International Journal of Systematic and Evolutionary Microbiology (2014), 64, 680–688
DOI 10.1099/ijs.0.059899-0
Morphological reports on two species of
Dexiotricha (Ciliophora, Scuticociliatia), with a note
on the phylogenetic position of the genus
Xinpeng Fan,1 Saleh A. Al-Farraj,2 Feng Gao3 and Fukang Gu1
Correspondence
Fukang Gu
[email protected]
1
School of Life Sciences, East China Normal University, Shanghai 200062, PR China
2
Zoology Department, College of Science, King Saud University, Riyadh 11451, Saudi Arabia
3
Laboratory of Protozoology, Institute of Evolution and Marine Biodiversity,
Ocean University of China, Qingdao 266003, PR China
Two Dexiotricha species (Dexiotricha elliptica nov. comb. and Dexiotricha cf. granulosa),
respectively isolated from soil north-west of Riyadh, Saudi Arabia, and freshwater in Shanghai,
eastern China, were investigated using standard methods. The species Loxocephalus ellipticus
Kahl, 1931 is reclassified here in the genus Dexiotricha and was characterized mainly by
constantly showing 16 somatic kineties, three post-oral kineties with the middle one shortened, a
contractile vacuole located subcaudally with an excretory pore near the posterior end of somatic
kinety 2 and single caudal cilia. A Dexiotricha granulosa-like organism having a subcaudally
located contractile vacuole and fewer somatic kineties was designated D. cf. granulosa. The
small-subunit rRNA gene (SSU rDNA) sequences of these two species were characterized and
their phylogenetic positions based on SSU rDNA sequences were revealed by means of Bayesian
inference and maximum-likelihood analysis. Phylogenetic analyses confirmed Dexiotricha as a
monophyletic genus and supported its assignment to the order Loxocephalida. However, its family
assignment remains unsupported.
INTRODUCTION
Scuticociliates are free-living organisms of limnetic and
marine ecosystems, and this group often draws more
public attention than other ciliates, mainly because some
are symbionts or opportunistic parasites of aquatic animals
(Fan et al., 2011a, b; Foissner et al., 1994; Lobban et al.,
2011; Pan et al., 2013). In this age of ‘refinement’, taking a
phylogenetic view of this group with molecular analyses
plays an equal role with exploring species diversity and
providing standard descriptions of previously unclear
species or novel species (Fan et al., 2009; Pan et al., 2011,
2013; Zhang et al., 2010, 2011).
The order Loxocephalida was proposed by Jankowski
(1980) to include those genera with morphological characteristics of both scuticociliates (scutica and an apical
plate) and hymenostomes (obliquely orientated oral membranelles and post-oral kineties). Based on available data,
recent studies have reconsidered the phylogenetic status
of the order and confirmed it as paraphyletic group, and
Abbreviations: BI, Bayesian inference; ML, maximum-likelihood; PK,
post-oral kinety; SK, somatic kinety.
The GenBank/EMBL/DDBJ accession numbers for the SSU rDNA
sequences of Dexiotricha elliptica and Dexiotricha cf. granulosa are
respectively KF878932 and KF878931.
680
members of the group share no analogous morphogenetic
pattern (Gao et al., 2013; Li et al., 2010; Long et al., 2007;
Song et al., 2005; Zhang et al., 2010, 2011). However, since
the data used by these analyses were quite limited, and
the inter-relationship of the order was still indeterminate,
in order to characterize this order fully, more sequences
are needed, along with morphological/morphogenetic descriptions of suggested members (Gao et al., 2013; Lynn,
2008; Zhang et al., 2010, 2011). Dexiotricha, also a candidate member of the order, has not been subjected to a
molecular phylogenetic analysis. It is a confusing taxon in
terms of its taxonomy, because many species were assigned
to other genera, e.g. Loxocephalus, and several species have
been confirmed to be synonyms (Foissner et al., 1994;
Peck, 1974; Wilbert, 1986). Thus, a detailed morphological
description and the sequences of marker genes, e.g. the
small-subunit rRNA gene (SSU rDNA), will contribute not
only to the taxonomy of the genus but also to the further
understanding of the phylogenetic status of the order
Loxocephalida.
In our study, the morphological description of two members of Dexiotricha, isolated from soil and freshwater, is
documented based on observations of specimens in vivo,
after protargol staining and scanning electron microscope
(SEM) sample preparation. In addition, the SSU rDNA of
059899 G 2014 IUMS Printed in Great Britain
Morphology and phylogeny of two Dexiotricha species
the two species was sequenced for the first time, and phylogenetic analyses were performed to assess the systematic
position of the genus Dexiotricha in the order Loxocephalida.
METHODS
Sample collection, observation and identification. Dexiotricha
elliptica (Kahl, 1931) nov. comb. was collected from farmland at Zulfi
city, north west of Riyadh, Saudi Arabia (26u 139 220 N 44u 519 430 E).
The soil sample was taken directly, and was then processed with the
non-flooded Petri dish method in the laboratory (Foissner, 1987).
Dexiotricha cf. granulosa (Kent, 1881) Foissner et al., 1994 was collected from a freshwater pond in Changfeng Park, Shanghai, China
(31u 139 300 N 121u 239 560 E). Cells were isolated and observed in vivo
using differential interference contrast microscopy. Protargol staining
(Wilbert, 1975) was performed in order to reveal the infraciliature.
Drawings were made with the help of a camera lucida. Measurements
were made under 6100–1250 magnification. Samples for SEM were
prepared as described by Gu & Ni (1993) and were observed under
a Hitachi S-4800 scanning electron microscope with accelerating
voltage 10.0 kV. Terminology and classification mainly follow
Jankowski (1980) and Lynn (2008).
DNA extraction and gene sequencing. Extraction of genomic
DNA was carried out according to the methods described by Gong
et al. (2007). To minimize PCR amplification errors, high-fidelity
TaKaRa ExTaq was used to amplify the SSU rDNA gene using universal oligonucleotide primers (forward, 59-GAAACTGCGAATGGCTC-39 or 59-AATCTGGTTGATTTTGCCAGT-39; reverse, 59TGATCCTTCTGCAGGTTCACCTAC-39) designed by Medlin et al.
(1988) and Elwood et al. (1985). Cloning and sequencing were performed as reported by Zhang et al. (2011).
Phylogenetic analyses. Other than the SSU rDNA sequences of the
two species, sequences used in the present analyses were obtained
from the NCBI GenBank database. Sequences were first aligned using
the GUIDANCE algorithm (Penn et al., 2010a) with default parameters
in the GUIDANCE web server (Penn et al., 2010b). Ambiguous columns
in the alignment below the confidence score of 0.6 calculated by
GUIDANCE were removed. The final alignment including 1720 sites and
74 taxa was used to reconstruct phylogenetic trees using the methods
described below. Coleps nolandi, Tiarina fusa and Prorodon teres
were chosen as the outgroup taxa. The most appropriate model for
phylogenetic analyses was selected by both Modeltest version 3.4
(Posada & Crandall, 1998) and MrModeltest version 2.2 (Nylander,
2004). Maximum-likelihood (ML) analyses were carried out using
RAxML-HPC2 version 7.2.8 (Stamatakis et al., 2008) on the CIPRES
Science Gateway (Miller et al., 2010) using the GTR+G model as the
selected option. Support came from a majority rule consensus tree
of 1000 bootstrap replicates. Bayesian inference (BI) analysis was
performed using MrBayes 3.1.2 (Ronquist & Huelsenbeck, 2003) on
the CIPRES Science Gateway using the GTR+I+G model as selected
by MrModeltest version 2.2 according to the Akaike information
criterion. Markov chain Monte Carlo simulations were run with two
sets of four chains for 4 000 000 generations with a sample frequency
of 100 generations, and the first 10 000 trees were discarded as burnin. All remaining trees were used to calculate posterior probabilities
using a majority rule consensus.
RESULTS
Family Loxocephalidae Jankowski, 1964
Genus Dexiotricha Stokes, 1885
Dexiotricha elliptica (Kahl, 1931) nov. comb. (Figs
1 and 2; Table 1)
Basionyms: Loxocephalus ellipticus Kahl, 1931; Dexiotricha
media sensu Small & Lynn, 1985.
M1-3
PM
(b)
Sc
PK
SK1
SKn
Ma
Mi
CVP
(a)
(c)
(d)
(e)
Fig. 1. D. elliptica (Kahl, 1931) nov. comb. from life (a, c) and after protargol staining (b, d, e). (a) Ventral-lateral view (from
present work); arrow marks the caudal cilia. (b) D. media sensu Small & Lynn, 1985. (c) Ventral views of L. elliptica sensu Kahl,
1931 (from Kahl, 1931). (d, e) Ventral (d) and dorsal (e) views of infraciliature; arrowhead in (d) marks the densely arranged
basal bodies in the anterior part of SK1 and the arrow depicts the anteriorly shortened SKn. The arrowhead in (e) marks the two
dikinetids in each somatic kinety. CVP, Contractile vacuole pore; M1–3, membranelles 1–3; Ma, macronucleus; Mi,
micronucleus; PK, post-oral kineties; PM, paroral membrane; Sc, scutica; SK1, n, somatic kinety 1, n. Bars, 25 mm.
http://ijs.sgmjournals.org
681
X. Fan and others
(f)
(e)
(a)
(b)
(d)
(c)
(g)
M1-3
(h)
PM
PK
Sc
Ma
(i)
(j)
(k)
(l)
Fig. 2. D. elliptica (Kahl, 1931) nov. comb. from life (a–h) and after protargol staining (i–l). (a, e) Lateral-ventral views of slightly
compressed individuals; the arrow in (a) indicates the buccal field and the arrow in (e) refers to the contractile vacuole. (b)
Anterior portion, showing the buccal cavity (arrow) and the food vacuoles (arrowheads). (c, d) Individual in free swimming (c)
and slightly compressed (d); the arrow marks the caudal cilium, and arrowheads indicate the extrusomes. (f) Surface of the cell,
to show the kinety rows (arrowheads). (g) Posterior portion, showing the small granules in the cytoplasm (arrowheads). (h)
Dorsal view to show the somatic cilia. (i) Ventral view of infraciliature; the arrowhead shows the anteriorly shorted SKn and the
arrow marks the contractile vacuole pore. (j) Showing the buccal apparatus and part of the somatic kineties; the arrowhead
marks the densely arranged kinetids in the anterior part of SK1 and the arrow indicates the posterior end of SK2. (k) Showing
the buccal argyrome (arrow). (l) Dorsal view of infraciliature. See legend to Fig. 1 for abbreviations. Bars, 25 mm.
Detailed study and a description have never been given
for this species; thus, an improved diagnosis is supplied
here.
of Protozoology, East China Normal University, with
registration number FXP-2012-10-7-1.
with protargol specimens is deposited in the Laboratory
Cells 45–55620–25 mm in vivo, with
length : width about 2 : 1. Body shape long ellipsoidal, with
ventral side straight, dorsal side gently curved (Figs 1a and
2c). Buccal cavity is small with inconspicuous buccal cilia
(Figs 1a and 2a, b). Cytoplasm colourless and containing
numerous granules of about equal size (Figs 1a and 2d, g).
Food vacuoles approx. 5 mm in diameter present when well
fed (Fig. 2a, b). Pellicle non-concave, with inconspicuous
ridges along cilia rows (Fig. 2f). Extrusomes uneasy to
detect, about 4 mm long (Figs 1a and 2d). Contractile
vacuole located posteriorly near left of meridian, approx.
6 mm in diameter; contracting interval about 10 s (Figs 1a
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International Journal of Systematic and Evolutionary Microbiology 64
Description.
Improved diagnosis. Cells in vivo usually measure 506
25 mm, body claviform. Three membranelles composed of
three- or four-rowed kineties. Scutica comprising three
dikinetids. Constantly 16 somatic kineties, three post-oral
kineties with middle one shortened. Contractile vacuole
subcaudally located, with excretory pore near posterior end
of somatic kinety 2 (SK2). Single macronucleus and caudal
cilium.
Deposition of voucher specimens. One voucher slide
Morphology and phylogeny of two Dexiotricha species
Table 1. Morphometric characterization of D. elliptica (Kahl,
1931) nov. comb. (first line) and D. cf. granulosa (second line)
vacuole pore detectable after protargol staining, located
near the end of SK2 (Figs 1a and 2i).
Data are based on protargol-impregnated specimens. 2, Not detected.
Character
Body, length (mm)
Body, width (mm)
Somatic kineties (n)
Post-oral kineties (n)
Basal bodies of
SKn (n)
Macronucleus,
length (mm)
Macronucleus,
width (mm)
Min. Max. Mean
SD
CV
(%)
n
35
50
24
25
16
28
3
2
12
55
72
35
35
16
30
3
4
14
48.0
59.6
28.6
30.4
16.0
28.4
3.0
3
12.9
6.1
7.8
3.4
3.3
0
0.9
0
0.7
0.5
12.8
13.1
11.7
11.0
0
3.1
0
23.6
3.8
13
11
13
11
13
5
10
5
10
2
7
2
14
2
11.2
2
3.4
2
20.0
2
13
10
7
19
12
13.0
9.5
3.2
2.2
24.3
15.8
11
13
7
14
10.0
2.6
26.5
11
and 2e). Single macronucleus, located near mid-body,
about 11610 mm in size (Figs 1a, e and 2l). Length of
somatic cilia about 8 mm (Figs 1a and 2h). Caudal cilium
flexible and extremely long, 20–40 mm (Figs 1a and 2d).
Infraciliature as shown in Figs 1(d, e) and 2(i–l). Constantly 16 somatic kineties, which mostly consist of monokinetids extending along the entire length of the cell.
SK2 to SKn21 consist of two pairs of dikinetids in their
anterior end (Figs 1d and 2i, l). Anterior six or seven
kinetids of SK1 densely arranged, SKn anteriorly shorter
and starting at level of membranelle 1, consisting only of
monokinetids (Figs 1d and 2i, j). Three post-oral kineties
(PK), each consisting of one pair of dikinetids in its
anterior end. PK1 starting anteriorly below posterior end of
paroral membrane, nearly extending to the end of the cell,
PK3 shorter, starting anteriorly near cytostome, extending
over half of body, PK2 very short, comprising only four
kinetids, anteriorly near scutica (Figs 1d and 2i, j). SK2
posteriorly shorter and ending near contractile vacuole
pore (Figs 1d and 2j). Buccal apparatus consists of three
transversely positioned oral membranelles (M) as follows.
M1 extends obliquely toward posterior from right to left,
consisting of three parallel rows and a curved row around
anterior end of three rows. First row is two kinetids shorter
than the other two. M2 forms a 30u intersection angle with
M1 and consists of two rows of five to seven kinetids, and
some additional kinetids on its right side. M3 is parallel to
M2, smaller than M1 and M2. Paroral membrane is slightly
curved, located on right margin of buccal cavity, terminating anteriorly from level of M1 and M2. Scutica composed of three pairs of dikinetids located near posterior
end of paroral membrane (Figs 1d and 2j). About nine oral
ribs converge towards cytostome (Fig. 2k). Contractile
http://ijs.sgmjournals.org
Dexiotricha cf. granulosa (Kent, 1881) Foissner
et al., 1994 (Figs 3 and 4; Table 1)
Description. Cells measure 50–70615–25 mm in vivo,
oval shaped with anterior end narrowed and posterior
end broadly rounded (Figs 3a and 4a). Cytoplasm contains
numerous granules, distinctly ring-like and with a regular
size of about 1–2 mm (Figs 3a and 4a, b). Food vacuoles
present in well-fed individuals (Figs 3a and 4b). Contractile
vacuole located near caudal end about 6 mm in diameter,
with single excretory pore (Figs 3a and 4a, e, g, h); buccal
membranelles inserted in inner wall of buccal cavity, about
6 mm long (Figs 3a and 4f). Somatic cilia and caudal cilium
about 8 and 15–20 mm long, respectively (Figs 3a and 4a, e, g).
Infraciliature as shown in Figs 3(b, c) and 4(c, d, i, j).
Twenty-eight to thirty somatic kineties, mostly comprising
monokinetids extending along the whole body (Fig. 3b, c).
Somatic kineties near buccal field curved along shape
of paroral membrane (Fig. 4c). About eight kinetids in
anterior part of SK1 densely arranged (Figs 3b and 4i). SKn
anteriorly shorter than others and having just monokinetids (Fig. 3b). From left side of SKn, about 12 kineties
contain only one dikinetid in the anterior end, while the
remaining somatic kineties contain two pairs of dikinetids
in the anterior end; on the left of SKn, 12 kineties (Fig. 3b,
c). Two to four post-oral kineties: PK1, the rightmost one,
comprises one dikinetid in the anterior end and monokinetids in the remaining part; PK2 contains only three
to five monokinetids. The number of post-oral kineties on
the right of PK2 varies from zero to two (Figs 3b and 4c, e).
Buccal apparatus consists of three transversely positioned
oral membranelles: M1 and M2 three-rowed, M3 tworowed (Figs 3b and 4i). Paroral membrane distinctly curved
in mid-portion, starting anteriorly from level between
M1 and M2 and curving posteriorly around M2 and M3
(Figs 3b and 4i). About 11 oral ribs subtend the buccal area
(Fig. 4g). Scutica located near posterior end of PM, possibly
comprising three pairs of dikinetids (Fig. 3b, j).
SSU rDNA sequences and phylogenetic analyses
The SSU rDNA sequences of the two species have been
deposited in GenBank with the length, DNA G+C content
and accession number as follows: Dexiotricha elliptica,
1683 bp, 44.83 mol%, KF878932; D. cf. granulosa, 1769 bp,
46.30 mol%, KF878931. Pairwise comparison reveals that
D. elliptica and D. cf. granulosa differ from each other in 96
positions, with 94.32 % sequence identity. They differ from
the former Dexiotricha sp. MD-2012 (GenBank accession
no. JQ723963) in 105 and 15 positions (sequence identity
of 93.70 and 99.13 %), respectively.
The resulting topologies generated using two algorithms
(BI and ML) are generally concordant; therefore, a single
topology is presented with support values from both
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X. Fan and others
M1-3
PM
Sc
SK1
PK
SKn
(a)
(b)
(c)
Fig. 3. D. cf. granulosa (Kent, 1881) Foissner et al., 1994 from life (a) and after protargol staining (b, c). (a) Ventral-lateral view
of a typical individual; the arrow marks the contractile vacuole, and the arrowhead indicates the caudal cilia. (b) Ventral side of
the infraciliature; the arrowhead indicates the somatic kineties on the left of SKn, which comprise only one dikinetid. (b) Dorsal
side of infraciliature; the arrow marks the bare anterior end. See legend to Fig. 1 for abbreviations. Bars, 25 mm.
algorithms indicated on the branches (Fig. 5). In the phylogenetic trees, Loxocephalida is not monophyletic, with
most of its members forming a sister clade to the core
Ma
scuticociliates (25 % ML, 0.90 BI), while Cinetochilum
and Dexiotrichides group in other subclasses. All three
Dexiotricha species form a monophyletic group with full
Ma
(b)
(a)
(f)
(e)
(c)
(d)
(h)
M1-3
(i)
(g)
684
(j)
Fig. 4. D. cf. granulosa (Kent, 1881) Foissner
et al., 1994 from life (a, b), after protargol
staining (c, d, i, j) and after SEM preparation
(e–h). (a) Lateral-ventral view of a typical
individual; the arrowhead marks the caudal
cilium, and the arrow indicates the contractile
vacuole. (b) Showing the ring-like granules in
the cytoplasm. (c, d) Ventral (c) and dorsal (d)
view of the infraciliature; arrowheads mark the
post-oral kineties and the arrow refers to the
two dikinetids anterior of each somatic kinety.
(e) Ventral view; arrow indicates the shortened
post-oral kinety which is composed of only
three kinetids, the arrowhead marks the anteriorly shorted SKn and the double arrowhead
shows the contractile vacuole pore. (f) Details
of buccal field; arrowhead marks the oral
fibres. (g) Detail of contractile vacuole pore.
(h) Right-lateral view; the arrow indicates the
dikinetids in the anterior part and the arrowhead refers to the contractile vacuole. (i, j)
Infraciliature of buccal field; arrowhead marks
the single kinetids in the anterior of somatic
kineties at the left side of buccal cavity and the
arrow indicates the scutica. See legend to Fig.
1 for abbreviations. Bars, 25 mm.
International Journal of Systematic and Evolutionary Microbiology 64
Morphology and phylogeny of two Dexiotricha species
SSU-rDNA
64/1.00
37 core scuticociliates
SCUTICOCILIATIA
ML/BI
Paratetrahymena wassi GQ292767
99/1.00 Paratetrahymena sp. EU744176
0.05
25/0.90
Paratetrahymena parawassi FJ876969
78/1.00 Paratetrahymena wassi JX310019
Cardiostomatella vermiforme AY881632
9/Loxocephalida
Dexiotricha sp. JQ723963
9/0.81
Dexiotricha cf. granulosa KF878931
Dexiotricha elliptica KF878932
Sathrophilus holtae FJ868188
Sathrophilus planus FJ868186
13/0.80
Pseudoplatynematum denticulatum JX310020
93/1.00 Eudrilophrya complanata HQ446280
79/1.00
Paraclausilocola constricta HQ446275
Njinella prolifera HQ446276
ASTOMATIA
Metaracoelophrya sp. HQ446277
Almophrya bivacuolata HQ446281
Anoplophrya marylandensis AY547546
59/0.54
Hyalophysa lwoffi EU503538
Vampyrophrya pelagica EU503539
APOSTOMATIA
97/1.00
53/1.00
Gymnodinioides pitelkae EU503534
Cinetochilum ovale FJ870103
Loxocephalida
Dexiotrichides pangi AY212805
99/1.00
70/0.55
Glaucoma chattoni X56533
Tetrahymena pyriformis M98021
HYMENOSTOMATIA
Colpidium striatum HM030739
41/0.80
Ichthyophthirius multifiliis U17354
98/1.00 Ophryoglena catenula U17355
Vorticella campanula AF335518
64/0.98
70/0.76
Ophrydium versatile AF401526
PERITRICHIA
Epistylis chrysemydis AF335514
Zoothamnium alternans DQ868352
Paramecium tetraurelia AB252009
Frontonia tchibisovae DQ883820
PENICULIA
Lembadion bullinum AF255358
Coleps nolandi AM292313
Tiarina fusa FJ858217
PROSTOMATEA
Prorodon teres X71140
Fig. 5. ML tree inferred from SSU rDNA sequences, showing the position of D. elliptica and D. cf. granulosa. Nodal support for
branches in the ML/BI trees is indicated; ‘–’ indicates bootstrap value disagreement between the ML tree and the reference BI
tree at a given node. Bar, 5 substitutions per 100 nucleotide positions. Coleps nolandi, Tiarina fusa and Prorodon teres are the
outgroup taxa.
support, though the relationships within loxocephalids are
unsolved due to low support.
with Loxocephalus species. Consequently, a new combination Dexiotricha elliptica nov. comb. is proposed. Due to
the feminine gender of the genus, the species name must
be changed accordingly.
DISCUSSION
Small & Lynn (1985) reported a population of D. media
and illustrated the infraciliature without description (Fig.
1b). However, D. media sensu Peck, 1974 apparently has
more somatic kineties than the population of Small & Lynn
(1985) (31–34 compared with about 16, counted from the
figure). Moreover, Foissner et al. (1994) had synonymized
D. media Peck, 1974 with D. granulosa. According to the
great similarities in terms of the details of the infraciliature
and the subcaudally located contractile vacuole, we believe
that D. media sensu Small & Lynn, 1985 is conspecific with
D. elliptica.
Remarks on Dexiotricha elliptica
Kahl (1931) reported the species Loxocephalus ellipticus
from a freshwater habitat. According to his description
on two populations, cells of L. ellipticus measured about
40–55 mm in vivo, with eight or nine somatic kineties on
one side and coarse granules in the cytoplasm; moreover,
the contractile vacuole was located near the posterior end
(Fig. 1c). The abovementioned description corresponds
well with our organism, especially for the subcaudally
located contractile vacuole. Thus, we believe our population is conspecific with Loxocephalus ellipticus Kahl, 1931.
However, both the original description and the present
description showed that this organism has no similarities
http://ijs.sgmjournals.org
The genus Dexiotricha comprises only six species after
Foissner et al. (1994) synonymized D. media and Dexiotricha
plagia with D. granulosa. Considering the location of the
contractile vacuole and its excretory pore, D. elliptica is most
685
X. Fan and others
similar to Dexiotricha colpidiopsis, but differs by having
fewer somatic kineties (16 vs 24–28) (Fig. 6e, f; Table 2).
D. elliptica differs from D. granulosa, the type of the genus, in
having fewer somatic kineties (16 vs 30–38) and a posteriorly
located contractile vacuole and in the absence of ring-like
granules (Foissner et al., 1994; Fig. 6g–j; Table 2).
According to Jankowski (1964), D. elliptica differs from
Dexiotricha raikovi in having fewer somatic kineties (16 vs
20–22) and fewer basal bodies in each kinety (about 16 vs
20–23) and by the subcaudally located (vs near mid-body)
contractile vacuole (Fig. 6k, l; Table 2).
Dexiotricha tranquilla differs from D. elliptica in having
more kineties (20–24 vs 16) and two post-oral kineties/
fragments (Fig. 6a–c; Table 2).
Though no report of its infraciliature is available,
Dexiotricha polystyla can be easily distinguished from D.
elliptica in having multiple caudal cilia (Fig. 6d; Table 2).
previous reports on the general infraciliature and most
characters from living cells. The prominent difference is the
location of the contractile vacuole: in all previous reports,
the contractile vacuole of D. granulosa is located equatorially, while, in our population, it is located subcaudally
(five individuals in vivo), and the contractile vacuole pore
revealed by SEM is also located near the posterior 1/5 (three
individuals). Moreover, the number of somatic kineties
is fewer than in previous reports (28–30 vs 30–38). The
location of the contractile vacuole or excretory pore is
considered to be a stable feature and a very important
character to distinguish species, and the number of somatic kineties has also been taken into consideration for
classification (Fan et al., 2009, 2011a, b; Foissner et al.,
1994). However, in this case, considering the similarities of
all other characters, especially the general infraciliature and
ring-like granules, we are inclined to designate our organism
as D. cf. granulosa for the time being. More information
based on multiple populations along with sequences of the
SSU rDNA are obviously needed.
Remarks on Dexiotricha cf. granulosa
The species D. granulosa has been numerously reported
under several synonyms, and Foissner et al. (1994) synonymized D. plagia and D. media (Fig. 6g–j; Table 2). Considering the great similarities in body size, distinct ring-like
granules, the location of the contractile vacuole and the
infraciliature, the present study supports the synonymy of
the three species. Our population also corresponds well with
(a)
(b)
(g)
(h)
Phylogenetic analysis of the genus Dexiotricha
The order Loxocephalida is not monophyletic, since Cinetochilum clusters with Apostomatia and Dexiotrichides
clusters with the clade comprising Hymenostomatia and
Peritrichia, which corresponds well with previous reports
(Gao et al., 2013; Zhang et al., 2010, 2011). Three Dexiotricha species/populations form a monophyletic group with
(d)
(c)
(i)
(j)
(e)
(k)
(f)
(l)
Fig. 6. Various Dexiotricha species in vivo (a, d, g, k) and after staining (b, c, e, f, i, j, l). (a–c) D. tranquilla (Kahl, 1926) Augustin
& Foissner, 1992 (from Augustin & Foissner, 1992). (d) D. polystyla Foissner, 1987 (from Foissner, 1987). (e, f) D. colpidiopsis
(Kahl, 1926) Jankowski, 1964 (from Fauré-Fremiet, 1968). (g–j) D. granulosa (Kent, 1881) Foissner et al. 1994 [taken from
Foissner et al. (1994) (g), Wilbert (1986) (h, i) and Peck (1974) (j)]. (k, l) D. raikovi Jankowski, 1964 (from Jankowski, 1964).
Bars, 25 mm.
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International Journal of Systematic and Evolutionary Microbiology 64
Morphology and phylogeny of two Dexiotricha species
Table 2. Comparison of Dexiotricha species
CC, Caudal cilia; CV, contractile vacuole; Ma, macronucleus; PK, post-oral kineties; SK, somatic kineties;
Species
D.
D.
D.
D.
D.
D.
granulosa
granulosa
granulosa
cf. granulosa
colpidiopsis
colpidiopsis
D. media
D. raikovi
D. tranquilla
D. polystyla
D. elliptica
*Data from:
CH,
NA,
data not available.
Body size (mm)*
SK (n)
PK (n)
Ma, CV position
CC (n)
61627 (V?)
65–80625–35 (V)
40–80615–30 (V)
45–50615–20 (V)
51624 (V)
51624 (CH)
48624 (P)
55629 (CH)
45 long (P)
50623 (V?)
30–44612–19 (P)
30–60618–25 (V)
50–70 long (V)
45–55620–25 (V)
38
30–35
30–38
28–30
24
26–28
3
3
NA
Mid-body
Mid-body
Mid-body
Subcaudal
Subcaudal
Subcaudal
1
1
1
1
1
1
Jankowski (1964)
Wilbert (1986)
Foissner et al. (1994)
This study
Jankowski (1964)
Peck (1974)
31–34
2
Mid-body
1
Peck (1974)
20–22
20–24
NA
Mid-body
Mid-body
1
1
Jankowski (1964)
Augustin & Foissner (1992)
Mid-body
Subcaudal
Multiple
1
NA
2–4
3
2
NA
NA
16
2–4
Source
Foissner (1987)
This study
Chatton–Lwoff-prepared specimen; P, protargol-prepared specimen; V, specimen observed in vivo.
full support, which then clusters with Cardiostomatella
and Paratetrahymena, suggests the validity of assigning
Dexiotricha to Loxocephalida based on their morphological
characters, e.g. having oblique oral membranelles, post-oral
kineties and scutica (Li et al., 2006; Lynn, 2008). However,
the node supports are quite low, which indicates that
phylogenetic relationships within this order are still
indeterminate. Dexiotricha, Paratetrahymena, Cardiostomatella and Dexiotrichides belong to the family Loxocephalidae (Lynn, 2008), but such an assignment is not well
supported by phylogenetic analyses, since Dexiotrichides
is outside the clade that contains the other three genera.
Moreover, previous morphogenesis studies rejected the
familial assignment, because there is not much similarity
among the morphogenesis of the three genera (data for
Cardiostomatella are not available): Paratetrahymena was
reported to have a ‘monoparakinetal’ pattern, quite like a
hymenostome (Li et al., 2010); Dexiotricha was defined
to have a ‘scuticobuccokinetal’ pattern, but the parental
paroral membrane was not involved in the formation
of the opisthe’s oral primordial (Song et al., 2005); in
contrast, Peck (1974) reported that, in D. media, the
parental paroral membrane took part in the formation
of the opisthe’s oral primordia. Thus, greater taxonomic
sampling as well as more molecular evidence is needed in
order to investigate further the systematic relationships
of the order Loxocephalida and relationships among its
members.
Great thanks are due to the anonymous reviewers for their valuable
comments.
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