Deuterostome Phylogeny and the Sister Group of the Chordates:
Evidence from Molecules and Morphology
J. McClintock Turbeville, Joseph R. Schulz, 1 and RudoljA. Raff
Department
of Biology and Institute for Molecular and Cellular Biology, Indiana University
Complete coding regions of the 18s rRNA gene of an enteropneust hemichordate and an echinoid and ophiuroid
echinoderm were obtained and aligned with 18s rRNA gene sequences of all major chordate clades and four
outgroups. Gene sequences were analyzed to test morphological character phylogenies and to assess the strength
of the signal. Maximum-parsimony
analysis of the sequences fails to support a monophyletic Chordata; the urochordates form the sister taxon to the hemichordates, and together this clade plus the echinoderms forms the sister
taxon to the cephalochordates plus craniates. Decay, bootstrap, and tree-length distribution analyses suggest that
the signal for inference of deuterostome phylogeny is weak in this molecule. Parsimony analysis of morphological
plus molecular characters supports both monophyly of echinoderms plus enteropneust hemichordates and a sister
group relationship of this clade to chordates. Evolutionary parsimony does not support chordate monophyly.
Neighbor-joining, Fitch-Margoliash, and maximum-likelihood analyses support a chordate lineage that is the sister
group to an echinoderm-plus-hemichordate
lineage. The results illustrate both the limitations of the 18s rRNA
molecule alone for high-level phylogeny inference and the importance of considering both molecular and morphological data in phylogeny reconstruction.
Introduction
The quest to find the closest relatives of the chordates has been one of the classic problems in systematic
biology. Although embryological and anatomic characters support monophyly of a taxon that includes the
Chordata, Hemichordata, and Echinodermata, the relationships among these deuterostomes remain controversial (fig. 1). However, the favored hypothesis inferred
with morphological characters from extant taxa indicates
that the echinoderms are the sister group of a hemichordate-plus-chordate clade (fig. 1). It is important to
note that chaetognaths (phylum Chaetognatha) traditionally have been considered deuterostomes, but their
relationships remain uncertain based on morphological
and molecular data (Brusca and Brusca 1990, pp. 882,
889; Bone et al. 199 1; Telford and Holland 1993), and
they are not included in this investigation.
Hypotheses of deuterostome phylogeny have only
recently been evaluated with analyses of gene sequence
Key words: bilateria, chordates, deuterostomes,
phylogeny, 18s rRNA.
morphology,
Address for correspondence and reprints: J. M. Turbeville, Department of Biology, University of Michigan, Ann Arbor, Michigan
48109-1048.
1. Present address: Department of Biology, University of California, San Diego, La Jolla.
Mol. Bid. Evol. 11(4):648-655.
1994.
0 1994 by The University of Chicago. All rights reserved.
0737-4038/94/l
lOS-0009$02.00
648
data. Analyses of partial 18s rRNA sequences (- 1,000
nucleotides) from echinoderms, a urochordate, a cephalochordate, and vertebrates by distance-matrix (Field
et al. 1988) and parsimony (Patterson 1989) analyses
did not support deuterostome monophyly. Analyses with
evolutionary parsimony supported monophyly of the
deuterostomes, although relationships among deuterostome taxa were not resolved (Lake 1990). A recent
analysis (Holland et al. 199 1) compared -500 bp of
the 18s rRNA genes of an enteropneust hemichordate,
echinoderm, and two vertebrates. The analysis weakly
supported a hemichordate-plus-vertebrate clade, but very
few informative sites are present in the region sequenced.
An integrated approach, combining morphological and
molecular characters, has not been utilized to infer deuterostome phylogeny. We report here the first test of
phylogenetic relationships of deuterostomes with complete coding regions of 18s rRNA genes from all major
deuterostome phyla and include an assessment of deuterostome phylogeny with both molecular and morphological characters.
Material and Methods
The complete coding regions of 18s genes of the
sea urchin S’trongylocentrotus purpuratus (Echinodermata, Echinoidea), the brittle star Ophiopholis aculeata
(Echinodermata, Ophiuroidea), and the acorn worm
Deuterostome
Phylogeny
649
ECHINODERMATA
HEMICHORDATA
FIG. 1.-Hypotheses
of deuterostome
relationships. A, Traditional hypothesis. The Echinodermata
is the sister taxon of a hemichordateplus-chordate
clade (Maisey 1986; Schaeffer 1987). B, Hypothesis indicating monophyly
of Hemichordata
plus Echinodermata
(see Grobben
1908; Schaeffer 1987, p. 190). C, Phylogeny indicating that the Echinodermata
is the sister group of the Chordata (Jefferies 1986, p. 323). This
phylogeny is dependent on the hypothesis that the extinct Stylophorans are not echinoderms but possess features of echinoderms and chordates.
The systematic position of these fossils continues to be disputed (Jollie 1982; Jefferies 1986, pp. 345-358; Parsley 1988). Common to all
hypotheses is chordate monophyly. Deuterostome morphological synapomorphies
(among others) are as follows: anus forms near site of blastopore,
and mouth forms secondarily elsewhere (Ax 1989; Schaeffer 1987); and protocoel pore(s) (Balser and Ruppert 1990). Only characters from
extant taxa relevant to the conflicts are plotted and, except where indicated, are from Maisey (1986) and Schaeffer (1987). 1 = Axial complex
(heart/glomerulus
in hemichordates;
Ruppert and Balser 1986; Balser and Ruppert 1990); 2 = gill clefts; 3 = dorsal hollow nerve cord with
neuropore (Bateson 1884); 4 = neurenteric
canal (Siewing 1969, p. 189; Schaeffer 1987); 5 = notochord
(the hemichordate
stomochord
is
commonly
interpreted as a unique feature of the group, but homology of the stomochord
and the chordate notochord
is still disputed; see
Balser and Ruppert 1990); 6 = postanal somatic tail; 7 = endostyle; 8 = vertebrate-like
actins (Vandekerckhove
and Weber 1984; Kovilur et
al. 1993); 9 = equivalence of fate maps; IO = somites; and 11 = differentiation
of neural tube into ependymal and synaptic layers. For additional
synapomoprhies
of the cephalochordates
plus craniates and for craniate synapomorphies,
see (Maisey 1986; Schaeffer 1987). Homology of the
neural gland to Hatschek’s pit and craniate adenohypophysis
is doubtful (Ruppert
1990) and was excluded. For disputed fossil characters
defining nodes marked by an asterisk (*) in phylogeny C, see the work of Jefferies (1986, p. 323). A comprehensive
comparison
of phylogeny
C would require inclusion of fossil characters and taxa; however, although critical reevaluation
of the conflicting interpretation
of the fossil
characters is prerequisite and necessary, it is beyond the scope of this contribution.
Blackened blocks represent characters defining the branch;
and unblackened
blocks represent hypothesized losses. Outgroups are not shown. For recent alternatives that include taxa for which sequences
are not yet available, see the work of Brusca and Brusca (1990, pp. 872-873) and Schram (1 991).
Saccoglossus
kowalevskii (Hemichordata,
Enteropneusta) were amplified by the polymerase chain reaction
(PCR) from genomic DNA templates with eukaryoticspecific 18s primers (2-22 forward, S-ACCTGGTTGATCCTGCCA-3’,
1865- 1847 reverse, S-TGATCCATCTGCAGGTTC-3’;
adapted from Medlin et al. 1988).
The amplified genes were ligated into a T/A vector (Invitrogen) prior to sequencing both coding and noncoding
strands. Direct sequencing of asymmetrically
amplified
PCR product was initially attempted for S. purpuratus,
but sequencing the cloned gene was more consistent. In
addition to the M 13 plasmid primers, five forward and
six reverse internal primers were used,.whose sequences
are as follows (numbers correspond to positions on the
human sequence): 430-447, S-CGGAGARGGAGCCTGAGA-3’; 6 1 1-626,5’-GTGCCAGCMGCCGCGG-3’;
1024-1038, 5’-ATCAAGAACGAAAGT-3’;
1324-1338,
5’-GGTGGTGCATGGCCG-3’;
1488- 1503, 5’-CAGGTCTGTGATGCYC-3’;
445-429, 5’-TCAGGCTCCCTCTCCGG-3’;
632-6 15, 5’-GWATTACCGCGCGGCKGCTG-3’;
880-865, 5’-CCGAGGTCCTATTCCA-3’;
120 1- 1187, 5’-ATTCCTTTRAGTTTC-3’;
1503- 1488,
5’-GGGCATCACAGACCTG-3’;
and 1708- 1692, 5’ACGGGCGGTGTGTRC-3’.
Sequences were aligned using the SeqApp manual
aligner for Macintosh (D. Gilbert, Indiana University).
Alignments were refined, where possible, by using a secondary-structure
model of Xenopus laevis (Atmadja et
al. 1984). Analyses were limited to “reliably”
aligned
regions (Turbeville
et al. 1992; m-65%-70% similarity
and greater; see Hillis and Dixon 199 1). Data were analyzed using the maximum-parsimony
program
of
PAUP 3.0s and 3.1.1 (D. Swofford, Smithsonian
Institution),
the DNADIST,
FITCH,
DNAML,
and
NEIGHBOR
programs of Phylip 3.4 and 3.5~ (J. Felsenstein, University of Washington),
and the evolutionary-parsimony
program (Lake 1987) as implemented
in
the PAUP package. Analyses with evolutionary
parsimony were carried out as described by Turbeville et al.
( 199 1, 1992). For parsimony analyses, the HEURISTIC
search was used with random stepwise addition. Gaps
were treated as missing data in all analyses. The Kimura
correction to estimated distances was employed in distance analyses. One hundred bootstrap replicates were
performed for maximum-likelihood
analysis, and 500
were run for all other analyses. The tree-length distribution test was carried as described by Hillis and Huelsenbeck ( 1992).
650
Turbeville et al.
Results
Maximum parsimony (MP) analyses of 18s sequences alone do not support monophyly of the Chordata, a result that is at variance with strong morphological evidence (fig. 1). The urochordates form the sister
taxon to the hemichordate, and together this clade plus
the echinoderms form the sister taxon to the cephalochordates plus craniates (fig. 2A). Support is weak for
this arrangement, as indicated by bootstrap percentages
and decay values (fig. 2A). The poorly supported artifactual placement of the long-branched urochordates
may be the result of their sequences converging with the
most divergent nonchordate deuterostome, the enteropneust hemichordate
Saccoglossus. Chordata was
monophyletic in only 16% of 500 bootstrap replicates.
A monophyletic Chordata is found as three of nine
equally parsimonious trees that require four additional
substitutions. Two correspond to morphological phylogeny B (fig. 1). Morphological hypotheses A and C (fig.
1) are among 45 equally parsimonious trees saved at 11
extra substitutions. Evaluation of 100 trees saved that
are within 1% of the length of the shortest tree suggests
further that signal for reliable phylogeny inference
among the major deuterostome taxa is weak in this molecule (fig. 2B).
The tree-length distribution test (Hillis and Huelsenbeck 1992) was utilized to assess the strength of the
signal in these sequence data. According to Hillis and
Huelsenbeck ( 1992), data sets containing phylogenetic
signal result in left-skewed tree-length distributions, and
those containing random noise result in nearly symmetrical distributions.
Tree-length distribution of
100,000 trees including all taxa is significantly left skewed
(gl = -0.794; fig. 3A). However, when all strongly supported clades are constrained (those with decay indices
of five or greater and with bootstrap values 88% and
greater), the distribution of the remaining trees is nearly
symmetrical (gl = 0.031; fig. 3B). A similar result is
obtained if only a single representative of each clade is
included in the analysis. Thus, this test suggests that the
signal is limited across the major deuterostome taxa.
Topological variation was observed when individual ingroup or outgroup taxa were removed or substituted and analyses were rerun. For example, the hemichordate-plus-urochordate
clade was found as the sister
group to the cephalochordate-plus-craniate
clade when
Placopecten and Eurypelma alone were used as outgroups. When the short-branched (as inferred from the
tree with all taxa) echinoderm Strongylocentrotus was
removed, two of three equally parsimonious trees were
equivalent to morphological hypothesis B (fig. 1). The
other tree is identical to the shortest tree found including
all taxa (fig. 2A). Thus, topological instability is apparently not related solely to the longest-branched taxa
(Styela, Herdmania, and Myxine), and their removal
has no effect on the placement of the other taxa.
When the 1 molecular and 11 morphological characters listed in figure 1 are combined with the sequence-
3
G
1
9
XWlOpJS
sebastolobus
SqUallJS
Lathha
Myhe
.
Bmnchioskma CEPHALOCHORDATA
2
a
0
2
Petromyzon
8
Brandliostofna
0
Stvh
4
Hwdman&
S8ccoglossu8
Ophlophollr
Stmngylocontrotus
Placopecten
Limicolaria
FIG. 2.-Phylogenies inferred by parsimony analyses of 18s rRNA gene sequences. A, Minimal length tree found by analyzing all reliably
aligned positions. The HEURISTIC search was employed with random stepwise addition. Two hundred replicates were run. There are 304
phylogenetically informative (parsimony) sites of 1,63 1 analyzed. Tree length is 1,155 and consistency index is 0.656. If uninformative sites are
excluded, tree length is 876 and consistency index is 0.547. Numbers above nodes are bootstrap percentiles for 500 replicates. Numbers in
boldface (i.e., “decay indices”) indicate the number of extra steps, within five, at which the clade descending from that node is no longer
supported (see Bremer 1988; Donoghue et al. 1992). Branch lengths are proportional to the number of nucleotide substitutions. Scale bar = 50
substitutions. B,Strict consensus of 100 trees within 1% length of the shortest (885~step) tree.
Deuterostome Phylogeny
65 1
(ML) trees are similar in major respects to neighborjoining trees (fig. 6C and D and table 1). A chordate
lineage is weakly supported, as suggested by bootstrap
results (66% with FM and 45% with ML). Both FM and
ML find a poorly supported echinoderm-plus-enteropneust hemichordate group (47% with FM and 58%
with ML) as the sister group of chordates (fig. 6C and D).
Tree lengths
FIG. 3.-Tree-length distributions. A, Distribution of a random
sample of 100,000 trees including all taxa. B,Distribution of the remaining 3 15 trees when all clades with decay indices >5 or bootstrap
values >88% are removed with the constraint option of PAUP 3.1.1.
The EXHAUSTIVE search was employed.
data matrix, the shortest tree indicates a monophyletic
Chordata that is the sister group to an echinoderm-plushemichordate clade (fig. 4). This tree is equivalent to
one of nine suboptimal trees of length 880, when the
rRNA data alone are used. The phylogeny inferred
with the combined data set is equivalent to phylogeny B
(fig. 1). The combined analysis also indicates where the
signal is weak, since only five chordate morphological
synapomorphies are required to find a monophyletic
Chordata.
Evolutionary parsimony does not find support at
the 5% significance level for a monophyletic Chordata;
support for inclusion of the urochordates in a chordate
clade is not significant (fig. 5). The method favors a
hemichordate-plus-echinoderm
clade, .but not significantly (fig. 5).
Neighbor joining (NJ) supports a chordate lineage,
whose sister taxon is a Saccoglossus-plus-echinoderm
group. It also indicates that the cephalochordate is the
closest chordate relative of the craniates. Support for
these relationships as assessed by bootstrapping is weak
(fig. 6A). This method failed to find a monophyletic
Chordata when mollusks alone were included as outgroups and the longest branched ingroup taxon was deleted; the position of the urochordates is unresolved (fig.
6B). Fitch-Margoliash (FM) and maximum-likelihood
Discussion
The results of these analyses raise cautions regarding
reliability of phylogenies reconstructed with rRNA gene
sequences. Taxa chosen for the ingroup and outgroup
may radically alter the topology, regardless of length, as
previously recognized (see Swofford and Olsen 1990;
Turbeville et al. 199 1; Hillis and Huelsenbeck 1992).
The variation in topology observed in this study when
particular taxa, long-branched or otherwise, are deleted
and the analyses are rerun suggests that there is conflict
in the existing signal. Artifactual grouping of taxa with
long branches (see Felsenstein 1978) is also a recurrent
problem. Increasing the number of both ingroup and
outgroup taxa could potentially reduce these problems,
because their inclusion might reveal previously undetected homoplasy (Sanderson 1990; Swofford and Olsen
1990).
Another serious pitfall of sequence comparisons
concerns the inference of positional homology, or alignment (see Swofford and Olsen 1990; Hillis and Dixon
199 1). Although we have limited our analyses to,regions
Branchiostoma
S=Wllo-u~
HEMICHORDATA
Ophlopholls
ECHINODERMATA
Strongylocentrotu8
1
1
E”~~‘
1 ma
;;;?$@?I”
Ten&ii
MOLLUSCA
ARTHROPODA
FIG. 4.-Strict consensus of two equally parsimonious trees found
in a combined parsimony analysis of 18s rDNA sequence data, the
morphological and single molecular character listed in fig. 1. All characters were weighted equally. A monophyletic Chordata is found.
Numbers above nodes are bootstrap percentiles. Numbers in boldface
below nodes are clade decay values. Length of trees is 1,172 (893 when
uninformative sites are ignored). The overall confidence index is 0.656
and. when uninformative sites are excluded.I 0.549.
652
Turbeville et al.
Xenopus
Latimeria
sebastolobus
squaius
Myxine
Petromyzon
BraflChiO6t~
XLatimeda
sebastolokrs
Sty&
Herdmania
tiww
Myxine
PelKMnyron
Branch&stoma
)pz0.*4(
Saccoglorsur
>--
Ophiopholla
Strongybcontrotur
PLacoQedm
LimkAaria
Saccoglor8ur
(
Ophlophdis
Strongyiocontrotur
EUrypelM
Tenekio
A
B
FIG.5.-Representative
analyses with evolutionary parsimony. A, Test of urochordate relationships. The favored tree from combining
results for 6 craniates and the cephalochordate X 2 urochordates X hemichordate x 2 echinoderms (28 quartets) is shown. The P value for the
expected tree (linking urochordates and craniates and the cephalochordate) is 0.72. The P value for the third alternative is 0.69. B, Test of
deuterostome relationships. The favored tree from a combination of 6 craniates and the cephalochordate X hemichordate X 2 echinoderms
X the 2 mollusks and 2 arthropods (56 quartets) is shown. The favored tree links echinoderms and hemichordates, although not significantly.
The P values for trees linking the hemichordate and chordates and the echinoderms and chordates are P = 0.6 and P = 0.38, respectively.
of the sequence that are well conserved (Hillis and Dixon
199 1), it is probable that there are improperly
aligned
positions contributing
to inferred noise (homoplasy),
in
addition to homologous sites that have incurred multiple
hits. It should be noted that somewhat different topologies resulted when a computer-generated
alignment
(Clustal V; Higgins et al. 1992) was analyzed; only FM
and ML found a chordate lineage (authors’ unpublished
observations).
Nonindependent
changes may also lead to incorrect
phylogenies, although this may be difficult to assess (see
Wheeler and Honeycutt
1988; Hillis and Dixon 199 1;
Dixon and Hillis 1993). Substitution
bias also has been
implicated
in incorrect phylogeny inference (Lockhart
et al. 1992; Marshall 1992). Our preliminary
observations (authors’ unpublished
data) reveal no obvious bias
that might account for the placement of the urochordates
outside the chordates by parsimony analysis.
The expectation
has been that gene phylogenies
would allow reliable inference of the phylogenetic
position of organisms that cannot be placed on the basis
of morphological
comparisons.
But because optimal
trees reconstructed
from sequence data alone often conflict with well-supported morphological
phylogenies (e.g.,
chordate monophyly),
it will be essential to compare
phylogenies obtained from several appropriate genes.
The additional information
present in complete 18s
rDNA sequences does not provide significantly
greater
resolving power for inference of deuterostome
relationships than do previously
published partial sequences.
The data still contain minimal signal and sufficient noise
to prevent unambiguous
resolution
of the conflicting
hypotheses.
It is significant that one of the hypotheses (fig. 1B)
is supported by NJ, FM, and MP when morphological
and molecular data are combined (ML differs only in
the placement of the urochordates)
(fig. 4 and table 1).
The phylogeny inferred from total evidence by MP may
be considered the current best combined hypothesis of
deuterostome
relationships.
However, the combined
analysis of molecular and morphological
data, although
justifiable (IUuge 1989; Donoghue and Sanderson 1992;
Jones et al. 1993), remains controversial (Swofford 199 1;
de Queiroz 1993). A major concern is that that the different data sets may not be homogeneous
(Bull et al.
1993). The limitations
of sequence data previously discussed also illustrate the value of initially analyzing sequence data apart from other characters, as these problems might otherwise go undetected (Swofford 199 1).
If the phylogeny inferred from total data is confirmed, it will suggest that the axial complex (heart-glomerulus) of hemichordates
and echinoderms
is a synapomorphy (see Balser and Ruppert 1990) of the clade and
that two complex morphological
characters-gill
slits
and dorsal hollow nerve cord-shared
by enteropneust
hemichordates
and chordates are homoplasies
rather
than shared-derived homologues (see Schaeffer 1987; fig.
1B and table 1). One interpretation
of character evolution in the combined
hypothesis is that the ancestral
deuterostome
possessed gill slits and a dorsal hollow
nerve cord and that these features were lost in the stem
lineage of the echinoderms
(fig. 1B). If the combined
Deuterostome
Phylogeny
653
1
5
5
f
Petru4nyzorl
Branchiostoma
c
UROCHORDATA
_
-
0
,_j-sstyela
Herdmania
Saazoglossus
Ophlophotls
Strongyloconbotus
P-m
ARTHROPODA
Limicdaria
1
B
A
1
YpJfw_ 100
- 07
57
PWOmyZOfl
98
bhwia
sebastolobus
Gqal~
47
Myxine
Myxine
66.
Bmnchiostoma
4!3
loo
100f-SW
Herdmania
Branchiioma
Strongylocwtrotus
1
4
C
D
FIG. B.-Distance
and likelihood trees. A, Phylogeny inferred by neighbor-joining
analyses of 18s sequences. Branch lengths are proportional
to the number of nucleotide substitutions.
B,Neighbor-joining tree resulting after the longest-branch ingroup (Myxine) and arthropod outgroups
were removed. C, Fitch-Margoliash
tree. D,Maximum-likelihood
tree. Scale bars = 20 substitutions/l,000
nucleotide positions. Numbers at
nodes are bootstrap percentiles; there were 500 replicates for A and C and 100 for D.
hypothesis is confirmed,
it will also strengthen the necessity of reevaluating
the extinct stylophorans.
Jefferies
(1986, pp. 19 l-238) interprets these fossils as chordates
possessing a calcite skeleton and asymmetry/dexiothetism (reduction
of structures on the right side). These
two characters occur in echinoderms and are interpreted
by Jefferies ( 1986, p. 323; also see Cripps 199 1; fig. 1C)
as synapomorphies
of an echinoderm-plus-chordate
clade. Other paleontologists
interpret stylophorans
as
extinct echinoderms
(see Parsley 1988; Sprinkle 1992),
a hypothesis in accord with the provisional results presented here.
An exhaustive evaluation of character evolution will
require a strongly supported hypothesis of deuterostomes
relationships,
and we advocate further testing of deuterostome
phylogeny.
This will require
additional
molecular
data from all deuterostome
taxa, including
pterobranch
hemichordates,
in combination
with a
reassessment
of deuterostome
phological characters.
relationships
with mor-
Sequence Availability
New sequences are available in GenBank.
Accession numbers are L28054, L28055, and L28056 for Saccoglossus, Ophiopholis, and Strongylocentrotus, respectively. The alignment is available from EMBL (accession
number DS 169 14) by anonymous
FTP or E-mail and
from the senior author (J.M.T.) via E-mail: “[email protected]”.
Note added in proof: While this paper was in press,
an assessment of deuterostome
phylogeny with complete
18s rDNA sequences appeared (Wada and Satoh 1994).
The results of this study, which employed NJ and ML
and included additional sequences, are only partly congruent with those presented here. A chordate lineage
was not found.
654
Turbeville et al.
Table 1
Summary of Molecular and Morphological Data Analyses
BRUSCA,
R. C., and G. J. BRUSCA. 1990. Invertebrates. Sinauer,
Sunderland, Mass.
BULL, J. J., J. P. HUELSENBECK, C. W. CUNNINGHAM, D. L.
MORPHOLOGICAL
PHYLOGENY~
ANALYSIS~
A
B
MPC......
MP”.......
EPd. ......
NJ. .......
FM. .......
ML. ......
-
+
-
+
+
+
C
-
“MPC, Maximum parsimony analysis of molecular plus morphological
characters. Other analyses with sequencesalone: MP, maximum parsimony;
EP, evolutionary parsimony; NJ, neighbor joining, FM, Fitch-Margoliash;
ML, maximumlikelihood.Withinthe Chordata,MPC, NJ, andFM linkcephalochordatesandcraniates.ML linksurochordatesandcraniates.
bA plus sign (+) indicatesthat topology was found; and a minus sign (-)
indicatesthat it was not found. CH = Chordata; HE = Hemichordata;and EC
= Echinodermata.
‘See text andfig. 2.
dSee text.
SWOFFORD, and P. J. WADDELL. 1993. Partitioning and
combining data in phylogenetic analysis. Syst. Biol. 42 : 384397.
CRIPPS, A. P. 199 1. A cladistic analysis of the cornutes (stem
cornutes). Zool. J. Linnean Sot. 102:333-336.
DE QUEIROZ, A. 1993. For consensus (sometimes). Syst. Biol.
42:368-372.
DIXON, M. T., and D. M. HILLIS. 1993. Ribosomal RNA secondary structure: compensatory mutations and implications
for phylogenetic analysis. Mol. Biol. Evol. 10:256-267.
DONOGHUE, M. J., R. G. OLMSTEAD, J. F. SMITH, and J. D.
PALMER. 1992. Phylogenetic relationships of Dipsacales
based on rbcL sequences. Ann. MO. Bot. Gardens 79:333345.
DONOGHUE, M. J., and M. J. SANDERSON. 1992. The suitability
of molecular and morphological evidence in reconstructing
plant phylogeny. Pp. 340-368 in P. S. SOLTIS, D. E. SOLTIS,
and J. J. DOYLE, eds. Molecular systematics of plants.
Chapman & Hall, New York and London.
FELSENSTEIN, J. 1978. Cases in which parsimony or compatibility methods will be positively misleading. Syst. Zool. 27:
401-410.
FIELD, K. G., G. J. OLSEN, D. J. LANE, S. J. GIOVANNONI,
Acknowledgments
We thank Jessica Kissinger, John Logsdon, and
Gavin Naylor for helpful suggestions and Greg Wray for
Ophiopholis sperm samples. This research was supported
by NSF grant BSR-88 18044.
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SIMON EASTEAL, reviewing
Received
January
Accepted
February
10, 1994
3, 1994
editor