Sipunculan Larvae and ‘‘Cosmopolitan’’ Species
Anja Schulze,1,* Anastassya Maiorova,†,‡Laura E. Timm* and Mary E. Rice§
*Department of Marine Biology, Texas A&M University at Galveston, P.O. Box 1675, Galveston, TX 77553, USA;
†Laboratory of Productive Biology, A.V. Zhirmunsky Institute of Marine Biology, Far East Branch of Russian Academy of
Sciences, Vladivostok 690059, Russia;‡Interdepartmental Laboratory ‘‘Biology of Marine Invertebrates,’’ Far Eastern
Federal University, Vladivostok 690091, Russia;§Smithsonian Marine Station at Fort Pierce, 701 Seaway Drive,
Fort Pierce, FL 34949, USA
From the symposium ‘‘Dispersal of Marine Organisms’’ presented at the annual meeting of the Society for Integrative and
Comparative Biology, January 3–7, 2012 at Charleston, South Carolina.
Synopsis Sipuncula is a relatively small taxon with roughly 150 recognized species. Many species are geographically
widespread or ‘‘cosmopolitan.’’ The pelagosphera larvae of some species are estimated to spend several months in the
plankton. However, recent molecular evidence suggests that many of the ‘‘cosmopolitan’’ species actually represent
species-complexes, some not even monophyletic. Herein, we present data on three sipunculan species with different
developmental modes that occur both in the Sea of Japan and in the Northeast Pacific. The development of the three
species—Phascolosoma agassizii, Thysanocardia nigra, and Themiste pyroides—is exceptionally well studied in both regions
of the Pacific. Significant differences have been observed between the two regions with respect to egg size, developmental
mode, and developmental timing. In general, eggs are larger and development slower in the Northeast Pacific when
compared with the Sea of Japan. These differences have been explained as a result of phenotypic plasticity exhibited
under different environmental conditions, in particular temperature, but we show that the populations of all three species
are also remarkably distinct genetically and that gene flow between the two regions is extremely unlikely.
In Thysanocardia nigra, we even found two very distinct genetic lineages within the same location in the Northeast
Pacific. The amount of genetic divergence between populations from the Sea of Japan and those from the Northeast
Pacific is not correlated with developmental mode. Themiste pyroides, the species with the most abbreviated development,
actually has the least degree of genetic divergence between the regions. Analyses of molecular variance show that the
majority of the observed variation in all three species is between the regions. We conclude that all three ‘‘cosmopolitan’’
species actually represent complexes of cryptic or pseudo-cryptic species. These examples demonstrate that a solid
taxonomic framework based on molecular and morphological evidence is a prerequisite for evaluating relationships
between dispersal capabilities, species’ ranges, and the connectivity of populations.
species’ ranges, and population structure in marine
organisms is subject to ongoing discussion (e.g.,
Lester and Ruttenberg 2005; Hedgecock et al. 2007;
Shanks 2009; Weersing and Toonen 2009). In recent
years, novel approaches to studying dispersal, span-
ning genetics, isotope signatures, and biophysical
modeling have greatly increased our understanding
of the connectivity of marine populations (reviewed
However, the success of all these methods hinges
on a solid taxonomic framework for the taxa under
question. By definition, studies of connectivity
require some prior knowledge of species’ ranges,
and a shaky taxonomy can seriously violate the im-
plicit assumptions of these studies. Herein, we pre-
sent a few cases of the Sipuncula.
Sipuncula, also known as peanut worms or star
worms, are theoretically very suitable for studying
marine biogeography, phylogeography, and popula-
tion genetics because many species are reportedly
widespread and abundant enough to be collected in
sufficient quantities for studies at the population
Integrative and Comparative Biology
Integrative and Comparative Biology, volume 52, number 4, pp. 497–510
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level. On the other hand, sipunculans are notoriously
difficult to identify as they have a relatively sim-
ple body plan that is conserved throughout the
group, limiting the number of taxonomically useful
The body of an adult sipunculan is divided into an
unsegmented trunk and a retractable introvert with
an array of tentacles at its tip. Commonly used
externally visible taxonomic
arrangement and number of the tentacles, the
arrangement and structure of introvert hooks, and
the distribution and shape of epidermal papillae.
Size ratios, such as that of the length-of-introvert:-
length-of-trunk and the trunk’s length:width, are
usually cited but can be problematic if the animal
is contracted—which tends to be the case unless the
specimen is carefully relaxed before fixation. Many
species require dissection for proper identification.
Important internal characters are the number and
degree of fusion of the introvert retractor muscles;
their point of insertion in the body wall, the arrange-
ment of the musculature of the body wall, the shape
of the nephridia, and the presence or absence of
contractile vessel villi, among others.
Stephen and Edmonds published a first mono-
graph on sipunculans and echiurans in 1972 that
recognized 320 sipunculan species (Stephen and
Edmonds 1972). Over the next 2 decades, Cutler
and co-authors performed a series of generic revi-
sions as summarized by Cutler (1994) who reduced
the number of species to less than half that of
Stephen and Edmonds. Of Cutler’s roughly 150 spe-
cies, many are ‘‘cosmopolitan.’’ Note that the term
‘‘cosmopolitan’’ is loosely applied here for any spe-
cies that have ranges spanning at least the width of
an oceanic basin.
Why are so many sipunculan species so widespread?
Or are they? There are four alternative hypotheses that
would explain the reported distributions:
(1) Efficient long-distance dispersal: geographically
disparate populations are connected through
(2) Cryptic speciation: disparate populations repre-
sent distinct genetic lineages, but the morphol-
ogy is so conserved that they cannot easily be
distinguished morphologically (‘‘sibling species’’
sensu Mayr 1948);
(3) Taxonomic ‘‘lumping’’: disparate populations
are genetically and morphologically distinct, but
taxonomists judge the morphological differ-
ences not worthy of delineating species. The
result is a complex of pseudo-cryptic species
(or ‘‘pseudo-sibling species’’ sensu Knowlton
(4) Inadequate morphological information: disparate
populations are genetically and morphologically
distinct, but the morphological differences have
not been detected or documented yet. This
would be another case of pseudo-sibling species
but for different reasons than in 3.
Only the first hypothesis justifies the claim of true
cosmopolitanism. The other three imply that the
reported distributions are artifacts, and the real dis-
tributions are a lot smaller.
Support for the first hypothesis comes primarily
from the work of Scheltema and Hall (1975), who
studied the presence of sipunculan pelagosphera
larvae in plankton tows throughout the Atlantic
Ocean in the 1960s and 1970s. Based on known cur-
rent speeds in the North Atlantic Gyre and distance
to the closest possible shallow-water habitat from
which these larvae could have originated, Scheltema
and Hall estimated that some of the larvae caught in
the middle of the Atlantic were 3–4-months old.
They concluded that ‘‘zoogeographical evidence con-
firms the hypothesis that many tropical and warm-
temperate, shoal-water species of sipunculans are
widely dispersed. The mechanism for this dispersal
is the transport of pelagosphera larvae on ocean cur-
rents’’ (Scheltema & Hall 1975, p. 114). Other pela-
gosphera larvae had been kept in laboratory cultures
for up to 7 months without metamorphosing (Rice
1967), providing further support for the possibility
of long-distance dispersal.
On the other hand, many recent studies on the
relationship between the pelagic larval duration
(PLD) and the connectivity of populations in a va-
riety of other taxa have found that many species with
long PLDs show stronger population structure than
predicted on the basis of passive dispersal of the
larvae by oceanic currents. This holds true even if
more realistic Lagrangian models of larval transport
are applied (Knowlton and Keller 1986; Cowen et al.
2000; Shanks 2009).
Phylogenetic studies have provided some support
for the presence of cryptic or pseudo-cryptic species
complexes in sipunculans. The analyses by Maxmen
et al. (2003), Schulze et al. (2007), and Kawauchi
et al. (2012) included multiple individuals of some of
the ‘‘cosmopolitan’’ species from different geographic
regions. Commonly, each was a clearly distinct lineage,
and the species often did not appear as monophyletic.
because the same taxonomists identified the samples,
using the most current key (Cutler 1994).
A.Schulze et al.
by guest on December 23, 2015
species. A more extensive phylogenetic analysis of
Thysanocardia and related genera would clarify how
far the three species are separated in the phylogenetic
tree. The two phylogenetic trees of P. agassizii and
other congeners show that the two lineages are not
sister taxa and probably diverged relatively early in
the radiation of the genus, although neither COI nor
16S resolve the basal relationships among the
Phascolosoma species with high support. It is inter-
esting to note that the hook morphologies of those
species that fall between the northeastern Pacific and
Sea of Japan clades of P. agassizii in the tree (Fig. 3)
are quite distinct from that of P. agassizii. Evidently,
hook morphology either evolved convergently in the
two branches of P. agassizii or it represents an an-
cestral condition in the genus. The two populations
of T. pyroides, on the other hand, appear as mono-
phyletic to the exclusion of T. dyscrita, T. minor, and
T. lageniformis and probably diverged relatively
If the populations from Sea of Japan and from the
NE Pacific are separated at the species level, pheno-
typic plasticity in response to different environmental
conditions need not be invoked to explain the
differences in egg size, developmental mode, or
developmental timing between the two regions.
Phenotypic plasticity in development may still exist,
but this is a different issue that should be investi-
gated under carefully controlled conditions.
Having rejected true cosmopolitanism for all the
three species, the question still remains whether we
are dealing with cryptic or pseudo-cryptic species.
Our preliminary observations of the specimens did
reveal some differences between the two regions. In
P. agassizii, we observed that the populations from
the NE Pacific have dark bands on the introvert;
these are missing in populations from the Sea of
Japan. The populations of T. nigra seem to differ
with respect to insertion of the introvert retractor
muscles in the body wall, and differences in the ar-
rangement of the tentacles were observed in T. pyr-
oides. More detailed morphological studies will
determine how well the morphological characteristics
whether the lineages represent species that are new
These examples of cryptic or pseudo-cryptic spe-
cies in sipunculans demonstrate that a combination
of approaches is required to properly evaluate the
relationship between dispersal capabilities, species’
ranges, and population connectivity. A solid taxo-
nomic framework is necessary for interpreting popu-
lation-level genetic data, but genetic data are also
crucial for establishing a taxonomic framework.
Data on developmental timing are necessary to
form initial hypotheses about the connectivity of
populations but without a proper understanding of
regional variation they can also be misleading.
‘‘Dispersal of Marine Species.’’ We are also grateful
to Gustav Paulay (Florida Museum of Natural
History), Louise Page (University of Victoria), and
the captain of the R/V Centennial for contributing to
sample collection. This publication is Smithsonian
Marine Station Contribution #877. Comments from
two anonymous reviewers greatly improved the qual-
ity of the manuscript.
thankthe organizersofthe symposium
Our participation in the symposium was supported
by the American Microscopical Society, Society for
Integrative and Comparative Biology Divisions of
Evolutionary Developmental Biology, Ecology &
National Science Foundation Grant IOS-1148884 to
Sara M. Lindsay. The research was funded by a col-
laborative grant from the Far East Branch of the
Russian Academy of Science and Civilian Research
and Development Foundation (CRDF Global) to
A.S. and A.M. (RUB1-2996VL-11), by National
Science Foundation Assembling the Tree of Life
grant DEB-1036186 to A.S. Further funding was pro-
vided by the Far East Branch of the Russian
Academy of Sciences (grants 12-HHC-007, 12-I-0-
10132-? and Ministry of Education and Science of
Russian Federation (grant 11.G34.31.0010).
Adrianov A, Maiorova A. 2010. Reproduction and develop-
ment of common species of peanut worms (Sipuncula)
from the Sea of Japan. Russian J Mar Biol 36:1–15.
Carr CM,HardySM, Brown
Hebert PDN. 2011. A tri-oceanic perspective: DNA barcod-
ing reveals geographic structure and cryptic diversity in
Canadian polychaetes. PLoS One 6:e22232.
Cowen RK, Lwiza KMM, Sponaugle S, Paris CB, Olson DB.
2000. Connectivity of marine populations: open or closed?
Cowen RK, Sponaugle S. 2009. Larval dispersal and marine
population connectivity. Ann Rev Mar Sci 1:443–66.
Cutler EB. 1994. The Sipuncula. Their systematics, biology
and evolution. Ithaca, NY: Cornell University Press.
Cutler EB, Cutler NJ, Gibbs PE. 1983. A reconsideration of
(Sipuncula). Proc Biol Soc Wash 96:669–94.
Golfingiella and Siphonoides
by guest on December 23, 2015
Du X, Chen Z, Deng Y, Wang Q, Huang R. 2008. Genetic
diversity and population structure of the peanut worm
(Sipunculus nudus) in Southern China as inferred from mi-
tochondrial 16S rRNA sequences. Isr J Aquacul-Bamid
Excoffier L, Lischer HEL. 2010. Arlequin suite ver 3.5: a
new series of programs to perform population genetics
analyses under Linux and Windows. Mol Ecol Res
Excoffier L, Smouse PE, Quattro JM. 1992. Analysis of molec-
ular variance inferred from metric distances among DNA
haplotypes: application to human mitochondrial DNA
restriction data. Genetics 131:479–91.
Folmer O, Black M, Hoeh W, Lutz R, Vrijenhoek R. 1994.
DNA primers for amplification of mitochondrial cyto-
chrome c oxidase subunit I from diverse metazoan inverte-
brates. Mol Mar Bio Biotechnol 3:294–5.
Hebert PDN, Ratnasingham S, DeWaard JR. 2003. Barcoding
animal life: cytochrome c oxidase subunit 1 divergences
among closely related species. Proc R Soc B Biol Sci
Hedgecock D, Barber PH, Edmands S. 2007. Genetic
Kawauchi GY, Giribet G. 2010. Are there true cosmopolitan
sipunculan worms? A genetic variation study within
Phascolosoma perlucens. Mar Biol 157:1417–31.
Kawauchi GY, Sharma PP, Giribet G. 2012. Sipunculan phy-
logeny based on six genes, with a new classification and
description of two new families. Zool Scripta 41:186–210.
Keferstein W. 1867. Untersuchungen uber einige amerika-
nische Sipunculiden. Z wiss Zool 17:44–55.
Kitamura A, Takano O, Takata H, Omote H. 2001. Late
Pliocene–early Pleistocene paleoceanographic evolution of
the Sea of Japan. Palaeogeogr Palaeoclimatol Palaeoecol
Knowlton N. 1993. Sibling species in the sea. Ann Rev Ecol
Knowlton N, Keller BD. 1986. Larvae which fall far short of
their potential: highly localized recruitment in an alpheid
shrimp with extended larval development. Bull Mar Sci
Lester SE, Ruttenberg BI. 2005. The relationship between pe-
lagic larval duration and range size in tropical reef fishes: a
synthetic analysis. Proc R Soc B Biol Sci 272:585–91.
Levin LA. 2006. Recent progress in understanding larval dis-
persal: new directions and digressions. Int Comp Biol
MaxmenAB, KingBF, Cutler
Evolutionary relationships within the protostome phylum
Sipuncula; a molecular analysis of ribosomal genes and his-
tone H3 sequence data. Mol Phylogenet Evol 27:489–503.
Mayr E. 1948. The bearing of the new systematics on genetic
problems. Adv Genet 2:205–37.
Palumbi SR. 1996. Nucleic acids II: the polymerase chain
reaction. In: Hillis DM, Moritz C, Mable BK, editors.
Molecular systematics. Sunderland: Sinauer.
Rambaut A. 2006–2009. FigTree v.1.3.1. Tree figure drawing
tool. University of Edinburgh, Edinburgh, UK: Institute of
Rice ME. 1967. A comparative study of the development of
P. agassizii, Golfingia pugettensis, and Themiste pyroides
with a discussion of developmental patterns in the
Sipuncula. Ophelia 4:143–71.
Rice ME. 1976. Larval development and metamorphosis in
Sipuncula. Amer Zool 16:563–71.
Rice ME. 1978. Morphological and behavioral changes at
metamorphosis in the Sipuncula. In: Rice ME, Chia FS,
editors. Settlement and metamorphosis of marine inverte-
brate larvae. New York: Elsevier North-Holland Biomedical.
Scheltema RS, Hall JR. 1975. The dispersal of pelagosphera
larvae by ocean currents and the geographical distribution
of sipunculans. In: Rice ME, Todorovic M, editors.
Proceedings of the International Symposium on the biology
of the Sipuncula and Echiura. Kotor: Naucno Delo.
Schulze A, Cutler EB, Giribet G. 2007. Phylogeny of sipuncu-
lan worms: a combined analysis of four gene regions and
morphology. Mol Phylogenet Evol 42:171–92.
Shanks AL. 2009. Pelagic larval duration and dispersal
distance revisited. Biol Bull 216:373–85.
Staton J, Rice ME. 1999. Genetic differentiation despite tele-
planic larval dispersal: allozyme variation in sipunculans of
the Apionsoma misakianum species complex. Bull Mar Sci
Stephen AC, Edmonds SJ. 1972. The phyla Sipuncula and
Echiura. London: Trustees Brit Mus Nat Hist.
Tamura K, Nei M, Kumar S. 2004. Prospects for inferring
very large phylogenies by using the neighbor-joining
method. Proc Natl Acad Sci USA 101:11030–5.
Tamura K, Peterson D, Peterson N, Stecher G, Nei M,
Kumar S. 2011. MEGA5: molecular evolutionary genetics
analysis using maximum likelihood, evolutionary distance,
and maximum parsimony
Weersing K, Toonen RJ. 2009. Population genetics, larval dis-
persal, and connectivity in marine systems. MEPS 393:1–12.
Zhang YP, Ryder OA. 1993. Mitochondrial DNA evolution in
the Artoidea. Proc Nat Acad of Sci USA 90:9557–61.
methods.Mol Biol Evol
A.Schulze et al.
by guest on December 23, 2015