The origins and evolution of the Ecdysozoa

Ecdysozoa is a clade composed of eight phyla, three of which—arthropods, tardigrades, and onychophorans—share segmentation and have appendages, and the remaining five—nematodes, nematomorphs, priapulids, kinorhynchs, and lor- iciferans—are worms with an anterior proboscis or introvert. Ecdysozoa contains the vast majority of animal species and there is a great diversity of body plans among both living and fossil members. The monophyly of the clade has been called into ques- tion by some workers, based on analyses of whole- genome data sets, and we review the evidence that now conclusively supports the unique origin of these phyla. Relationships within Ecdysozoa are also controversial and we discuss the molecular and morphological evidence for several monophy- letic groups within this superphylum.


8.1 Introduction

The Ecdysozoa is a widely accepted clade that encompasses the Euarthropoda (Insecta, Crustacea, Myriapoda, and Chelicerata), the arthropod-like Onychophora and Tardigrada, and five phyla of introvert bearing worms: the Nematoda, Nematomorpha, Priapulida, Kinorhyncha, and Loricifera. In terms of species numbers and niche diversity, the Ecdysozoa is far and away the most significant clade of animals ever to have existed, with over a million described species and an esti- mated total of more than 4.5 million living spe- cies (Chapman, 2005). The extraordinary number of insects is well known—there are estimated to be more than 10 times as many species of insects
than there are of all the deuterostomes and lopho- trochozoans put together—yet even if the founder of the insect lineage had been eaten by a passing frog, the nematodes and the rest of the arthropods (myriapods, chelicerates, and crustaceans) would still easily outnumber all other living animals by close to a quarter of a million species (Chapman,
2005). Their characteristic tough cuticle also means that ecdysozoans are well represented in the fos- sil record, adding further wonderful forms to the diversity of the clade.
Despite the huge number of species and great niche diversity, the basic body plans of the Ecdysozoa are rather conservative, being either insect-like with a segmented body and jointed appendages or worm-like with an anterior cir- cum-oesophageal nerve ring and a terminal mouth usually found on an introvert. All groups lack a primary larva as generally conceived and possess a moulted cuticle with concomitant lack of locomotory cilia. The periodic moulting or ecdy- sis of the cuticle gives the assemblage its name of Ecdysozoa. Although the morphological diver- sity of ecdysozoan phyla may be seen as fairly restrained when compared with the diversity of shapes seen among Lophotrochozoa, for example, these two ecdysozoan body plans happen to manifest themselves in the two most intensively studied invertebrates on the planet, the nematode Caenorhabditis elegans and the fruitfly Drosophila melanogaster.
Prior to 1997, the prevalent view of arthropod relationships linked them, via the onychophorans, to the annelid worms. This annelid–arthropod


71

72 AN I M AL EV O L UTI O N



clade is called Articulata in recognition of the prin- cipal character uniting these phyla: a segmented body. Articulata was generally thought to be part of a larger assemblage of animal phyla linked by the possession of a coelomic cavity and called Coelomata. Although the concept of a relation- ship between arthropods and pseudocoelomate worms such as nematodes and priapulids existed much earlier (discussed in Schmidt-Rhaesa, 1998), the first support from molecular sequence data for such a relationship, and indeed the first reference to the Ecdysozoa, date to a paper by Aguinaldo et al., (1997).
Our discussion is predicated on the assump- tion that the Ecdysozoa is a natural, monophyletic group; however, the existence of the Ecdysozoa is not yet universally accepted and so we will con- sider the evidence that has amassed in support of the monophyly of this group in the decade since the paper by Aguinaldo et al. The relationships among the introvertan worms, their position rela- tive to Panarthropoda (Onychophora, Tardigrada, and Euarthropoda), and several aspects of the phyl- ogeny within Panarthropoda and Euarthropoda themselves are all still controversial and we will consider recent arguments concerning each of these.


8.2 Ecdysozoa is a monophyletic group

The initial support for the Ecdysozoa came from a study of small-subunit (18S) ribosomal RNA (SSU rRNA) genes, that specifically addressed a com- mon problem of phylogeny reconstruction—long branch attraction (LBA; Felsenstein, 1978). This sys- tematic error is encountered when using molecular data derived from C. elegans and many other nema- todes (Aguinaldo et al., 1997) and stems from the fact that these genomes have evolved rapidly rela- tive to those of most other animals. This instance of LBA would tend to cause the branch leading to the fast-evolving nematodes to be shifted towards the root of a tree. The use of short-branched nema- todes in the analysis of Aguinaldo et al., resulted in the nematodes moving from their position close to the root of the bilaterian tree (one also supported by consideration of their morphology, in particular
their lack of a coelomic cavity), to a close relation- ship with the arthropods and priapulid worms in a clade which the authors named the Ecdysozoa (Aguinaldo et al., 1997).
Subsequent analyses of rRNA genes have confirmed this result and extended membership of the Ecdysozoa beyond Nematoda and Priapulida to include three further phyla of worms— Nematomorpha, Kinorhyncha, and Loricifera. The contribution of pseudocoelomate worms to the ecdysozoan clade had been anticipated by various authors who had already linked these five worm phyla in a group called the Cycloneuralia (Ahlrichs,
1995) or Introverta (Nielsen, 2001).
The finding of monophyletic Ecdysozoa has
been replicated by other taxonomically well-sam-
pled data sets, including combined small- and
large-subunit (LSU) rRNAs (Mallatt and Winchell,
2002; Mallatt and Giribet, 2006) and myosin heavy-
chain sequences (Ruiz-Trillo et al., 2002), as well as
Hox gene signature peptides (de Rosa et al., 1999)
and the (somewhat puzzling) shared presence in
the nervous system of all studied ecdysozoans
of an unidentified antigen recognized by the
anti-horseradish peroxidase (anti-HRP) antibody
(Haase et al., 2001). The multimeric beta-thymosin
gene found in flies and nematode worms (Manuel
et al., 2000) has been shown not to be a reliable syn-
apomorphy of the Ecdysozoa (Telford, 2004).
Despite these congruent results, there exists
a powerful series of papers arguing against the
close relationship of nematodes and arthropods
and supporting instead the traditional view of
the monophyletic Coelomata linking arthropods
such as D. melanogaster to humans rather than to
nematode worms (Blair et al., 2002; Wolf et al., 2004;
Philip et al., 2005; Ciccarelli et al., 2006; Rogozin
et al., 2007a,b, 2008). This specific phylogenetic
question has the attraction of being approachable
with the largest possible molecular data sets: the
completely sequenced genomes of flies, worms,
and humans. What almost all studies that have
used this approach have found is that the evidence
is strongly in favour of the Coelomata hypothesis
and against Ecdysozoa.
The counter argument, naturally, is that these
whole-genome studies suffer from precisely the
problem that the Aguinaldo et al. paper addressed;

ECD YS OZ OA N E VO LU T I O N 73



the systematic artefactual attraction of the nema- tode branch towards the root of the Bilateria due to LBA. This contention does seem to be borne out by a number of publications in the past few years. Copley et al., (2004) compared the presence or absence of 1712 genes or distinct combinations of protein domains specific either to flies and humans or to flies and nematode worms. There were many more of the former, giving apparently strong sup- port to the Coelomata hypothesis. However, they were able to show that this strong signal was an artefact resulting from a strong tendency towards secondary loss of genes in the nematode, a fea- ture of its high rate of genomic evolution (Copley et al., 2004). In parallel, Philippe et al., (2005a) used large ‘phylogenomic’ data sets (whole genomes combined with data from expressed sequence tag projects and hence having much broader taxon sampling) and showed that experiments designed to reduce potential long-branch effects—using less distant outgroups, selecting slowly evolving nema- todes, and discarding the more unevenly evolving genes—supported Ecdysozoa while Coelomata was supported without these efforts. Finally, a similar approach has been used (Irimia et al., 2007; Roy and Irimia, 2008a,b) to show that claims of an excess of identical, rarely changing, amino acids and of spe- cifically located introns shared by flies and humans and lacking in nematode worms (e.g. Rogozin et al.,
2007a,b, 2008) are biased by the use of distant out- groups and by the rapid evolution of C. elegans. They show that, when these biases are accounted for, there is significantly more support for Ecdysozoa than for Coelomata from this source of evidence (Irimia et al., 2007; Roy and Irimia 2008a,b).
The phylogenomic approach has recently been extended to the slowly evolving Priapulida that are strongly supported as ecdysozoans (Webster et al., 2006) and for Nematomorpha (T. Juliusdottir, R. Jenner, MJT, and R. Copley 2007, unpublished). This result was further strengthened by analyses of the very arthropod-like mitochondrial genome of Priapulus caudatus. Perhaps even more strikingly, the priapulid mitochondrial gene order can be rec- onciled with that of the arthropods by a single inversion (Webster et al., 2006).
We would also like to highlight a further
very convincing synapomorphy supporting the
monophyly of Protostomia, and hence, we believe, definitively ruling out the Coelomata hypothesis. Papillon et al., (2004) used the presence of a dozen rarely changing amino acids in the mitochondrial nad5 gene of protostomes as a striking indication that the chaetognaths were protostomes and not deuterostomes as traditionally believed. The sig- nature constitutes a very complex, conserved, derived character defining a monophyletic group of Protostomia. We have extended this analysis of the nad5 gene, which appears to have undergone a significant burst of evolution within the lineage leading to the protostomes. Almost all of these sig- nature amino acids are found in nematodes and priapulids as well as in other controversial proto- stomes, including rhabditophoran and catenulid flatworms and lophophorates. A monophyletic Protostomia, while not specifically proving the existence of Ecdysozoa, is clearly incompatible with a monophyletic group of coelomate animals and therefore contradicts the results from whole- genome studies supporting Coelomata.
We strongly support the notion of a mono- phyletic Ecdysozoa and feel that the only oppos- ing evidence—the whole genome support for Coelomata—is flawed by systematic error, which has been addressed successfully by much better taxon sampling, in particular the use of a close out- group (Philippe et al., 2005a; Dunn et al., 2008). In addition to the molecular systematic support, the monophyly of Ecdysozoa is supported by a number of morphological synapomorphies including ecdy- sis of a trilayered cuticle (consisting of epi-, exo-, and endocuticle), lack of locomotory cilia, lack of primary larva, terminal mouth, the HRP antigen in the nervous system, and conserved mitochon- drial gene order that have been mentioned (see also Schmidt-Rhaesa, 1998).


8.3 Cycloneuralia, Introverta, Scalidophora, and Nematoida

While we do not have an equivalent of the nad5 rare genomic change to support the monophyly of the Ecdysozoa within the Protostomia, as we have seen, we do have strong evidence from phy- logenomic data sets of tens to hundreds of genes for the monophyly of Arthropoda plus Nematoda

74 AN I M AL EV O L UTI O N



and Priapulida (Philippe et al., 2005a; Webster et al.,
2006). More recently, data from Kinorhyncha and
Nematomorpha have been added, and these phyla
too were found to be part of the Ecdysozoan radi-
ation (Dunn et al., 2008). These worm phyla had
previously been linked to one further phylum, the
Loricifera, in a group collectively known as the
Cycloneuralia (Ahlrichs, 1995). The name refers to
their collar-shaped, circum-oral brain; something
similar is seen in Gastrotricha which are, how-
ever, lophotrochozoans not ecdysozoans (Telford
et al., 2005; Todaro et al., 2006). These phyla (but not
Gastrotricha) also share an eversible anterior end,
or introvert, which terminates in the mouth and
gives the alternative name of Introverta (Nielsen
1995, 2001), although the introvert is only seen
in the larvae of Nematomorpha and in isolated
examples of Nematoda.
What we still do not have is much reliable infor-
mation on the relationships between these phyla or
their relationships to the Panarthropoda. This may
be explained in part by the difficulty in working
on the minute and hard to study Kinorhyncha and
Loricifera. The two groupings that do seem cred-
ible are a close relationship between Nematoda
and Nematomorpha and between Priapulida and
Kinorhyncha. Nematodes and nematomorphs share
a number of characters, including the reduced cir-
cular muscles in the body wall, the cloaca seen in
both sexes, the aflagellate sperm, the cuticle (col-
lagenous not chitinous), and the ectodermal ven-
tral and dorsal nerve cords and were grouped
by Nielsen (1995, 2001) and named Nematoida by
Schmidt-Rhaesa (1998). This clade has weak sup-
port from SSU rRNA gene analyses (Peterson and
Eernisse, 2001), combined analyses of LSU and SSU
rRNA (Mallatt et al., 2004), and from a recent phyl-
ogenomic analysis (Dunn et al., 2008).
Morphologists have also united Priapulida,
Kinorhyncha, and Loricifera in the Scalidophora
(Schmidt-Rhaesa, 1998) or Cephalorhyncha
(Nielsen, 1995, 2001), on the basis of an introvert
with scalids and the presence of two rings of
retractor muscles on the introvert. The close rela-
tionship between priapulids and kinorhynchs at
least seems to hold up (Dunn et al., 2008), but of the
two existing phylogenetic studies that include data
from the Loricifera, one showed their position to be
ambiguous (Park et al., 2006) and the other linked them to the Nematomorpha (Sørensen et al., 2008). Early studies gave no strong indication that the Cycloneuralia is a monophyletic group, and in fact most evidence pointed to Priapulida (and there- fore Kinorhyncha too) as being the earliest branch and the Nematoida as being the sister group of the Panarthropoda (e.g. Mallatt and Giribet 2006; Webster et al., 2006). In the phylogenomic analysis of Dunn et al., (2008), however, the Cycloneuralia are monophyletic. These alternative scenarios clearly have important consequences for the evo- lution of the arthropods, as the former implies either convergent evolution of complex structures such as segments and coeloms in the arthropods and annelids or parallel losses of such structures in independent clades of paraphyletic cyloneuralians. The latter result—monophyletic cycloneuralia—is a more parsimonious view of the evolution of morphology but is less informative regarding the
origins of the Arthropoda.


8.4 Panarthropoda: Euarthropoda, Tardigrada, and Onychophora

The monophyly of the Euarthropoda plus Onychophora and Tardigrada seems, on the face of it, uncontentious. They are linked by a number of features, the most important of which are the segmentally repeated limbs with terminal claws; Onychophora translates as ‘claw bearer’. The limbs in all three groups straddle parasegmental bound- aries marked by the expression of the segment polarity gene engrailed (Patel et al., 1989; Gabriel and Goldstein, 2007). Their segmental paired, sac- cate nephridia (reduced in number and function- ing as excretory organs in euarthropods) and open circulatory system also seem to be valid synapo- morphies, although both are missing in the mini- aturized tardigrades (Hejnol and Schnabel, 2005). The circulatory system is characteristically formed as a fusion of both the coelomic cavities and the primary body cavity/embryonic blastocoel (i.e. a mixocoel), and there is a dorsal heart with charac- teristic openings (ostia) into the open circulatory system (Nielsen, 1995, 2001).
Despite the characters in common with
Panarthropoda, not all molecular studies support

ECD YS OZ OA N E VO LU T I O N 75



their monophyly, some grouping Tardigrada with the nematodes rather than with Euarthropoda (Dunn et al., 2008). This relationship between tar- digrades and nematodes is biologically implausible and LBA is a strong contender for an explanation of this result. If we assume that Panarthropoda (including Tardigrada) is monophyletic, three trees could unite the euarthropods, tardigrades, and onychophorans: Euarthropoda with either (1) Tardigrada or (2) Onychophora, or (3) Tardigrada and Onychophora as sister groups. The branching order of these three taxa is not resolved by mol- ecules or morphology. If we assume that the small size of tardigrades is derived and accounts for the absence of mixocoel, heart, and nephridia (Schmidt- Rhaesa, 2001), characteristics found in Onychophora and Euarthropoda), we suggest that the similarities of cuticle, ganglionated ventral nerve chord, and limbs in tardigrades and Euarthropoda may indi- cate a sister-group relationship.


8.5 Euarthropoda: Myriochelata versus Mandibulata

The relationships of the four euarthropod clades— Chelicerata, Myriapoda, Crustacea, and Hexapoda (Hexapoda = Insecta, plus the basally branching groups Diplura, Protura, and Collembola) have long been disputed. A decade or so ago there were even serious arguments over the single ver- sus multiple origins of arthropodization, and therefore over the monophyly versus polyphyly of euarthropods (Fryer, 1997). Molecular analyses have emphatically supported the monophyly of euarthropods and a unique origin of their cuticu- larized body and jointed appendages, and in the past years attention has been focused more on the relationships between these four groups. One com- mon feature of morphology-based interpretations of arthropod phylogeny was the close relationship between Myriapoda and Hexapoda in a clade called the Atelocerata (which means malformed horns and refers to their common lack of a pair of second antennae) defined additionally by unbranched (uniramous) appendages, Malpighian tubules, and tracheal breathing (Telford and Thomas, 1995). According to the polyphyleticists, the Atelocerata are grouped with the Onychophora in a clade
called the Uniramia. Arguably, the clearest result to date in arthropod phylogeny shows that the insects are not most closely related to the myria- pods but to the crustaceans (Boore et al., 1995) and, in all likelihood, constitute a subgroup within the Crustacea. This clade of crustaceans plus insects has been referred to as the Pancrustacea or as the Tetraconata due to the tetrapartite crystalline cones of the ommatidia (Dohle 1997, 2001; Harzsch 2002,
2004; Harzsch et al., 2005).
More controversial, though, is the true position
of Myriapoda which share numerous similarities
of head organization not only with the insects (as
discussed) but also with crustaceans, most notably
the presence of a mandible on the third, appendage-
bearing head segment and maxillae on the fourth
and fifth head segments. The common head struc-
ture of myriapods, crustaceans, and insects with
two pairs of antennae (at least primitively), paired
gnathobasic mandibles, and two pairs of maxillae
strongly supports their monophyly. This group is
named the Mandibulata, reflecting the particular
importance of detailed similarities seen between
the mandibles of Pancrustacea and Myriapoda in
terms of segmental identity, positioning relative to
other body parts, gene expression, detailed simi-
larities in terminal differentiation, and, of course,
in function (Scholz et al., 1998; Edgecombe et al.,
2003; Harzsch et al., 2005). Surprisingly, a number
of molecular studies using rRNA genes, nuclear
protein-coding genes, and complete mitochondrial
genome sequences do not support the mandibu-
late clade, instead linking the myriapods to the
chelicerates in a group called the Paradoxopoda
or the Myriochelata (Mallatt et al., 2004; Negrisolo
et al., 2004; Pisani, 2004). An analysis of nuclear
protein-coding genes (Regier et al., 2005), however,
did not find strong support for either Myriochelata
or Mandibulata, suggesting that there is uncer-
tainty over the affinity of myriapods. Recognizing
that the distinction between the two possibilities
comes down to the position of the root of the
euarthropod tree, we have reanalysed the com-
plete mitochondrial genome sequences of various
arthropods using a priapulid as a short-branched,
phylogenetically close relative of the arthropods
(Rota-Stabelli and Telford, 2008). We find that, in
contrast to previous studies that had used more

76 AN I M AL EV O L UTI O N



distant outgroups (lophotrochozoans), our mito- chondrial tree narrowly supports Mandibulata over Myriochelata (Figure 8.1; see also Pisani,
2004). We have also analysed a number of nuclear protein-coding genes and have reached the same conclusion (Bourlat et al., 2008; Figure 8.2). While our bias in support of a return to the Mandibulata is probably obvious, it is clear that this ques- tion remains to be resolved one way or the other. While there are specific characteristics shared by myriapods and chelicerates but absent from Pancrustacea (Dove and Stollewerk, 2003; Kadner and Stollewerk, 2004; Stollewerk and Simpson,
2005), it is difficult to demonstrate these as syn- apomorphies as we have insufficient data from an outgroup and the suspicion is that the Myriapoda/ Chelicerata character state may be plesiomorphic and uninformative (Harzsch, 2004; Harzsch et al.,
2005). While the same criticism may be made of some of the characters supporting Mandibulata, the chelicerate homologue of the mandible (the first walking leg; see Telford and Thomas, 1998) seems likely to represent the plesiomorphic condition as it strongly resembles adjacent, serially homologous,

dible itself is a shared derived character uniting the mandibulates.
(a)










(b)










(c)








Lophotrochozoa

Phylogenetically distant outgroups: Paradoxopoda








Nematoda

Long branch Ecdysozoa: unresolved Euarthropoda








Priapulida

Short branch Ecdysozoa: Mandibulata



8.6 Pycnogonids are chelicerates

The pycnogonids, or sea spiders, have long been associated with the chelicerates due to the shared character of chelicerae (chelifores in pycnogonids) on the first limb-bearing segment. This phylogen- etic link was questioned recently both by studies of their nervous systems and by molecular systematic analyses. The larval nervous system of a pycnogo- nid from the genus Anoplodactylus was studied and the chelifore appeared to be innervated from the frontmost portion of the brain (the protocerebrum), suggesting that this appendage was therefore not homologous to the chelicerae of other chelicerates which is innervated from the second portion of the brain (the deutocerebrum; Budd and Telford, 2005; Maxmen et al., 2005). This tied in with a molecu- lar phylogenetic study placing the Pycnogonida at the base of Euarthropoda and not with Chelicerata (Regier et al., 2005). Subsequent analysis of Hox
Figure 8.1 Different outgroups support different positions of the root of the Euarthropoda. (a) Support for Myriochelata (Myriapoda + Chelicerata) is strong using mitochondrial genome data when the Euarthropod tree is rooted using phylogenetically distant lophotrochozoans. (b) Support is equivocal using long branch but phylogenetically closer ecdysozoan nematodes (unresolved Euarthropoda). (c) The tree switches to supporting
a monophyletic Mandibulata (Myriapoda with Crustacea + Hexapoda) when using the phylogenetically close and short- branched priapulid as an outgroup (Rota-Stabelli and Telford,
2008).


expression patterns have disproved the protocer- ebral position of the chelifores, showing that they are indeed in the same deutocerebral position as chelicerae (Jager et al., 2006) and most molecu- lar data imply that the Chelicerata including Pycnogonida is a monophyletic group and that the contrary result was most likely derived from the rapid evolutionary rate of the Pycnogonida (Mallatt and Giribet, 2006).

ECD YS OZ OA N E VO LU T I O N 77




1.00/97
Arachnida


0.96/50



1.00/91
Xiphosaura

Pycnogonida

Chilopoda
1.00/100
Diplopoda


1.00/89
Crustacea



1.00/91 1.00/95

0.70/47

1.00/88
Thysanura


Apis


1.00/100
Anopheles









1.00/70
Drosophila

Priapulida


1.00/100


Nematomorpha





Clade III Nematoda

Caenorhabditis
Figure 8.2 Phylogeny of the Ecdysozoa. Bayesian analysis using small-subunit (SSU) and large-subunit (LSU) ribosomal RNA sequences, complete mitochondrial genomes, and eight nuclear protein-coding genes (vacuolar ATP- synthase, enolase, glyceraldehyde 3-phosphate dehydrogenase, carnitine palmitoyl transferase,
Na/K ATPase, RNA Pol II, dyskerin, and EF1-alpha). Some taxa with missing data have been merged into composite sequences. Support values are shown as Bayesian posterior probabilities and non-parametric

0.1 substitutions/site
bootstrap. Data are from Bourlat et al., (2008).




8.7 The position of the Hexapoda within the Pancrustacea

The support for the monophyly of Crustacea + Hexapoda, which came most emphatically from the evidence of a shared mitochondrial genome rearrangement, has been bolstered by numer- ous subsequent molecular phylogenetic analyses (Friedrich and Tautz, 1995; Hwang et al., 2001; Delsuc et al., 2003; Nardi et al., 2003; Regier et al.,
2005). Consideration of various aspects of morph- ology, in particular of nervous system ontogeny and structure, gives further weight to the integ- rity of this clade. Harzsch (2004) and Harzsch et al., (2005) list neuroblasts, two pairs of serotonergic neurons per hemineuromere, a fixed number of excitatory motor neurons per limb muscle, and aspects of lateral eye ultrastructure in support of the Crustacea + Hexapoda clade, which they
term Tetraconata. More controversial has been the placement of Hexapoda within Crustacea and the monophyly versus polyphyly of Hexapoda, with a number of studies separating the Collembola from the Insecta. While it is generally agreed that Crustacea is paraphyletic rather than being the sis- ter group of the Hexapoda (and that Hexapoda is in effect a terrestrial group of crustaceans), the closest crustacean sister group of the hexapods has been debated. Ignoring for the moment the little-studied Cephalocarida and Remipedia, there are two con- tenders among the main crustacean classes: the Malacostraca, that includes familiar species such as crabs and mantis shrimps, and Branchiopoda such as Artemia the brine shrimp and Daphnia the water flea. The Hexapoda–Malacostraca clade is supported by various features of brain anat- omy; specifically, members of these two groups share the presence of three brain neuropils joined

78 AN I M AL EV O L UTI O N



by chiasmata where other crustaceans have two
neuropils linked by parallel fibres (Harzsch,
2002). Analyses of complete mitochondrial gen-
ome sequences on the other hand support a mono-
phyletic Malacostraca and Branchiopoda clade
as a sister group to Hexapoda (Cook et al., 2005).
Most other molecular analyses, however, support
a sister-group relationship between Hexapoda
and Branchiopoda (Regier et al., 2005; Mallatt and
Giribet, 2006).
We have recently gathered all available data
from rRNAs, mitochondrial genomes, and various
nuclear protein-coding genes, and our analyses
support the close relationship between Hexapoda
and Branchiopoda (Economou, 2008). This relation-
ship is of great interest to the many workers inter-
ested in the evolution of the insects as it shows that
Daphnia, a crustacean with a completely sequenced
nuclear genome, is a relatively close sister group
of insects. Our analyses also include data from
Cephalocarida and Remipedia, and the placement
of these two groups is less certain. Both taxa are
atypical in terms of numbers of substitutions.
While the remipedes consistently groups close to
the hexapods, the position of the cephalocarids
is very unstable (Economou, 2008). Although the
relationships within the Hexapoda are beyond
the scope of this discussion, the controversy over
the placement of the Collembola is worth men-
tioning. While rRNA and nuclear protein-coding
gene phylogenies recover the expected monophyly
of the hexapods (Insecta, Diplura, Protura, and
Collembola), analyses using complete mitochon-
drial genomes recover a diphyletic Hexapoda with
the Insecta separated from the Collembola (Nardi
et al., 2003); mitochondrial sequences for Diplura
and Protura were not available. The basis of this
result has been questioned by subsequent authors,
and one must conclude that although monophyly of
Hexapoda ultimately seems the most likely result,
this needs to be tested with larger data sets.


8.8 Conclusion

In Figure 8.3 we summarize our best current esti- mate of ecdysozoan phylogeny. The first thing that is obvious from this tree and from the pre- ceding discussion is that while it seems clear that
Ecdysozoa is a monophyletic group, the relation- ships between phyla and major classes within the clade are often uncertain. While the pattern of rela- tionships of the Ecdysozoa has its own great intrin- sic interest, the phylogeny should also be viewed as the basis for a further understanding of the evo- lution of the Ecdysozoa. The mapping of charac- ter states onto a phylogeny allows us to go beyond the relationships of organisms to the evolution of characters and ultimately a fuller understanding of the process of evolution. The characteristics of the common ancestor of Ecdysozoa is of particu- lar interest, and it can be safely assumed to have possessed the synapomorphies of the group; Budd (2001a) has tentatively reconstructed the common ancestor as a large worm-like form with a terminal mouth, and to these characteristics we can add the shared characters discussed previously. The monophyly versus paraphyly of the Cycloneuralia becomes important now as, if paraphyletic, then their common ancestor becomes synonymous with the ecdysozoan ancestor, and suggests that it also possessed a cycloneuralian brain (not unreasonable considering the similar situation seen in onycho- phorans; Eriksson et al., 2003), and an introvert.
More controversial is the possibility that the ecdysozoan ancestor was segmented. While the kinorhynch metameres are generally referred to as zonites rather than segments, this seems a rather pointless distinction and is one indication that seg- mentation may be primitive in the group (Müller and Schmidt-Rhaesa, 2003; Schmidt-Rhaesa and Rothe, 2006). The similar deployment of homolo- gous genes (‘segment polarity’ or ‘pair rule’ genes) in arthropods and kinorhynchs would be a more direct indication of homology, and hence com- mon ancestry of segmentation within the group, as would the demonstration that arthropod seg- mentation can be convincingly homologized with that of annelids (Prud’homme et al., 2003) or even vertebrates (Damen, 2007).
Through comparison of the completely sequenced genomes of D. melanogaster and C. elegans, there is also the theoretical possibility of learning some- thing about the genome of the ecdysozoan com- mon ancestor, or perhaps something close to it depending on the position of the Arthropoda/ Nematoida common ancestor. One significant con-

ECD YS OZ OA N E VO LU T I O N 79


Deuterostomia Lophotrochozoa Nematoda
Nematomorpha Loricifera Priapulida Kinorhyncha
Onychophora

Tardigrada

Chelicerata


Pycnogonida Myriapoda Crustacea Hexapoda

Figure 8.3 The phylogeny of the Ecdysozoa espoused in this chapter. Names of probable monophyletic groups are given for each box. Unresolved portions of the tree are shown as multifurcations. We have shown Cycloneuralia, Mandibulata (including Myriapoda), and Panarthropoda (including Tardigrada) as monophyletic groups, despite some uncertainty, as we feel the morphological evidence is particularly convincing for these clades.




clusion from comparative genomics to date has been the secondary loss of large numbers of genes in the two model ecdysozoans (Copley et al., 2004; Putnam et al., 2007). The problem, of course, is that the two model species appear to have very derived genomes making comparisons particularly difficult to interpret—are they different from other animals due to common ecdysozoan gene losses or through convergent gene losses in these two derived mod- els? The ecdysozoan genome projects ongoing or recently announced, in particular that of the pri- apulid Priapulus caudatus, are very exciting for the
purpose of reconstructing the ancestral ecdyso- zoan genome, and should also add further to our understanding of the evolutionary relationships of this huge, diverse, and fascinating group.


8.9 Acknowledgements

Research in the laboratory was supported by the BBSRC and by the Marie Curie RTN ZOONET (MRTN-CT-2004–005624). We are very grateful for careful reviews by Andreas Schmidt-Rhaesa and Davide Pisani.