.subfusca\ T = type specimens of C. teneraeformis 116 ,13 Figuro 14 Figure 15. Projection of ’typical’ group ellipses ofpachystachya C. onto first two principal component axes 19 22 Figure 15 Figure 16. Projection of group ellipses of C.pachystacnya onto first two principal component axes. OTU’s of known chromosome number are shown as # : n = 41; A: n = 39; ■ : n = 38; O: « = 37 120 22 19i 14 Figure 16 Figure 17. Distribution of C.integra 122 123 Figure 17 Figure 18. Distribution of C.abrupta 124 125 % Figure 18 Figure 19. Distribution of C.subbracteata ssp. subbracleata 126 127 Figure 19 Figure 20. Distribution of C.subbracteata ssp. gracilior 128 129 Figure 20 Figure 21. Distribution of C.harfordii 130 131 Figure 21 Figure 22. Distribution of C.mariposana 132 133 Figure 22 Figure 23. Distribution of C.subfusca 134 135 Figure 23 Figure 24. D istribution of C. pachystachya ssp. pachystachya. Closed circles represent specimens examined in this study, open circles are collections cited in Whitkus (1981). 136 137 O o O • Figure 24 Figure 25. D istribution of C. pachystachya ssp. compacta. Closed circles represent specimens examined in this study, open circles are collections cited in Whitkus (1981). 138 139 Figure 25 SUMMARY This Study has demonstrated that the morphological complexity seen in the C. pachystachya complex is mirrored by similar complex patterns in chromosomal and genetic relationships. The data strongly suggest a recent origin of the group, 20,000 years or less, to account for such complex patterns. In addition, the evolution must have been rapid to allow enough morphological variation to have accumulated for the recognition of seven taxonomic species and two subspecies, with at least one subspecies being genetically and chromosomally distinct enough to be considered a biological species. Finally, a possible mechanism for the rapid evolution has been shown by seeing how agmatoploidy, combined with chromosomal structural rearrangements and inbreeding, allows reproductive isolation to develop rapidly. These isolated populations then can become fixed for different isozyme alleles, and probably other alleles, resulting in rapid genetic divergence. 140 PLEASE NOTE: Copyrighted materials in this document have not been filmed at the request of the author. They are available for consultation, however, in the author's university library. These consist of pages: 141-148 University M icrofilms Internationa! 300 N. ZEES RD.. ANN ARBOR, Ml 48106 13131 761-4700 APPENDIX B DETERMINATION OF MAXIMUM OBSERVED AFFINITY PAIRING FORMULA or. 37 21 22II 3III 3IV IV IVI 31 26II 4III IV IVI 41 19II 2III 5IV 2V 31 24II 3III 3IV IVI 1. Subtract largest multivalent (IVI) from all configurations. present. use next largest multivalent and univalents. 21 22II 3III 3IV IV 31 26II 4III IV 31 19II 2III 5IV IV 31 24II 3III 3IV 2. Repeat Step Iwith IV. 21 22II 31II 3IV 31 26II 4III 31 19II 2III 5IV 21 24II 3III 2IV 3. Repeat Step 1 with 5IV. Use bivalents if necessary. 22II n i l 11 23II 31 19II 2III 11 23II 4. Repeat Step 1with 2III. Again, use bivalents if necessary. 1 T II 20II 31 19II 11 20IÎ 5. Pick out largest number of bivalents and fewest univalents, i.e.: II 20II. 6. MOAP = II 20II 2III 5IV IV IVI. 149 APPENDIX C TABLE OF ALLELIC FREQUENCIES O ooo o oo oo oo oo oo ooo o ooo o o o O o o o ooo oo oo ooo oo oo oo o o o o o op oo o oo o o oo o oo oo o ooo o o o oo ooo o o o o o o o o o o• • o o z rvj AJ oo ooo o o oo oo o oo o o o o o oo o o ro oo o o o o o oo oo oo oo o o o o o o o eo ooooo o ooo ij o o o oo o o oo o o o o o o dod d o d o o o o o o d^ o o o z M Kl Kl N* N o Kî to Kl Kl Kl sO o o o oo oo oo o oo o ooo oo o ooo o o o rj o oo o ooo o o oo oo oo o o o oo o o 00 o oo ooo ooo o oo o ooo oo ooo o o o o oo d od o o o oo o o d o d o z 9^ 9. O' 'O o owo oo ooo o ooo o o oo o oo o o K oo o o o o oo oo oo o oo o o o o oo o o N o oo ooo o o ooo oo ooooo ooooo o o o o oo o o o o o oo o oo o z lA lA LA Kl to s. N. o o oo o oo o o oo ooo oo o ooo o o o 00 oo oo oo oo o o oo o oo o o o oo o o o o N ooooo o o oo o o oo ooo o ooooo o o o o o oo o oo o o oo oo oo o ^ o o ZN N tw O' O' A i OJ o oo o o o o o oo oo o o oo oo ooo %» ooo o o ooo oo oo ooo o oo oo o i n oo o o o o o oo o o oo o ooo ooo o ill oo ooo oo o oo o oo oo o z Kl Kl Kl o 'O LA N. o o o o o o o oo ooo o ooo oo oooo o o o o o o oo o o o o o oo ooo o o o i n o o o o o oo oo o o o ooo oo o o o oo o o o o o o od o o «J o o oo o od o 2 Sf o oo o o o o oo oo oo o o ooooo o o o o oooo o o oo oo o o o ooooo o o o 9» o o o o o ooo o oo o oo oooo oo oo o o o od d od oo o o oo od o o o Z O o o AJ A i o* t n LA LA o oooo o o oo oo o o o oo o oo oC5 o o o S. o ooo o ooo o o o oo o ooo o o o o « ooo oo o oo o o o o oo o o oooo g g g oo d od d o o o o o d dd d «Jd z Kl Kl Kl Kl Kl LA oooo oo c oo o oo o oo ooo ooo o N. ooo o o ooo o o o cooooo o o o o o to o w w c o oo ©o Q o ooo o o ws G Ô o “ o d od d d d «J d d d d «J d o d d d •- o o z OJ r\i rvj s . h— '9 'O ooo o o oo o o o o oooo o oo oo o o o o oooo o o ooo o oo o oo o o o o o o o o mo os oLA LA LA LA o o o oooo o ooo o o o ood d d o o o oo o o ovZd o do o^ ^ o o z Kl Kl Kl ooo oo o o o o o oo o oo o o o_ _ o_ o_ opoo o o o o o oo oo o o o oooo oo o oo o «f ôôôôôôo oo o o oo ôôôo o o ôôo oôô o ooôôôôôo o oo oooo o ^0d— — d— d— d— o — — d— o — V-— — o—' d d do ^ o ^o «Jd « - 0 0 z 00 <0 co 'O N o o ooo oo oo o oo oo o oo o ooo o o o o fM ooo ooo ooo o o o oo ooo oo o o o o O o o ooo ooo oo coAJ o o oo o oLA o o o o (NJ oo o o o o o o oo o o o oo o Z 9s O' O' o o O' t tu O n ALLELE N 2197 N 2528 N 2440 N 2360 M 2469 N 2284 N 2276 N 2172 N 3076 N 2343 N 2344 N 3905 N 4418 LAP1 16 19 40 37 19 19 22 19 0 20 19 0 0 A 0.000 0.000 0.000 0.050 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.950 0.650 0.380 0.000 0.320 0.360 0.000 0.000 1.000 0.000 0.000 0.000 C 1.000 0.050 0.350 0.570 1.000 0.680 0.640 1.000 0.000 0.000 1.000 0.000 0.000 D 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 E 0.000 O.OCO 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 F 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 LAP2 16 19 40 37 19 19 22 19 0 20 19 0 0 A 0.000 0.000 0.100 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.310 0.000 0.030 0.000 0.000 0.000 0.000 0.000 0.000 0.000 1.000 0.000 0.000 C 0.690 1.000 0.870 1.000 1.000 1.000 1.000 1.000 0.000 1.000 0.000 0.000 0.000 LAP3 16 19 40 0 0 19 22 19 0 0 0 0 0 A 0.380 1.000 0.080 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.620 0.000 0.920 0.000 0.000 1.000 1.000 1.000 0.000 0.000 0.000 0.000 0.000 IDM1 0 0 0 0 0 0 0 0 0 0 0 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 O.OCO 0.000 0.000 0.000 0.000 0.000 0.000 1DH2 19 19 40 37 19 23 24 19 19 20 19 19 19 A 0.350 0.000 0.000 0.000 0.000 0.000 0.000 1.000 0.000 0.000 0.000 0.000 0.000 B 0.650 0.050 0.200 0.550 0.000 0.440 0.000 0.000 0.000 0.000 0.000 0.000 0.000 C 0.000 0.950 0.700 0.450 0.740 0.560 1.000 0.000 0.000 0.000 1.000 0.000 0.000 B 0.000 0.000 0.100 0.000 0.260 0.000 0.000 0.000 1.000 1.000 0.000 0.000 0.000 E 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 c.ooo 0.000 0.000 1.000 1.000 MDII1 19 19 40 37 19 23 24 19 19 20 19 19 19 A 0.370 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0 0 0 0.000 0.000 0.000 C 0.630 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 D 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 H0II2 19 19 40 37 19 23 24 19 19 20 19 19 19 A 1.000 1.000 0.050 1.000 1.000 1.000 1.000 1.000 1.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.950 0.000 0.000 0.000 0.000 0.000 0.000 1.000 1.000 1.000 1.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 MO M3 19 19 40 0 0 0 0 0 0 0 0 0 0 en A 1.000 1.000 1.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 ADHI 0 0 40 0 0 23 24 19 19 0 19 0 0 A 0.000 0.000 1.000 0.000 0.000 1.000 1.000 1.000 1.000 0.000 1.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 A0H2 0 0 40 37 19 23 24 19 19 20 19 19 19 A 0.000 0.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 AAT1 16 19 40 37 19 23 25 19 19 20 19 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.630 0.900 1.000 1.000 1.000 1.000 0.920 1.000 1.000 1.000 1.000 0.000 0.000 C 0.370 0.100 0.000 0.000 0.000 0.000 0.080 0.000 0.000 0.000 0.000 0.000 0.000 AAT2 16 19 40 37 19 23 25 19 19 20 19 0 0 A 1.000 0.160 0.950 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 C 0.000 0.840 0.050 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 est 19 19 40 17 19 12 24 19 19 0 19 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 1.000 1.000 0.970 1.000 1.000 0.750 0.920 1.000 1.000 0.000 1.000 0.000 0.000 C 0.000 0.000 0.030 0.000 0.000 0.250 0.080 0.000 0.000 0.000 0.000 0.000 0.000 6PGDH 19 19 40 37 19 23 25 19 19 20 19 19 19 A 1.000 1.000 0.950 1.000 1.000 0.830 0.920 1.000 1.000 0.000 0.000 0.050 0.950 B 0.000 0.000 0.050 0.000 0.000 0.170 0.080 0.000 0.000 1.000 1.000 0.950 0.050 PGM 19 19 40 37 19 23 25 19 19 20 19 19 19 A 0.000 0.000 0.000 0.000 0.130 0.170 0.080 0.000 0.000 0.000 0.000 0.000 0.000 B 1.000 1.000 1.000 1.000 0.870 0.830 0.920 1.000 1.000 1.000 0.000 1.000 1.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 1.000 0.000 0.000 PGI 19 19 40 37 19 23 25 19 19 20 19 19 19 A 0.000 0.000 0.050 0.000 0.000 0.000 0.000 0.000 0.000 1.000 1.000 0.000 1.000 B 1.000 1.000 0.950 1.000 1.000 1.000 1.000 1.000 1.000 0.000 0.000 1.000 0.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 GDH 0 0 40 20 0 23 24 19 19 20 19 19 19 A 0.000 0.000 1.000 1.000 0.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 soo 19 19 40 37 19 13 25 19 19 20 19 0 0 OO A 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 0.0000.000 G6PDH 0 0 40 37 19 23 25 19 19 20 19 0 0 A 0.000 O.OCO 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 0.000 0.000 B 0.000 O.OCO 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 SKDH 19 19 40 37 19 23 25 19 19 0 19 19 19 A 0.000 0.000 0.000 0.000 0.000 0.000 0.400 0.000 0.000 fl.OOO 0.000 0.0000.000 B 1.000 1.000 1.000 1.000 1.000 1.000 0.600 1.000 1.000 0.000 1.000 1.000 1.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 0.000 ALD 19 19 40 37 19 23 25 19 0 20 19 0 0 A 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 0.000 1.000 1.000 0.0000.000 LOCUS- INTE INTE INTE INTE ABRU ABRU ABRU ABRU HARF HARF HARF HARF HARF ALLELE N 3805 N 3191 N 4097 N 3988 N 3971 H 4230 N 3891 N 3784 N 2781 N 2918 N 2950 N 3074N 2911 LAP1 19 19 0 0 0 0 0 0 39 0 19 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 0.000 C 0.050 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.900 0.000 1.0000.000 0.000 D 0.840 0.210 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 0.000 0.000 0.000 E 0.110 0.790 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 F 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.100 0.000 0.0000.000 0.000 LAP2 19 19 0 0 0 0 0 0 0 0 19 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 0.000 B 0.470 0.160 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 1.000 0.000 0.000 C 0.530 0.840 0.000 0.000 0.000 Ü.OOO 0.000 0.000 0.000 0.000 0.000 0.000 0.000 LAP3 0 0 0 0 0 0 0 0 0 0 19 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.300 0.000 0.000 0.000 0.000 0.000 1.000 0.0000.000 IDHI 0 0 0 0 0 0 0 0 0 0 0 0 0 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 O.OCO 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.0000.000 IDH2 19 19 19 19 19 19 6 4 39 19 38 13 6 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.003 0.000 0.000 o.ooc 0.000 0.000 0.000 0.000 C 0.000 0.000 0.000 0.000 0.900 0.840 0.830 0.000 1.000 0.000 0.790 1.000 1.000 D 1.000 1.000 1.000 1.000 0.100 0.160 0.000 1.000 0.000 1.000 0.000 0.000 0.000 L/1 VO E 0.000 0.000 0.000 0.000 0.000 0.000 0.170 0.000 0.000 0.000 0.210 0.000 0.000 HDH1 19 19 19 19 19 19 6 4 39 19 38 13 6 A 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 C 0.320 0.000 0.130 0.290 1.000 1.000 0.830 1.000 1.000 1.000 1.000 1.000 1.000 D 0.680 1.000 0.8 7 0 0.710 0.000 0.000 0.1 7 0 0.000 0.000 0.000 0.000 0.000 0.000 H0H2 19 19 19 19 19 19 6 4 39 19 38 13 6 A 1.000 1.000 1.000 1.000 0.900 0.840 1.00C 0.000 1.000 1.000 0.760 1.000 1.000 B 0.000 0.000 0.000 0.000 0.100 0.160 0.000 1.000 0.000 0.000 0.240 0.000 0.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 HDH3 19 19 19 19 0 0 0 0 0 0 0 0 0 A 1.000 1.000 1.000 1.000 0.000 0.000 0.0 0 0 0.000 0.000 0.000 0.000 0.000 0.000 B 0.000 0.000 O.OOC 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 ADHI 19 19 19 19 19 19 6 4 0 19 38 0 0 A 0.950 1.000 1.000 1.000 0.900 1.000 1.000 1.000 0.000 1.000 1.000 0.000 0.000 B 0.050 0.000 o.ooc 0.000 0.100 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 ADH2 19 19 19 19 19 19 6 4 39 19 38 13 6 A 1.000 1.000 1.00C 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 B 0.000 0.000 o.ooc 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 AAT1 19 19 19 19 19 0 0 0 39 19 38 13 6 A 0.000 0.000 O.OOC 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 6 1.000 1.000 1.000 1.000 1.000 0.000 0.000 0.000 1.000 1.000 1.000 1.000 1.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 AAT2 19 19 19 19 19 0 0 0 39 19 38 13 6 A 1.000 1.000 1.000 1.000 0.840 0.000 0.000 0.000 1.000 1.000 1.000 1.000 1.000 B 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 C 0.000 0.000 0.000 0.000 0.160 0.000 0.000 o.ôoo 0.000 0.000 0.000 0.000 0.000 EST 19 19 0 0 13 0 0 0 39 15 38 13 6 A 0.000 0.000 0.000 0.000 1.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0 1.000 1.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 1.000 1.000 0.000 0.000 C o.ooc 0.000 0.000 0.000 0.000 0.000 0.000 0.000 1.000 0.000 0.000 1.000 1.000 6PGDH 19 19 19 19 19 19 6 4 39 19 38 13 6 A 0.000 0.000 0.000 0.110 0.900 0.840 0.830 0.000 1.000 1.000 1.000 1.000 1.000 n 1.000 1.000 1.000 0.890 0.100 0.160 0.170 1.000 0.000 0.000 0.000 0.000 0.000 a\ o PGM 19 19 19 19 19 19 6 4 39 19 38 13 6 A 0.450 0.000 0.000 0.740 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 0.550 1.000 1.000 0.260 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 C 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 PGI 19 19 19 19 19 19 6 4 39 19 19 13 6 A 0.760 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 O.OCO 0.000 0.000 0.000 B 0.240 1.000 1.000 1.000 0.900 0.840 1.000 0.000 1.000 1.000 1.000 1.000 1.000 C 0.000 0.000 0.000 0.000 0.100 0.160 0.000 1.000 0.000 0.000 0.000 0.000 0.000 GOH 19 19 19 19 19 19 6 4 39 19 38 13 6 A 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 1.000 SOD 19 19 19 19 19 0 0 0 39 19 19 13 6 A 1.000 1.000 1.000 1.000 1.000 0.000 0.000 0.000 1.000 1.000 1.000 1.000 1.000 G6P0H 0 0 19 19 0 19 6 4 20 0 38 0 0 A 0.000 0.000 1.000 0.000 0.000 1.000 1.000 1.000 1.000 0.000 1.000 0.000 0.000 0 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 SKDH 19 19 19 19 19 19 6 4 39 19 38 13 6 A 0.000 0.000 0.000 0.050 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 0.000 B 1.000 1.000 1.000 0.210 1.000 1.000 1.000 1.000 1.000 1.000 0.900 1.000 1.000 C 0.000 0.000 0.000 0.740 0.000 0.000 0.000 0.000 0.000 0.000 0.100 0.000 0.000 ALD 19 19 0 0 13 0 0 0 20 16 0 0 0 A 1.000 1.000 0.000 0.000 1.000 0.030 0.000 0.000 1.000 1.000 0.000 0.000 0.000 LOCUS- HARF SUBB ALLELE N 2769 N 3035 N LAF'1 0 0 A 0.000 0.000 B 0.000 0.000 C 0.000 0.000 D 0.000 0.000 E 0.000 0.000 F 0.000 0.000 LAP2 0 0 A 0.000 0.000 Q 0.000 0.000 C 0.000 0.000 LAP3 0.000 0.000 0.000 0.000 IDHI 0 0 0.000 0.000 0.000 0.000 IDH2 19 199 0.000 0.000 0.000 1.000 1.000 0.000 0.000 0.000 0.000 0.000 HD HI 19 19 0.000 0.000 0.000 0.000 1.000 1.000 0.000 0.000 HDH2 19 19 1.000 1.000 0.000 0.000 o.ouo 0.000 H0II3 19 19 1.000 0.950 0.000 0.050 ADHI 19 19 1.000 1.000 0.000 0.000 ADH2 19 19? 1.000 1.000 0.000 0.000 AAÏ1 19 199 0.000 0.000 1 .0 0 0 1 .0 0 0 0.000 0.000 0\ to AAT2 19 19 A 1.000 1.000 B 0.000 0.000 C 0.000 0.000 EST 19 19 A 0.320 1.000 B 0.680 0.000 C 0.000 0.000 6PG0H 19 19 A 1.000 1.0c. B 0.000 0.000 PGM 19 19 A 0.000 0.000 B 1.000 1.000 C 0.000 0.000 PGI 19 19 A 0.000 0.000 B 1.000 1.000 C 0.000 0.000 H)H 19 19 A 1.000 1.000 SOO 19 19 A 1.000 1.000 G6P0H 19 19 A 1.000 1.000 B 0.000 0.000 SKDH 19 19 A G.OOO 0.000 B 1.000 1.000 C 0.000 0.000 ALD 19 19 A 1.000 1.000 o\ w LITERATURE CITED Bruederle, L. P. and D. E. Fairbrothers. 1986. Allozyme variation in populations of theCarex crinita complex (Cyperaceae). Syst. Bot. 11:583- 594. Cardy, B. J., C. W. Stuber, and M. M. Goodman. 1981. Techniques for starch gel electrophoresis of enzymes from maize {Zea mays L.). Institute of Statistics Mimeograph Series No. 1317. North Carolina State Univ., Raleigh. Cayouett, J. and P. Morisset. 1985. Chromosome studies on natural hybrids between maritime species of Carex (section Phacocystis and Cryptocarpae) in north eastern North America, and their taxonomic implications. Can. J. Bot. 63:1957-1982. Cayouett, J. and P. Morisset. 1986a. 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