European Journal of Taxonomy 191: 1-18
http://dx.doi.org/10.5852/ejt.2016.191
BY
This work is licensed under a Creative Commons Attribution 3.0 License.
ISSN 2118-9773
www. europeanj ournaloftaxonomy. eu
2016 • Pan X. et al.
Research article
urn:lsid:zoobank.org:pub:A36F7A67-A688-4D5B-AD3E-112344E3D3EA
Taxonomy and morphology of four “oph rys- rel a ted ”
scuticociliates (Protista, Ciliophora, Scuticociliatia), with the
description of a new genus, Paramesanophrys gen. nov.
Xuming PAN >, Xinpeng FAN 2 *, Saleh A. AL-FARRAJ 3 , Shan GAO 4 & Ying CHEN 5 -*
15 College of Life Science and Technology, Harbin Normal University, Harbin 150025, China.
12 School of Life Sciences, East China Normal University, Shanghai, 200062, China.
3 Zoology Department, King Saud University, Riyadh 11451, Saudi Arabia.
4 Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao 266003, China,
and Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine
Science and Technology, China.
* Corresponding authors: [email protected] (Xinpeng Fan); [email protected] (Ying Chen)
1 E-mail: [email protected]
3 E-mail: salfarrai@hotmail. com
4 E-mail: [email protected]
1 urn: lsid: zoobank, ore: author :B43 8F 4F 6-95CD-4E3F -BD95-527616F C27C3
2 urn:lsid:zoobank.org:author:AC458497-30FF-411C-8724-D8297B3BE5EA
3 urn:lsid:zoobank.org:author:BA12A34C-2A08-4493-97DA-0BCCF6B7ED36
4 urn:lsid:zoobank.org:author:527DECF 1-6523-4213-B33F-1369F8602C02
5 um:lsid:zoobank.org:author:4FB3E509-lD9C-41EA-906A-lF5BCBC6EDD6
Abstract. Generally, ‘Yp/zr^-related” scuticociliates belong to a specialised group of ciliated protozoa
that may act as commensals or pathogens of fishes and crustaceans. In the present study, four “ ophrys-
taxa” scuticociliates, i.e., Paramesanophrys typica gen. et sp. nov., Mesanophrys carcini (Groliere
& Leglise, 1977) Small & Lynn in Aescht, 2001, Metanophrys sinensis Song & Wilbert, 2000, and
Metanophrys similis Song et al, 2002, were collected from Chinese coastal waters or mariculture ponds
and investigated. Paramesanophrys gen. nov. is assigned to the family Orchitophryidae and differs from
its other genera ma inl y by the position of the paroral membrane relative to membranelle 1-3, i.e., the
membrane extends anteriorly to the posterior end of membranelle 3. The type species P. typica gen.
et sp. nov., is defined by an elongated body with the posterior end depressed where the caudal cilium
is located; 20 or 21 somatic kineties; double-rowed membranelle 1 with eight to ten basal bodies in
each kinety; irregularly multi-rowed membranelle 2 and membranelle 3; scutica comprising c. seven
or eight kinetosome pairs; a single macronuclear nodule; and marine habitat. The redescription of the
three previously known species can be summarized as follows: 1) improved diagnosis is provided for
Metanophrys sinensis Song & Wilbert, 2000 based on the original description and the present study;
2) some population-dependent characteristics of our new Mesanophrys carcini isolate are presented;
3) Metanophrys similis , collected from the South China Sea, resembles the original Qingdao population.
1
European Journal of Taxonomy 191: 1-18 (2016)
Keywords. Scuticociliates, Paramesanophrys gen. nov., Paramesanophrys typica gen. et sp. nov.,
Mesanophrys, Metanophrys.
Pan X., Fan X., Al-Farraj S.A., Gao S. & Chen Y. 2016. Taxonomy and morphology of four "ophrys-vdated”
scuticociliates (Protista, Ciliophora, Scuticociliatia), with the description of a new genus, Paramesanophrys gen.
nov. European Journal of Taxonomy 191: 1-18. http://dx.doi.org/10.5852/eit.2016.191
Introduction
Ciliates in the subclass Scuticociliatia, commonly found in ecosystems worldwide, exhibit a great
biological diversity and play important roles in marine ecosystems (Thompson & Kaneshiro 1968;
Foissner & Wilbert 1981; Cawthorn et al. 1996; Lynn & Struder-Kypke 2005; Fan et al. 2011a, b,
2014; Pan et al. 2013a, b; Castro et al. 2014; Foissner et al. 2014; Zhan et al. 2014). Many of them
are common pathogens of fishes and invertebrates and can cause severe disease or even death in
economically important aquaculture animals (Perez-Uz & Song 1995; Song & Wilbert 2002; Fan et
al. 2009, 2010; Mallo et al. 2014; Ofelio et al. 2014). However, due to their small body size and a high
degree of similarity in ciliature, the taxonomy of this group of organisms remains difficult and confusing
(Thompson 1964; Agatha et al. 1993; Song 2000; Song & Wilbert 2000; Pan et al. 2010). Recent
investigations in Chinese seas have shown a high diversity of scuticociliates, and the discovery of new
Fig. 1 . Sampling sites. A. Coastal waters of the Yellow Sea at Qingdao, Shandong province. B. A coastal
mariculture-region in Zhanjiang, Guangdong province. C. Coastal waters of Daya Bay, Guangdong
province.
2
PAN X. et al. , Descriptions of four marine scuticociliates
Table 1 . Comparison of collection data for four species.
Paramesanophrys
typica gen. et sp. nov.
Mesanophrys
carcini
Metanophrys
similis
Metanophrys
sinensis
Collecting site
Daya Bay, Guangdong
Qingdao, Shandong
Zhanjiang, Guangdong
Zhanjiang, Guangdong
Collecting date
21 Apr. 2011
26 Feb. 2010
6 Nov. 2011
6 Nov. 2011
Habitat
marine
marine
brackish water
brackish water
Temperature (°C)
19
11
21
20
Salinity (%o)
30
31
23
23
pH
7.5
7.3
6.7
6.8
Relative abundance
Low
Low
Low
High
taxa has highlighted the necessity to conduct further studies on this group (Wang et al. 2008a, b, 2009;
Gao et al. 2010, 2012a, b, 2013; Pan etal. 2011, 2015a, b).
The “ophrys- taxa” scuticociliates include species of Mesanophrys Puytorac et al., 1974, Metanophrys
Small & Lynn, 2001 and Paranophrys Thompson & Berger, 1965. Noticeably, they share many
common morphological characteristics, e.g., body usually elongate, oval or cylindrical with a pointed
anterior end but no apical plate; cytostome positioned at, or anterior to, the mid-body; buccal apparatus
comprising a paroral membrane (PM) and three Parauronema-likQ membranelles, membranelle 1 (Ml)
and membranelle 2 (M2) each composed of two or more rows of kinetids. Most species are opportunistic
parasites (Noland 1937; Borror 1963; Groliere & Leglise 1977; Striider & Wilbert 1992; Song &
Wilbert 2000; Song et al. 2002, 2003, 2009; Budino et al. 2011). Among these genera, Metanophrys
and Mesanophrys are the most closely related (body slender, with apical plate absent, cytostome in
the anterior half of the body and three Parauronema-likQ membranelles), with their main difference
being the position of the PM relative to M2 (PM extending anteriorly to the middle portion of M2 in
Metanophrys vs. to the posterior end of M2 in Mesanophrys ) (Small & Lynn 1985; Striider & Wilbert
1992; Song & Wilbert 2000).
In the present study, a new genus, Paramesanophrys gen. nov., is established and detailed morphological
information is provided for four scuticociliates, including one new species, Paramesanophrys typica
gen. et sp. nov., and three nominal “ ophrys ” species.
Material and methods
Paramesanophrys typica gen. et sp. nov. was sampled on 21 Apr. 2011 from the coastal waters of
Daya Bay near Huizhou (22°66'23" N, 114°65'09" E), China (Fig. 1C). Mesanophrys carcini
was collected on 26 Feb. 2010 from the coastal waters off Olympic Sailing Center harbour of
Qingdao (36°06'45" N, 120°39'78" E), China (Fig. 1A). Metanophrys similis and M. sinensis were
collected on 6 Nov. 2010 from the surface water of a coastal shrimp-culturing pond off Zhanjiang
(21°15'01" N, 110°44'04" E), C hin a (Fig. IB). Detailed collection information is given in Table 1. After
isolation, cells were maintained in the laboratory as a uniprotistan culture (Pan et al. 2013a, b).
Cells were observed in vivo using an oil immersion objective with brightfield and Nomarski differential
interference contrast optics. Mixtures of a saturated mercury dichloride solution and Bouin’s fluid were
used to fix samples. The protargol silver staining method (Wilbert & Song 2008; Pan et al. 2013a) was
applied to reveal the infraciliature. Measurements were performed at magnifications of 100-1250x.
Drawings were produced with the help of a camera lucida. Systematics and terminology are mainly used
in accordance with Lynn (2008) and Small & Lynn (1985).
3
European Journal of Taxonomy 191: 1-18 (2016)
We failed to extract DNA from Paramesanophrys typica gen. et sp. nov. due to the low number of
specimens of this species. If possible, we will try to acquire sequence data from it in the future.
Results
Subclass Scuticociliatia Small, 1967
Order Philasterida Small, 1967
Family Orchitophryidae Cepede, 1910
Genus Paramesanophrys gen. nov.
um:lsid:zoobank.org:act:29EF1135-5A4C-4E60-8977-DAEB6EF21370
Diagnosis
Orchitophryidae with cytostome above mid-body; buccal apparatus consisting of three Parauronema-
like membranelles; PM with zigzag structure, extending anteriorly to posterior end of M3; Ml composed
of two rows of kinetids; scutica comprising basal body pairs arranged in a line parallel to somatic
kineties; single caudal cilium.
Type species
Paramesanophrys typica gen. et sp. nov.
Etymology
The generic epithet, Paramesanophrys , refers to the similarity of the oral apparatus to that of the genus
Mesanophrys.
Paramesanophrys typica gen. et sp. nov.
urn:lsid:zoobank.org:act:7B31578B-6C9A-4F80-BAA9-86C9ED9DC735
Figs 2-3, 4A; Table 2
Diagnosis
Size in vivo about 90-100 x 25-35 pm, elongate body, with pointed anterior end and narrowly rounded
caudal end; posterior end distinctly depressed where caudal cilium located; buccal field approximately
40% of body length; 20 or 21 somatic kineties; Ml with 8-10 basal bodies in each kinety; M2 and M3
irregularly multi-rowed; scutica comprising c. seven kinetosome pairs; single macronuclear nodule;
contractile vacuole caudally positioned; marine habitat.
Etymology
The epithet of this new species, typica (Greek, the type/typical, gender feminine), refers to the fact that
it is the type of the new genus, Paramesanophrys gen. nov.
Type locality and ecological features
Coastal waters of Daya Bay (22°66'23" N, 114°65'09" E), Guangdong Province, China, with pH 8.0,
salinity 31%o and water temperature about 16 °C.
Type slides
A protargol slide with the holotype specimen encircled in black ink is deposited in the Laboratory
of Protozoology, Ocean University of China (PXM-2011042101). A paratype slide is deposited in the
Natural History Museum, London, UK (2016.3.10.1).
4
PAN X. et al ., Descriptions of four marine scuticociliates
Description
Size 90-100 x 25-35 pm in vivo , body elongate, spindle-shaped, with pointed anterior end (Figs 2A-B,
3A-D). Posterior end narrowly rounded and distinctly depressed in middle of caudal margin at bottom
of caudal cilium (Figs 2A, 3A, F). Buccal field approximately 40% of body length; shape of buccal
cavity frequently changed from “falcate-shaped” to oval to circular, then conversed (Figs 2C, 3G, I-N).
Pellicle slightly indented at bases of cilia (Figs 2F, 3E, H). Extrusomes spindle-shaped, c. 2-4 pm long
(Fig. 2F). Cytoplasm colourless to greyish, containing several to many large ( c . 5 pm across) food
vacuoles filled with bacteria, often concentrated in anterior and posterior ends of body (Figs 2A, E, 3E,
H). Single ellipsoid to spherical macronucleus, c. 15 pm across, no micronucleus observed (Fig. 3R).
Contractile vacuole caudally located, approximately 8 pm across during diastole, pulsating at intervals
of approximately 30 s (Figs 2A, 3F). Somatic cilia, approximately 10 pm long, densely arranged; single
caudal cilium approximately 30 pm long (Figs 2A, 3E-F). Movement by swimming while rotating about
long body axis without pause or by gliding on substrate (Fig. 2D).
Twenty or 21 somatic kineties, extending entire length of body and consisting of dikinetids in most of
body and monokinetid in rest of body (Figs 2G-H, 3S). Buccal apparatus (Figs 21, 30-Q) consisting of
PM and three Parauronema-like membranelles. Ml composed of two rows of kinetids with 8-10 basal
bodies each (Figs 21, 3Q). M2 and M3 irregularly multi-rowed. M3 much shorter than M2 (Fig. 30-Q).
Fig. 2. Paramesanophrys typica gen. et sp. nov., from life (A-F) and after protargol staining (G-I).
A. Ventral view of a representative individual. B. Different body shapes. C. Changing shapes of buccal
field of the same individual. D. Movement trace. E. Food granules. F. Part of pellicle, to show extrusomes.
G-H. Ventral (G) and dorsal (H) views of the same specimen (holotype), showing infraciliature and
nuclear apparatus. I. Detailed structure of the buccal area. Abbreviations: Ml-3 = membranelles 1, 2
and 3; Ma = macronucleus; PM = paroral membrane; Sc = scutica. Scale bars: A = 30 pm; B = 40 pm.
5
European Journal of Taxonomy 191: 1-18 (2016)
Table 2. Morphometric characterization of Paramesanophrys typica gen. et sp. nov. (Typ), Mesanophrys
carcini (Groliere & Leglise, 1977) Small & Lynn in Aescht, 2001 (Car), Metanophrys sinensis Song &
Wilbert, 2000 (Sin) and Metanophrys similis Song et al, 2002 (Sim).
Character
Species
Min
Max
Mean
M
SD
CV
n
Body length (pm)
Typ
96
113
105.1
107
9.7
9.2
20
Car
42
74
56.9
60
6.7
12.1
25
Sin
33
59
43.6
41
14.4
27.1
23
Sim
47
75
61.7
57
5.2
16.4
23
Body width (pm)
Typ
32
38
34.7
35
5.8
3.9
20
Car
40
50
43.1
43
5.8
11.7
25
Sin
16
24
19.9
20
1.9
9.9
23
Sim
35
52
44.7
45
7.3
17.6
23
Number of somatic kineties
Typ
20
21
20.4
20
1.3
6.1
18
Car
10
11
10.6
11
0.8
7.8
22
Sin
10
11
10.1
10
0.6
5.9
20
Sim
17
17
17
17
3.5
20.6
21
Length of buccal field (pm)
Typ
37
41
38.4
39
3.7
9.8
18
Car
21
24
23.1
23
1.2
9.4
25
Sin
15
26
19.6
19
1.6
8.7
23
Sim
34
43
37.3
38
1.3
3.4
19
Macronucleus, length (pm)
Typ
13
16
14.7
15
2.8
10.5
18
Car
10
14
12.8
12
1.5
12.6
24
Sin
5
6
5.7
6
0.3
5.8
21
Sim
8
11
9.5
9
1.9
21.6
22
Macronucleus, width (pm)
Typ
12
15
13.4
13
1.4
10.9
18
Car
11
13
12.1
12
0.8
6.8
24
Sin
6
7
6.3
6
1.3
20.4
21
Sim
7
10
8.6
9
0.4
2.8
22
Number of basal bodies in somatic kinety 1* *
Typ
22
24
23.2
23
1.1
4.8
17
Car
35
40
37.4
37
4.6
9.9
24
Sin
32
36
33.9
34
8.1
24.4
20
Sim
23
26
24.1
24
4.9
2.1
22
Number of basal bodies in membranelle 1
Typ
8
10
8.6
9
1.2
12.4
14
Car
7
9
8.0
8
0.4
5.0
19
Sin
7
10
8.4
8
1.3
16.6
15
Sim
6
6
6.0
6
0
0
14
Number of scutia pairs
Typ
7
8
7.4
7
0.6
8.5
14
Car
4
4
4.0
4
0
0
19
Sin
4
5
4.4
4
0.2
5.2
15
Sim
5
7
6.1
6
0.3
5.1
14
Abbreviations: CV = coefficient of variation in %; M = median; Max = maximum; Mean = arithmetic mean; Min
= minimum; n = number of individuals examined; SD = standard deviation.
* Basal body pairs counted as single units.
6
PAN X. et al ., Descriptions of four marine scuticociliates
PM with paired basal bodies organized in zigzag pattern, extending anteriorly to posterior end of M3
(Figs 2G, I, 30, Q). Scutica located at posterior end of PM, comprising c. seven or eight kinetosome
pairs aligned in line parallel to somatic kineties (Figs 2G, I).
Mesanophrys carcini (Groliere & Leglise, 1977) Small & Lynn in Aescht, 2001
Figs 4B, 5; Table 2
Small & Lynn in Aescht (2001) did not formally combine this species with Mesanophrys Small &
Lynn in Aescht, 2001. However, since they fixed it as the type species, they automatically produced the
combination.
Fig. 3. Paramesanophrys typica gen. et sp. nov., from life (A-N) and after protargol staining (O-S).
A. Ventral view of a representative individual. B-E. Different individuals; arrowhead in B shows
caudal cilium, arrowheads in E mark somatic cilia. F. Posterior region of cell; arrow shows contractile
vacuole and arrowhead marks caudal cilium. G. Anterior region of cell; arrowhead marks buccal region.
H. Ventral view, showing food vacuoles (arrowhead). I-N. Ventral views, to show various shapes
of buccal regions (arrowheads). O-Q. Detailed infraciliature of buccal area (P from holotype).
R. Macronucleus. S. Posterior region; arrowheads show dikinetids of somatic kineties. Abbreviations:
Ml-3 = membranelles 1,2 and 3; Ma = macronucleus; PM = paroral membrane. Scale bars: A,E = 40 pm;
B-D = 70 pm; G, M-N =10 pm.
7
European Journal of Taxonomy 191: 1-18 (2016)
Some characteristics, e.g., a larger body size and fewer somatic kineties, were found in the Qingdao
population. Hence, a description of the Qingdao population as well as a comparison between different
populations are supplied.
Par ante sanophrys Mesanophrys
Uronema
c
Uronemella Metanophrys
Anophryoides
F
Philasterides
G
Paranophrys
*
. 1
i
t
t
H
Paralembus
Cohnilembus
\
i :• % *
i j
Fig. 4. Comparisons among different buccal apparatus patterns of Paramesanophrys gen. nov. and some
related genera; arrows in A-J show different positions to which PM extends anteriorly and highlighted
structures in A-J mark M2. A. Parame sanophrys typica gen. et sp. nov. (from the present work).
B. Mesanophrys carcini (Groliere & Leglise, 1977) Small & Lynn in Aescht, 2001 (from Song &
Wilbert 2000). C. Uronema marinum Dujardin, 1841 (from Song et al. 2009). D. Uronemella filificum
(Kahl, 1931) Song & Wilbert, 2002 (from Song & Wilbert 2002). E. Metanophrys sinensis Song &
Wilbert, 2000 (from Song & Wilbert 2000). F. Anophryoides haemophila Cawthorn et al., 1996 (from
Cawthorn et al. 1996). G. Philasterides armatalis Song, 2000 (from Song 2000). H. Paranophrys marina
Thompson & Berger, 1965 (from Song et al. 2002). I. Paralembus digitiformis Kahl, 1931 (from Song
& Wilbert 2000). J. Cohnilembus verminus (Muller, 1786) Kahl, 1933 (from Song 2000). Abbreviations:
Ml-3 = membranelles 1, 2 and 3; PM = paroral membrane; Sc = scutica.
8
PAN X. et al ., Descriptions of four marine scuticociliates
Description of Qingdao population
Body size 45-65 x 15-25 pm in vivo , spindle-shaped to long fusiform, with pointed anterior end and
narrowly rounded caudal end (Fig. 5A-C). Body shape variable, likely due to nutritional conditions or
stage in life cycle: from slender, spindle-like to pyriform (Fig. 5C-D). Buccal field short and narrow,
with length of about 30% of body (Fig. 5B). Somatic cilia densely arranged and about 6-8 pm long
(Fig. 5B). Pellicle thin and smooth, with no distinguishable extrusomes. Cytoplasm colourless to
slightly greyish, containing several to many differently-sized (3-5 pm) refringent granules (Fig. 5A, D).
Single caudal cilium about 15 pm in length (Fig. 5B, arrow) and one large, spherical, centrally located
macronucleus; one micronucleus closely associated with macronucleus. Contractile vacuole small
(5 pm across), terminally positioned and pulsating at intervals of approximately 30 s (Fig. 5A, arrow).
Movement by continuous swimming in water without pause or gliding slowly on substrate.
Ten or 11 somatic kineties, consisting of dikinetids in anterior two-thirds and monok in etid in posterior
third of body (Fig. 5G, arrow). Ml slightly separated from apex, composed of two rows of kinetids with
Fig. 5. Mesanophrys carcini Small & Lynn in Aescht, 2001, in vivo (A-D) and after protargol staining
(E-G). A. Ventral view of a representative individual; arrow shows contractile vacuole. B-D. Ventral
views of four individuals; arrow in B shows caudal cilium and arrow in D marks food vacuole.
E-F. Ventral views, detailed structure of buccal area. G. Dorsal view; arrow indicates somatic kinety.
Abbreviations: Ml—3 = membranelles 1, 2 and 3; Ma = macronucleus; PM = paroral membrane;
Sc = scutica. Scale bars: A-D = 30 pm; E-G = 5 pm.
9
European Journal of Taxonomy 191: 1-18 (2016)
7-9 basal bodies each (Fig. 5E-F). M2 composed of five or six longitudinal rows, each containing about
6-8 basal bodies (Fig. 5E-F). M3 located close to M2, much shorter than M2 and composed of three
short, irregularly arranged rows of kinetosomes (Fig. 5E-F). PM extending anteriorly to posterior end
of M2. Scutica Y-shaped, with c. four pairs of kinetosomes (Fig. 5E-F).
Ecological features
Salinity 32%o, pH 7.9 and water temperature about 11 °C.
Metanophrys sinensis Song & Wilbert, 2000
Figs 4E, 6; Table 2
This species was described by Song & Wilbert (2000) in detail based on their Qingdao population. In the
current work, it is reported for the first time from South China Sea. An improved diagnosis is provided
herein based on all these data; the improved parts are highlighted in bold.
Improved diagnosis
Slender to elongated oval body shape; in vivo about 25-50 x 10-20 pm with pointed anterior end;
buccal field about 30%-50% of body length; mostly ten somatic kineties, of which somatic kinety 1
consists of c. 35 basal pairs; Ml composed of two rows, each with 7-10 kinetosomes, longer than M2;
two-rowed M2; contractile vacuole pore located near posterior end of kinety 2; extrusomes present;
marine habitat.
Description of Zhanjiang population
Body 25-30 x 10-15 pm in vivo , usually elongate-oval in outline, with anterior end distinctly pointed
and posterior rounded (Fig. 6A-B). Body asymmetrical in outline when viewed ventrally, with anterior
end slightly curved sideways (Fig. 6A-B). Ventral side almost straight, while dorsal side convex. Buccal
field 2 /5 to l A of body length, with cytostome located anterior to equatorial plane of body (Fig. 6C). Cilia
densely packed, about 7-8 pm long. Caudal cilium about 15 pm in length (Fig. 6B). Pellicle thin and
slightly notched, with extrusomes about 2-3 pm long and dense beneath cortex (Fig. 6D). Endoplasm
colourless to greyish, containing several food vacuoles and bar- or dumbbell-like crystals, which are
usually 3 pm long and located in anterior and posterior regions of body (Fig. 6A-B, G). One large round
to oval macronucleus approximately centrally located, with many small, irregularly shaped nucleoli
on surface. Contractile vacuole about 5 pm in diameter and caudally positioned near ventral side
(Fig. 6B). Movement with no special features, including swimming moderately fast, sometimes
continuously swimming in water without pause.
Ten somatic kineties arranged longitudinally, and dikinetids about 3 A of length of each in anterior part
(Fig. 6J). Buccal apparatus consists of three Parauronema -like membranelles (Fig. 6E-F). Ml slightly
below apex and composed of two rows of kinetids with 7-10 basal bodies each, and longer than M2.
M2 two-rowed, containing about five basal bodies in each row. M3 located close to M2, normally with
three short, obliquely arranged rows of basal bodies. Scutica Y-shaped, with several pairs of kinetosomes
(Fig. 61). Silverline system in quadrangular mesh-pattem (Fig. 6H). Contractile vacuole pore located
near posterior end of kinety 2.
Ecological features
Salinity 21%o, pH 7.3 and water temperature 26 °C.
10
PAN X. et al ., Descriptions of four marine scuticociliates
Metanophrys similis Song et al ., 2002
Fig. 7; Table 2
This species is reported for the first time from the South China Sea. The morphological and behavioural
characteristics of the Zhanjiang population closely resemble those of the Qingdao population; therefore,
only the morphometric characterisation and important features are supplied.
Fig. 6. Metanophrys sinensis Song & Wilbert, 2000, in vivo (A-D, G) and after protargol (E-F, I-J) or
silver nitrate (H) staining. A. Ventral view of a typical individual. B. Ventral view of another individual;
arrowheads mark somatic cilia. C. Ventral view; arrowhead exhibits buccal field. D. Notched pellicle
(arrowhead). E. Detailed structure of buccal area. F. Individual in morphogenesis, to show buccal
apparatus. G. Ventral view, showing bar-shaped crystal (arrowhead). H. Detail of somatic kinetids.
I. Dikinetids of scutica (arrowheads). J. Posterior region; arrowheads show monokinetids of so¬
matic k in eties Abbreviations: Ml-3 = membranelles 1, 2 and 3; PM = paroral membrane. Scale bars:
A-B =15 pm.
11
European Journal of Taxonomy 191: 1-18 (2016)
Description of Zhanjiang population
Body in vivo about 35-40 x 20-25 pm, plump pyriform, tapering anteriorly and rounded posteriorly,
and no apical plate formed (Fig. 7A-C). Ventral side almost straight, while dorsal side slightly convex
(Fig. 7A). Length: width ratio approximately 2:1 (Fig. 7A-C). Buccal field occupies about 40% of total
body length, with buccal cilia about 5-8 pm in length. Pellicle thick and strongly notched. Somatic
cilia about 7-8 pm long and densely arranged (Fig. 7D, arrowheads). Single caudal cilium about 15 pm
in length (Fig. 7F). Extrusomes, about 2 pm in length, arranged in rows between somatic kineties.
Endoplasm colourless to greyish and contains abundant food vacuoles (Fig. 7E). One large, spherical
to ovoid macronucleus centrally located (Fig. 7C, J). Contractile vacuole about 5 pm in diameter and
caudally positioned near ventral side (Fig. 7A).
Fig. 7. Metanophrys similis Song et al ., 2002, in vivo (A-F) and after protargol staining (G-J). A. Ventral
view of a typical individual. B-D. Ventral views of three individuals; arrowheads in D mark somatic
cilia. E. Food vacuole (arrow). F. Posterior region; arrow shows caudal cilium. G, I. Ventral views, to
show detailed structure of the buccal area. H. Dorsal view; arrow shows monokinetids, arrowhead marks
dikinetids. J. Macronucleus. Abbreviations: Ml-3 = membranelles 1, 2 and 3; Ma = macronucleus;
PM = paroral membrane. Scale bars: A-D = 30 pm.
12
PAN X. et al ., Descriptions of four marine scuticociliates
Locomotion by swimming moderately fast, sometimes continuously without pause, or by crawling on
substrates.
Twelve somatic kineties with dikinetids arranged in approximately anterior half of each row and
monokinetids positioned posteriorly (Fig. 7H). Ml positioned near apex and comprised of three
longitudinal rows of kinetids with six basal bodies each (Fig. 7G, I). M2 three-rowed, as long as Ml and
also composed of about six basal bodies in each longitudinal row (Fig. 7G, I). M3 located close to M2
and normally comprised of three short, parallel arranged rows of basal bodies (Fig. 7G, I). PM extends
to about anterior third of body (Fig. 7G). Scutica, with about 5-7 basal bodies, arranged in long line.
Ecological features
Salinity 21%o, pH 7.3 and water temperature 26 °C.
Discussion
About Paramesanophrys gen. nov. and P. typica gen. et sp. nov.
The family Orchitophryidae is characterised as follows: small- to medium-sized body; ovoid-shaped;
caudal cilium often present; oral region in anterior A to % of body; scutica aligned along midventral
postoral region; bacterivorous and histophagous; marine habitats, always as facultative parasites of
crustaceans, asteroids, fish and free-swimming (Lynn 2008). Paramesanophrys gen. nov. should be
assigned to Orchitophyridae based on its morphological characters and habitat.
Hitherto, five genera have been assigned to Orchitophryidae according to Lynn (2008), namely
Anophryoides de Puytorac & Groliere, 1979, Mesanophrys Small & Lynn in Aescht, 2001, Metanophrys
Puytorac et al ., 1974, Orchitophrya Cepede, 1907 and Paranophrys Thompson & Berger, 1965.
Compared with these related genera, Paramesanophrys gen. nov. has a unique oral apparatus, with the
PM extending anteriorly to the posterior end of M3 (vs. PM extending anteriorly to the anterior end/
middle portion/posterior end of M2; Fig. 4) (Small & Lynn 1985; Striider & Wilbert 1992; Cawthorn et
al. 1996; Song & Wilbert 2000).
Besides having a unique Paramesanophrys-typQ PM, Paramesanophrys typica gen et sp. nov. also has
the scutica comprising c. seven or eight kinetosome pairs aligned in a line parallel to the somatic kineties
and a conspicuous pellicle depression in the middle of caudal margin. This combination of features
clearly separates it from all known scuticociliates at the species level.
Mesanophrys carcini (Groliere & Leglise, 1977) Small & Lynn in Aescht, 2001
The main characteristics that aid in identifying this species are the slender body, short buccal field,
oral apparatus and somatic infraciliature (Song & Wilbert 2000). The characteristics of the Qingdao
population are different from those of the population reported by Song & Wilbert (2000) in having a
larger body size (on average 55 x 20 pm vv 40 x 12 pm) and a variable number of somatic kineties
(10 or 11 vv constantly 11; Table 3). These variations are considered population-dependent (Song &
Wilbert 2000).
Metanophrys sinensis Song & Wilbert, 2000
Our population is virtually identical to the two Qingdao populations (Song & Wilbert 2000; Ma &
Song 2003), that is, they agree in body size and shape, habitat, infraciliature, silverline system and
marine habitat, except the proportion of buccal field length to body length (40-50% in the present study
vs. ca. 30—40% in the previous studies) and the presence of extrusomes (vs. not observed in previous
descriptions) (Song & Wilbert 2000; Ma & Song 2003; Table 3). Nevertheless, it is believed that they are
conspecific because of their close similarity in other living characteristics and infraciliature.
13
European Journal of Taxonomy 191: 1-18 (2016)
Table 3. Morphometrical comparison of known ophrys- species populations. Abbreviations: QD,
Qingdao or Qingdao population; ZJ, Zhanjiang or Zhanjiang population.
Mesanophrys
carcini
Mesanophrys
carcini QD
Metanophrys
similis
Metanophrys
similis ZJ
Metanophrys
sinensis
Metanoph/ys
sinensis QD
Metanophrys
sinensis ZJ
Sample location
shrimp culture
coastal waters,
coastal waters,
maricultural
molluscan culture
molluscan culture
mariculture
pond, QD
QD
QD
waters, ZJ
pond, QD
pond, QD
waters, ZJ
Body size in vivo
(pm)
30-55 x 10-15
45-65 x 15-25
30-45 x 10-12
35-40 x 20-25
30-50 x 10-20
30-50 x 10-20
25-30 x 10-15
Somatic kineties
(no.)
11 or 12
10 or 11
11 or 12
12
10
10 or 11
10
Buccal length/
body length (%)
25-33
30
40-50
40
40
35-40
40-50
Extrusome
not observed
not observed
present
present
not observed
not observed
present
Data source
Song & Wilbert
(2000)
present study
Song et al.
(2002)
present study
Song & Wilbert
(2000)
Ma & Song
(2003)
present study
Metanophrys similis Song et al ., 2002
The Zhanjiang population is identical to the original description (Song et al. 2002) according to the
body size, ciliature and habitat; hence, the identity of this species is not in doubt. Compared with the
original description, the population described in this paper has a different body shape (plump pyriform
vs*, slender body shape in Song et al. 2002; Table 3), which may be due to different nutritional conditions
(Song et al. 2002).
Acknowledgments
This work was supported by the Natural Science Foundation of China (project numbers: 31471973,
31501844, 31470064). The authors extend their sincere appreciation to the Deanship of Scientific
Research at King Saud University for its funding of this Prolific Research Group (PRG-1436-24). Many
thanks are given to the reviewers for their helpful suggestions to this manuscript.
References
Agatha S., Spindler M. & Wilbert N. 1993. Ciliated protozoa (Ciliophora) from Arctic sea ice. Acta
Protozoologica 32: 261-268.
Aescht E. 2001. Catalogue of the Generic Names of Ciliates (Protozoa, Ciliophora). Biologiezentrum
des Oberosterreichischen Tandesmuseum, Tinz, Austria.
Borror A.C. 1963. Morphology and ecology of some uncommon ciliates from Alligator Harbor, Florida.
Transactions of the American Microscopical Society 82: 125-131. http://dx.doi.org/10.2307/3223987
Budino B., Tamas J., Pata M., Arranz J., Sanmartin MX. & Leiro J. 2011. Intraspecific variability in
several isolates of Philasterides dicentrarchi (syn. Miamiensis avidus), a scuticociliate parasite of farmed
turbot. Veterinary Parasitology 175: 260-272. http://dx.doi.org/doi: 10.1016/j.vetpar.2010.10.011
Castro T.A., Kiippers G.C., Schweikert M., Harada M.L. & Paiva T.S. 2014. Ciliates from eutrophized
water in the northern Brazil and morphology of Cristigera hammeri Wilbert, 1986 (Ciliophora,
Scuticociliatia). European Journal of Protistology 50: 122-133. http://dx.doi.org/10.1016/i.
eiop.2014.01.005
14
PAN X. et al. , Descriptions of four marine scuticociliates
Cawthorn R.J., Lynn D.H., Despres B., MacMillan R., Maloney R., Loughlin M. & Bayer R. 1996.
Description of Anophryoides haemophila n. sp. (Scuticociliatida: Orchitophryidae), a pathogen of
American lobsters Homarus americanus. Diseases of Aquatic Organisms 24: 143-148. http://dx.doi.
org/10.3354/dao024143
Fan X., Miao M., Al-Rasheid K.A.S. & Song W. 2009. Anew genus of marine scuticociliate (Protozoa,
Ciliophora) from northern China, with a brief note on its phylogenetic position inferred from small
subunit ribosomal DNA sequence data. Journal of Eukaryotic Microbiology 56: 577-582. http://dx.doi.
org/10.1111/i. 1550-7408.2009.00436.x
Fan X., Chen X., Song W., Al-Rasheid K.A.S. & Warren A. 2010. Two new marine scuticociliates,
Sathrophilus planus n. sp. and Pseudoplatynematum dengi n. sp., with improved definition of
Pseudoplatynematum (Ciliophora, Oligohymenophora). European Journal of Protistology 46: 212-220.
http://dx.doi.Org/10.1016/i.eiop.2010.03.002
Fan X., Hu X., Al-Farraj S.A., Clamp J.C. & Song W. 2011a. Morphological description of three marine
ciliates (Ciliophora, Scuticociliatia), with establishment of a new genus and two new species. European
Journal of Protistology 47: 186-196. http://dx.doi.Org/10.1016/i.eiop.2011.04.001
Fan X., Lin X., Al-Rasheid K.A.S., Al-Farraj S.A., Warren A. & Song W. 2011b. The diversity of
scuticociliates (Protozoa, Ciliophora): a report on eight marine forms found in coastal waters of China,
with a description of one new species. Acta Protozoologica 50: 219-234. http://dx.doi.org/10.4467/168
90027AP.1L 021,0021
Fan X., Al-Farraj S.A., Gao F. & Gu F. 2014. Morphological reports on two species of Dexiotricha
(Ciliophora, Scuticociliatia), with a note on the phylogenetic position of the genus. International Journal
of Systematic and Evolutionary Microbiology 64: 680-688. http://dx.doi.Org/10.1099/ijs.0.059899-0
F oissner W. & Wilbert N. 1981. A comparative study of the infraciliature and silverline system of the fresh¬
water scuticociliates Pseudocohnilembus putrinus (Kahl, 1928) nov. comb., P. pusillus (Quennerstedt,
1869) nov. comb., and the marine form P. marinus Thompson, 1966. Journal of Protozoology 28: 291—
297. http://dx.doi.Org/10.llll/i.1550-7408.1981.tb02853.x
Foissner W., Jung J.H., Filker S., Rudolph J. & Stoeck T. 2014. Morphology, ontogenesis and molecular
phylogeny of Platynematum salinarum nov. spec., a new scuticociliate (Ciliophora, Scuticociliatia)
from a solar saltern. European Journal of Protistology 50: 174-184. http://dx.doi.org/10.1016/j.
eiop.2013.10.001
Gao F., Fan X., Yi Z., Striider-Kypke M. & Song W. 2010. Phylogenetic consideration of two
scuticociliate genera, Philasterides and Boveria (Protozoa, Ciliophora) based on 18S rRNA gene
sequences. Parasitology International 59: 549-555. http://dx.doi.Org/10.1016/i.parint.2010.07.002
Gao F., Katz L.A. & Song W. 2012a. Insights into the phylogenetic and taxonomy of philasterid ciliates
(Protozoa, Ciliophora, Scuticociliatia) based on analyses of multiple molecular markers. Molecular
Phylogenetics and Evolution 64: 308-317. http://dx.doi.Org/10.1016/i.vmpev.2012.04.008
Gao F., Striider-Kypke M., Yi Z., Miao M., Al-Farraj S.A. & Song W. 2012b. Phylogenetic analysis
and taxonomic distinction of six genera of pathogenic scuticociliates (Protozoa, Ciliophora) inferred
from small-subunit rRNA gene sequences. International Journal of Systematic and Evolutionary
Microbiology 62: 246-256. http://dx.doi.Org/10.1099/ijs.0.028464-0
Gao F., Katz L.A. & Song W. 2013. Multigene-based analyses on evolutionary phylogeny
of two controversial ciliate orders: Pleuronematida and Loxocephalida (Protista, Ciliophora,
15
European Journal of Taxonomy 191: 1-18 (2016)
Oligohymenophorea). Molecular Phylogenetics and Evolution 68: 55-63. http://dx.doi.org/10.1016/j.
ympev.2013.03.018
Groliere C.A. & Leglise M. 1977. Paranophrys carcini n. sp., Cilie Philasterina recolte dans
Fhemolymphe du crabe Cancer pagurus Linne. Protistologica 13: 503-507.
Lynn D.H. 2008. The Ciliated Protozoa. Characterization, Classification and Guide to the Literature.
Springer Verlag, Dordrecht.
Lynn D.H. & Striider-Kyplce M. 2005. Scuticociliate endosymbionts of echinoids (phylum
Echinodermata): phylogenetic relationships among species in the genera Entodiscus, Plagiopyliella ,
Thyrophylax , and Entorhipidium (phylum Ciliophora). The Journal of Parasitology 91: 1190-1199.
http://dx.doi.org/10.1645/GE-445R. 1
Ma H. & Song W. 2003. Stomatogenesis of the marine ciliate Metanophrys sinensis (Protozoa:
Ciliophora: Scuticociliatida). Journal of the Marine Biological of Association of the United Kingdom
83: 407M10. http://dx.doi.org/10.1017/S00253154030Q7252h
Mallo N., Lamas J., Piazzon C. & Leiro J.M. 2014. Presence of a plant-like proton-translocating
pyrophosphatase in a scuticociliate parasite and its role as a possible drug target. Parasitology 142:
449M62. http://dx.doi.org/10.1017/S00311820140Q1267
Noland L.E. 1937. Observations on marine ciliates of the Gulf Coast of Florida. Transactions of the
American Microscopical Society 56: 160-171. http://dx.doi.org/10.2307/3222944
r
Ofelio C., Blanco A., Roura A., Pintado J., Pascual S. & Planas M. 2014. Isolation and molecular
identification of the scuticociliate Porpostoma notata Moebius, 1888 from moribund reared
Hippocampus hippocampus (L.) seahorses, by amplification of the SSU rRNA gene sequences. Journal
of Fish Diseases 37: 1061-1065. http://dx.doi.org/10.llll/jfd.12207
Pan H., Huang J., Hu X., Fan X., Al-Rasheid K.A.S. & Song W. 2010. Morphology and SSU rRNA
gene sequences of three marine ciliates from Yellow Sea, China, including one new species, Uronema
heteromarinum nov. spec. (Ciliophora, Scuticociliatida). Acta Protozoologica 49: 45-49.
Pan X., Shao C., Ma H., Fan X., Al-Rasheid K.A.S., Al-Farraj S.A. & Hu X. 2011. Redescriptions of
two marine scuticociliates from China, with notes on stomatogenesis in Parauronema longurn
(Ciliophora, Scuticociliatida). Acta Protozoologica 50: 301-310. http://dx.doi.org/10.4467/16890Q27
API! ,027.0064
Pan X., Bourland W.A. & Song W. 2013a. Protargol synthesis: An in-house protocol. Journal of
Eukaryotic Microbiology 60: 609-614. http://dx.doi.org/10.llll/jeu.12067
Pan X., Gao F., Liu W., Fan X., Warren A. & Song W. 2013b. Morphology and SSU rRNA gene sequences
of three Frontonia species, including a description of F. subtropica spec. nov. (Ciliophora, Peniculida).
European Journal of Protistology 49: 61-11. http://dx.doi.Org/10.1016/j.eiop.2012.05.002
Pan X., Huang J., Fan X., Ma H., Al-Rasheid K.A.S., Miao M. & Gao F. 2015a. Morphology and
phylogeny of four marine scuticociliates (Protista, Ciliophora), with descriptions of two new species:
Pleuronema elegans spec. nov. and Uronema orientalis spec. nov. Acta Protozoologica 54: 31-43.
http://dx.doi.org/10.4467/16890027AP15.003.219Q
Pan X., Yi Z., Li J., Ma H., Al-Farraj S.A. & Al-Rasheid K.A.S. 2015b. Biodiversity of marine
scuticociliates (Protozoa, Ciliophora) from China: description of seven morphotypes including a new
species, Philaster sinensis spec. nov. European Journal of Protistology 51: 142-157. http://dx.doi.
org/10.1016/i.eiop.2015.02.005
16
PAN X. et al ., Descriptions of four marine scuticociliates
Perez-Uz B. & Song W. 1995. Uronema gallicum sp. n. (Protozoa: Ciliophora) a new marine scuti-
cociliate from the coastal area of Calais. ActaProtozoologica 34: 143-149.
Small E.B. & LynnD.H. 1985. Phylum Ciliophora Doflein, 1901. In: Lee J.J., Hunter S.H. & Bovee E C.
(eds) An Illustrated Guide to the Protozoa : 393-575. Society of Protozoologists, Lawrence, Kansas.
Song W. 2000. Morphological and taxonomical studies on some marine scuticociliates from China Sea,
with description of two new species, Philasterides armatalis sp. n. and Cyclidium varibonneti sp. n.
(Protozoa: Ciliophora: Scuticociliatida). Acta Protozoologica 39: 295-322.
Song W. & Wilbert N. 2000. Redefinition and redescription of some marine scuticociliates from
China, with report of a new species, Metanophrys sinensis nov. spec. (Ciliophora, Scuticociliatida).
Zoologischer Anzeiger 239: 45-74.
Song W. & Wilbert N. 2002. Reinvestigations of three “well-known” marine scuticociliates: Uronemella
filificum (Kahl, 1931) nov. gen., nov. comb., Pseudocohnilembus hargisi Evans & Thompson, 1964 and
Cyclidium citrullus Cohn 1865, with description of the new genus Uronemella (Protozoa, Ciliophora,
Scuticociliatida). Zoologischer Anzeiger 241: 317-331. http://dx.doi.org/10.1078/0044-5231-00Q75
Song W., Shang H., Chen Z. & Ma H. 2002. Comparison of some closely-related Metanophrys- taxa with
description of a new species Metanophrys similis nov. spec. (Ciliophora, Scuticociliatida). European
Journal of Protistology 38: 45-53. http://dx.doi.org/10.1078/0932-4739-0Q848
Song W., Zhao Y., Xu K., Hu X. & Gong J. 2003. Pathogenic Protozoa in Mariculture. Science Press,
Beijing.
Song W., Warren A. & Hu X. 2009. Free-living Ciliates in Bohai and Yellow Sea, China. Science Press,
Beijing.
Striider M. & Wilbert N. 1992. Contribution to the taxonomy of the family Paranophryidae Jankowski
in Small & Lynn, 1985. Acta Protozoologica 31: 33-37.
Thompson Jr. J.C. 1964. A redescription of Uronema marinum, and a proposed new family Uronematidae.
The Virginia Journal of Science 15: 80-87.
Thompson Jr. J.C. & Kaneshiro E.S. 1968. Redescriptions of Uronema filificum and U. elegans. Journal
of Protozoology 15: 141-144. http://dx.doi.org/10. Ill 1/i. 1550-7408.1968.tb02099.x
Wang Y., Hu X., Long H., Al-Rasheid K.A.S., Al-Parraj S. & Song W. 2008a. Morphological studies
indicate that Pleuronema grolierei nov. spec, and P. coronatum Kent, 1881 represent different sections
of the genus Pleuronema (Ciliophora: Scuticociliatida). European Journal of Protistology 44: 131-140.
http://dx.doi.Org/10.1016/j.ejop.2007.08.008
Wang Y., Song W., Hu X., Warren A., Chen X. & Al-Rasheid K.A.S. 2008b. Descriptions of two new
marine species of Pleuronema, P. czapikae sp. n. and P. wiackowskii sp. n. (Ciliophora: Scuticociliatida),
from the Yellow Sea, North China. Acta Protozoologica 47: 35—45.
Wang Y., Song W., Warren A., Al-Rasheid K.A.S., Al-Quraishy S., Al-Larraj S.A., Hu X. & Pan H.
2009. Descriptions of two new marine scuticociliates, Pleuronema sinica n. sp. and P. wilberti n. sp.
(Ciliophora: Scuticociliatida), from the Yellow Sea, China. European Journal of Protistology 45: 29-37.
http://dx.doi.Org/10.1016/i.eiop.2008.06.001
Wilbert N. & Song W. 2008. A further study on littoral ciliates (Protozoa, Ciliophora) near King George
Island, Antarctica, with description of a new genus and seven new species. Journal of Natural History
42: 979-1012. http://dx.doi.org/10.1080/0022293070187754Q
17
European Journal of Taxonomy 191: 1-18 (2016)
ZhanZ., StoeckT., DunthomM. &XuK. 2014. Identification of the pathogenic ciliat e Pseudocohnilembus
persalinus (Oligohymenophorea: Scuticociliatia) by fluorescence in situ hybridization. European
Journal of Protistology 50: 16-24. http://dx.doi.Org/10.1016/j.ejop.2013.09.004
Submitted: 30 September 2015
Accepted: 2 January 2016
Published: 18 April 2016
Topic editor: Rudy Jocque
Desk editor: Danny Eibye-Jacobsen
Printed versions of all papers are also deposited in the libraries of the institutes that are members of the
EJT consortium: Museum national d’Histoire naturelle, Paris, France; Botanic Garden Meise, Belgium;
Royal Museum for Central Africa, Tervuren, Belgium; Natural History Museum, London, United
Kingdom; Royal Belgian Institute of Natural Sciences, Brussels, Belgium; Natural History Museum of
Denmark, Copenhagen, Denmark.
18