American Journal of Botany 86(7): 1027–1037. 1999.

PHYLOGENETIC RELATIONSHIPS OF THE

HAMAMELIDACEAE INFERRED FROM SEQUENCES OF
INTERNAL TRANSCRIBED SPACERS (ITS) OF NUCLEAR
RIBOSOMAL DNA1
JIANHUA LI,2,5 A. LINN BOGLE,3 AND ANITA S. KLEIN4
2Harvard University Herbaria, 22 Divinity Avenue, Cambridge, Massachusetts 02138;

3
Department of Plant Biology, University of New Hampshire, Durham, New Hampshire 03824; and
4Department of Biochemistry and Molecular Biology, University of New Hampshire, Durham, New Hampshire 03824

Intergeneric relationships in the Hamamelidaceae have long been controversial. In this study, sequences of the internal
transcribed spacers of nuclear ribosomal DNA were used to reconstruct the phylogeny for the Hamamelidaceae. Three major
clades were recognized in the ITS-based phylogenetic tree: (1) Mytilaria-Exbucklandia-Rhodoleia, (2) Disanthus, and (3)
the Hamamelidoideae. Within the Hamamelidoideae there were three well-supported lineages: (1) Corylopsis-Loropetalum-
Tetrathyrium-Maingaya-Matudaea, (2) Eustigmateae sensu Endress, plus Molinadendron-Dicoryphinae, and (3) Hamamelis-
Fothergilleae sensu Endress, excluding Matudaea and Molinadendron. The Exbucklandioideae sensu Endress were not
monophyletic, nor were the tribes in the Hamamelidoideae in their current circumscriptions except for the Corylopsideae.
Strap-shaped petals, apetaly, and wind pollination have evolved three times independently in the Hamamelidaceae s.s.
(Hamamelidaceae minus Altingioideae), suggesting that homoplasy should be considered in future classifications of the
family.

Key words: Hamamelidaceae; ITS DNA sequences; homoplasy; phylogeny.

There are 31 genera and .140 species in the Hama- Hemsl., Semiliguidambar Chang, and Shaniodendron
melidaceae (Endress, 1993; Zhang and Lu, 1995). This (Chang) Deng, Wei et Wang can be found only in small
family is known for its broad geographic distribution, areas of China. However, there are two genera in this
high number of endemics, monotypic or oligotypic gen- family whose species are widely and disjunctly distrib-
era, and diverse morphology (Bogle, 1968; Endress, uted in several continents. One of them is Liquidambar
1989a, b, c, 1993). L., which can be found in western Asia, southeastern
The geographic range includes eastern and southern Asia, southeastern North America, and Central America
Africa, Madagascar, northeastern Australia, western, cen- (Bogle, 1968, 1986); the other disjunct genus is Hama-
tral, and southeastern Asia, eastern North America, Cen- melis L., which is composed of species distributed in
tral America, and northern South America (Gentry, 1993; southeastern Asia, eastern North America, and northern
Ulloa and Jørgensen, 1993; Zhang and Lu, 1995; Lozano- Mexico (Mione and Bogle, 1990). In the Hamamelida-
Contreras, 1996; Rakotobe, 1996). Whereas most of the ceae, while many genera are either monotypic or oligo-
genera occur in the Northern Hemisphere, five genera are typic, there are several genera that contain more than ten
distributed only in the Southern Hemisphere, including species, such as Corylopsis Sieb. et Zucc., Dicoryphe,
Trichocladus Pers., Ostrearia Baill., Neostrearia Smith, and Distylium Sieb. et Zucc. (Chang, 1979; Endress,
Noahdendron Endress, Hyland et Tracey, and Dicoryphe 1993; Rakotobe, 1996).
Du Petit-Thouars. More than half of the genera in the Morphological characters in the Hamamelidaceae are
Hamamelidaceae are endemic to very restricted areas. For highly diverse. For example, leaves are persistent or de-
instance, Ostrearia, Neostrearia, and Noahdendron are ciduous, simple and pinnately veined, or palmately lobed
distributed only in the rainforests of northeastern Queens- and veined. Most species are bisexual, but some are an-
land, Australia; Maingaya Oliv. is endemic to northeast- dromonoecious, and still others are monoecious. Flowers
ern Malaysia; Fortunearia Rehd. et Wils., Sinowilsonia are complete and five-merous in most genera, four-mer-
ous in several genera, and variable in others; a few genera
1 Manuscript received 3 April 1998; revision accepted 26 March
have an incomplete perianth, or are naked. Flowers are
1999. insect-, or bird-, or wind-pollinated (Bogle, 1970; En-
The authors thank P. K. Endress, T. D. Omar, Z.-C. Luo, R. Saunders, dress, 1989a; Li, 1997).
L. G. Saw, and Y.-L Qiu for providing leaf material or DNA for this
study; The Arnold Arboretum (MA), the Missouri Botanical Garden Most of the traditional classification systems place the
(MO), the Morris Arboretum (PA), and Longwood Gardens (PA) for Hamamelidaceae in the ‘‘Lower’’ Hamamelidae, which
granting them permission to collect material. The research was partially includes Cercidiphyllaceae, Tetracentraceae, Trochoden-
supported by the University of New Hampshire Graduate Student Re- draceae, Daphniphyllaceae, Platanaceae, Myrothamna-
search Enhancement Fund to JLI and by the Howard and Dorothy Pow- ceae, and Eupteleaceae (Cronquist, 1981; Takhtajan,
ers Fund to ALB.
This is a portion of the doctoral thesis of the first author presented
1997). Endress (1977) emphasized this family as a con-
to the Graduate School of the University of New Hampshire, Durham. necting taxon between the ‘‘Lower’’ and the ‘‘Higher’’
5 Author for correspondence, current address: Arnold Arboretum of Hamamelidae (e.g., Betulaceae, Fagaceae, and Juglan-
Harvard University, 125 Arborway, Jamaica Plain, MA 02130. daceae). Some members of the Hamamelidaceae have
1027
1028 AMERICAN JOURNAL OF BOTANY [Vol. 86

also been considered as linking taxa between the ‘‘lower’’ this region is informative in resolving subfamilial and
hamamelids and some basal elements of rosids and as- tribal relationships (Shi et al., 1998). However, more than
terids (Hufford, 1992; Chase et al., 1993; Endress, 1993; half of the recognized genera were missing in Shi et al.’s
Morgan and Soltis, 1993). A more comprehensive study data set, and a study with a much broader sampling is
(more taxa and more sources of data) is needed to further needed to address the intergeneric relationships in the Ha-
assess the systematic position of the Hamamelidaceae. mamelidaceae.
Nevertheless, the Hamamelidaceae (including the Altin- Therefore, the objectives of this study were as follows:
gioideae) appears to be monophyletic based on both mor- (1) to reconstruct phylogenetic relationships of the Ha-
phological (Hufford, 1992) and DNA sequence data (Li, mamelidaceae using ITS sequence data, and (2) to eval-
Bogle, and Klein, unpublished data). uate previous classification systems of the family in terms
The name Hamamelidaceae dates back to 1818 when of the monophyly of the subfamilies, tribes, and subtribes
Brown recognized it as a natural group with four genera, of the Hamamelidaceae.
Hamamelis, Dicoryphe, Dahlia Thunb. (5 Trichocladus
Pers.), and Fothergilla. Since then, more than half a doz- MATERIALS AND METHODS
en classification systems have been proposed for the Ha-
Plant material—Thirty-two species were sampled in this study (Table
mamelidaceae (see review in Tong, 1930; Bogle, 1968; 2), representing 28 genera in the Hamamelidaceae. Three genera were
Chang, 1979; Endress, 1989c). However, the most com- not available for analysis: Chunia, Semiliquidambar, and Embolanthera.
prehensive classification system for the Hamamelidaceae Vouchers for the sampled species are deposited in the Hodgdon Her-
was put forward by Harms (1930). As shown in Table 1, barium of the University of New Hampshire (NHA). Sequences of both
based on morphological and anatomical characteristics, ITS-1 and ITS-2 have been submitted to the GenBank, and the acces-
Harms recognized five subfamilies, the largest of which, sion numbers are given in Table 2. The aligned sequences are available
the Hamamelidoideae, was further divided into five from the first author and American Journal of Botany’s website.
tribes. However, he did not classify the two then little-
known genera Mytilaria Lecomte and Ostrearia. Schul- Molecular techniques—Total genomic DNAs were extracted from
ze-Menz (1964) transferred Sinowilsonia from the tribe fresh or silica-gel-dried leaves or buds following the protocol of Doyle
Distylieae into the Corylopsideae, which had included and Doyle (1987). Polymerase chain reactiony (PCR) was conducted
Corylopsis and Fortunearia. Chang (1973, 1979) erected in 0.2-mL thin-walled microcentrifuge tubes following the procedures
a new subfamily Mytilarioideae for Mytilaria and Chunia described in Li et al. (1997) and Li, Bogle, and Klein (1998). The PCR
Chang. primers were universal ITS4 and ITS5 of White et al. (1990).
Endress (1989c) reviewed the previous classification The PCR products were purified following Li et al. (1997). The re-
systems, especially those of Harms and Chang, for the sulting PCR product was used directly as a sequencing template. Se-
Hamamelidaceae, and recognized four subfamilies: Altin- quencing reactions were carried out using Cycle Sequencing Kits and
following the manufacturer’s protocols (Applied Biosystems, Foster
gioideae, Exbucklandioideae, Rhodoleioideae, and Ha-
City, California).
mamelidoideae (Table 1). The Exbucklandioideae are a
The sequencing primers were ITS2, ITS3, ITS4 and ITS5 of White
combination of the Exbucklandioideae Harms, Disan- et al. (1990). The cycle sequencing products were then separated on
thoideae Harms, and the Mytilarioideae Chang. For the 6% polyacrylamide gel using an Automated Sequencer 373A (Applied
largest subfamily Hamamelidoideae, Endress (1989b, c) Biosystems, Foster City, California) in the Sequencing Facility Center
united the tribes Fothergilleae sensu Harms and Disty- of the University of New Hampshire (UNH).
lieae sensu Harms, forming the tribe Fothergilleae sensu For Liquidambar, Tetrathyrium, and Maingaya, 0.8% DMSO (di-
Endress. Another major revision in Endress’s system is methylsulfoxide) was added to both the PCR and cycle sequencing re-
that Fortunearia and Sinowilsonia were grouped with actions.
Eustigma Gardn. et Champ., thus establishing a more in- The chromatograms were analyzed using the SEQED program (Ap-
clusive Eustigmateae. Furthermore, Endress (1989c) plied Biosystems, Foster City, California). Also, in order to assure cor-
treated the five Southern Hemisphere genera (Dicoryphe, rect base-calling, we overlapped sequences generated from adjacent
Neostrearia, Noahdendron, Ostrearia, Trichocladus) as primers of either the same or opposite directions. The boundaries of
one of the three subtribes in the tribe Hamamelideae. ITS-1 and ITS-2 were determined by comparing sequences of the 39
As can be seen from the comparison of the major clas- 18S and 5.8S and the 59 end of the 26S ribosomal genes of Canella
sification systems, fundamental questions concerning the winterana (GenBank accession number GBAN-L03844; the prefix
systematics of the Hamamelidaceae still remain. That is, GBAN- has been added for linking online version of American Journal
are the subfamilies, tribes, and subtribes monophyletic? of Botany to GenBank but is not part of the actual accession number).
In other words, do the classification systems reflect nat-
Sequence alignment—The ITS sequences within a genus and among
ural relationships in the Hamamelidaceae?
closely related genera were easily aligned by sight. However, ITS se-
Nucleotide sequences of a chloroplast gene, rbcL, have
quences among genera of different subfamilies were not readily align-
been extensively used to examine plant phylogenies at able, thus resulting in some ambiguous regions. These ambiguous sites,
higher taxonomic levels (Chase et al., 1993; Qiu et al., as identified by alignability by eye, can be eliminated from the data
1998). However, to resolve phylogenetic relationships matrix before a phylogenetic analysis is conducted (Downie and Katz-
among closely related genera, a fast-evolving DNA frag- Downie, 1996). This ‘‘culled’’ method tends to create clades where
ment, the nrDNA ITS region, has proven to be more use- internal relationships are not well resolved (Wheeler, Gatesy, and De-
ful (Baldwin, 1992; Baldwin et al., 1995; Bogler and salle, 1995; Soltis, Johnson, and Looney, 1996).
Simpson, 1996; Downie and Katz-Downie, 1996; Kron Another way of dealing with alignments with ambiguous sites is to
and King, 1996; Schilling and Panero, 1996; Soltis, John- select a so-called optimal alignment. This approach involves consisten-
son, and Looney, 1996). A phylogenetic analysis of the cy indices of the aligned sequences and the phylogenetic trees produced
Hamamelidaceae using nrDNA ITS data has shown that based on the data. Bogler and Simpson (1996) used this ‘‘optimality’’
 July 1999]

TABLE 1. Comparison of two major classification systems of the Hamamelidaceae (? 5 uncertain, N/A 5 unapplicable).

Harms (1930) Endress (1989c)

Genus Subfamily Tribe Subfamily Tribe Subtribe

Disanthus Maxim. 1. Disanthoideae III. Exbucklandioideae

LI

Hamamelis L. 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae I. Hamamelideae I. Hamamelidinae

Loropetalum R. Br. ex Reichb. 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae I. Hamamelideae II. Loropetalinae
Tetrathyrium Benth. 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae I. Hamamelideae II. Loropetalinae
Embolanthera Merr. 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae I. Hamamelideae II. Loropetalinae
Maingaya Oliv. 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae 1. Hamamelideae II. Loropetalinae
Dicoryphe Du Petit-Thouars 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae I. Hamamelideae III. Dicoryphinae
Trichocladus Pers. 2. Hamamelidoideae 1. Hamamelideae IV. Hamamelidoideae I. Hamamelideae III. Dicoryphinae
Ostrearia Baill. ? ? IV. Hamamelidoideae I. Hamamelideae III. Dicoryphinae
Neostrearia L. S. Smith N/A N/A IV. Hamamelidoideae I. Hamamelideae III. Dicoryphinae
Noahdendron Endress, Hyland & Tracey N/A N/A IV. Hamamelidoideae I. Hamamelideae III. Dicoryphinae
Eustigma Gard. & Champ. 2. Hamamelidoideae 2. Eustigmateae IV. Hamamelidoideae II. Eustigmateae
Fortunearia Rehd. & Wils. 2. Hamamelidoideae 3. Corylopsideae IV. Hamamelidoideae II. Eustigmateae
Sinowilsonia Hemsl. 2. Hamamelidoideae 5. Distylieae IV. Hamamelidoideae II. Eustigmateae
Corylopsis Sieb. & Zucc. 2. Hamamelidoideae 3. Corylopsideae IV. Hamamelidoideae III. Corylopsideae
ET AL.—PHYLOGENETICS OF

Molinadendron Endress 2. Hamamelidoideae 4. Fothergilleae IV. Hamamelidoideae IV. Fothergilleae

Fothergilla Murray 2. Hamamelidoideae 4. Fothergilleae IV. Hamamelidoideae IV. Fothergilleae
Parrotiopsis Schneider 2. Hamamelidoideae 4. Fothergilleae IV. Hamamelidoideae IV. Fothergilleae
Parrotia C. A. Mey. 2. Hamamelidoideae 5. Distylieae IV. Hamamelidoideae IV. Fothergilleae
Sycopsis Oliv. 2. Hamamelidoideae 5. Distylieae IV. Hamamelidoideae IV. Fothergilleae
Distyliopsis Endress N/A N/A IV. Hamamelidoideae IV. Fothergilleae
Distylium Sieb. & Zucc. 2. Hamamelidoideae 5. Distylieae IV. Hamamelidoideae IV. Fothergilleae
Matudaea Lundell N/A N/A IV. Hamamelidoideae IV. Fothergilleae
Rhodoleia Champ. ex Hook. 3. Rhodoleioideae II. Rhodoleioideae
Exbucklandia R. W. Br. 4. Exbucklandioideae III. Exbucklandioideae
HAMAMELIDACEAE

Chunia Chang N/A N/A III. Exbucklandioideae

Mytilaria Lecomte ? ? III. Exbucklandioideae
Altingia Nor. 5. Altingioideae I. Altingioideae
Semiliquidambar Chang N/A N/A I. Altingioideae
Liquidambar L. 5. Altingioideae I. Altingioideae
1029
1030 AMERICAN JOURNAL OF BOTANY [Vol. 86

TABLE 2. Species sequenced for nrDNA ITS for this analysis.

Taxon GenBank
Species Abbr. Collector and voucher Source accessionsa

Altingia sp. ALT Y.-L. QIU China GBAN-AF015417-18

Corylopsis pauciflora S.& Z. CPA J.-H. LI 01 Arnold Arboretum, MA. GBAN-U65462
Corylopsis sinensis Hemsl. CSI J.-H. LI 02 Arnold Arboretum, MA. GBAN-U65461
Corylopsis spicata S.& Z. CSP J.-H. LT 03 Arnold Arboretum, MA. GBAN-U65463
Dicoryphe stipulacea Janume St. Hil. DIC A. Randrianasolo 543 Tulear, Madagascar GBAN-AF015419-20
Disanthus cercidifolius Max. DIS A. L. BOGLE Woodlanders, Inc. SC. GBAN-AF015421-22
Distyliopsis tutcheri Endress DOT A. L. BOGLE Woodlanders, Inc. SC. GBAN-AF019231-2
Distylium myricoides Hemsl. DIM A. L. BOGLE Woodlanders, Inc. SC. GBAN-U65464
Distylium racemosum S.& Z. DIR A. L. BOGLE Woodlanders, Inc. SC. GBAN-U65465
Eustigma oblongifolium Gardn. & EUS N.-J. CHUNG Taiwan GBAN-U65466
Champ.
Exbucklandia populnea (R.Br.) R.W. EXB A. L. BOGLE Manuka State Roadside Park, Hawaii GBAN-AF015423-24
Br.
Fortunearia sinensis R.& W. FOR J.-H. LI 04 Arnold Arboretum, MA. GBAN-U65467
Fothergilla major Lodd. FOT J.-H. LI Univ. of New Hampshire campus GBAN-AF015425-26
Hamamelis virginiana L. HVI J.-H. LI Univ. of New Hampshire campus GBAN-AF015656-57
Hamamelis vernalis Sarg. HVE J.-H. LI Arnold Arboretum, MA. GBAN-AF015655
Liquidambar styraciflua L. LIQ A. L. BOGLE Univ. of New Hampshire greenhouse. GBAN-AF055886
Loropetalum chinense (R.Br) Oliv. LOR A. L. BOGLE Missouri Bot. Gard. GBAN-AF015427-28
Maingaya malayana Oliv. MAI L. G. SAW Kepong, Malaysia GBAN-AF022241
Matudaea trinervia Lund. MAT P. K. ENDRESS Botanical Garden of Zurich, Switzerland GBAN-AF015437
Molinadendron guatemalense Endress MOL P. K. ENDRESS Botanical Garden of Zurich, Switzerland GBAN-AF015438
Mytilaria laosensis Lec. MYT Z.-C. LUO Guangxi, China GBAN-U65469
Neostrearia fleckeri Smith NEO P. K. ENDRESS Botanical Garden of Zurich, Switzerland GBAN-AF015439
Noahdendron nicholasii Endress, Hy-
land & Tracey NOA P. K. ENDRESS Botanical Garden of Zurich, Switzerland GBAN-AF015440
Ostrearia australiana Baill. OST P. K. ENDRESS Botanical Garden of Zurich, Switzerland GBAN-AF015441
Parrotia persica C.A.Mey PAR A. L. BOGLE Univ. of New Hampshire greenhouse GBAN-AF015443
Parrotiopsis jacquemontiana Rehd. PPS A. L. BOGLE Harvard Univ. campus GBAN-AF015442
Rhodoleia championii Hook. f. RHO A. L. BOGLE Lyon Arboretum, Hawaii GBAN-AF015429-30
Shaniodendron subaequale Deng, Wei
& Wang SHA Y.-L. QIU Jiangsu, China GBAN-AF015431-32
Sinowilsonia henryi Hemsl. SIN J.-H. LI 05 Arnold Arboretum, MA GBAN-U65468
Sycopsis sinensis Oliv. SYC A. L. BOGLE Woodlanders, Inc. SC. GBAN-AF015433-34
Tetrathyrium subcordatum Benth. TET R. Saunders Hong Kong GBAN-AF022242
Trichocladus crinitus Pers. TRI A. L. BOGLE Longwood Gardens, PA. GBAN-AF015435
GBAN-AF019233
a The prefix GBAN- has been added for linking the version of American Journal of Botany to GenBank but is not part of the actual GenBank

accession number.

method to analyze the phylogenetic relationships of the Agavaceae. All characters and their states were equally weighted in the parsimony
Many indices can be used to assess the optimality of sequence align- analyses. Sequence divergence was analyzed using the pairwise differ-
ment, including the number of trees generated, the Consistency Index ence obtained from PAUP*. Due to the size of the data set and the
(CI), Retention Index (RI), and Rescaled Consistency Index (RC). Bog- limitation of computer memory, the heuristic search option was used to
ler and Simpson (1996), however, implied that the higher the RC was, find the shortest trees with TBR (Tree Bisection and Reconnection)
the better the alignment. This seems to be reasonable because the RC branch swapping, MULPARS on, and STEEPEST DESCENT off.
index excludes characters that do little to the ‘‘fit’’ of the tree but inflate It became impossible to produce reasonable sequence alignments
the CI (Wiley et al., 1991). when we tried to use taxa outside the Hamamelidaceae as outgroups.
In this study, the optimality method was employed to generate a data Therefore, in the parsimony analysis, Altingia and Liquidambar were
matrix of the ITS sequences of the Hamamelidaceae. The alignment used as outgroups because: (1) phylogenetic analyses using ITS data
that created trees with the highest RC index was considered as optimal (Shi et al., 1998) and evidence from morphology and matK gene se-
for both ITS-1 and ITS-2. Individual alignments were conducted using quences (Li, 1997) have revealed that these two genera form a clade
the CLUSTAL option of the MEGALIGN program of DNA* software sister to the clade containing the rest of the Hamamelidaceae, and (2)
package (DNA* Inc., Madison, Wisconsin). the fossil record has shown that these genera are the most ancient mem-
bers in the Hamamelidaceae (Zhang and Lu, 1995). As a result, we
Phylogenetic analysis—The resulting data matrices from the opti- focus our analysis and discussion on the Hamamelidaceae s.s. (Hama-
mality alignments were imported into the beta-test version of PAUP* melidaceae minus Altingioideae).
4.0d62 computer program for phylogenetic analyses, written by David Both bootstrap and decay analyses were conducted using the PAUP*
L. Swofford (1997) at the Smithsonian Institution, with permission. program to test the relative strength of putative clades (Felsenstein,
Indels were treated as missing data because this coding strategy re- 1985; Bremer, 1988; Donoghue et al., 1992).
tains information about substitutions that occur in other taxa in the indel MacClade 3.03 (Maddison and Maddison, 1992) was used to trace
region. However, it does not convey the information regarding the evo- unambiguous changes along branches and to compare competing hy-
lutionary event involved in the insertion or deletion (Platnick, Griswold, potheses concerning the relationships.
and Coddington, 1991; Wojciechowski et al., 1993). Thus, a parsimony Both the skewness test (Huelsenbeck, 1991) and the permutation test
analysis was also conducted treating gaps as the fifth character state. (Faith and Cranston, 1991) were conducted to evaluate the phylogenetic
July 1999] LI ET AL.—PHYLOGENETICS OF HAMAMELIDACEAE 1031

TABLE 3. Characteristics of the most parsimonious trees obtained by TABLE 4. Sequence characteristics of the two internal transcribed spac-
PAUP* heuristic searches of ITS-1 (1st line) and ITS-2 (2nd line) ers, separated and combined, in 32 species of the Hamamelidaceae.
sequence data sets using different combination of gap and gap
length penalties. Abbreviations: Consistency Index (CI), Retention Combined
ITS-1 ITS-2 (ITS-1 and ITS-2)
Index (RI), Rescaled Consistency Index (RC), number of steps
(Steps), number of trees found (Trees), Number of characters in the Length range (bp) 236–277 224–249 471–526
matrix (Nchar). The alignment considered optimal is in boldface Length mean (bp) 266.9 235 502.5
for ITS-1 and underlined for ITS-2. Aligned length (bp) 295 257 552
G1C content range (%) 57.5–69 61.3–68.2 60.8–68.3
Penalty G1C content mean (%) 63 64.9 64
(Gap/gap
length) Steps Trees Nchar CI RI RC Sequence divergence (%) 0.7–39.3 0.9–33.9 1.4–33.8
Number of indels 49 47 96
10/10 574 180 291 0.613 0.690 0.423 Number of variable sites 204 (69.2%) 159 (61.9%) 363 (65.8%)
438 126 259 0.564 0.628 0.354 Number of potentially in-
15/8 581 1008 291 0.608 0.689 0.419 formative sites 146 (49.5%) 117 (45.5%) 263 (47.6%)
439 270 259 0.563 0.629 0.354 Number of constant sites 91 (30.8%) 98 (38.1%) 189 (34.2%)
20/20 658 117 284 0.565 0.658 0.372 Transitions (average) 334 261 604
479 162 251 0.572 0.631 0.361 Transversions (average) 208 192 401
20/8 586 312 289 0.602 0.690 0.416 Transition/transversion 1.6 1.4 1.5
452 162 257 0.573 0.637 0.365 Skewness of tree length
distribution (g1 value
8/15 572 268 289 0.614 0.696 0.427 for 10 000 random
450 126 255 0.562 0.628 0.353 trees) 20.9 20.9 20.9
8/20 579 858 288 0.605 0.696 0.421
462 459 252 0.569 0.633 0.360
8/8 541 180 295 0.623 0.695 0.433
411 81 262 0.562 0.610 0.343 tween the two genera of Altingioideae and the Hama-
melidaceae s.s. was ;30% for both spacers (Table 5).

Phylogenetic relationships—The parsimony analysis

information contained in the ITS data matrix. The skewness test was
with gaps as missing data using the optimality data of
implemented using the Random tree option of PAUP*, and 10 000
random trees were examined. The permutation test, as used in Plunkett,
ITS-1 and ITS-2 yielded 90 shortest trees of 1005 steps,
Soltis, and Soltis (1997), was performed using the Permutation option with a consistency index of 0.59. Figure 1 is the strict
of PAUP* with 100 replicates and heuristic searches. consensus tree. Mytilaria, Exbucklandia, and Rhodoleia
formed the basal clade, while Disanthus was sister to the
RESULTS clade containing the sampled genera of the Hamameli-
doideae. In the Hamamelidoideae, three major clades
Sequence alignment—Among the seven pairs of were recognized: the Corylopsis clade, Hamamelis clade,
alignment parameters, Gps (Gap Penalty)/GLPs (Gap and the Trichocladus clade, among which the Corylopsis
Length Penalty), including 8/8, 8/15, 10/20, 10/10, 15/8, clade was basal. Both Corylopsis and Hamamelis clades
20/8, and 20/20, the pair of 8/8 resulted in the alignment were strongly supported, whereas the Trichocladus clade
of the highest RC index for ITS-1 and that of 20/8 pro- was weakly supported (Fig. 1).
duced the optimal alignment for ITS-2 (Table 3). The Corylopsis clade consisted of two subclades, the
first of which was composed of three species of Cory-
Sequence characteristics—The lengths of ITS-1 and lopsis; the second included Loropetalum, Tetrathyrium,
ITS-2 ranged from 236 to 277 bp (base pairs) and from Maingaya, and Matudaea. Both subclades were strongly
224 to 249 bp, respectively, in the Hamamelidaceae, and supported (Fig. 1).
the means were 267 bp for ITS-1 and 236 for ITS-2. In the Trichocladus clade, four branches were recog-
Several genera had roughly equal lengths of ITS-1 and nized: Dicoryphe, Trichocladus, the Eustigmateae sensu
ITS-2, including Fortunearia, Eustigma, Molinadendron, Endress (1989b) Plus Molinadendron, and the three Aus-
and Sinowilsonia. GC content ranged from 57 to 69% tralian genera. Among the three Australian genera, Neos-
with an average of 63% in ITS-1 and from 61 to 68% trearia was basally sister to the latter two genera. In the
with an average of 65% in ITS-2 (Table 4). Statistical clade of the Eustigmateae sensu Endress (1989b) plus
analysis (chi-square test) showed that the differences of Molinadendron, Molinadendron and Sinowilsonia were
nucleotide composition in both ITS-1 and ITS-2 across bound with a bootstrap value of 77% and a decay index
taxa, and between the two spacers were not significant of three steps (Fig. 1).
(P . 0.9). The Hamamelis clade was essentially the Fothergilleae
The percentages of potentially informative sites were sensu Endress (1989c), plus Hamamelis. There were four
49.5 and 45.5% for ITS-1 and ITS-2, respectively. The clades whose relationships were not well resolved in the
alignment of the sequences required 49 and 47 indels in phylogeny, including Hamamelis, Fothergilla, Parrotiop-
ITS-1 and ITS-2, respectively (Table 4). In both spacers, sis, and the well-supported Distylium group consisting of
;60% of the indels were a single base in length. There Distylium, Distyliopsis, Parrotia, Shaniodendron, and Sy-
were three indels of ten or more bases in the ITS region: copsis (Fig. 1).
two indels (12 and 26 bases) were in ITS-1, and one (ten The phylogenetic analysis with gaps as the fifth char-
bases) in ITS-2. Pairwise sequence divergence in ITS-1 acter state generated 46 shortest trees of 1266 steps with
was generally higher than that in ITS-2, with averages of a consistency index of 0.6. The tree topology was the
18.5 and 16.6%, respectively. Pairwise divergence be- same as the phylogeny in Fig. 1. with several exceptions
1032 AMERICAN JOURNAL OF BOTANY [Vol. 86

TABLE 5. Sequence divergence of ITS-1 and ITS-2 (below and above diagonal) in the Hamamelidaceae. The values are derived from pairwise
distances, calculated in PAUP* (mean distance 3 100), and gaps as missing data. Taxon abbreviations are as in Table 1.

Taxon CPA CSI CSP DIS DIC DIM DIR DOT EUS EXB FOR FOT HVE HVI LOR

CPA — 3 3.9 21.3 16.7 15.4 15.8 15.1 15.1 19.5 14.1 15 12 13.3 19
CSI 4.1 — 0.9 20.3 15.7 14 14 13.3 14.1 18.5 13.6 14 11.5 12.8 18
CSP 3.7 2.2 — 20.9 16.6 14.9 14.9 14.1 15 19 14.4 14.9 12.4 13.7 18.9
DIS 24.4 26.3 26.2 — 21.1 19.4 19.4 18.5 21.4 24.9 21.2 19.4 19.4 19.6 23.4
DIC 14.9 16.8 17.5 24.3 — 11.6 10.7 9.8 8.1 22.9 5.9 10.7 10.3 10 17.2
DIM 15.3 17.3 17.2 23.6 11.2 — 1.7 2.6 9.9 19.4 8.2 6.4 4.3 4.7 19.1
DIR 15.3 17.2 17.2 24 12 1.1 — 1.7 9.1 19.7 8.6 5.5 5.2 4.7 18.1
DOT 15.6 17.6 17.5 23.7 12 0.7 0.4 — 8.3 20.3 8.6 4.7 4.3 3.9 16.7
EUS 10.9 13.7 13.6 23.2 10.8 8 8.5 8.4 — 21.1 6 8.2 8.6 8.2 17.5
EXB 30.6 32.8 32.7 31.9 30.1 30.4 30.6 30.8 28.6 — 20.6 20.7 20.3 20.8 27.5
FOR 12.9 14.9 14.9 22.7 9.4 8 8.4 8.4 3.4 28.1 — 9.4 8.1 7.8 15.9
FOT 14 16.4 16 22.9 11.9 5.3 5.7 5.7 7.6 30.4 8.4 — 4.7 4.3 20.2
HVE 14.5 16.5 16.4 22.1 11.1 3.7 4.1 4.1 7.7 30.5 7.7 3.7 — 2.1 19.5
HVI 15.6 17.9 17.9 23.6 11.9 4.5 4.9 4.9 8.5 30.9 8.5 4.5 1.5 — 19.1
LOR 11.5 12.5 12.8 26.4 18 17.7 17.7 17.7 15.7 30.7 16.8 17.5 17.2 18.3 —
MAI 15.2 15.8 15 27.1 17.4 18.3 18.6 18.7 17.5 31.7 16.8 18.2 18.2 19.1 11.6
MAT 13.7 13.5 13.5 26 18.7 19.6 19.6 19.6 17 30.8 17.3 19 18.4 19.6 9.7
MOL 12.2 15 14.9 24.4 10.2 8.4 8.4 8.4 3.4 27.7 5.5 7.6 8.1 8.9 16.9
MYT 30 32.2 32.1 29.6 28.8 28.5 27.9 28.2 26 25.9 23.8 28.9 27.1 26.7 30.5
NEO 13.8 17.2 16.8 25.1 13.1 11.9 12.3 12.3 6.4 31 7.1 11.2 10.5 11.9 19.2
NOA 12.9 15.7 15.3 24.8 10.7 11.2 11.5 11.5 7.3 30.9 8 9.6 9.6 10.3 18.2
OST 11.6 14.9 14.5 25.1 10.4 10.4 10.8 10.8 7.7 32.5 8.5 9.2 9.2 10 17.6
PAR 16.8 18.7 18.6 24.8 12 1.9 3 2.6 8.6 29.7 8.4 5.7 3.4 4.1 18.4
PPS 16 17.9 17.9 23.6 13 6.3 7.1 7.1 8.5 29 9.3 6.4 5.6 6.3 18.7
RHO 28.7 30.8 31.5 28.8 28.2 30.1 29.4 29.7 27.2 19.1 25.7 28.8 28.2 28.6 29.3
SHA 16 18.3 18.3 23.6 10.8 2.2 3.3 3 9.4 30.8 8.5 6.1 3 4.5 18
SIN 13 15.9 15.8 26.1 10.7 8.8 9.2 9.2 3.8 30.9 5.1 9.3 8.5 9.3 17.4
SYC 17.5 19.4 19.3 26.5 13.5 3.4 4.1 3.7 9.8 31.5 9.8 7.5 5.2 5.6 18.7
TET 11.4 13.2 12.8 25.7 16.7 16.9 17.2 17.2 15.4 27.9 16.9 17.1 17.2 18.4 5.6
TRI 15.6 16.4 16.4 25.1 11.5 11.6 11.6 11.6 9.4 32.2 9.8 11.1 10.7 11.5 18.3
LIQ 31.9 32.6 32.6 33.2 34.1 30.4 29.6 30.4 27.2 34.1 27.2 32 30.9 31.2 33.2
ALT 35.5 37.4 36.9 36 35.6 32.7 32 32.8 31.4 39.3 30.5 34 32.9 32.9 37

(Fig. 2). In the Trichocladus clade, Trichocladus was the Polyphyletic Exbucklandioideae sensu Endress—The
basal taxon, followed by Dicoryphe, the paraphyletic Exbucklandioideae sensu Endress (1989c) includes four
Australian genera, and the monophyletic Eustigmateae genera, Chunia, Disanthus, Exbucklandia, and Mytilaria.
sensu Endress (plus Molinadendron). In the Hamamelis The morphological synapomorphies of this subfamily in-
clade, the clade of Hamamelis species was sister to the clude large, persistent stipules, palmate venation, and 4–
Fothergilleae sensu Endress (minus Matudaea and Mol- 6 ovules per carpel. However, the phylogenetic analysis
inadendron), in which Fothergilla was the basal taxon. based on the ITS DNA sequences provided a different
picture of this taxon (Chunia was not available for this
DISCUSSION study). Exbucklandia and Mytilaria were grouped with
Rhodoleia, while Disanthus formed its own clade (Fig.
Sequence characteristics—In the angiosperms ITS-1 1). This indicates that the Exbucklandioideae sensu En-
and ITS-2 are each less than 300 bp long: ITS-1 ranges dress (1989c) is polyphyletic. Furthermore, when the
from 187 to 298 bp and ITS-2 from 187 to 252 bp (Bald- three genera (Disanthus, Exbucklandia, and Mytilaria)
win et al., 1995; Downie and Katz-Downie, 1996; Padg- were forced into a monophylectic group, 35 more steps
ett, 1997). The length of the two spacers in the Hama-
were required.
melidaceae fell within these ranges, but ITS-1 sequences
in most of the species sampled were in the high end of Mytilaria, Exbucklandia, and Rhodoleia have been
the range, except for the four genera (Eustigma, Fortu- previously recognized as subfamilies Mytilarioideae, Ex-
nearia, Molinadendron, and Sinowilsonia) that shared bucklandioideae, and Rhodoleioideae, respectively
two unique deletions (12 and 26 bp). The ITS-2 in the (Harms, 1930; Chang, 1948, 1973, 1979; Tahktajan,
Hamamelidaceae was relatively less variable in length, 1997). In the ITS phylogeny, Mytilaria was allied with
mostly in the range of 224–249 bp, which agrees with a the clade of Exbucklandia and Rhodoleia, suggesting a
generalization that ITS-2 is more conservative than ITS- close relationship among the three genera. Exbucklandia
1 (Hershkovitz and Lewis, 1996; Liston et al., 1996). and Rhodoleia also shared 32 unambiguous base substi-
A permutation test for the data set from the optimality tutions (Fig. 1). Interestingly, this pattern agrees with an
alignment resulted in a probability of 0.01, indicating that earlier taxonomic treatment (Reinsch, 1889) and a rbcL
the ITS data contain significant phylogenetic signal (Faith phylogeny (Chase et al., 1993; Qiu et al., 1998). Never-
and Cranston, 1991). This is consistent with the skewness theless, these three genera differ greatly in morphology.
of tree length distribution (Table 4). Mytilaria and Exbucklandia share palmate venation (vs.
July 1999] LI ET AL.—PHYLOGENETICS OF HAMAMELIDACEAE 1033

TABLE 5. Continued.

MAI MAT MOL MYT NEO NOA OST PAR PPS RHO SHA SIN SYC TET TRI LIQ ALT

11.2 12.1 15.3 23.6 17.6 14 15 14.6 13.6 21.9 15.7 18.2 15.9 12.5 14.4 31.2 30.9
9.1 10.8 13.5 24.3 16.6 13 14.4 12.7 12.7 20 13.9 16.4 14.1 12 14.8 31.8 31.5
9.9 11.7 14.4 23.8 17.5 13.9 15.3 13.6 13.5 20.4 14.8 17.3 15 12.9 15.7 32.3 31.9
23.2 22 20.2 27.6 22.2 20.2 21.2 18 19.4 26.9 18.5 23.2 19.3 19.9 20 29.3 30.5
15.1 14.8 8.9 25.1 9.4 8.5 8.1 10.3 9.6 24.4 9.9 9.3 10.7 16.8 9.3 34 33.1
16.9 17.9 10.3 25.7 13.4 11.6 11.1 2.1 4.8 23.9 2.6 12.9 3.4 18.6 11.2 32.1 33.1
15.6 17 9.8 25.2 12.5 10.7 10.2 1.3 3.9 23 1.7 12.9 2.5 17.2 11.1 31.6 32
15.6 17.1 10.8 24.8 12.5 9.9 9.4 1.3 3 23.1 1.7 12.1 0.4 17.3 10.3 31 31.5
15.6 16.5 8.5 25.8 11.2 7.7 7.7 8.6 8.4 23.6 9.1 9.4 9.1 17.7 10.3 30.1 31.9
21.3 23.8 19.7 27.2 24.5 24.5 21.9 19.3 20.6 15.9 20.5 24.7 20.7 22.4 23.7 30 31
14.2 18.2 5.9 26.9 9.8 6.8 8.1 9 7.9 23 9.4 7.6 9.4 17.5 10.2 30 31.2
16.7 17.4 11.5 24.6 12.4 10.2 9.8 4.3 3.9 24.4 5.5 11.9 5.1 18.5 11.6 32.3 32.7
16.4 17.1 12 26 12.9 10.3 9.8 3.9 3.5 24 5.2 12 5.2 17.8 9.9 33.1 33.6
16.1 16.8 11.6 26.2 13 10.4 9.5 3.4 3 23.7 4.7 12.1 4.7 17.4 10.3 32.9 33.3
19.9 11.2 15.8 32.2 17.3 14.5 17.6 17.6 18.2 28 17.1 19.6 17.5 14.1 19 29.7 30.8
— 14.7 14.5 24.4 16.9 14 15.5 15.1 14.6 22.7 16.3 18.8 16.4 15.1 17.5 29.9 30.9
11.2 — 14.7 27.2 17.5 14.7 15.1 16.6 15.4 23.7 17 17.2 17.9 9.4 17.7 28.1 28.5
18.2 17.8 — 27.9 12 10.2 9.7 10.3 11.3 23 10.7 7.6 11.6 16.7 11 30.8 32
32.7 31 25.6 — 26 25.8 25.8 25.1 24.6 30.4 24.4 29.1 25.6 25.1 25.6 32.3 30.1
21 21.1 6.8 30 — 9.4 8.1 12.9 12.7 24.4 12.5 13.6 13.3 15.5 12.4 30.1 30.6
18.9 20.1 7.6 30.5 6.7 — 5.5 10.3 9.6 24.3 10.7 10.6 10.7 14.4 11 30.8 31.7
18.3 19.2 8.1 30.8 7.3 5 — 9.8 10 23.4 10.2 10.6 10.2 15 10.2 29.4 30.4
19.5 20 8.9 27.5 12 11.9 11.2 — 2.6 22.6 1.3 12.4 2.1 16.8 10.7 31.7 32.1
20.5 20.3 8.9 28.8 12.2 12.5 11.5 5.2 — 23.7 3.9 11.7 3.9 16.8 10.9 30.4 30.8
29.5 28.3 25.8 23.8 29.1 30.3 28.6 29.1 27 — 23.3 24.9 23.9 22.8 22.7 33.8 33.1
18.3 19.2 9.8 27.9 12.3 12.3 11.6 1.8 6.3 29.1 — 12.8 2.5 16.2 11.1 31.4 31.4
18.7 19.2 4.7 26.8 8 9 9.4 9.3 10.1 29.6 10.2 — 12.9 20.5 11.9 33.9 34.7
19.5 21.1 9.4 29.1 14.3 13 12.7 3.4 7.9 30.5 3.7 10.6 — 18.1 11.1 31.8 32.3
11.2 10.5 17 28.7 19.9 18.9 18.3 16.9 18.3 28.5 16.5 17.9 18.4 — 18.4 30.1 30.2
18.5 19.1 8.9 30.4 11.5 9.2 9.6 13.1 12.2 30.5 13.5 11.1 13.4 18.2 — 32.5 33.1
33.2 31.8 26.8 31.7 33.5 31.8 31.2 30.6 31.1 31.2 31 29.5 31 32.9 32.4 — 5.4
36.5 36.3 31.3 36.8 36.1 33.8 34.3 32.9 34 36.7 33.7 31.8 34.7 37 35.1 15.5 —

pinnate venation in Rhodoleia), but differ in nodal anat- 1993; matK—Li, 1997; trn noncoding regions—Li et al.,
omy (multilacunar vs. trilacunar; Bogle, 1990), inflores- unpublished data). The difference is probably due to
cence type (spadix vs. head), and chromosome base num- long-branch attractions resulting from the absence of My-
ber (x 5 13 vs. x 5 8; Pan and Yang, 1994). Rhodoleia tilaria or from the use of Corylus (Betulaceae, higher
is distinct from Mytilaria and Exbucklandia in a suite of Hamamelidae) as the outgroup in Shi et al.’s analysis. In
characters, including bird-pollinated, asymmetric flowers, addition, we found DNA sequence differences for the
and a chromosome base number of x 5 12. In the ITS same species sampled in both this study and Shi et al.’s
region, these three genera diverged greatly from their (1998).
common ancestor (Table 5, Fig. 1). It therefore seems
reasonable to continue to treat them as belonging to in- Monophyly of the Hamamelidoideae—It has been
dividual subfamilies (Harms, 1930; Chang, 1973, 1979; widely recognized that the Hamamelidoideae is a mono-
Bogle, 1989; Endress, 1989b, c). phyletic group, characterized by a ballistic seed dispersal
Disanthus was the immediate sister taxon to the Ha- mechanism and one mature seed per carpel (Bogle, 1968;
mamelidoideae in the phylogenetic tree (Fig. 1), reminis- Endress, 1989a, b, c; Hufford and Crane, 1989). The
cent of the conclusions from previous morphological present analysis adds strong evidence to this hypothesis
analyses (Hufford and Crane, 1989; Pan, Lu, and Wen, because the clade of this subfamily received a 100%
1991). The intermediate systematic position of Disanthus bootstrap value and a decay index of more than five steps,
between the Exbucklandia-Rhodoleia-Mytilaria clade and and was further supported by 36 unambiguous base sub-
the Hamamelidoideae in the ITS-based tree does not pro- stitutions (Fig. 1). This result is consistent with the pre-
vide evidence for the hypothesis that this genus is most vious ITS phylogeny (Shi et al., 1998).
primitive in the Hamamelidaceae, as has been suggested The intergeneric relationships within the Hamameli-
by some authors (Reinsch, 1889; Harms, 1930; Tong, doideae, however, have been contentious (Harms, 1930;
1930; Cronquist, 1981; Pan, Lu, and Wen, 1991; Takh- Schulze-Menz, 1964; Endress, 1989b, c). The relation-
tajan, 1997). ships among the five tribes recognized by Harms (1930)
In the previous nrDNA ITS phylogeny (Shi et al., and more recently revised by Endress (1989c) and others
1998), Exbucklandia is sister to the Hamamelidoideae, are discussed below.
whereas Disanthus and Rhodoleia form a clade. This re-
sult does not agree with morphological analysis (Hufford Corylopsideae—The concept of the tribe Corylopsi-
and Crane, 1989), or the ITS analysis of this study, or deae has been revised several times in the past several
the chloroplast gene phylogenies (rbcL—Chase et al., decades. Harms (1930) delimited Corylopsideae as in-
1034 AMERICAN JOURNAL OF BOTANY [Vol. 86

Fig. 1. Strict consensus of 90 shortest trees of 1005 steps, generated

using nrDNA ITS sequences of the Hamamelidaceae treating gaps as
missing data. Taxon abbreviations are as in Table 2. Numbers above
and below branches are bootstrap percentages and decay values/number
of unambiguous changes, respectively. CI 5 0.59, and RC 5 0.39.
Grouping on the right side follow Endress (1989c). ALToideae 5 Al-
tingioideae, COReae 5 Corylopsideae, DICinae 5 Dicoryphinae, EU- Fig. 2. Strict consensus of 48 shortest trees of 1266 steps, generated
Seae 5 Eustigmateae, EXBoideae 5 Exbucklandioideae, FOTeae 5 using nrDNA ITS sequences of the Hamamelidaceae treating gaps as
Fothergilleae, HAMeae 5 Hamamelideae, HAMoideae 5 Hamameli- the fifth character state. CI 5 0.6, RC 5 0.4. Numbers above and below
doideae, LORinae 5 Loropetalinae, RHOideae 5 Rhodoleioideae. branches are bootstrap percentages and decay values, respectively.

genera in the Hamamelidoideae that have strap-shaped

cluding Corylopsis and Fortunearia based on their sim- petals were placed in the tribe Hamamelideae and no fur-
ilarity of leaf morphology, while Schulze-Menz (1964) ther subdivisions were proposed. Endress (1989c), how-
added Sinowilsonia to this tribe considering similar floral ever, recognized three subtribes in the Hamamelideae, in-
structures of the latter to Fortunearia. However, Endress cluding the Hamamelidinae, Loropetalinae, and the Di-
(1989b, c) considered Corylopsideae as containing a sin- coryphinae. The Hamamelidinae contained a single ge-
gle genus, Corylopsis, characterized by the orbicular pet- nus, Hamamelis, characterized by the four-merous flower;
al. the Dicoryphinae included the five Southern Hemisphere
In the ITS-based phylogenies (Figs. 1, 2), the three genera (Dicoryphe, Trichocladus, Ostrearia, Neostrearia,
species of Corylopsis sampled formed a well-supported and Noahdendron) and was defined by the distinct anther
clade and were sister to the branch comprising Lorope- dehiscence pattern; the Loropetalinae contained the re-
talum, Tetrathyrium, Maingaya, and Matudaea, whereas maining four genera, Loropetalum, Embolanthera, Main-
Fortunearia and Sinowilsonia were phylogenetically dis- gaya, and Tetrathyrium.
tant from Corylopsis. Therefore, Corylopsis is not closely The analysis of the ITS data placed the genera of the
related to either Fortunearia or Sinowilsonia, as has al- Hamamelideae into three different clades (Fig. 1), sug-
ready been shown in previous ITS analyses (Li, Bogle, gesting that the tribe is polyphyletic (as shown by Shi et
and Klein, 1998; Shi et al., 1998). Floral ontogeny and al., 1998). Maingaya, Tetrathyrium, and Embolanthera
embryological study also support the separation of Cor- have been suggested to be the ancestral stock of the Aus-
ylopsis from Fortunearia and Sinowilsonia (Li and Bo- tralian hamamelids (Raven and Axelrod, 1974). However,
gle, 1998). this study does not support that proposition because in
the phylogeny (Fig. 1) the monophyletic Australian gen-
Hamamelideae—In Harms’s system (1930), all of the era were distant from the lineage containing Maingaya
July 1999] LI ET AL.—PHYLOGENETICS OF HAMAMELIDACEAE 1035

and Tetrathyrium (Embolanthera not available for this Molinadendron). Interestingly, this pattern finds support
study). The exclusive Southern Hemisphere distribution from leaf venation (Harms, 1930; Chang, 1979; Li and
and the unique pattern of anther dehiscence suggest a Hickey, 1988). The systematic position of Hamamelis
close relationship of the three Australian and two African was not resolved in the ITS phylogeny using gaps as
genera (Endress, 1989a, b, c). However, in the strict con- missing data (Fig. 1); however, in the phylogenetic tree
sensus trees (Figs. 1, 2), the relationships of the Austra- using gaps as the fifth character state (Fig. 2), the clade
lian and African genera were not well resolved, even of the two Hamamelis species was sister to the Fother-
though these genera formed a clade in the 50% majority gilleae sensu Endress. Also, Hamamelis differs greatly
consensus tree (tree not shown). More evidence is needed from the Fothergilleae in a group of floral characters,
to resolve relationships of the Southern Hemisphere gen- including showy, strap-shaped petals, bisexuality, and
era and thus to elucidate their biogeography. nectariferous phyllomes (Endress, 1989a). Therefore,
Hamamelis should be isolated from the Fothergilleae in
Eustigmateae—The delimitation of the Eustigmateae taxonomic treatments.
has expanded from monogeneric (Harms, 1930) to tri- The evolutionary relationships shown in the phyloge-
generic (Endress, 1989c). Eustigma is distinct in its small, netic trees (Figs. 1, 2) have significant implications for
auriculate petals and greatly enlarged stigmatic surfaces character evolution and biogeography. For example, apet-
(Harms, 1930). The similarities of Eustigma with For- aly and wind-pollination have evolved independently
tunearia in other morphological characters such as ped- more than three times in the Hamamelidaceae. The Amer-
icellate, lenticellate fruits, and covering sepals led En- ican Matudaea (Central and South America) and Moli-
dress (1989c) to the conclusion that Eustigmateae should nadendron (Central America) were suggested to be close-
include Fortunearia and Sinowilsonia. ly related to Asian Maingaya and Sinowilsonia, respec-
The ITS-based tree (Fig. 1) offers strong support for tively. The Southern Hemisphere genera were shown to
Endress’s hypothesis and further corroborates the impli- be a potentially monophyletic group by the 50% majority
cation (Endress, 1967) that Molinadendron and Sinowil- consensus tree. However, we believe that a well-resolved
sonia are closely related. Interestingly, these four genera phylogeny based on multiple sources of data and detailed
shared nine unambiguous base substitutions (Fig. 1) and studies of the fossil record are needed to reveal a com-
two unique deletions totaling 38 bp (Li, Bogle, and Klein, plete picture of character evolution and biogeography of
1998). the Hamamelidaceae.
In conclusion, at the subfamily level, this study rec-
Combining the Distylieae and Fothergilleae—In the ognized the monophyly of the Exbucklandioideae Harms,
phylogeny (Fig. 1), the clade of two Hamamelis species Mytilarioideae Harms, Rhodoleioideae Harms, Disan-
was sister to a branch containing the Distylieae (Disty- thoideae Harms, and the Hamamelidoideae. Furthermore,
lium, Distyliopsis, and Sycopsis) and Fothergilleae sensu it suggested close relationships of Exbucklandia, Rho-
Harms (1930) (Fothergilla, Parrotia, and Parrotiopsis). doleia, and Mytilaria and the paraphyly of Exbucklan-
Neither the Distylieae nor the Fothergilleae formed a dioideae sensu Endress (1989c). At the tribal and subtri-
monophyletic clade. Instead, a close but unresolved re- bal levels, the monophyletic groups were the corylopsi-
lationship was shown among Sycopsis, Parrotia, and deae sensu Endress (1989c); Eustigmateae sensu Endress
Shaniodendron. This result agrees with the finding that (1989c), but expanded to include Molinadendron; Foth-
Sycopsis and Parrotia are interfertile, giving rise to the ergilleae sensu Endress (1989c), expanded to include
hybrid taxon, Sycoparrotia (Endress and Anliker, 1968). Hamamelis; Dicoryphinae; and Loropetalinae, expanded
Thus, the Fothergilleae sensu Endress (1989c), a merger to include Matudaea. The Hamamelideae, however, were
of Distylieae Harms and Fothergilleae Harms, is sup- polyphyletic. The ITS phylogeny also revealed that some
ported. morphological characteristics commonly used to classify
Fothergilla and Parrotiopsis each formed their own the Hamamelidoideae have evolved several times inde-
clades in the ITS phylogeny (Fig. 1). This is consistent pendently. These characteristics included strap-shaped
with the showy specialized structures they have evolved petals, apetaly, and wind pollination. Thus homoplasy
for insect pollination: long, white, clavate stamen fila- needs to be taken into account in future classifications of
ments in Fothergilla and large, white subfloral bracts in the Hamamelidaceae so that natural relationships can be
Parrotiopsis (Bogle, 1970; Endress, 1989a). These spe- reflected.
cializations do not occur in any other members of the
Fothergilleae sensu Endress (1989c) or in the Hamame- LITERATURE CITED
lidaceae.
The apetalous genera of the Hamamelidoideae have BALDWIN, B. G. 1992. Phylogenetic utility of the internal transcribed
spacers of nuclear ribosomal DNA in plants: an example from the
long been considered as monophyletic (Harms, 1930; En- Compositae. Molecular Phylogenetics and Evolution 1: 3–16.
dress, 1989c), but the ITS phylogeny (Fig. 1) suggested ———, M. J. SANDERSON, J. M. PORTER, M. F. WOJCIECHOWSKI, C. S.
that loss of petals in this subfamily has evolved at least CAMPBELL, AND M. J. DONOGHUE. 1995. The ITS region of nuclear
three times independently: Matudaea in the Corylopsis ribosomal DNA: a valuable source of evidence on angiosperm phy-
clade, Molinadendron in the Trichocladus clade, and the logeny. Annals of the Missouri Botanical Garden 82: 247–277.
Fothergilleae in the Hamamelis clade. Therefore, the BOGLE, A. L. 1968. Floral vascular anatomy and the nature of the
hamamelidaceous flowers. Ph.D. dissertation, University of Min-
apetalous genera do not belong to a single monophyletic nesota, Minneapolis, MN.
group, and apetaly is homoplasious in this subfamily. ———. 1970. Floral morphology and vascular anatomy of the Ha-
It is a novel relationship that Hamamelis was grouped mamelidaceae: the apetalous genera of Hamamelidoideae. Journal
with Fothergilleae sensu Endress (excl. Matudaea and of the Arnold Arboretum 51: 310–366.
1036 AMERICAN JOURNAL OF BOTANY [Vol. 86

———. 1986. The floral morphology and vascular anatomy of the Ha- value of the rDNA-ITS region. Molecular Biology and Evolution
mamelidaceae: subfamily Liquidambaroideae. Annals of the Mis- 13: 1276–1295.
souri Botanical Garden 73: 325–347. HUELSENBECK, J. P. 1991. Tree-length distribution skewness: an indi-
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