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Light microscopical observations on two marine dysteriid ciliates from Japan, including a description of Dysteria yagiui nov. spec. (Ciliophora, Cyrtophorida)
ARTICLE IN PRESS European Journal of
PROTISTOLOGY European Journal of Protistology 41 (2005) 29–36 www.elsevier.de/ejop
Light microscopical observations on two marine dysteriid ciliates from Japan, including a description of Dysteria yagiui nov. spec. (Ciliophora, Cyrtophorida) Xiaozhong Hua,b,, Toshikazu Suzukia a
Faculty of Fisheries, Nagasaki University, Bunkyo Machi 1-14, Nagasaki 852-8521, Japan Laboratory of Protozoology, Ocean University of China, Qingdao 266003, P.R. China
b
Received 24 May 2004; accepted 31 August 2004
Abstract Two cyrtophorid ciliates, Dysteria yagiui nov. spec. and D. derouxi, were isolated from fish-farming waters at the New Fishing Port of Nagasaki, Japan. Their morphology and infraciliature were investigated using observation of living cells and protargol impregnation. D. yagiui nov. spec. is a large marine species distinguished from its congeners by the presence of 13 ventral kineties in the right field, of which six extend apically and 6–8 short fragments of kineties in the left equatorial field. D. derouxi has the characteristics previously described except that ciliates of this population possess nine ventral kineties in the right field, of which four extend apically. r 2004 Published by Elsevier GmbH. Keywords: Cyrtophorida; Dysteria derouxi; Dysteria yagiui nov. spec.; Infraciliature; Marine ciliate; Morphology
Introduction The cyrtophorid genus Dysteria comprises highly specialized ciliates, which have been found worldwide in the periphyton, or ectocommensally (Borror 1972; Dragesco 1966; Kahl 1931). Unlike most other cyrtophorids, Dysteria is bilaterally compressed and hence its ventral side is constricted into a narrow ciliated groove between two lateral plates. Most of the somatic cilia lie in parallel rows oriented lengthwise within the ventral groove. These comprise: (1) several fragment-like, densely spaced, left equatorial Corresponding author. C/o Dr. Toshikazu Suzuki, Faculty of Fisheries, Nagasaki University, Bunkyo Machi 1-14, Nagasaki 8528521, Japan. E-mail addresses: [email protected] (X. Hu), [email protected] (T. Suzuki).
0932-4739/$ - see front matter r 2004 Published by Elsevier GmbH. doi:10.1016/j.ejop.2004.08.002
kineties; (2) one short equatorial fragment; (3) several ventral kineties of different length at the right side, whose kineties are fragmented into groups, each group being formed by several basal bodies. Two or more right ventral kineties extend to the apical groove. In addition, there is one terminal fragment positioned at the anterior end of the ventral kineties extending apically (Agamaliev 1983; Deroux 1976; Dragesco and Dragesco-Kerne´is 1986; Faure´-Fremiet 1965; Petz et al. 1995; Song and Packroff 1997). As confirmed by recent studies concerning members of Dysteria, the pattern of infraciliature, especially the ventral ciliary structure, is highly stable and hence is considered as one of the most significant criteria for species separation (Gong et al. 2002; Gong and Song 2003; Song and Wilbert 2002). During a survey on ciliate fauna in the coastal waters of Nagasaki, Japan in 2003, two large periphyton dysteriids were found. One was found to be a new
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species of the genus Dysteria and the other was the recently described D. derouxi.
Material and methods Samples were collected from the fish-farming water in the New Fishing Port of Nagasaki (321480 N; 1291460 E), Japan on March 20 and July 17, 2003 using the Polyurethane Foam Unit (PFU) method. The PFU, which were immersed in seawater for about 2 weeks, were used as artificial substrata for collecting ciliates. Isolated specimens were maintained in the laboratory for about 1 week as raw cultures in Petri dishes for observation of living cells and silver impregnation. Living organisms were observed with a light microscope equipped with differential interference contrast. The protargol impregnation method according to Wilbert (1975) was used to reveal the infraciliature. Measurements were made at 1000 magnification; drawings of stained cells were performed with the help of a camera lucida. Terminology is mainly according to Corliss (1979).
Results and comparison Dysteria yagiui nov. spec. (Figs. 1–3, 5, 8–28; Table 1) Diagnosis: Large marine Dysteria, about 180 75 mm in vivo; cell rectangular in outline with two ventrally located contractile vacuoles; consistently 13 ventral kineties in right field, of which six extend dorso-apically; 6–8 short fragments of kineties in the left equatorial field. Dedication: We dedicate this species to Professor R. Yagiu to express our respect for his contributions to ciliate studies in Japan, and thanks for his help in supplying important literature. Type location: New Fishing Port of Nagasaki, Japan (321480 N; 1291460 E). Type specimens: One holotype and one paratype of the new species as protargol-impregnated slides have been deposited in Department of Zoology, the Natural History of Museum, London, UK (registration number: 2004.6.2.2) and Laboratory of Protozoology, Ocean University of China, Qingdao, P.R. China (registration number: 2003.7.17.04), respectively. Ecological features: Found once (with low abundance) on July 17, 2003. Water temperature 25.5 1C, pH 8.2, salinity 33.5 psu, and dissolved oxygen 8.15 mg l1. Description: Size range 140–200 60–100 mm in vivo, body bilaterally flattened. Body elongate-rectangular in outline when viewed from side but with bluntly rounded
anterior and posterior margins (the ratio of length to width: 2–3:1; Figs. 1, 9, 10); ventral side straight, dorsal side conspicuously convex. Podite relatively large, about 20–30 mm in length, distal end pointed, emerges from posterior ventral side (Figs. 1, arrow; 13, arrowhead; 5, 8, 10). Cilia about 20 mm long. As in its congeners, somatic kineties restricted to ventral and apical grooves between left and right plates (Figs. 9, 10, 13). Conspicuous transverse groove at mid-body on lateral plates, as shown also in D. derouxi (Figs. 32, arrow; 33, arrowhead). A curved stick-like structure always present between the right-most two ventral kineties in the anterior apical groove (Figs. 1, 8; 2, 16, arrow; 11, arrowhead). Cytoplasm grayish to dark-brown, food vacuoles about 5–15 mm in diameter. Cytostome in anterior 1/5–1/6 of body and ventrally located. Two ventrally located contractile vacuoles, one in anterior third, the other in posterior third of cell (Fig. 1). Locomotion without peculiarity, by crawling slowly and irregularly over surfaces with podite extending slowly back and forth. Infraciliature as shown in Figs. 2, 3, 5, 8, 15–19, 21, 27, 28. The ventral field occupied from right to left by one short equatorial fragment comprising 8–24 basal bodies (Figs. 3, 17, arrowhead; 19, arrow), 13 long ciliary rows and 6–7 short kineties. The six right-most of the 13 kineties are longer and extend anteriorly into the apical groove, the longest row having more than 300 basal bodies (Figs. 2, 8; 11, arrow; 16, arrowhead). The remaining seven of these kineties are shortened, their anterior ends terminating at different levels within the groove, while their posterior ends are positioned far behind the podite except for the left-most kinety, which is about 30 mm shorter than the longest row (Figs. 5, 18, arrow; 24, double-arrowhead). All these ventral kineties fragmented, and formed by groups of 4–7 basal bodies. Terminal fragment dorsally positioned and adjacent to distal end of longest ventral kinety, comprised of ca. 30 basal bodies and forming a hook-shaped structure (Figs. 1, 2, 8); some cilia of terminal fragment positioned in the ear-shaped process (Figs. 1; 12, arrow). At mid-body, about 6–8 short rows of densely packed kinetosomes form the left equatorial field (Figs. 3, 17, 21, arrow; 28, arrowhead). One argyrophilic glandule-like structure, shaped like a bent sausage, with each arm about 12–18 mm long, subterminally present near the ventral margin (Figs. 5, 24, arrowhead; 8, 22, arrow); 10–15 kinetosome-like granules invariably present near the base of the podite (Figs. 5, double-arrowhead; 18, arrowhead; 24, arrow). Macronucleus elongated ellipsoid, 40–99 13–26 mm in size after impregnation, positioned at mid-body, characterically heteromerous, some parts being more argentophilic than others; micronucleus not easily stained with protargol. Oral structures as in other members of the genus Dysteria (Figs. 2, 8, 15, 16): they consist of two parallel
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Figs. 1–8. Morphology and infraciliature of D. yagiui nov. spec. (1–3, 5, 8) and D. derouxi (4, 6, 7) from life (1) and after protargol impregnation (2–8). 1. Left lateral view, showing two ventrally located contractile vacuoles, arrow to mark podite. 2. View of the anterior part of the cell, noting ventral kineties, teeth and cytopharyngeal rods, arrow to indicate curved stick-like structure, arrowhead to mark the left frontal kineties. 3. Views of the middle part of the cell to show equatorial fragment (arrowheads) and short rows of kinetosomes in left equatorial field (arrows). 4. View of the anterior part of the cell, arrow to indicate curved stick-like structure, arrowhead marking the short ventral kinety extending apically. 5. Detail from the posterior part of the cell to show the posterior end of left-most ventral kinety (arrow), glandule-like structure (arrowhead) and kinetosome-like granules at the base of the podite (double-arrowhead). 6,7. Infraciliature in different cells, arrow marks the glandule-like structure (6) and micronuclei (7); arrowhead to indicate kinetosome-like granules (6) and excretory pore of contractile vacuole (7); double-arrowhead to demonstrate podite (6) and short rows of kinetosomes in left equatorial field (7). 8. Infraciliature and macronucleus; arrow marks the glandulelike structure, arrowhead indicates the equatorial fragment. Co—circumoral kineties; CR—cytopharyngeal rods; CVP—excretory pore of contractile vacuole; Ef—equatorial fragment; Lf—left frontal kineties; Ma—macronucleus; P—podite; Pr—preoral kinety; T—teeth; Tf—terminal fragment; VK—ventral kineties. Scale bars—40 mm (6); 50 mm (1, 7); 80 mm (8).
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Figs. 9–19. Photomicrographs of D. yagiui nov. spec. from life (9–14) and after protargol impregnation (15–19). 9,10. Left lateral views of living cells, arrow marks cytopharyngeal rod, arrowhead indicates lateral plate. 11,12. Views of the anterior part of the cell, arrowhead shows curved stick-like structure, arrow marks ventral kineties at apical groove (11) and ear-shaped process dorsally located, where some cilia of terminal fragment are seen (12). 13. Left lateral view of the cell at higher magnification, showing cytopharyngeal rods (arrow) and podite (arrowhead). 14,15. Details of the oral area, 14 showing teeth (arrow) and cytopharyngeal rods (arrowhead), 15 showing preoral kinety (arrow) and left frontal kineties (arrowhead). 16. View of the anterior part of the cell, showing curved stick-like structure (arrow), ventral kineties extending apically (arrowhead) and circumoral kineties (doublearrowhead). 17. View of the middle part of the cell, showing the equatorial fragment (arrowhead) and short rows of kinetosomes in left equatorial field (arrow). 18. View of the posterior part of the cell, arrow to show the posterior end of the left-most ventral kinety, arrowhead to mark kinetosome-like granules near the base of podite, double arrowhead points to glandule-like structure. 19. Arrow to show equatorial fragment with only eight basal bodies. Scale bars=80 mm.
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Figs. 20–28. Photomicrographs of D. yagiui nov. spec. after protargol impregnation. 20,23,26. Details of the cytopharynx, showing tooth (arrow in 20 and arrowhead in 26) and cytopharyngeal rod (arrowhead in 20 and arrow in 23 and 26). 21. View of the middle part of the cell to indicate short rows of kinetosomes in left equatorial field (arrow). 22. Details from the posterior part of the cell, arrow to depict glandule-like structure. 24. Region of the podite, showing kinetosome-like granules near the base of podite (arrow), glandule-like structure (arrowhead) and the shortened left-most ventral kinety (double-arrowhead). 25. Macronucleus with DNA replication band giving the appearance of an apparent constriction (arrowhead), arrow to show concentration of chromatin. 27. Anterior region of the cell, arrow to indicate ventral kineties extending apically, arrowhead to mark hook-like terminal fragment. 28. Middle region of the cell, showing excretory pore of contractile vacuole (arrow) and short rows of kinetosomes in left equatorial field (arrowhead).
circumoral kineties and a single-rowed preoral kinety, both of which are composed of dikinetids and positioned anterior to cytostome, and three parallel left frontal kineties composed of monokinetids. Cytopharynx diagonally oriented and extending ca. 30% of cell length, composed of cytopharygeal rods (Figs. 2, 8; 13, 23, 26, arrow; 14, 20, arrowhead), each with a tooth (Figs. 2; 14, 20, arrow; 26, arrowhead). Comparison: This new species may be distinguished from other Dysteria species by its huge size, rectangular body shape, two ventrally located contractile vacuoles and the presence of 13 ventral ciliary rows of somatic kineties in the right field. Among Dysteria species with large cell size and from marine habitat, only three morphologically similar
congeners, namely D. magna, D. antarctica and D. oblongnucleata, are reported to exceed 120 mm in length (Gong et al. 2002; Gong and Song 2003; Ozaki and Yagiu 1943). Compared with D. yagiui, D. magna and D. antarctica can be clearly separated by fewer right ventral kineties (8 and 5, respectively vs. 13) and ellipsoidal macronucleus (vs. long elliptical). Additionally, D. magna is broader than this new species (ratio of length to width ca. 1.5:1 vs. 2–3:1). Regarding body size and its occurrence in the coastal waters of Japan, Dysteria oblongnucleata is very similar to this new species. However, the former can be separated from the latter by its roughly elliptical body shape (vs. rectangular), single contractile vacuole (vs. 2) and sub-triangular podite (vs. distal end pointed) as well
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Table 1. Morphometric characteristics of Dysteria yagiui nov. spec. (upper line) and D. derouxi (lower line) from Nagasaki New Fishing Port Character
Min
Max
Mean SD
n
Body length
140 73
193 110
177.9 93.1
15.10 10.29
16 16
Body width
54 35
94 51
75.3 40.5
9.02 3.76
16 16
Ventral kineties, number
13 9
13 9
13 9
0 0
16 16
Ventral kineties extending apically, number
6 4
6 4
6 4
0 0
16 16
Left equatorial kineties, 6 number 7
8 9
7.3 7.9
0.79 0.57
16 16
Basal bodies in terminal fragment, number
26 13
32 16
28.7 14.5
1.71 1.15
15 16
Basal bodies in equatorial fragment, number
8 4
24 17
15.8 8.9
4.68 4.50
14 11
Macronucleus length
40 36
99 50
72.3 43.4
17.09 4.40
15 16
Macronucleus width
13 13
26 15
20.5 14.1
3.07 0.85
15 16
Gland, diameter
6 5
10 6
8.2 5.1
1.33 0.34
16 16
Podite length
18 10
28 14
23.3 11.1
3.13 1.09
15 16
Cytopharynx length
40 —
65 —
55.8 —
6.37 —
16 —
Arm of glandule-like structure, length
12 10
18 11
14.6 —
1.78 —
12 3
Kinetosome-like granules near the base of podite, number
10 9
15 13
12.2 —
1.47 —
12 6
Distance of the 28 posterior end of the 7 left-most kinety from that of the longest one
35 10
32.8 8.8
2.04 1.13
12 16
Data are based on protargol-impregnated specimens. Measurements in mm. Abbreviations: Max, maximum; Mean, arithmetic mean; Min, minimum; n, number of specimens examined; and SD, standard deviation.
as the presence of a short row of 5–6 strong cilia behind the podite (vs. absence in D. yagiui), although its infraciliature remains unknown. Another large form, Dysteria brasiliensis is characterized by its conspicuously slender body shape and in
having a dorsal spine (vs. absence in D. yagiui) and a lower number of ventral kineties in the right field (5 vs. 13) (Faria et al. 1922; Ozaki and Yagiu 1943; Song and Packroff 1997).
Dysteria derouxi (Figs. 4, 6, 7, 29–38; Table 1) Since this species was recently described from Chinese waters by Gong and Song (2004), our redescription from the Japanese population will concentrate on those features that differ from the original description, or are more clearly illustrated from new preparations. Redescription: The body shape, dimensions, podite, contractile vacuoles and oral structures are similar in the Japanese and Chinese populations and are not redescribed here, although morphometric data of Nagasaki specimens are given in Table 1, and some features are illustrated. Japanese specimens have conspicuous transverse groove at mid-body on lateral plates both in vivo and after protargol impregnation (Figs. 32, arrow; 33, arrowhead), and longitudinal striations on the lateral plates on protargol-stained slides. A curved stick-like structure is always present between the right-most two ventral kineties in anterior-apical groove (Fig. 4, arrow); this was not shown in the microphotographs because adequate contrast could not be obtained. Cilia are about 15 mm long. There were some differences in details of the somatic infraciliature and nuclei of the Nagasaki specimens compared with those from China, which will be described and shown in Figs. 4, 6, 7, 30, 31, 35, 38. Right field occupied by one short equatorial fragment composed of 4–17 basal bodies (Figs. 7, 38); nine long ciliary rows (Figs. 6, 7, 30, 35, 38), the four right-most of which extend anteriorly to the apical groove, the leftmost kinety being shorter than the other three (Figs. 4, 30, 35, arrowhead). Posteriorly these nine ventral kineties end at almost the same level as the podite except for the left-most kinety, which terminates anterior to the podite and about 7–10 mm shorter than the longest row (Fig. 30, arrow). At mid-body, 7–9 short rows of kinetosomes form the left equatorial field (Figs. 7, 30, double-arrowhead; 38, arrow). Terminal fragment comprised of 13–16 basal bodies forming a hook-like structure (Figs. 4, 7, 35). Macronucleus elongate and ellipsoid with posterior end narrowed and more or less pointed after fixation (Figs. 6, 7, 36, 37), with constriction at anterior 2/5 (Figs 31, 36, arrowhead), 36–50 13–15 mm in size after impregnation; micronucleus not argentophilic, and only observed in two cells (1 and 3 in number, respectively, ellipsoid, about 2–3 mm long; Figs. 7, 36, 37, arrows). Nine to 13 kinetosome-like and irregularly arranged granules present near the base of podite (Figs. 6, 38, arrowhead). Ahead of it, a
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Figs. 29–38. Photomicrographs of D. derouxi from life (29, 32) and after protargol impregnation (30, 31, 33–38). 29,32. Left lateral view at different focal plane to show living appearance, arrow to show podite (29) and transverse groove in lateral plate (32). 30,38. Views of the general ciliary pattern, arrow in 30 to show the posterior end of left-most ventral kinety, arrowhead to mark the short ventral kinety extending apically (30) and kinetosome-like granules (38), double-arrowhead in 30 and arrow in 38 to demonstrate short rows of kinetosomes in left equatorial field, double-arrowheads in 38 to indicate excretory pore of contractile vacuole. 31,35. Views of the anterior part of the cell, arrow to show cytopharyngeal rods (31) and circumoral kineties (35), arrowhead to indicate constriction in macronucleus (31) and the short ventral kinety extending apically (35). 33,34. Views of the middle part of the cell, arrowhead in 33 to show transverse groove in lateral plate, arrow in 33 to mark longitudinal striae in lateral plate, arrows in 34 to show the excretory pore of the contractile vacuole. 36,37. To show macronucleus and micronuclei (arrow in 36 and arrows in 37), Co—circumoral kineties; Cy—cytostome; Ef—equatorial fragment; Lf—left frontal kineties; Pr—preoral kinety; Tf—terminal fragment; and VK—ventral kineties.
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glandule-like structure, bent-sausage-shaped, with each arm 10–11 mm long (Figs. 6, arrow; 7). Ecological features: Found once (with low abundance) on March 20. Water temperature 15 1C, pH 8.2, salinity 34 psu. Comparison: Members of the Nagasaki population correspond well with the Qingdao population although they differ slightly in the number of ventral kineties (consistently 9 in the former vs. 8). In spite of these small differences, we are confident that our species identification is correct. We were fortunate to obtain some improved protargol preparations, which add details to, but generally confirm, the species description given by Gong and Song (2004). Ozaki and Yagiu (1943) misidentified a morphotype under the name of Dysteria cristata. The form they described resembles D. derouxi in body size and shape in vivo, two ventrally located contractile vacuoles, elongated elliptical macronucleus and apparently four ciliary rows extending apically (from illustration); thus it could very likely have been D. derouxi.
Acknowledgements This work was supported by ‘‘JSPS Postdoctoral Fellowship for Foreign Researcher’’ and the ‘‘Natural Science Foundation of China (project number: 40206021)’’.
References Agamaliev, F.G., 1983. Ciliates of the Caspian Sea: Systematics Ecology, Zoogeography. Nauka, Leningrad (in Russian). Borror, A.C., 1972. Tidal marsh ciliates (Protozoa): morphology, ecology, systematics. Acta Protozool. 10, 29–71. Corliss, J.O., 1979. The Ciliated Protozoa: Characterization, Classification, and Guide to the Literature, second ed. Pergamon Press, Oxford. Deroux, G., 1976. Le plan cortical des Cyrtophorida. III. Les structures diffe´renciatrices chez les Dysteriina. Protistologica 12, 505–538.
Dragesco, J., 1966. Observations sur quelques cilie´s libres. Arch. Protistenkd. 109, 155–206. Dragesco, J., Dragesco-Kerne´is, A., 1986. Cilie´s libres de l’ Afrique intertropicale. Faune Trop. 26, 1–559. Faria, J.G., da Cunha, A.M., Pinto, C., 1922. Estudos sobre protozoarios do mar. Mem. Inst. Oswaldo Cruz 15, 186–208. Faure´-Fremiet, E., 1965. Morphologie des Dysteriidae (Ciliata Cyrtophorina). Compt. Rend. Acad. Sci. Paris 260, 6679–6684. Gong, J., Song, W., 2003. Morphology and infraciliature of two marine benthic ciliates, Dysteria procera Kahl, 1931 and Dysteria magna nov. spec. (Protozoa, Ciliophora, Cyrtophorida), from China. Eur. J. Protistol. 39, 301–309. Gong, J., Song, W., 2004. Description of a new marine cyrtophorid ciliate, Dysteria derouxi nov. spec., with an updated key to 12 well-investigated Dysteria species (Ciliophora, Cyrtophorida). Eur. J. Protistol. 40, 13–19. Gong, J., Song, W., Warren, A., 2002. Redescription of two marine cyrtophorid ciliates, Dysteria cristata (Gourret and Roeser, 1888) Kahl, 1931 and Dysteria monostyla (Ehrenberg, 1838) Kahl, 1931 (Protozoa, Ciliophora, Cyrtophorida), from China. Eur. J. Protistol. 38, 213–222. Kahl, A., 1931. Urtiere oder Protozoa I: Wimpertiere oder Ciliata (Infusoria). 2. Holotricha. Tierwelt Dtl. 21, 181–398. Ozaki, Y., Yagiu, R., 1943. Studies on the marine ciliates of Japan, mainly the Setonaikai (the Inland Sea of Japan), I. J. Sci. Hiroshima Univ. Ser. B, Div. 1 (10), 21–52. Petz, W., Song, W., Wilbert, N., 1995. Taxonomy and ecology of the ciliate fauna (Protozoa, Ciliophora) in the endopagial and pelagial of the Weddell Sea, Antarctica. Stapfia 40, 1–233. Song, W., Packroff, G., 1997. Taxonomische Untersuchungen an marinen Ciliaten aus China mit Beschreibungen von zwei neuen Arten, Strombidium globosaneum nov. spec. und S. platum nov. spec. (Protozoa, Ciliophora). Arch. Protistenkd. 147, 331–360. Song, W., Wilbert, N., 2002. Faunistic studies on marine ciliates from the Antarctic benthic area, including descriptions of one epizoid form, 6 new species and, 2 new genera (Protozoa, Ciliophora). Acta Protozool. 41, 23–61. Wilbert, N., 1975. Eine verbesserte Technik der Protargolimpra¨gnation fu¨r Ciliaten. Mikrokosmos 64, 171–179.
PROTISTOLOGY European Journal of Protistology 41 (2005) 29–36 www.elsevier.de/ejop
Light microscopical observations on two marine dysteriid ciliates from Japan, including a description of Dysteria yagiui nov. spec. (Ciliophora, Cyrtophorida) Xiaozhong Hua,b,, Toshikazu Suzukia a
Faculty of Fisheries, Nagasaki University, Bunkyo Machi 1-14, Nagasaki 852-8521, Japan Laboratory of Protozoology, Ocean University of China, Qingdao 266003, P.R. China
b
Received 24 May 2004; accepted 31 August 2004
Abstract Two cyrtophorid ciliates, Dysteria yagiui nov. spec. and D. derouxi, were isolated from fish-farming waters at the New Fishing Port of Nagasaki, Japan. Their morphology and infraciliature were investigated using observation of living cells and protargol impregnation. D. yagiui nov. spec. is a large marine species distinguished from its congeners by the presence of 13 ventral kineties in the right field, of which six extend apically and 6–8 short fragments of kineties in the left equatorial field. D. derouxi has the characteristics previously described except that ciliates of this population possess nine ventral kineties in the right field, of which four extend apically. r 2004 Published by Elsevier GmbH. Keywords: Cyrtophorida; Dysteria derouxi; Dysteria yagiui nov. spec.; Infraciliature; Marine ciliate; Morphology
Introduction The cyrtophorid genus Dysteria comprises highly specialized ciliates, which have been found worldwide in the periphyton, or ectocommensally (Borror 1972; Dragesco 1966; Kahl 1931). Unlike most other cyrtophorids, Dysteria is bilaterally compressed and hence its ventral side is constricted into a narrow ciliated groove between two lateral plates. Most of the somatic cilia lie in parallel rows oriented lengthwise within the ventral groove. These comprise: (1) several fragment-like, densely spaced, left equatorial Corresponding author. C/o Dr. Toshikazu Suzuki, Faculty of Fisheries, Nagasaki University, Bunkyo Machi 1-14, Nagasaki 8528521, Japan. E-mail addresses: [email protected] (X. Hu), [email protected] (T. Suzuki).
0932-4739/$ - see front matter r 2004 Published by Elsevier GmbH. doi:10.1016/j.ejop.2004.08.002
kineties; (2) one short equatorial fragment; (3) several ventral kineties of different length at the right side, whose kineties are fragmented into groups, each group being formed by several basal bodies. Two or more right ventral kineties extend to the apical groove. In addition, there is one terminal fragment positioned at the anterior end of the ventral kineties extending apically (Agamaliev 1983; Deroux 1976; Dragesco and Dragesco-Kerne´is 1986; Faure´-Fremiet 1965; Petz et al. 1995; Song and Packroff 1997). As confirmed by recent studies concerning members of Dysteria, the pattern of infraciliature, especially the ventral ciliary structure, is highly stable and hence is considered as one of the most significant criteria for species separation (Gong et al. 2002; Gong and Song 2003; Song and Wilbert 2002). During a survey on ciliate fauna in the coastal waters of Nagasaki, Japan in 2003, two large periphyton dysteriids were found. One was found to be a new
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species of the genus Dysteria and the other was the recently described D. derouxi.
Material and methods Samples were collected from the fish-farming water in the New Fishing Port of Nagasaki (321480 N; 1291460 E), Japan on March 20 and July 17, 2003 using the Polyurethane Foam Unit (PFU) method. The PFU, which were immersed in seawater for about 2 weeks, were used as artificial substrata for collecting ciliates. Isolated specimens were maintained in the laboratory for about 1 week as raw cultures in Petri dishes for observation of living cells and silver impregnation. Living organisms were observed with a light microscope equipped with differential interference contrast. The protargol impregnation method according to Wilbert (1975) was used to reveal the infraciliature. Measurements were made at 1000 magnification; drawings of stained cells were performed with the help of a camera lucida. Terminology is mainly according to Corliss (1979).
Results and comparison Dysteria yagiui nov. spec. (Figs. 1–3, 5, 8–28; Table 1) Diagnosis: Large marine Dysteria, about 180 75 mm in vivo; cell rectangular in outline with two ventrally located contractile vacuoles; consistently 13 ventral kineties in right field, of which six extend dorso-apically; 6–8 short fragments of kineties in the left equatorial field. Dedication: We dedicate this species to Professor R. Yagiu to express our respect for his contributions to ciliate studies in Japan, and thanks for his help in supplying important literature. Type location: New Fishing Port of Nagasaki, Japan (321480 N; 1291460 E). Type specimens: One holotype and one paratype of the new species as protargol-impregnated slides have been deposited in Department of Zoology, the Natural History of Museum, London, UK (registration number: 2004.6.2.2) and Laboratory of Protozoology, Ocean University of China, Qingdao, P.R. China (registration number: 2003.7.17.04), respectively. Ecological features: Found once (with low abundance) on July 17, 2003. Water temperature 25.5 1C, pH 8.2, salinity 33.5 psu, and dissolved oxygen 8.15 mg l1. Description: Size range 140–200 60–100 mm in vivo, body bilaterally flattened. Body elongate-rectangular in outline when viewed from side but with bluntly rounded
anterior and posterior margins (the ratio of length to width: 2–3:1; Figs. 1, 9, 10); ventral side straight, dorsal side conspicuously convex. Podite relatively large, about 20–30 mm in length, distal end pointed, emerges from posterior ventral side (Figs. 1, arrow; 13, arrowhead; 5, 8, 10). Cilia about 20 mm long. As in its congeners, somatic kineties restricted to ventral and apical grooves between left and right plates (Figs. 9, 10, 13). Conspicuous transverse groove at mid-body on lateral plates, as shown also in D. derouxi (Figs. 32, arrow; 33, arrowhead). A curved stick-like structure always present between the right-most two ventral kineties in the anterior apical groove (Figs. 1, 8; 2, 16, arrow; 11, arrowhead). Cytoplasm grayish to dark-brown, food vacuoles about 5–15 mm in diameter. Cytostome in anterior 1/5–1/6 of body and ventrally located. Two ventrally located contractile vacuoles, one in anterior third, the other in posterior third of cell (Fig. 1). Locomotion without peculiarity, by crawling slowly and irregularly over surfaces with podite extending slowly back and forth. Infraciliature as shown in Figs. 2, 3, 5, 8, 15–19, 21, 27, 28. The ventral field occupied from right to left by one short equatorial fragment comprising 8–24 basal bodies (Figs. 3, 17, arrowhead; 19, arrow), 13 long ciliary rows and 6–7 short kineties. The six right-most of the 13 kineties are longer and extend anteriorly into the apical groove, the longest row having more than 300 basal bodies (Figs. 2, 8; 11, arrow; 16, arrowhead). The remaining seven of these kineties are shortened, their anterior ends terminating at different levels within the groove, while their posterior ends are positioned far behind the podite except for the left-most kinety, which is about 30 mm shorter than the longest row (Figs. 5, 18, arrow; 24, double-arrowhead). All these ventral kineties fragmented, and formed by groups of 4–7 basal bodies. Terminal fragment dorsally positioned and adjacent to distal end of longest ventral kinety, comprised of ca. 30 basal bodies and forming a hook-shaped structure (Figs. 1, 2, 8); some cilia of terminal fragment positioned in the ear-shaped process (Figs. 1; 12, arrow). At mid-body, about 6–8 short rows of densely packed kinetosomes form the left equatorial field (Figs. 3, 17, 21, arrow; 28, arrowhead). One argyrophilic glandule-like structure, shaped like a bent sausage, with each arm about 12–18 mm long, subterminally present near the ventral margin (Figs. 5, 24, arrowhead; 8, 22, arrow); 10–15 kinetosome-like granules invariably present near the base of the podite (Figs. 5, double-arrowhead; 18, arrowhead; 24, arrow). Macronucleus elongated ellipsoid, 40–99 13–26 mm in size after impregnation, positioned at mid-body, characterically heteromerous, some parts being more argentophilic than others; micronucleus not easily stained with protargol. Oral structures as in other members of the genus Dysteria (Figs. 2, 8, 15, 16): they consist of two parallel
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Figs. 1–8. Morphology and infraciliature of D. yagiui nov. spec. (1–3, 5, 8) and D. derouxi (4, 6, 7) from life (1) and after protargol impregnation (2–8). 1. Left lateral view, showing two ventrally located contractile vacuoles, arrow to mark podite. 2. View of the anterior part of the cell, noting ventral kineties, teeth and cytopharyngeal rods, arrow to indicate curved stick-like structure, arrowhead to mark the left frontal kineties. 3. Views of the middle part of the cell to show equatorial fragment (arrowheads) and short rows of kinetosomes in left equatorial field (arrows). 4. View of the anterior part of the cell, arrow to indicate curved stick-like structure, arrowhead marking the short ventral kinety extending apically. 5. Detail from the posterior part of the cell to show the posterior end of left-most ventral kinety (arrow), glandule-like structure (arrowhead) and kinetosome-like granules at the base of the podite (double-arrowhead). 6,7. Infraciliature in different cells, arrow marks the glandule-like structure (6) and micronuclei (7); arrowhead to indicate kinetosome-like granules (6) and excretory pore of contractile vacuole (7); double-arrowhead to demonstrate podite (6) and short rows of kinetosomes in left equatorial field (7). 8. Infraciliature and macronucleus; arrow marks the glandulelike structure, arrowhead indicates the equatorial fragment. Co—circumoral kineties; CR—cytopharyngeal rods; CVP—excretory pore of contractile vacuole; Ef—equatorial fragment; Lf—left frontal kineties; Ma—macronucleus; P—podite; Pr—preoral kinety; T—teeth; Tf—terminal fragment; VK—ventral kineties. Scale bars—40 mm (6); 50 mm (1, 7); 80 mm (8).
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Figs. 9–19. Photomicrographs of D. yagiui nov. spec. from life (9–14) and after protargol impregnation (15–19). 9,10. Left lateral views of living cells, arrow marks cytopharyngeal rod, arrowhead indicates lateral plate. 11,12. Views of the anterior part of the cell, arrowhead shows curved stick-like structure, arrow marks ventral kineties at apical groove (11) and ear-shaped process dorsally located, where some cilia of terminal fragment are seen (12). 13. Left lateral view of the cell at higher magnification, showing cytopharyngeal rods (arrow) and podite (arrowhead). 14,15. Details of the oral area, 14 showing teeth (arrow) and cytopharyngeal rods (arrowhead), 15 showing preoral kinety (arrow) and left frontal kineties (arrowhead). 16. View of the anterior part of the cell, showing curved stick-like structure (arrow), ventral kineties extending apically (arrowhead) and circumoral kineties (doublearrowhead). 17. View of the middle part of the cell, showing the equatorial fragment (arrowhead) and short rows of kinetosomes in left equatorial field (arrow). 18. View of the posterior part of the cell, arrow to show the posterior end of the left-most ventral kinety, arrowhead to mark kinetosome-like granules near the base of podite, double arrowhead points to glandule-like structure. 19. Arrow to show equatorial fragment with only eight basal bodies. Scale bars=80 mm.
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Figs. 20–28. Photomicrographs of D. yagiui nov. spec. after protargol impregnation. 20,23,26. Details of the cytopharynx, showing tooth (arrow in 20 and arrowhead in 26) and cytopharyngeal rod (arrowhead in 20 and arrow in 23 and 26). 21. View of the middle part of the cell to indicate short rows of kinetosomes in left equatorial field (arrow). 22. Details from the posterior part of the cell, arrow to depict glandule-like structure. 24. Region of the podite, showing kinetosome-like granules near the base of podite (arrow), glandule-like structure (arrowhead) and the shortened left-most ventral kinety (double-arrowhead). 25. Macronucleus with DNA replication band giving the appearance of an apparent constriction (arrowhead), arrow to show concentration of chromatin. 27. Anterior region of the cell, arrow to indicate ventral kineties extending apically, arrowhead to mark hook-like terminal fragment. 28. Middle region of the cell, showing excretory pore of contractile vacuole (arrow) and short rows of kinetosomes in left equatorial field (arrowhead).
circumoral kineties and a single-rowed preoral kinety, both of which are composed of dikinetids and positioned anterior to cytostome, and three parallel left frontal kineties composed of monokinetids. Cytopharynx diagonally oriented and extending ca. 30% of cell length, composed of cytopharygeal rods (Figs. 2, 8; 13, 23, 26, arrow; 14, 20, arrowhead), each with a tooth (Figs. 2; 14, 20, arrow; 26, arrowhead). Comparison: This new species may be distinguished from other Dysteria species by its huge size, rectangular body shape, two ventrally located contractile vacuoles and the presence of 13 ventral ciliary rows of somatic kineties in the right field. Among Dysteria species with large cell size and from marine habitat, only three morphologically similar
congeners, namely D. magna, D. antarctica and D. oblongnucleata, are reported to exceed 120 mm in length (Gong et al. 2002; Gong and Song 2003; Ozaki and Yagiu 1943). Compared with D. yagiui, D. magna and D. antarctica can be clearly separated by fewer right ventral kineties (8 and 5, respectively vs. 13) and ellipsoidal macronucleus (vs. long elliptical). Additionally, D. magna is broader than this new species (ratio of length to width ca. 1.5:1 vs. 2–3:1). Regarding body size and its occurrence in the coastal waters of Japan, Dysteria oblongnucleata is very similar to this new species. However, the former can be separated from the latter by its roughly elliptical body shape (vs. rectangular), single contractile vacuole (vs. 2) and sub-triangular podite (vs. distal end pointed) as well
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Table 1. Morphometric characteristics of Dysteria yagiui nov. spec. (upper line) and D. derouxi (lower line) from Nagasaki New Fishing Port Character
Min
Max
Mean SD
n
Body length
140 73
193 110
177.9 93.1
15.10 10.29
16 16
Body width
54 35
94 51
75.3 40.5
9.02 3.76
16 16
Ventral kineties, number
13 9
13 9
13 9
0 0
16 16
Ventral kineties extending apically, number
6 4
6 4
6 4
0 0
16 16
Left equatorial kineties, 6 number 7
8 9
7.3 7.9
0.79 0.57
16 16
Basal bodies in terminal fragment, number
26 13
32 16
28.7 14.5
1.71 1.15
15 16
Basal bodies in equatorial fragment, number
8 4
24 17
15.8 8.9
4.68 4.50
14 11
Macronucleus length
40 36
99 50
72.3 43.4
17.09 4.40
15 16
Macronucleus width
13 13
26 15
20.5 14.1
3.07 0.85
15 16
Gland, diameter
6 5
10 6
8.2 5.1
1.33 0.34
16 16
Podite length
18 10
28 14
23.3 11.1
3.13 1.09
15 16
Cytopharynx length
40 —
65 —
55.8 —
6.37 —
16 —
Arm of glandule-like structure, length
12 10
18 11
14.6 —
1.78 —
12 3
Kinetosome-like granules near the base of podite, number
10 9
15 13
12.2 —
1.47 —
12 6
Distance of the 28 posterior end of the 7 left-most kinety from that of the longest one
35 10
32.8 8.8
2.04 1.13
12 16
Data are based on protargol-impregnated specimens. Measurements in mm. Abbreviations: Max, maximum; Mean, arithmetic mean; Min, minimum; n, number of specimens examined; and SD, standard deviation.
as the presence of a short row of 5–6 strong cilia behind the podite (vs. absence in D. yagiui), although its infraciliature remains unknown. Another large form, Dysteria brasiliensis is characterized by its conspicuously slender body shape and in
having a dorsal spine (vs. absence in D. yagiui) and a lower number of ventral kineties in the right field (5 vs. 13) (Faria et al. 1922; Ozaki and Yagiu 1943; Song and Packroff 1997).
Dysteria derouxi (Figs. 4, 6, 7, 29–38; Table 1) Since this species was recently described from Chinese waters by Gong and Song (2004), our redescription from the Japanese population will concentrate on those features that differ from the original description, or are more clearly illustrated from new preparations. Redescription: The body shape, dimensions, podite, contractile vacuoles and oral structures are similar in the Japanese and Chinese populations and are not redescribed here, although morphometric data of Nagasaki specimens are given in Table 1, and some features are illustrated. Japanese specimens have conspicuous transverse groove at mid-body on lateral plates both in vivo and after protargol impregnation (Figs. 32, arrow; 33, arrowhead), and longitudinal striations on the lateral plates on protargol-stained slides. A curved stick-like structure is always present between the right-most two ventral kineties in anterior-apical groove (Fig. 4, arrow); this was not shown in the microphotographs because adequate contrast could not be obtained. Cilia are about 15 mm long. There were some differences in details of the somatic infraciliature and nuclei of the Nagasaki specimens compared with those from China, which will be described and shown in Figs. 4, 6, 7, 30, 31, 35, 38. Right field occupied by one short equatorial fragment composed of 4–17 basal bodies (Figs. 7, 38); nine long ciliary rows (Figs. 6, 7, 30, 35, 38), the four right-most of which extend anteriorly to the apical groove, the leftmost kinety being shorter than the other three (Figs. 4, 30, 35, arrowhead). Posteriorly these nine ventral kineties end at almost the same level as the podite except for the left-most kinety, which terminates anterior to the podite and about 7–10 mm shorter than the longest row (Fig. 30, arrow). At mid-body, 7–9 short rows of kinetosomes form the left equatorial field (Figs. 7, 30, double-arrowhead; 38, arrow). Terminal fragment comprised of 13–16 basal bodies forming a hook-like structure (Figs. 4, 7, 35). Macronucleus elongate and ellipsoid with posterior end narrowed and more or less pointed after fixation (Figs. 6, 7, 36, 37), with constriction at anterior 2/5 (Figs 31, 36, arrowhead), 36–50 13–15 mm in size after impregnation; micronucleus not argentophilic, and only observed in two cells (1 and 3 in number, respectively, ellipsoid, about 2–3 mm long; Figs. 7, 36, 37, arrows). Nine to 13 kinetosome-like and irregularly arranged granules present near the base of podite (Figs. 6, 38, arrowhead). Ahead of it, a
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Figs. 29–38. Photomicrographs of D. derouxi from life (29, 32) and after protargol impregnation (30, 31, 33–38). 29,32. Left lateral view at different focal plane to show living appearance, arrow to show podite (29) and transverse groove in lateral plate (32). 30,38. Views of the general ciliary pattern, arrow in 30 to show the posterior end of left-most ventral kinety, arrowhead to mark the short ventral kinety extending apically (30) and kinetosome-like granules (38), double-arrowhead in 30 and arrow in 38 to demonstrate short rows of kinetosomes in left equatorial field, double-arrowheads in 38 to indicate excretory pore of contractile vacuole. 31,35. Views of the anterior part of the cell, arrow to show cytopharyngeal rods (31) and circumoral kineties (35), arrowhead to indicate constriction in macronucleus (31) and the short ventral kinety extending apically (35). 33,34. Views of the middle part of the cell, arrowhead in 33 to show transverse groove in lateral plate, arrow in 33 to mark longitudinal striae in lateral plate, arrows in 34 to show the excretory pore of the contractile vacuole. 36,37. To show macronucleus and micronuclei (arrow in 36 and arrows in 37), Co—circumoral kineties; Cy—cytostome; Ef—equatorial fragment; Lf—left frontal kineties; Pr—preoral kinety; Tf—terminal fragment; and VK—ventral kineties.
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glandule-like structure, bent-sausage-shaped, with each arm 10–11 mm long (Figs. 6, arrow; 7). Ecological features: Found once (with low abundance) on March 20. Water temperature 15 1C, pH 8.2, salinity 34 psu. Comparison: Members of the Nagasaki population correspond well with the Qingdao population although they differ slightly in the number of ventral kineties (consistently 9 in the former vs. 8). In spite of these small differences, we are confident that our species identification is correct. We were fortunate to obtain some improved protargol preparations, which add details to, but generally confirm, the species description given by Gong and Song (2004). Ozaki and Yagiu (1943) misidentified a morphotype under the name of Dysteria cristata. The form they described resembles D. derouxi in body size and shape in vivo, two ventrally located contractile vacuoles, elongated elliptical macronucleus and apparently four ciliary rows extending apically (from illustration); thus it could very likely have been D. derouxi.
Acknowledgements This work was supported by ‘‘JSPS Postdoctoral Fellowship for Foreign Researcher’’ and the ‘‘Natural Science Foundation of China (project number: 40206021)’’.
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