Phylogeny of the Gnathostomulida Inferred from a Combined
Approach of Four Molecular Loci and Morphology
Martin V. Sørensen1, Wolfgang Sterrer2 and Gonzalo Giribet1
1
Department of Organismic and Evolutionary Biology, Harvard University,
Cambridge, MA, USA
2Bermuda
Natural History Museum, Flatts FLBX, Bermuda
Corresponding author: Martin V. Sørensen. Current address: Ancient DNA and
Evolution Group, Department of Geophysics, University of Copenhagen, Juliane
Maries Vej 30, 2100 Copenhagen, Denmark. Email: mvsorensen@bi.ku.dk
Morphological characters
General body plan
1) Animals long and vermiform, body index > 25: 0=absent, 1=present.
The filospermoid taxa generally have a much higher body index than the
bursovaginoids (Fig. 1). The body index is defined as the body length to
maximum body width ratio (Sterrer, 1972).
2) Rostrum long and pointed, rostrum index > 3: 0=absent, 1=present.
The filospermoid rostrum (the preoral part of the head) is likewise longer and
more pointed, compared to the bursovaginoids (Fig. 1). The rostrum index is
defined as the rostrum length to maximum rostrum width ratio (Sterrer, 1972).
3) Body divided into head, trunk and foot: 0=absent, 1=present.
This character refers to the general rotifer body plan, here used as outgroups.
4) Animals with sessile feeding stages with ciliated buccal funnel:
0=absent, 1=present.
1
This character refers to the cycliophoran feeding stages (Funch and Kristensen,
1997) for one of the outgroup taxa.
5) Triradial pharynx: 0=absent, 1=present.
Gastrotrichs have a characteristic triradial pharynx not observed in any of the
other taxa represented in our matrix.
Epidermis
6) Ciliation of epidermis: 0=monociliated cells, 1=multiciliated cells.
The character refers to the number of cilia on the epidermal cells. All
gnathostomulids have exclusively monociliated epidermal cells, whereas most of
the outgroup taxa have multiciliated epidermis. The gastrotrich genus
Tetranchyroderma has multiciliated epidermal cells, whereas they are multi- or
monociliated in Megadasys and Urodasys (Rieger, 1976).
7) Epidermis with intraskeletal lamina: 0=absent, 1=present.
This character refers to the presence of a protein lamina located in the cytoplasm
close to the apical membrane of the epidermal cells. It is absent in
gnathostomulids, but is present in all rotifers and on the dorsal side of
Limnognathia maerski.
8) Epidermal cilia covered with cuticle: 0=absent, 1=present.
In gastrotrichs the cuticle invests the epidermal cilia (Ruppert, 1991).
9) Epithelium red-pigmented: 0=absent, 1=present.
The character refers to the conspicuous red-pigmented epidermis in
Haplognathia rosea and H. ruberrima (Fig. 2).
Sensory organs
10) Occipitalia: 0=absent, 1=present.
2
Occipitalia are single, stiff, non-unified sensory cilia that form a medio-dorsal row
on tip of rostrum. They are probably present in all gnathostomulids, but their
presence in Chirognathia dracula and Problognathia minima are not yet
confirmed.
11) First pair of apicalia: 0=absent, 1=present.
Apicalia are single, stiff sensory cilia located on the tip of the rostrum (Fig. 1).
They are present in most bursovaginoids, but have apparently never been found
in Labidognathia longicollis even though the species is well-studied (Riedl,
1970a; Sterrer, 1998, 2001).
12) Second pair of apicalia: 0=absent, 1=present.
An additional, more medial positioned pair of apicalia, present in
Austrognathiidae, i.e., Austrognathia, Austrognatharia, and Triplignathia (Farris,
1973, 1977; Sterrer, 1991a).
13) Frontalia: 0=absent, 1=present.
Paired, unified, sensory cilia, located on the rostrum lateral to the apicalia.
Frontalia are present in most bursovaginoids.
14) Ventralia: 0=absent, 1=present.
Paired sensoria, located ventral to the frontalia.
15) Dorsalia present: 0=absent, 1=present.
Paired, dorsally displaced sensoria, located caudal to the frontalia.
16) Lateralia: 0=absent, 1=present.
Paired sensoria located caudal to the dorsalia.
17) Paired lateral ciliary pits 0=absent, 1=present.
Paired ciliary inclusions on rostrum near dorsalia and lateralia.
3
18) Paired lateral ciliary pits bipartite: 0=absent, 1=present.
Sterrer (1976) described each of the paired ciliary pits in Tenuignathia rikerae as
bipartite, divided into a large posterior pit and a smaller anterior one. This
bipartition is present in Mesognatharia remanei and Labidognathia longicollis as
well (Sterrer, 1966a, 1998).
19) Unpaired ciliary pit on tip of rostrum: 0=absent, 1=present.
In addition to the paired ciliary pits, an additional single ciliary pit may be present
on the tip of the rostrum (Sterrer, 1976, 1997, 1998).
20) Lateral system: 0=absent, 1=present.
A pair of elongated tissue stripes of unknown function is located between the
epidermis and the gut in several Bursovaginoidea.
21) Locomotory cilia able to perform backward stroke: 0=absent,
1=present.
The epidermal cilia are used for locomotion, and some bursovaginoid taxa have
the capability of reversing the ciliary active stroke, and hence swim backwards.
Reproductive system
22) Bursa: 0=absent, 1=present.
This character refers to the presence of a bursa (Figs. 1, 3A); a conspicuous
element in the female organs of Bursovaginoidea. The bursa is a sack- or bellshaped organ that posteriorly is connected with the smaller prebursa. It probably
functions as a sperm storage organ, but its functional aspects are not yet
understood (Sterrer, 1972; Mainitz, 1989).
23) Substructure of bursa: 0=soft, 1=cuticularised.
4
The bursa may either be cuticularized or soft. All members of the family
Austrognathiidae have a soft bursa, whereas it is cuticularized in the remaining
bursovaginoids, the Scleroperalia.
24) Bursa with conspicuous constriction near distal end: 0=absent,
1=present.
Riedl (1971a) described and compared the detailed morphology of the
reproductive system in several species of Gnathostomula, and noted different
trends and character correlations within the genus. Based on his observations,
the Gnathostomula bursa can be subdivided into different types and one of them,
the “constriction” type, is characterized by a constriction near the caudal end and
only present among certain Gnathostomula species.
25) Injectory penis: 0=absent, 1=present.
Filospermoids lack an injectory penis. It is assumed that during mating the
filospermoid penis papilla adheres to the partner’s integument where the sperm
is deposited. Afterwards the sperm cells penetrate the integuments actively
(Sterrer, 1972; Mainitz, 1989).
26) Penis stylet: 0=absent, 1=present.
The penises in the scleroperalian bursovaginoids are supported with a central,
tube-like penis stylet composed of concentrically arranged rods (Fig. 3B).
27) Proximal stylet sack overlaps stylet distally: 0=absent, 1=present.
The proximal stylet sack is a muscular structure, located anterior to the penis
(Fig. 3B). Mainitz (1979) investigated the reproductive system of several
gnathostomulids and suggested an evolutionary trend from having all parts of the
stylet and stylet sheath located posterior to the proximal stylet towards having the
proximal stylet sack overlapping the anterior part of the stylet apparatus.
28) Granulated ring surrounds proximal end of penis stylet: 0=absent,
5
1=present.
Riedl (1971a) furthermore divide the male reproductive system of Gnathostomula
into different types. One of these, the “belt-penis type”, is characterized by the
presence of a granulated belt that wraps around the proximal part of the penis
stylet.
29) Location of opening in proximal stylet sack in relation to stylet
orientation: 0=opening of stylet sack in same axis as stylet, 1=proximal
stylet sack opens ventral to stylet axis.
Mainitz (1977) described an evolutionary trend from having the opening of the
stylet sack located in same plane as the axis of the stylet, present in most
Scleroperalia, to having the stylet sack opening located in a plane perpendicular
to the stylet axis, present in Gnathostomula.
30) Testes: 0=paired, 1=unpaired.
The character refers to the condition of the testes that may be paired (in
Haplognathiidae and “Scleroperalia”) or unpaired (in Austrognathiidae).
31) Sperm type: filiform: 0=dwarf type, 1=conuli.
The character refers to the three different sperm types that are present within
Gnathostomulida: the flagellar filiform type present in Filospermoidea, the nonflagellar dwarf type found in “Scleroperalia” (Fig. 3D), and the giant conuli
present in the members of the suborder Conophoralia, comprising the
austrognathiid genera (Fig. 3C). All outgroup taxa with males are coded as
having filiform sperm. Following a narrow definition of this sperm type, it may be
problematic to consider it present in all outgroup taxa (see Jenner, 2004). It is,
however, likely to assume that the filiform sperm present in Filospermoidea are
closer to the sperm in the outgroup taxa, whereas the highly modified dwarf
sperm and conuli are derived forms.
Pharyngeal structures
6
32) Pharynx musculature tri-lobed: 0=absent, 1=present.
The character refers to the formation of one medial and two lateral muscle sacs
in the pharyngeal bulb.
33) Arrangement of pharyngeal musculature surrounding the hard parts:
0=musculature capsular, 1=musculature arranged in loose network.
Sterrer (1972) characterized the pharyngeal musculature in the Haplognathiidae
as a “loose network”, contrary to the more capsular muscles in Pterognathiidae
and the Bursovaginoidea (see also Sterrer, 1969; Tyler and Hooge, 2001;
Sørensen et al, 2003a). The rotiferan and micrognathozoan outgroup taxa have
very compact mastax musculature as well (see, e.g., Kristensen and Funch,
2000; Sørensen et al., 2003b), whereas outgroup taxa without pharyngeal hard
parts are coded as inapplicable for the character.
34) Prepharyngeal gland: 0=absent, 1=present.
The character refers to the presence or absence of prepharyngeal glands (Fig.
1). The glands are present in most gnathostomulids, except Gnathostomula and
Clausognathia (Sterrer, 1972, 1992).
35) Prepharyngeal glands unpaired or paired: 0=unpaired, 1=paired.
The character refers to the paired or unpaired condition of the prepharyngeal
glands.
Jaws
36) Jaws: 0=absent, 1=present.
This character refers to the presence of pharyngeal hard parts with similar
ultrastructure, namely a composition of longitudinally arranged rods with lucent
material surrounding an electron-dense core. The homology of the pharyngeal
elements in Gnathifera has been supported in various studies (Ahlrichs, 1995;
Rieger and Tyler, 1995, Herlyn and Ehlers, 1997; Kristensen and Funch, 2000;
7
Sørensen, 2003). Among the gnathostomulids, jaws are not present in the
members of the genus Agnathiella.
37) Composition of central parts of jaws: 0=homogeneous material,
1=exclusively rods.
As stated above, the homology of the pharyngeal hard parts in Gnathostomulida,
Rotifera and Micrognathozoa is well supported, hence it is obvious that the
gnathostomulid lamellae symphysis, rotifer rami (Fig. 4A) and the
micrognathozoan main jaw (Fig. 4B) can be considered homologous as well.
However, the general appearance of these elements varies between the groups.
In Micrognathozoa and all rotifers the ultrastructural composition of bundled rods
are not visible unless the jaws are sectioned, and when visualized with SEM the
jaw sclerites appear to be composed of a much more compact and homogenous
material (Fig. 4). In gnathostomulids however, the lamellae symphyses are solely
made up by the rods, and this is in particular conspicuous in Gnathostomula (Fig.
5C), but also recognizable in all other gnathostomulids (Fig. 5).
38) Jaws with isolated, paired, interlocking sclerites forming mallei:
0=absent, 1=present.
This character refers to the differentiation of paired unci and manubria in rotifers
(Fig. 4A).
39) Anterior parts of jaws forming anchoring suspensorium: 0=absent,
1=suspensorium hard and sclerotized, 2=suspensorium soft and porous.
The lamellae symphyses in the filospermoid taxa are equipped with a
suspensorium that anchors the jaws inside the pharyngeal wall. A similar
suspensorium is present in the bursovaginoid taxa, but it differs in being much
softer and porous, compared to the hard and compact filospermoid
suspensorium (compare Fig. 5A with 5B-C).
40) Suspensorium horizontally bipartite: 0=absent, 1=present.
8
The suspensorium in the genera Pterognathia (not included in analysis) and
Cosmognathia (Fig. 6C) is horizontally bipartite, which is one of the main
characteristics for the family Pterognathiidae (Sterrer, 1966b, 1969, 1970, 1972,
1991b, 1998).
41) Suspensorium with rostral apophyses: 0=absent, 1=present.
The character refers to the presence of rostral apophyses, viz., compact
apophyses that emerge from the rostral part of the suspensorium (Fig. 6). The
rostral apophyses are present in the filospermoid taxa and differ from the
apophyses in, e.g., Gnathostomula that are formed by much softer lamellae.
42) Appearance of rostral apophyses: 0=rostral apophyses pointing
caudally, 1=rostral apophyses pointing ventrally.
The character refers to the orientation of the distal ends of the rostral apophyses
(Fig. 6). Taxa without rostral apophyses are coded as inapplicable for this
character.
43) Suspensorium with caudal apophyses: 0=absent, 1=present.
The character refers to the presence of apophyses that structurally resemble
rostral apophyses but that are located more caudally (Fig. 6).
44) Suspensorium with strong cristae: 0=absent, 1=present.
Cristae are dorsolaterally extending crescentic apophyses, present in the genera
Pterognathia and Cosmognathia (Fig. 6C).
45) Jaws (exclusive teeth) reduced to extremely delicate lamellae:
0=absent, 1=present.
Gnathostomulid jaws are usually made of strong hard parts, but in the genera
Austrognathia, Austrognatharia, and Triplignathia all parts of the jaws, except the
teeth, are extremely delicate and almost invisible in SEM (Fig. 7).
9
46) Anterior part of suspensorium forms two well-defined chambers:
0=absent, 1=present.
Kristensen and Nørrevang (1977) described the presence of two windows or
chambers, fenestra lateralis and fenestra ventralis, in Rastrognathia
macrostoma. This observation was later confirmed with SEM (Sørensen, 2000),
and further studies revealed that similar chambers are present in
Vampyrognathia (not included in analysis), Valvognathia and Onychognathia
(Fig. 8) (Sørensen and Sterrer, 2002; Sørensen, unpubl. obs.).
47) Anterior part of suspensorium forms delicate shoulder lamellae:
0=absent, 1=present.
Shoulder lamellae are thin lamellae that extend laterally and then caudally from
the anterior part of the suspensorium. The character is easily observed in some
taxa, i.e., Labidognathia, and to some extent in Gnathostomula (Riedl, 1970a;
Sterrer, 1998, Sørensen et al., 2003a), but it has never been reported from any
onychognathiid taxa. However, recent SEM studies (Sørensen and Sterrer, 2002;
Sørensen, unpubl. obs.) show that shoulder lamellae are present in these taxa as
well, but that they are extremely delicate and hence invisible with LM (Fig. 8).
Shoulder lamellae are probably present in all bursovaginoid taxa, but this still
needs confirmation from a few taxa.
48) Caudal part of jaws extended laterally into anchoring symphysis:
0=absent, 1=present.
This character refers to the caudal symphysis present in all filospermoid taxa
(Fig. 5A).
49) Cauda surrounding symphysis: 0=absent, 1=present.
This character refers to the presence of a caudally extending element, the socalled cauda. Proximally it joints the symphysis of the lamellae symphyses, and
the paired or unpaired distal ends are connected to the pharyngeal wall, hence
10
serving as the caudal anchoring point for the jaws (Figs. 5C, A, D) (see Riedl
and Rieger, 1972; Sterrer, 1972; Sørensen and Sterrer, 2002).
50) Appearance of cauda: 0=unpaired, 1=paired.
This character refers to the condition of the cauda that may be either paired (Fig.
5C) or unpaired (Fig. 8A). Taxa without a cauda are coded as inapplicable for
this character.
51) Attachment point of cauda: 0=cauda encapsulates symphysis, 1=cauda
attaches on ventrocaudal corner of symphysis.
The attachment point of the cauda may be either terminal, almost encapsulating
the symphysis, or alternatively attached on the ventrocaudal corner of the
symphysis (Fig. 8A-D).
52) Appearance of teeth in the dentarium: 0=teeth large, lanceolate or
conical, 1=teeth short, pointed needles.
The anterior denticulated part of the jaws, the dentarium, may be equipped with
different kinds of teeth. Generally they can be divided into two types: either large
well-developed teeth, present in Bursovaginoidea (Fig. 9E-H), or short, pointed,
needle-like denticles, found in Filospermoidea (Fig. 9A-D). The teeth in the jaw
possessing outgroup taxa belong to the first type (Fig. 4).
53) Arrangement of needle-shaped denticles: 0=arranged in a cluster,
1=differentiated into one or two main teeth and several closely set rows,
2=arranged into one to four well-defined rows
The needle-like denticles can be arranged in different rows (Fig. 9B-D) or in a
simple cluster (Fig. 9A). Taxa without needle-like denticles are coded as
inapplicable for this character.
11
54) Larger teeth arranged in multiple rows: 0=teeth in three rows, 1=medial
row reduced to one terminal tooth, 2=dorsal and medial rows each reduced
to one terminal tooth. Additive.
In some bursovaginoid taxa the teeth are arranged in multiple rows. The genus
Triplignathia (not included in analysis) and all genera in Gnathostomulidae
possess three rows (Fig. 9G-H), whereas Austrognathia and Austrognatharia
have only two and one row, respectively (Fig. 7). Sterrer (1972) suggested that
three rows represented the basic pattern for these taxa and that Austrognathia
and Austrognatharia lost their rows through a series a reductions. To incorporate
this information, the character states have been coded as additive. Taxa without
teeth arranged in multiple rows are coded as inapplicable for this character.
55) Regular tooth rows supported by rows of finer teeth: 0=absent,
1=present.
SEM studies have revealed that the regular tooth rows in some taxa may be
supported by rows of much finer teeth (Fig. 9G-H).
56) Larger teeth emerging from a single transverse row: 0=absent,
1=present.
In most taxa with teeth in rows, the rows are arranged longitudinally, but in at
least two taxa, Mesognatharia remanei and Tenuignathia rikerae, the rows are
clearly transverse (Figs. 5B, 9E-F).
57) Larger teeth on ventral side of jaws short, stout, conical, arranged in
one longitudinal row, pointing perpendicular to main axis of jaws:
0=absent, 1=present.
The dentaria in the onychognathiid taxa follow some distinct patterns. The teeth
can generally be divided into a dorsal and a ventral part, and depending on their
location they are differentiated in different ways. This character refers to the
ventral teeth that in some taxa are rather short, stout and conical and arranged
perpendicular to the main axis of the jaws (Fig. 8B, D, F). This arrangement is
12
found in all onychognathiid genera and in the outgroup taxon Limnognathia
maerski as well (Kristensen and Funch, 2000; Sørensen, 2003).
58) Larger teeth emerging rostrally, forming a rostral basket: 0=absent,
1=present.
The dorsal and rostral teeth in the onychognathiid taxa, exclusive Goannagnathia
(not included in analysis), emerge from a lateral point in the dentarium and
extend rostrally and dorsally, forming a basket. The character is also coded as
present for Limnognathia maerski.
59) Dorsal teeth more loosely attached than other teeth: 0=absent,
1=present.
A conspicuous trait for the dentaria in some onychognathiids is the most dorsal
teeth that tend to be more loosely attached than the other teeth (Fig. 8A, C). The
trait is present in Onychognathia, Valvognathia, and Vampyrognathia, but not in
Rastrognathia or Goannagnathia.
Basal plate
60) Unpaired basal plate consisting of five alae (wings): 0=absent,
1=present.
This character refers the presence of an unpaired basal plate. The coding follows
the interpretation of Sørensen (2003) and not Kristensen and Funch (2000),
hence Limnognathia maerski is not coded as having a basal plate.
61) Shape of basal plate: 0=Triangular or droplet-shaped, 1=rod-shaped,
2=flattened, crescentic.
The basal plate in the bursovaginoid taxa is mostly crescentic or winged (Fig.
11), with distinct division into plates, whereas it is more irregularly shaped in
Filospermoidea (Fig. 10). In Haplognathia the basal plate is rather small and may
be triangular or more or less droplet-shaped (Fig. 10A). In Pterognathia and
13
Cosmognathia it is also shaped like a transverse rod (Fig. 10A-B), but it may be
equipped with different notches and extensions.
62) Differentiation of medial ala in crescentic basal plate: 0=not
differentiated, 1=medial ala conspicuously broader than other alae
According to Riedl and Rieger (1972) and Sterrer (1972), the bursovaginoid
basal plate was composed of five more or less equally sized plates in its
ancestral state. Afterwards the basal plate was modified through a series of
modifications of the different plates. This character refers to the lateral elongation
of the medial plate.
63) Differentiation of paramedial plates in crescentic basal plate: 0=not
different, 1=paramedial plates form conspicuous rostral wings
This character refers to the presence of rostral wings, alae rostralis, formed by
modification of the paramedial plates.
64) Differentiation of lateral plates in crescentic basal plate: 0=not different,
1= lateral plates form conspicuous lateral wings
The character refers to the presence of lateral wings, alae lateralis, formed by
modification of the lateral plates. The modification is conspicuous in members of
Gnathostomulidae. Riedl and Rieger (1972) suggest that the same modification
happened in Austrognathiidae, but SEM observations show that the
austrognathiid basal plate differs considerably from the type found in
Gnathostomulidae.
65) Caudal end of basal plate with transverse crest: 0=absent, 1=present.
The character refers to the presence of a caudal crest that is present in the basal
plate of some members of Gnathostomulidae.
14
66) Denticulation of basal plate: 0=needle-shaped denticles on various
parts of basal plate, 1=regular denticles arranged in one row on rim of
basal plate
The basal plate may either carry small needle-shaped denticles (Fig. 10), like
those in the filospermoid dentarium, or more regular teeth (Fig. 11).
67) Differentiation of denticles in row on rim of basal plate: 0=regular teeth;
1=denticles tripartite; 2=teeth appear like small scales.
The basal plate denticles in Bursovaginoidea appear in three different kinds. In
Gnathostomulidae they form elongated pointed teeth (Fig. 11C, E), whereas
several other bursovaginoid taxa only have small scale-like teeth (Fig. 11F). SEM
studies on basal plates in Austrognatharia and Austrognathia have revealed that
the denticles in these taxa are tripartite with one medial pointed tip, and two
smaller lateral tips (Fig. 11A-B) (Sørensen and Sterrer, 2002). The denticles in
Problognathia minima are clearly tripartite as well (Fig. 11D), even though the
medial tip does not differ from the lateral ones (Sørensen and Sterrer, 2002).
68) Denticles in second position from the lateral side and in the medial
position on basal plate considerably larger than other denticles: 0=absent,
1=present.
The character refers to the condition found in the austrognathiid taxa, where the
medial basal plate denticle and the second denticle counted from each side in
the row, are considerably larger than the rest (Fig. 11A-B).
69) Rostral margin of basal plate serrated: 0=absent, 1=present.
In some taxa the rostral rim of the basal plate bears a row of densely set serrulae
(Fig. 11C, E). Riedl and Rieger (1972) suggest that this trait is an important
synapomorphy for Gnathostomulidae. The serrulated margin has also been
found in Chirognathia, whereas the basal plate in Ratugnathia apparently lacks
all kinds of dentation and serrulation (Sterrer, 1991b).
15
70) Basal plate with strong medial notch: 0=absent, 1=present.
The characters refer to the conspicuous medial notch found on the basal plate of
Cosmognathia aquila (Fig. 10B) and C. arcus (Sterrer, 1970, 1991b, 1998).
71) Basal plate with three prominent rostral lobes: 0=absent, 1=present.
The character refers to the three rostral lobes that are present on the basal plate
in Austrognathia but not in Austrognatharia.
72) Crescentic jugum: 0=absent, 1=present.
The character refers to presence of an unpaired crescentic structure, the jugum,
which is located in the dorsal part of the pharyngeal wall. The jugum is usually
considered autapomorphic for the Gnathostomulidae, but it was not found in
Chirognathia, even though the taxon displays clear affinities to the family.
Kristensen and Nørrevang (1978) reported the presence of a jugum in
Valvognathia pogonostoma, but even though the species has been investigated
several times since then (Sørensen and Sterrer, unpupl. obs.), the presence of a
jugum has never been confirmed. Hence, V. pogonostoma is coded as uncertain
for this character.
Other
73) Ectoparasites on nephropid lobster mouth parts: 0=absent, 1=present.
This character refers to the special habitat of the ectoparasitic Cycliophora
(Funch and Kristensen, 1995, 1997; Obst et al. 2005).
16