Integrative and Comparative Biology Advance Access originally published online on May 3, 2006
Integrative and Comparative Biology 2006 46(3):255-268; doi:10.1093/icb/icj035
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Coloniality has evolved once in Stolidobranch Ascidians
* Department of Biology and Center for Developmental Biology, University of Washington Box 351800, Seattle, WA 98195-1800, USA
Friday Harbor Laboratories, University of Washington 620 University Road, Friday Harbor, WA 98250-9299, USA
Correspondence: 1E-mail: bjswalla{at}u.washington.edu
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Ascidians exhibit a rich array of body plans and life history strategies. Colonial species typically consist of zooids embedded in a common test and brood large, fully developed larvae, while solitary species live singly and usually free-spawn eggs that develop into small, undifferentiated larvae. Ascidians in the order Stolidobranchia include both colonial and solitary species, as well as several species with intermediate morphologies. These include social species, which are colonial but do not live completely embedded in a common test, and a few solitary species that brood embryos and larvae until they are competent to metamorphose. We examined how many times coloniality has evolved within the Stolidobranchia, with phylogenetic analyses using full-length 18S rDNA and partial cytochrome oxidase B sequences for taxa in the families Molgulidae, Styelidae, and Pyuridae. Tunicata orders Phlebobranchia and Stolidobranchia are sister groups, and the family Molgulidae is a monophyletic group and should be raised to the subordinal level, as shown previously by analyses from this lab with partial 18S sequences. In contrast to previous studies, styelids and pyurids are separated into monophyletic groups by ML and Bayesian analyses. We show a single clade within the family Styelidae that contains two colonial (compound) botryllid species, a Symplegma (colonial compound), a colonial (social) species Metandrocarpa taylori, as well as four solitary species, thus confirming that the botryllids are a subfamily of the Styelidae. These results suggest that the ancestor of the Stolidobranchia was solitary and that coloniality has evolved only once within this clade of ascidians. Further phylogenetic analyses of aplousobranch and phlebobranch ascidians will be necessary to understand the number of times that coloniality has evolved within the class Ascidiacea.
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Deuterostomes are a monophyletic group of animals (Cameron and others 2000
) and can be further subdivided into two clades (Fig. 1I and II). One clade includes the phyla Echinodermata, Hemichordata, and Xenoturbellida, while the other consists of the chordates (Cameron and others 2000; Peterson and Eernisse 2001; Bourlat and others 2003; Smith and others 2004; Blair and Hedges 2005; Zeng and Swalla 2005). Phylum Chordata was originally divided into three subphyla: the Tunicata, Cephalochordata, and Vertebrata, which share five morphological characters (Rychel and others 2006). However, it has been suggested that the Tunicata should be raised to the phylum level (Kozloff 1990; Cameron and others 2000; Zeng and Swalla 2005).
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Many phylogenetic and developmental studies suggest that Cephalochordata and Vertebrata are sister groups, more closely related to each other than either is to the Tunicata (Turbeville and others 1994
; Wada and Satoh 1994; Cameron and others 2000; Winchell and others 2002). However, recent genome analyses suggest that tunicates may be more closely related to vertebrates than cephalochordates (Blair and Hedges 2005; Philippe and others 2005; Delsuc and others 2006). Also, tunicates possess neural crest cells (Jeffery and others 2004) and placodes (Manni and others 2004; Bassham and Postlethwait 2005; Mazet and others 2005) that are lacking in cephalochordates. Tunicates typically have long branch-lengths, which confound phylogenetic analyses and create artifacts (Blair and Hedges 2005; Zeng and Swalla 2005; Delsuc and others, 2006). In summary, the placement of the tunicates within deuterostomes has been problematic (Winchell and others 2002; Blair and Hedges 2005; Zeng and Swalla 2005; Delsuc and others 2006), even though studies have shown that tunicates are monophyletic (Swalla and others 2000; Stach and Turbeville 2002; Winchell and others 2002).
Ascidians, or sea squirts, are members of the class Ascidiacea, within Tunicata, that exhibit diverse life history strategies (Satoh 1994; Burighel and Cloney 1997; Davidson and others 2004). Ascidian tadpoles have key chordate characteristics such as a notochord and a dorsal hollow nerve cord (Swalla 2004a, 2004b; Fig. 2A), but these traits are lost after metamorphosis. Adult ascidians may be solitary and sexual or colonial and alternating between sexual and asexual reproduction by budding (Fig. 2) (Berrill 1935, 1936; Nakauchi 1982; Burighel and Cloney 1997). Colonial ascidians (Fig. 2B and C) tend to be ovoviviparous, producing large eggs and releasing adultated larvae that stay in the water column for only a short period of time before settling and initiating metamorphosis into the adult form (Fig. 2A) (Berrill 1935, 1936; Jeffery and Swalla 1992; Burighel and Cloney 1997; Davidson and others 2004). Solitary ascidians (Fig. 2D–G) either release large numbers of relatively small eggs into the water column, where fertilization and subsequent development into tadpole larvae takes place, or brood large, highly differentiated larvae (Berrill 1935).
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Ascidians were originally divided into colonial and solitary species by taxonomists, but in the early part of the 20th century classification based on branchial sac and gonad morphology became universally accepted (Van Name 1945
; Berrill 1950; Nishikawa 1990; Kott 1998; Monniot F and Monniot C 2001; Monniot and others 2001; Lambert 2005). Recently, phylogenies based on DNA sequences have helped to clarify some evolutionary relationships among the tunicates, although most phylogenies are entirely consistent with the taxonomic relationships (Wada and others 1992; Hadfield and others 1995; Wada 1998; Cameron and others 2000; Swalla and others 2000; Stach and Turbeville 2002; Winchell and others 2002; Turon and López-Legentil 2004). Wada and colleagues (1992) examined the evolution of coloniality and concluded that coloniality has evolved several times within class Ascidiacea, but did not identify particular clades within orders.
The ascidian order Stolidobranchia contains three widely acknowledged families: Molgulidae, Pyuridae, and Styelidae. Recent studies have shown that solitary molgulids are a monophyletic group, but styelids and solitary pyurids have remained unresolved by previous analyses (Wada and others 1992; Swalla and others 2000; Stach and Turbeville 2002). Colonial species are currently distributed among several taxonomically separate groups within the family Styelidae (Berrill 1950; Kott 1985). Many Tunicata taxonomists presently include the colonial (compound) botryllids as a subfamily within styelids (Kott 1998; Monniot F and Monniot C 2001; Monniot and others 2001; Saito and others 2001), while Nishikawa (1990, 1995) considers Botryllidae a separate family, partially because of their coloniality.
The diversity of life histories and morphologies among the ascidians make them an excellent group in which to examine life history evolution. For example, some species within the families Molgulidae and Styelidae bypass the tadpole stage altogether and exhibit direct (anural) development (Jeffery and Swalla 1992; Hadfield and others 1995; Huber and others 2000). When the developmental mode is mapped onto a phylogeny, then it is clear that direct (anural) development has evolved more than once within the Molgulidae, suggesting the possibility that anural development may be mediated by a conserved switch that can be activated with relative ease in evolutionary time (Hadfield and others 1995; Swalla and Jeffery 1996; Huber and others 2000).
Kott (1989) has suggested an additional family within the Stolidobranchia of deep-sea tunicates, the Hexacrobylidae, while the Monniots have created a separate class, Sorberacea, for them (Monniot and others 1975; Monniot C and Monniot F 1990). Unfortunately, we have not been able to obtain any Sorberaceans for phylogenetic studies, but the described species are solitary and morphologically similar to the Molgulidae (Monniot and others 1975; Kott 1989; Monniot C and Monniot F 1990).
The family Styelidae is particularly interesting because it contains both colonial and solitary species as well as a number of species with intermediate morphologies (Figure 2, Table 1, Van Name 1945). For example, Metandrocarpa taylori and other social colonial species reproduce clonally, brood large larvae, and have reduced adult body size similar to other colonial species, but zooids are connected only by stolons rather than being completely embedded together in a common test (Fig. 2C; Van Name 1945). Styelidae also contain Dendrodoa grossularia, which does not reproduce clonally but does brood very large larvae and grows in large dense clusters that superficially resemble social colonies (Van Name 1945); however, the individuals in these clusters are not necessarily close relatives (Bishop and Ryland 1993). The styelid species Polycarpa pomaria has also been observed to brood larvae under laboratory conditions, although the larvae are small and it is unclear whether this normally occurs in nature (Berrill 1950; Svane and Young 1989). It is intriguing that the evolution of brooding and large offspring size may be related to the evolution of coloniality, but to date there has been no phylogenetic evidence for this hypothesis.
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Examination of relationships within botryllids, which are all colonial, are interesting in light of understanding allorecognition (Cohen and others 1998
) and population dynamics (Stoner and others 1997; Stoner and others 2002) as well as higher taxonomic level phylogenies. Our study uses complete 18S rDNA sequences and the partial mitochondrial cytochrome oxidase B (cob) gene for twenty-eight species in thirteen genera to construct a phylogeny of the Stolidobranchia (Table 1). We use this phylogeny to test the hypotheses that coloniality has evolved multiple times in the Stolidobranchia. We show that two different botryllid species and one Symplegma group closely together in phylogenetic analyses, suggesting that these colonial compound species are monophyletic. We show that coloniality is likely to have evolved only once within the Stolidobranchia because the colonial social species M. taylori falls as a sister group to the colonial compound clade. |
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Biological materials, DNA isolation, and DNA sequencing
M. taylori was collected on Tatoosh Island, WA or by dredging near San Juan Island and maintained in a tank of recirculating or running seawater. Boltenia villosa, Botrylloides violaceus, and Corella inflata were collected off the docks at Roche Harbor, WA, on San Juan Island. Styela gibbsii and Cnemidocarpa finmarkiensis were collected from the docks at Friday Harbor Laboratories in Friday Harbor, WA. Botryllus schlosseri was collected from the docks of Shilshole Marina in Seattle, WA. Dendrodoa grossularia was collected from rocks at Roscoff, France. Botryllus planus, Symplegma viride, and Polycarpa papillata were collected by BJS in Puerto Rico, while teaching an Evolution and Development course at University of Puerto Rico. For colonial ascidians, individual zooids were dissected by hand, taking care to discard parasites and food items in the colony. Solitary ascidians were dissected free of their tunics with similar care, and either gonad or mantle (in non-gravid individuals) was dissected out, macerated, and used for extraction. Genomic DNA was isolated according to Hadfield and colleagues (1995)
and amplified and purified according to Swalla and colleagues (2000). Primers used to amplify mitochondrial cob gene sequences from genomic DNA were CobF 5'-TGR GGN CAR ATG WSN TTY TG-3' and CobR 5'-GGR AAN ARR AAR TAY CAY TC-3' (Turon and others 2003). GenBank accession numbers for the mitochondrial cytochrome oxidase B sequences are DQ345907–DQ34592. Ciona intestinalis (GenBank accession no. NC004447; Gissi and others 2004), Ciona savignyi (GenBank accession no. NC004570; Yokobori and others 2003), and Doliolum nationalis (NC006627; Yokobori and others 2005) sequences were taken from the entire mitochondrial sequence. Sequencing was performed at the Biochemistry Sequencing Facility and the Biology Comparative Genomics Facility at the University of Washington.
Sequences, alignments, and phylogenetic analyses
The 18S rDNA sequences used for this study were mostly sequenced in our lab, but we also included a few additional species from GenBank (Table 1). Alignment of the ascidian 18S rDNAs was performed using Clustal W (Thompson and others 1994). There will be few gaps in the 18S rDNA alignment for tunicates if aplousobranch sequences are not included. Mitochondrial cytochrome oxidase B sequences were translated with an ascidian mitochondrial code in MacVector, and then aligned with Clustal W. Protein alignments were used to accurately align the nucleotide sequences. One Appendicularian, Oikopleura dioica, was included in the analysis. Sites containing gaps were excluded from phylogenetic analyses to reduce systematic errors. Alignments were analyzed with PAUP*4.0b2 (Swofford 1999) to produce bootstrap maximum parsimony (MP) trees and neighbor-joining (NJ) trees (Saitou and Nei 1987). NJ trees were built using a Kimura two-parameter model in PAUP (Kimura 1980). We used 
= 0.50 for the 
distribution model. A Minimum Evolution (ME) tree was produced by heuristic searches in PAUP* under the same models of nucleotide substitution described above for NJ tree. Bootstrap maximum parsimony was calculated with PAUP*. We used the program MODELTEST 3.06 to find the best model and parameters to build the Maximum Likelihood (ML) trees. Confidence in NJ, ME, and MP trees were determined by analyzing 1000 bootstrap replicates and ML was determined by analyzing 100 bootstrap replicates (Felsenstein 1985). We built the Bayesian trees using MrBayes with 1 x 106 generation repeats with the nucleotide model 4by4, Nst = 6, rates =invgamma, Ngammacat = 4, and Burnin = 155 (Huelsenbeck and Ronquist 2001; Ronquist and Huelsenbeck 2003; Ronquist and others 2005). Tree reliability was also determined by comparing trees based on the same data, but produced with different tree-making algorithms (NJ, ME, MP, ML, and Bayesian).
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Results |
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We constructed expanded molecular phylogenies of the Styelidae and Pyuridae in order to examine the evolution and speciation of colonial and solitary species within the ascidian order Stolidobranchia. We included two colonial (compound) botryllids, (Botrylloides, Botryllus), one colonial (compound) Symplegma, the colonial (social) M. taylori, and several new solitary species in our analyses (Fig. 2; Table 1). A total of forty-one tunicate species were analyzed in the present study (Table 1). Twenty-eight taxa are in order Stolidobranchia, including eleven species from suborder or family Molgulidae, twelve species from family Styelidae (including three botryllids) and five species from family Pyuridae; nine taxa are in the order Phlebobranchia, and four are tunicates from outside of class Ascidiacea, including three species from class Thaliacea and one from class Appendicularia (Fenaux 1993
; Table 1). The complete alignment of 18S rDNA of forty-seven chordate species with thirty-seven ascidians and six outgroup taxa (five vertebrate species and one cephalochordate species) contained 
1810 sites including gaps. This alignment is available on-line at http://www.treebase.org/treebase/. Full-length 18S rDNA sequences were subjected to phylogenetic analysis using MP, NJ, and Bayesian algorithms. The MP, NJ, ME, ML, and Bayesian trees were very similar and are available on-line (Supplementary Data). Fifteen equally parsimonious MP trees were recovered by using heuristic searches and 1000 bootstrap replicates with PAUP. The NJ analysis, calculated with Kimura two-parameter evolutionary distances and 1000 bootstrap pseudoreplicates was completely congruent with the tree resulting from a ME analysis with 1000 bootstrap pseudoreplicates. Figure 3 shows the combined tree resulting from the MP analysis with bootstrap values from MP, NJ, and ME with branches drawn to scale.
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50%. The 1000 replicates bootstrap ME tree has the same topology as the NJ tree. The bootstrap values of MP, NJ, and ME are put at each node of the tree, separated by slashes. Ascidian species' life histories are marked as compound, social, and solitary with icons.In this MP, NJ, and ME combined tree, Stolidobranchia contains two distinct clades, the suborder Molgulidae and the families Styelidae +Pyuridae. Colonial compound (B. violaceus, B. planus, S. viride) and colonial social (M. taylori) species group together with the solitary brooding species D. grossularia, the solitary occasional brooder P. pomaria, solitary P. papillata, and the solitary free spawner C. finmarkiensis. The other clade in family Styelidae is made up entirely of solitary free-spawning species. The node separating the two clades within the Styelidae is supported by 99, 100 and 100% bootstrap values, indicating high support for this node. The clustering of compound and social species within one clade of the Styelidae suggests a single evolutionary origin of coloniality within the Stolidobranchia. In MP, NJ, and ME trees, Appendicularia always falls as a sister group to the rest of the tunicates, although with low bootstrap support (52%/62%/58%). The MP tree shown (Fig. 3) is consistent with the NJ and ME trees with only minor differences (supplementary materials). In MP analyses, family Pyuridae is paraphyletic, but the Styelidae are a monophyletic group with high bootstrap support (supplementary materials). In contrast, trees constructed with NJ and ME algorithms recover family Pyuridae as a monophyletic group, with 68 and 80% bootstrap support (supplementary materials, see also Zeng and Swalla 2005
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Figure 4 shows a combined tree of ML and Bayesian analysis. The ML tree had 100 bootstrap replicates (Fig. 4 and supplementary material) with a reduced dataset (complete alignment, but only twenty-eight taxa). The Bayesian tree had a full dataset 1 x 106 generation repeats (Fig. 4 and supplementary material) and was mostly congruent within the stolidobranchs. The interesting placement of taxa in the ML and Bayesian trees is the position of class Appendicularia, as shown in the combined tree in Fig. 4. In NJ, ME, and MP analyses (Fig. 3), the Appendicularia fall as a sister group to the rest of the Tunicata, as suggested by previous analyses (Swalla and others 2000; Wada 1998; Stach and Turbeville 2002). In contrast, the ML and Bayesian trees place the Appendicularia as a sister group to the suborder Stolidobranchia, rather than a sister group to the rest of the tunicates, with bootstrap support of 85% and a posterior probability value of 67% (Fig. 4 and supplementary material). However intriguing, these results should be interpreted with caution because of the long branches of the Appendicularia taxa (Swalla and others 2000).
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The 18S rDNA was not divergent enough to resolve relationships well within the styelid clade that contains the colonial species (Figs 3 and 4). We first attempted to clarify relationships within this clade by sequencing the hypervariable D-loop of 28S rDNA, but there was much less variability within Styelidae than previous studies had shown within Molgulidae (Hadfield and others 1995
; Huber and others 2000). We found 591 total characters within the 28S rDNA fragment, 145 characters were constant, 326 variable characters were parsimony-uninformative, and only 120 were parsimony informative, so the trees are not shown. Mitochondrial genomes evolve much faster than the nuclear genome, so we sequenced part of the mitochondrial protein-coding gene, cytochrome oxidase B (cob). The sequenced fragment of cob is only 420 bp long, but it has 287 parsimony informative sites, approximately three times as much variation as the 28S rDNA gene fragment. Figure 5 shows the phylogenetic trees of stolidobranch families Styelidae (twelve species), Pyuridae (four species), phlebobranch family Cionidae (two species), and a single member of class Thaliacea using NJ and MP algorithms. We also built DNA and protein trees using Bayesian algorithms, but the results had almost the same topology as NJ and MP trees shown in Figure 5. NJ tree shows the result of 1000 bootstrap replicates and the MP tree generated by 1000 bootstrap repeat with 1000 random replication in PAUP* (Fig. 5). In these trees, the styelid and botryllid clades have the same topology, and the trees differ only by whether the pyurids come out monophyletic (MP) or paraphyletic (NJ) (Fig. 5). M. taylori (colonial, social), B. violaceus (colonial, compound), B. planus (colonial, compound), B. schlosseri (colonial, compound), S. viride (colonial, compound), P. papillata (solitary), and C. finmarkiensis (solitary) group together, while the solitary P. pomaria and D. grossularia form a second clade. In summary, whether the pyurids are monophyletic or paraphyletic is unresolved by these analyses, but the mitochondrial phylogenetic trees show a single clade within the Stolidobranchs that contains all the colonial species, both compound and social. These data strongly support the single origin of coloniality in Stolidobranch ascidians.
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18S rRNA genes have become popular tools for phylogenetic inferences because they are relatively easy to isolate, ubiquitous, and not prone to lateral gene transfer (Olsen 1988
). Questions have been raised about the suitability of 18S data for reconstructing deep phylogenetic relationships because 18S rRNA exhibits variation in evolutionary rates, both between taxa and between sites on the molecule itself (Abouheif and others 1997). Within the Tunicata, however, a conserved 1 kb portion of the 18S rRNA resolves relationships between ascidian families much better than mitochondrial genes (Hadfield and others 1995; Wada 1998; Swalla and others 2000; Stach and Turbeville 2002). This is likely due to large genetic distances within the Tunicata that have not been previously appreciated (Swalla and others 2000; Zeng and Swalla 2005). Relative rate tests have shown that the suborder Molgulida and class Appendicularia (Fenaux 1993) are evolving significantly faster than other tunicate clades (Huber and others 2000), but both of these clades are morphologically and molecularly distinct monophyletic groups. Since neither of these groups was used as an outgroup, this variation in rates is unlikely to substantively affect our results.
This study used the entire 18S rDNA molecule and partial mitochondrial cytochrome oxidase B sequences to examine phylogenetic relationships within one tunicate order, the Stolidobranchia. Including the more variable 5' and 3' regions of the 18S rDNA molecule increased the resolution of the analyses significantly relative to previous analyses using only a conserved 1 kb portion (Hadfield and others 1995; Swalla and others 2000; Stach and Turbeville 2002), but the Tunicata 18S sequences are easy to align, as there are few gaps, insertions, or deletions (see TreeBase http://www.treebase.org/treebase/) (study accession number S1440; matrix accession numbers M2591 and M2592). Our phylogenies show stolidobranchs to be monophyletic, although we would very much like to include the benthic deep-sea Hexacrobylidae (Kott 1989; Monniot C and Monniot F 1990) if specimens were available to us. All member of this class described so far are solitary, so the inclusion of this group of tunicates would not significantly affect the conclusions of this article.
In all analyses presented here, colonial (compound and social) species group together with the solitary brooder D. grossularia, the possibly brooding P. pomaria, and the solitary non-brooders P. papillata and C. finmarkiensis. Therefore, coloniality evolved only once within the Stolidobranchia and the stolidobranch common ancestor was likely to be solitary. M. taylori, the morphologically ambiguous social species, may be in the process of becoming less or more integrated as a colony. M. taylori buds from oozooids, (Watanabe and Newberry 1976), similar to colonial ascidians (Nakauchi 1982), so it is likely that the process of asexual reproduction is conserved between these species. Similarly, large larval size and brooding in D. grossularia and possibly brooding in P. pomaria may represent transitional states en route to the loss or gain of coloniality. Further phylogenetic efforts should focus on gaining better resolution of the "colonial" clade within the Stolidobranchia.
The solitary Molgulidae are a single monophyletic group that should be raised from the familial to the subordinal level. We propose renaming it Molgulida. The other recognized families of the Stolidobranchia are Styelidae (which includes botryllids for most taxonomists) and Pyuridae. There is some conflict in the data concerning the monophyly of the Pyuridae, but each analysis recovered Pyuridae + Styelidae as a monophyletic group with the botyllids coming out as a monophyletic group within the Styelidae, thus confirming its status as a subfamily (Kott 1998; Monniot F and Monniot C 2001; Monniot and others 2001; Saito and others 2001). The clade containing Styela gibbsii, Styela montereyensis, Styela plicata, and Pelonaia corrugata was recovered in all analyses, suggesting that this is a closely related monophyletic group of solitary species. It is an interesting grouping because P. corrugata has tailless larvae while the rest of the species in the clade have tailed larvae (Hadfield and others 1995).
Coloniality and the ability to reproduce asexually may be strongly selected for in certain environments (Nakauchi 1982; Satoh 1994). However, only a few phyla within the invertebrates contain colonial species (Davidson and others 2004). Shifting between colonial and solitary lifestyles involves alteration of a whole suite of life history characteristics, and it is possible that developmental or morphological constraints make that transition difficult to accomplish (Davidson and others 2004). Groups such as Styelidae, which contain a range of species from solitary to compound colonial, are particularly exciting from the point of view of an evolutionary developmental biologist.
Aplousobranchia, an order within the ascidians, are all colonial but have been particularly problematic to place phylogenetically (Stach and Turbeville 2002; Winchell and others 2002; Turon and López-Legentil 2004; Zeng and Swalla 2005). These species have long branches, and the zooids are small, leaving them prone to contamination artifacts (Stach and Turbeville 2002) as discussed in Yokobori and others (2006). There are three major hypotheses of tunicate evolution (Fig. 6). The first hypothesis (Fig. 6A) suggests that Appendicularia and Aplousobranchia are grouped together as a sister group of the rest of the tunicates, which is supported by Stach and Turbeville (2002). This relationship may be suspect because of the long-branch attraction of Appendicularia and Aplousobranchia 18S rDNA sequences. The second view (Fig. 6B) is supported by mitochondrial COI gene analysis (Turon and López-Legentil 2004) and suggests that Aplousobranchia is a sister group to Stolidobranchia, and then Phlebobranchia and Thaliacea are sister groups to them. Unfortunately, this analysis did not include any Appendicularia. A final hypothesis (Fig. 6C) shows Aplousobranchia grouped with Thaliacea and then grouped with Phlebobranchia, and collectively are a sister group to Stolidobranchia and Appendicularia. This hypothesis is partially supported by our analysis in the 18S rDNA Maximum Likelihood tree and Bayesian tree (Fig. 4), but the bootstrap and posterior probability values are not high enough (85 and 67%). These relationships are also supported by the recent trees published with Aplousobranchia 18S (Yokobori and others 2006). The position of Aplousobranchia and Appendicularia is still somewhat unresolved within the tunicates and needs further analysis. The placement of the colonial Aplousobranchia within the rest of the tunicates is critical to understand whether the tunicate ancestor was solitary or colonial (Zeng and Swalla 2005). When the entire Tunicate phylogeny is better resolved, detailed comparative analysis of development may begin to elucidate the critical steps in life history evolution that can lead to a switch between solitary and colonial lifestyles.
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Acknowledgements |
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This article is dedicated to the spirit of Dr Larry McEdward, the "Larval Marvel." Larry was a great colleague and a dear friend. A warm thanks to Dr Eduardo Rosa-Molinar and the Evo-Devo class of 2001 at the University of Puerto Rico for helping collect some of the species used in these analyses (BIOL 6999: Special Topics in Modern Biology; http://pisces.cnnet.clu.edu/erm-lab/). We would like to thank Dawn Vaughn, who contributed to this work during a rotation in the Swalla Lab in the fall of 2003, and J. Muse Davis, who helped with some of the initial sequencing. Bob Paine and Chris Harley are thanked for collecting samples of M. taylori on Tatoosh Island. Cory Bishop and Bryan Crawford are thanked for the beautiful underwater photos of adult ascidians shown in Figure 2B–E. We would like to thank Professor Ken Warheit, Professor Scott Edwards, and Chris Hess for their help in performing initial analyses in Molecular Evolution classes at the University of Washington. Dr Dennis Willows, former Director of Friday Harbor Laboratories, is thanked for his encouragement throughout the project. The FHL staff members, especially Scott Schwinge, Kathleen MacDanold, and Blanche Bybee, are thanked for lab space, housing, and supplies, respectively. Sequencing was performed in the Comparative Genomics Center in the Biology Department at the University of Washington, funded in part by Major Research Instrumentation Grant no. 2002236 from the Murdock Foundation. This work was supported by an International Graduate Fellowship to L.Z., an NSF graduate research fellowship to M.W.J., and by a Seaver Institute and UW Department of Biology grant to B.J.S. Conflict of interest: none declared.
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Footnotes |
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From the symposium "Complex Life-Histories of Marine Invertebrates" presented at the annual meeting of the Society for Integrative and Comparative Biology, January 4–8, 2005, San Diego, California.
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References |
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Abouheif, E, R Zardoya, A Meyer. 1997. Limitations of metazoan 18S rRNA sequence data: implications for reconstructing a phylogeny of the animal kingdom and inferring the reality of the Cambrian explosion. J Mol Evol 47:394–405.
Bassham, S and JH Postlethwait. 2005. The evolutionary history of placodes: a molecular genetic investigation of the larvacean urochordate Oikopleura dioica. Development 132:4259–72.
Berrill, NJ. 1935. Studies in tunicate development part III: differential retardation and acceleration. Philos Trans R Soc Lond B 225:256–326.
Berrill, NJ. 1936. Studies in tunicate development part V: The evolution and classification of ascidians. Phil Trans R Soc Lond B 226:43–70.
Berrill, NJ. 1950. The tunicata, with an account of the British species. London Ray Society.
Bishop, JDD and JS Ryland. 1993. Enzyme electrophoretic evidence for the prevalence of outcrossing in the hermaphroditic brooding ascidian Dendrodoa grossularia (Chordata, Urochordata). J Exp Mar Biol Ecol 168:149–65.[CrossRef]
Blair, JE and SB Hedges. 2005. Molecular phylogeny and divergence times of deuterostome animals. Mol Biol Evol 22:2275–84.
Bourlat, SJ, C Nielsen, AE Lockyer, DT Littlewood, MJ Telford. 2003. Xenoturbella is a deuterostome that eats mollusks. Nature 424:925–8.[CrossRef][Medline]
Burighel, P and RA Cloney. 1997. Urochordata: Ascidiacea. In Harrison, FW and EE Ruppert (Eds.). Microscopic anatomy of invertebrates 5: Hemichordata, Chaetognatha and the invertebrate Chordates NY Wiley-Liss pp. 221–347.
Cameron, C, J Garey, BJ Swalla. 2000. Evolution of the chordate body plan: new insights from Phylogenetic analyses of deuterostome phyla. Proc Natl Acad Sci USA 97:4469–74.
Cohen, CS, Y Saito, IL Weissman. 1998. Evolution of allorecognition in botryllid ascidians inferred from a molecular phylogeny. Evolution 52:746–56.[CrossRef][ISI]
Davidson, B, MW Jacobs, BJ Swalla. 2004. The individual as a module: Metazoan evolution and coloniality. In Schlosser, G and G Wagner (Eds.). Modularity in development and evolution Chicago, IL University of Chicago Press pp. 443–65.
Delsuc, F, H Brinkmann, D Chourrout, H Philippe. 2006. Tunicates and not cephalochordates are the closest living relatives of vertebrates. Nature 439:965–8.[CrossRef][Medline]
Felsenstein, J. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783–91.[CrossRef][ISI]
Fenaux, R. 1993. The classification of Appendicularia (Tunicata): History and current state. Mem Inst Oceanogr Monaco 17:123.
Gissi, C, F Iannelli, G Pesole. 2004. Complete mtDNA of Ciona intestinalis reveals extensive gene rearrangement and the presence of an atp8 and an extra trnM gene in ascidians. J Mol Evol 58:376–89.[CrossRef][ISI][Medline]
Hadfield, KA, BJ Swalla, WR Jeffery. 1995. Multiple origins of anural development in ascidians inferred from rDNA sequences. J Mol Evol 40:413–27.[CrossRef][ISI][Medline]
Huber, J, K Burke da Silva, W Bates, BJ Swalla. 2000. The evolution of anural larvae in molgulid ascidians. Cell Dev Biol 11:419–26.
Huelsenbeck, JP and F Ronquist. 2001. MRBAYES: Bayesian inference of phylogeny. Bioinformatics 17:754–5.
Jeffery, W and BJ Swalla. 1992. Evolution of alternate modes of development in ascidians. Bioessays 14:219–26.[CrossRef][ISI][Medline]
Jeffery, WR, AG Strickler, Y Yamamoto. 2004. Migratory neural crest-like cells form body pigmentation in a urochordate embryo. Nature 431:696–9.[CrossRef][Medline]
Kimura, M. 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–20.[CrossRef][ISI][Medline]
Kott, P. 1985. The Australian Ascidiacea: Part I, Phlebobranchia and Stolidobranchia. Mem Queensl Mus 23:1–440.
Kott, P. 1989. The family Hexacrobylidae Seeliger, 1906 (Ascidiacea, Tunicata). Mem Queensl Mus 27:517–34.
Kott, P. Hemichordata, Tunicata, Cephalochordata. In Wells, A and WWK Houston (Eds.). Zoological catalogue of AustraliaAustralia CSIRO Publishing pp. 51–252 259–61.
Kozloff, EN. Invertebrates 1990. Philadelphia Saunders College Publishing pp. 866.
Kurabayashi, A, M Okuyama, M Ogawa, A Takeuchi, Z Jing, T Naganuma, Y Saito. 2003. Phylogenetic position of a deep-sea ascidian, Megalodicopia hians, inferred from the molecular data. Zoolog Sci 20:1243–7.[CrossRef][ISI][Medline]
Lambert, CC. 2005. Historical introduction, overview, and reproductive biology of the protochordates. Can J Zool 83:1–7.[CrossRef]
Manni, L, NJ Lane, JS Joly, F Gasparini, S Tiozzo, F Caicci, G Zaniolo, P Burighel. 2004. Neurogenic and non-neurogenic placodes in ascidians. J Exp Zool B Mol Dev Evol 302:483–504.
Mazet, F, JA Hutt, J Milloz, J Millard, A Graham, SM Shimeld. 2005. Molecular evidence from Ciona intestinalis for the evolutionary origin of vertebrate sensory placodes. Dev Biol 282:494–508.[CrossRef][ISI][Medline]
Monniot, C and F Monniot. 1990. Revision of the class Sorberacea (benthic tunicates) with descriptions of seven new species. Zool J Linn Soc 99:239–90.
Monniot, F and C Monniot. 2001. Ascidians from the tropical western Pacific. Zoosystema 23:201–383.
Monniot, C, F Monniot, F Gaill. 1975. Les Sorberacea: une nouvelle classe de Tuniciers. Arch Zool Exp Gen 116:77–122.
Monniot, C, F Monniot, CL Griffiths, M Schleyer. 2001. South African ascidians. Ann S Afr Mus 108:1–141.
Nakauchi, M. 1982. Asexual development of ascidians: its biological significance, diversity, and morphogenesis. Am Zool 22:753–63.
Nishikawa, T. 1990. The ascidians of the Japan Sea. I. Publications of the Seto Marine. Biological Laboratory 34:73–148.
Nishikawa, T. 1995. Chapter Chordata (excluding vertebrates). In Nishimura, S (Ed.). Guide to seashore animals of Japan with color pictures and keys Osaka Japan: Hoikusha Publishing Company Ltd. pp. 573–610 (In Japanese).
Olsen, GJ. 1988. Phylogenetic analysis using ribosomal RNA. Methods Enzymol 164:793–812.[ISI][Medline]
Peterson, KJ and DJ Eernisse. 2001. Animal phylogeny and the ancestry of bilaterians: inferences from morphology and 18S rDNA gene sequences. Evol Dev 3:170–205.[CrossRef][ISI][Medline]
Philippe, H, N Lartillot, H Brinkmann. 2005. Multigene analyses of bilaterian animals corroborate the monophyly of ecdysozoa, lophotrochozoa, and protostomia. Mol Biol Evol 22:1246–53.
Ronquist, F and JP Huelsenbeck. 2003. MRBAYES 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19:1572–4.
Ronquist, F, JP Huelsenbeck, P van der Mark. 2005. MRBAYES 3.1 manual. (http://mrbayes.csit.fsu.edu/manual.php).
Rychel, AL, SE Smith, HT Shimamoto, BJ Swalla. 2006. Evolution and development of the chordates: Collagen and pharyngeal cartilage. Mol Biol Evol 23:541–9.
Saito, Y, M Shirae, M Okuyama, S Cohen. 2001. Phylogeny of botryllid ascidians. In Sawada, H, H Yokosawa, CC Lambert (Eds.). The biology of Ascidians Tokyo Springer-Verlag pp. 315–20.
Saitou, N and M Nei. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–25.[Abstract]
Satoh, N. Developmental Biology of Ascidians 1994. New York Cambridge University Press.
Smith, AB, KJ Peterson, G Wray, DTJ Littlewood. 2004. From bilateral symmetry to pentaradiality: The phylogeny of hemichordates and echinoderms. In Cracraft, J and MJ Donoghue (Eds.). Assembling the tree of life New York Oxford Press pp. 365–83.
Stach, T and JM Turbeville. 2002. Phylogeny of tunicata inferred from molecular and morphological characters. Mol Phylogenet Evol 25:408–28.[CrossRef][ISI][Medline]
Stock, DW and GS Whitt. 1992. Evidence from 18S ribosomal RNA sequences that lampreys and hagfishes form a natural group. Science 257:787–9.
Stoner, DS, JM Quattro, IL Weissman. 1997. Highly polymorphic microsatellite loci in the colonial ascidian Botryllus schlosseri. Mol Mar Biol Biotechnol 6:163–71.[ISI][Medline]
Stoner, DS, R Ben-Shlomo, B Rinkevich, IL Weissman. 2002. Genetic variability of Botryllus schlosseri invasions to the east and west coasts of the USA. Mar Ecol Prog Ser 243:93–100.
Svane, I and C Young. 1989. The ecology and behaviour of ascidian larvae. Oceanogr Mar Biol Annu Rev 27:45–90.
Swalla, BJ. 2004a. Procurement and culture of Ascidian embryos. In Ettenson, C, G Wessell, G Wray (Eds.). Methods in cell biology: experimental analysis of the development of sea urchins and other non-vertebrate Deuterostomes Elsevier Science/Academic Press pp. 115–41.
Swalla, BJ. 2004b. Protochordate Gastrulation: Lancelets and Ascidians. In Stern, C (Ed.). Gastrulation Cold Spring Harbor Cold Spring Harbor Press pp. 139–49.
Swalla, BJ and WR Jeffery. 1996. Requirement of the manx gene for expression of chordate features in a tailless ascidian larva. Science 274:1205–9.
Swalla, BJ, C Cameron, L Corley, J Garey. 2000. Urochordates are monophyletic within the deuterostomes. Syst Biol 49:52–64.[CrossRef][ISI][Medline]
Swofford, DL. PAUP*. Phylogenetic Analysis using parsimony (*and other methods). Version 4 1999. Sunderland, MA Sinauer Associates.
Thompson, JD, DG Higgins, TJ Gibson. 1994. Clustal W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–80.
Turbeville, JM, JR Schulz, RA Raff. 1994. Deuterostome phylogeny and the sister group of chordates: evidence from molecules and morphology. Mol Biol Evol 11:648–55.[Abstract]
Turon, X and S López-Legentil. 2004. Ascidian molecular phylogeny inferred from mtDNA data with emphasis on the Aplousobranchiata. Mol Phylogenet Evol 33:309–20.[CrossRef][ISI][Medline]
Turon, X, I Tarjuelo, S Duran, M Pascual. 2003. Characterising invasion processes with genetic data: an Atlantic clade of Clavelina lepadiformis (Ascidiacea) introduced into Mediterranean harbors. Hydrobiologia 503:29–35.[CrossRef]
Van Name, WG. 1945. The North and South American Ascidians. Bull Am Mus Nat Hist 84:1–476.
Wada, H and N Satoh. 1994. Details of the evolutionary history from invertebrates to vertebrates, as deduced from the sequences of 18S rDNA. Proc Natl Acad Sci USA 91:1801–4.
Wada, H. 1998. Evolutionary history of free swimming and sessile lifestyles in urochordates as deduced from 18S rDNA molecular phylogeny. Mol. Biol. Evol 15:1189–1194.[Abstract]
Wada, H, KW Makabe, M Nakauchi, N Satoh. 1992. Phylogenetic relationships between solitary and colonial ascidians, as inferred from the sequence of the central region of their respective 18S rDNAs. Biol Bull 183:448–55.[Abstract]
Watanabe, H and AT Newberry. 1976. Budding by oozoids in the polystyelid ascidian Metandrocarpa taylorii Huntsman. J Morphol 148:161–76.[CrossRef][ISI][Medline]
Winchell, CJ, J Sullivan, CB Cameron, BJ Swalla, J Mallatt. 2002. Evaluating hypotheses of deuterostome phylogeny and chordate evolution with new LSU and SSU ribosomal DNA data. Mol Biol Evol 19:762–76.
Yokobori, S, Y Watanabe, T Oshima. 2003. Mitochondrial genome of Ciona savignyi (Urochordata, Ascidiacea, Enterogona): Comparison of gene arrangement and tRNA genes with Halocynthia roretzi mitochondrial genome. J Mol Evol 57:574–87.[CrossRef][ISI][Medline]
Yokobori, S, T Oshima, H Wada. 2005. Complete nucleotide sequence of the mitochondrial genome of Doliolum nationalis with implications for evolution of urochordates. Mol Phylogenet Evol 34:273–83.[CrossRef][ISI][Medline]
Yokobori, S, A Kurabayashib, BA Neilanc, T Maruyamad, E Hirose. 2006. Multiple origins of the ascidian-Prochloron symbiosis: Molecular phylogeny of photosymbiotic and non-symbiotic colonial ascidians inferred from 18S rDNA sequences. Mol Phylogenet Evol (In Press).
Zeng, L and BJ Swalla. 2005. Molecular phylogeny of the protochordates: Chordate evolution. Can J Zool 83:24–33.[CrossRef]
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