ISSN 1995�0829, Inland Water Biology, 2011, Vol. 4, No. 3, pp. 287–292. © Pleiades Publishing, Ltd., 2011.Original Russian Text © V.A. Zolotarev, Z.M. Myl’nikova, A.P. Myl’nikov, 2011, published in Biologiya Vnutrennikh Vod, No. 3, 2011, pp. 15–21.

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INTRODUCTION

Taumatomastix Lauterborn, 1899, species arewidespread in sea and brackish water. They are rarelyfound in fresh water [10, 11]. This genus belongs to thethaumatomonads group (order ThaumatomonadidaShirkina, 1987, em. Karpov, 1990). This order consistsof seven genera (Bodopsis Lemmermann, 1914;Gyromitus Skuja, 1939; Hyaloselene Skuja, 1956; Pro�taspis Skuja, 1939; Rhizaspis Skuja, 1948; and Thau�matomonas De Saedeleer, 1931) and is characterizedby a thickened cell body, two heterodynamic flagella,and a ventricular furrow from which filose pseudopo�dia run [8]. One unique feature of these singled�celledanimals is the formation of somatic scales on the sur�face of mitochondria [2, 4, 5]. Inside the Thaumato�mastix genus, 17 species are known that are mainly dif�ferentiated by their form and scale size. Their scalescan be triangular, oval, or have a complex form withanabolies [10, 11]. The most detailed study was under�taken on the structure of Thaumatomonas and Gyromi�tus thaumatomonads [1–3, 9]. Among Thaumatomas�tix species, only fragmentary data about the structureof Th. (Reckertia) sagittifera Beech et Moestrup areavailable [7].

The goal of this research work is to make a detailedstudy of the thin cell structure of the Thaumatomastixrepresentative.

MATERIAL AND METHODS

Thaumatomastix sp. flagellates were found in sam�ples taken in October 2002 from the White Sea shore�land near the settlement of Kartesh (Karelia). Thesalinity of water was 22%. The clonal TM–5 culturewas examined. The flagellates were cultivated in theShmaltz�Pratt medium, the salinity of which was 20%.Pseudomonas fluorescens Migula bacteria were used asfood. A light microscopical investigation was donewith the help of the Biolam R microscope, which wasprovided with a phase�contrast device.

To perform an electron microscopic investigation,the flagellate suspension was condensed by centrifuga�tion and the gained cells were fixed in a mixture of 2%OsO4 solution and 0.6% glutaraldehyde solution (1 : 1),which were prepared in the Shmaltz�Pratt medium, for15–30 min under the temperature of 1°С. After dehy�dratation in a series of alcohol and anhydrous acetone,the material was put into a mixture of Araldite and Eponand examined under a JEM�1011 microscope (Japan).The sections were obtained with the help of LKB ultra�microtom (Sweden). The air�dried total preparationswere sprayed with tungsten oxide.

RESULTS

The cells are highly flattened. They have oval orovoid forms (Figs. 1a, 1b). The length of a cell reached9–15 µm and the width was 6–9 µm. Two unequal fla�

The Ultrafine Structure of Amoeboid Flagellate Thaumatomastix sp. (Thaumatomonadida (Shirkina) Karpov, 1990)

V. A. Zolotarev, Z. M. Myl’nikova, and A. P. Myl’nikovPapanin Institute for Biology of Inland Waters, Russian Academy of Sciences,

Borok, Nekouzskii raion, Yaroslavl oblast, 152742 Russiae�mail: noo@ibiw.yaroslavl.ru

Received January 27, 2010

Abstract—The ultrafine structure of amoeboid flagellate Thaumatomastix sp. is considered. The cell is sur�rounded by two�layered triangular scales. They are formed on the surface of mitochondria. Pseudopodiagrabbing bacteria run from ventricular furrow, which is armored with two longitudinal bands of microtubules.Heterodynamic flagella run from small flagellant pocket. Long back flagellum has thin mastigonemes. Prox�imal area of short flagellum is covered with flat oval scales. Transitional flagellant zone has no spiral or otheradditional elements. Transverse plate is localized above cell surface. Kinetosomes are parallel to each other.Bubblelike nucleus and Golgi apparatus have typical structure. Oval mitochondria contain tubular cristae.Within cytoplasm, urticant organelles (kinetocysts) containing amorphous material and capsule were found.The latter consists of muff and cylinder. Plasmodial and cystic phases of development have not been discov�ered. Contractile vacuole is absent. The resemblance between Thaumatomastix sp. and other thaumato�monads has been discussed.

Keywords: @@@

DOI: 10.1134/S1995082911020222

BIOLOGY, MORPHOLOGY, AND SYSTEMATICS OF HYDROBIONTS

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Fig. 1. General morphology of Thaumatomastix sp. cell: (a) external view of cell, light microscope; (b, c) total preparations of celland scales; (d, e) transverse sections of cells; and (f–h) flagella and flagellant pocket. (a) Light microscope and (b–h) transmis�sion microscope. (a.g) Golgi apparatus, (f.p) flagellant pocket, (f.s) flagellant scales, (b.f) back flagellum, (ks) kinetosome,(kc) kinetocyst, (m) mitochondrion, (mn) mastigonemes, (o.m) osmiophil material, (d.v) digestive vacuole, (f.f) front flagellum,(bs) bubbles, (s.s) somatic scales, and (n) nucleus.

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gella run from flagellant pocket (Fig. 1a). Short flagel�lum (1.5–2.0 µm) makes frequent vibrations back andforth. Long flagellum (18–20 µm) is directed back�wards and goes through a ventricular furrow. It is notattached to cell body and it can be easily seen whencell is turned around (Fig. 1b). Nucleus lies in front.Contractile vacuole was not found. While moving(crawling on substrate) cell back end is often a littleraised. When movement stops, pseudopodium isformed on the ventral side of cell. The flagellate eatsbacteria. Predation was not registered. Plasmodia(multinucleated amoeboid cells) and cysts are notfound.

Cell is covered by triangular scales (Fig. 1c). Thelatter were found almost on all cell areas, exceptpseudopodia, flagella, and flagellant pocket. Bubble�like nucleus, flagellant pocket, digestive vacuole, andcluster of submembranous bubbles (Fig. 1d) areobserved on transverse section of frontal cell end.Inside 3–4 µm nucleus there are chromatin cobs(Figs. 1d–1f, 2b). Nucleus is near kinetosomes of fla�gella. The kinetosomes are near the only dictyosomeof Golgi apparatus, which forms a pile of flattened cis�terns (Fig. 1e). The axes of flagella kinetosomes areparallel to each other (Fig. 1e, 2b, 2e). No fibrillarbridge was found between kinetosomes. Flagellum hastypical structure. Axoneme microtubules are orga�nized according to the 9 + 2 formula (Figs. 1g, 2d, 2e).Transitional flagellant zone has no additional ele�ments, such as cylinder or spiral (Fig. 1f). Inside theaxoneme of front flagellum, osmiophil cobs were reg�istered (Fig. 1g). Transverse plate is 0.3 µm raised overthe cell surface (Fig. 1f). Front flagellum in proximalarea is covered by thin one�layered scales (Figs. 1f–1h,2c). These scales consist of an oval plate with a longi�tudinal comb (thickening). The length of this comb is0.3 µm. On the dried preparations, such flagellantscales were not found. Back flagellum is even coveredby thin and coiled unicellular mastigonemes (flagel�lant hairs) with lengths of 0.8 µm (Figs. 1g, 2a). Theyfill the whole flagellant pocket. Mastigoneme rudi�ments were found in the derivatives of endoplasmicreticulum (Fig. 3c). No mastigonemes were found onthe dried preparations (Fig. 1c).

Flagellant pocket, the depth of which is 1.5–2.5 µmand diameter is 1.2–1.3 µm, is seen in the light micro�scope (Fig. 1a). It is armored with two bands consist�ing of six and eight microtubules lying parallel to theflagella and bands consisting of two microtubules(Figs. 2c–2e). The bands must be incurvated near thehole of flagellant pocket and then go along the sides(borders) of ventricular furrow (Fig. 2e). The furrowserves as a place where pseudopodia are formed. Thesepseudopodia contain small bubbles and are not coveredwith scales. No mitochondria were found insidepseudopodia. Bands consisting of 8–12 and 6–8 micro�tubules go along the furrow sides (Figs. 2f–2h). Thenumber of microtubules within bands decreases to thecell back end.

Oval mitochondria, sized 1.5…1.8 × 0.5…0.8 µm,are evenly interspersed in the cytoplasm (Figs. 1e, 2b).Somatic scales consist of two triangular plates that arelinked to each other by three hollow cylindric bridgeson the corners of these plates. The length of such aplate rib is 0.55–0.45 µm; the crura height is 0.25–0.35 µm (Figs. 3a, 3b). On the upper plate of scale,three holes can be observed. On cell surface, scalesmake up a smooth row. They are closely attached toeach other in such a way that one scale hooks on theedge of another (Fig. 3b). Scales are formed on thesurface of mitochondria (Fig. 1e). Triangular scalescan be seen on some sections (Fig. 3c).

Urticant organelles that are bubblelike kinetocystswith diameters of 0.60–0.65 µm lie under plasmale�mma (Figs. 1d, 2a, 2f). Inside the bubble there areasymmetrically arranged capsules and amorphousmaterials (Figs. 3d, 3e). On the longitudinal section,the capsule is oval, while it is round on the transversesection (Figs. 3d, 3e). Inside the capsule lies a cylindersurrounded by matrix (muff). The capsule size is0.34…0.40 × 0.25…0.28 µm. The rudiments of kineto�cysts are near the cisterns of Golgi apparatus (Fig. 3f).No discharged kinetocysts were discovered.

DISCUSSION OF THE RESULTS

The structure of TM–5 clone can be comparedwith the structure of two very similar genera: Thau�matomonas and Thaumatomastix. Representatives ofThaumatomonas are creepers. They live in freshwaterand soil. Representatives of Thaumatomastix are float�ing or sometimes creeping organisms that are typicalfor sea water. However, one Thaumatomastix specieswas found in fresh water [8]. The mentioned generahave the following common features: flattened cellbodies, two heterodynamic flagella (the front flagel�lum is often shorter), ventricular furrow and somaticscales formed on the surface of mitochondria, mas�tigonemes on back flagellum, and small flagellantscales [4, 6, 8, 10]. Triangular somatic scales are com�mon. Oval somatic scales are rare. Moreover, somerepresentatives of Thaumatomastix have additionalsomatic scales that look like spinelets or spicules.

In the research work [10] the classification ofThaumatomastix scales is given. According to the pic�tures, the form of scales in the TM�5 clone is similar tothe form of scales in Thaumatomastix formosa Thom�sen et al. [10]. Nevertheless, they have different edgeperforations, and their cylindric crura have dissimilarforms. The research work [11] shows that there are alsosimilar features between scales of the TM�5 clone andThaumatomastix sp. 4, but the latter has no edge per�forations. For the Thaumatomastix genus, small flat�tened scales on the front flagellum are typical [10, 11].Similar scales were found in the TM–5 clone. The setof external features gives evidence that the TM�5 clonebelongs to the Thaumatomastix genus according to thesimilarities in their habitats (sea water). Unlike therepresentatives of Thaumatomonas, which are found in

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Fig. 2. Flagellant pocket and furrow in Thaumatomastix sp.: (a) back flagellum, (b) nucleus and flagellant pocket, (c–e) flagellantpocket, (f) pseudopodium, and (g, h) transverse section of ventricular furrow; (fr) furrow, (m.b) microtubular band, and(ps) pseudopodium. Other symbols correspond to those used in Fig. 1.

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0.5 µm(a)

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Fig. 3. The structure of scales and kinetocysts in Thaumatomastix sp.: (a, b) somatic scales, (c) rudiment of somatic scale,(d, e) kinetocysts, and (f) kinetocyst and Golgi apparatus; (a.m) amorphous material. Other symbols correspond to those used inFigs. 1 and 2.

fresh water, the TM�5 clone has relatively big flagellantscales. In Thaumatomonas, these scales are repre�sented as small cone�shaped bodies [3]. It should bementioned that the division on such genera as Thau�matomonas and Thaumatomastix is quite relative, andnot all the investigators approve of it [8]. Particularly,both these genera have similar triangular and ovalscales with edge perforations [5, 10, 11].

There are only fragmentary data on the ultrafinestructure of Thaumatomastix representatives. Theinvestigations on Thaumatomonas genus are moreextensive [1, 2, 4, 6]. According to Moestrup’s data,[7] in Thaumatomastix (Reckertia) sagittifera, the for�mation of somatic and flagellant scales takes place onmitochondria surfaces that contain tubular cristae.The inside cell has one dictyosome, and kinetocystsare absent. Somatic scales lie in a row on the cell sur�

face. Small flat scales cover short flagellum. Mas�tigonemes cover long flagellum. Analogous structuresare found in the TM–5 clone.

The structure of the TM–5 clone’s cell is very sim�ilar to the cell structure of freshwater Thaumatomonaslauterborni de Saedeleer and Th. seravini Mylnikov etKarpov [1–4]. At the same time, in Th. seravini, like inthe TM�5 clone, somatic scales are triangular and haveedge perforations. Kinetosomes of the TM�5 cloneand Thaumatomonas lauterborni are parallel to eachother. However, in the latter they are not connected bya bridge. Ventricular furrows of the TM�5 clone andThaumatomonas have identical structures. They arearmored with two marginal bands of microtubules[1, 3, 4]. In the TM–5 clone these bands contain 8–12and 6–8 microtubules; in Thaumatomonas representa�tives their number is 5–6. The place where somatic

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scales are formed on the mitochondria surface is thesame both in the TM–5 clone and representatives ofThaumatomonas [2, 6, 8]. This is a distinct featureaccording to which all thaumatomonads differ fromother flagellates [1–4, 8].

Kinetocyst of TM�5, Thaumatomonas lauterborni,and Th. seravini have considerable resemblances.These organelles run from a bubble, inside which thereare capsule and muff with cylinder. Since the rudi�ments of kinetocysts are found near Golgi apparatus,it can be assumed that they are formed in the deriva�tives of this apparatus.

The differences in cell structures are the following.The transitional flagellant zone of the TM�5 clone hasno thin cylinder like Th. lauterborni and Th. seravinihave. Near the flagellant pocket’s wall in the TM�5clone, like in Th. seravini, there are at least three bandsof microtubules, whereas in Thaumatomonas lauter�borni four flagellant roots are found [1]. The cells ofthe TM�5 clone have one dictyosome of Golgi appara�tus, while Thaumatomonas cells have two dictyosomes[1, 3, 4]. Representatives of Thaumatomonas have twoalternately functioning contractile vacuoles [3]. Thesevacuoles are absent in TM–5, which is explained bythe fact that this organism lives in water. No cysts orplasmodia are discovered in TM–5 lifecycle, unlike inthe Thaumatomonas life cycle. However, plasmodiahave never been revealed among Thaumatomastix spe�cies [8, 10, 11].

Thus, the internal structure of the cell and the feed�ing trends in the TM–5 clone, representatives ofThaumatomonas, and the only fragmentarily studiedrepresentative of Thaumatomastix have substantialsimilarities.

The TM–5 clone resembles cells of Gyromitusplanktonic thaumatomonads in some cell structures[9]. They have similar tubular cristae of mitochondria,a similar place of scale formation (atypical for otherthaumatomonads) or plates with the beam, a similarfurrow position on cell back end, and an identical fla�gella length.

The sizes of scales, kinetocysts, and mitochondriaare almost the same. Two microtubular bands go insidethe walls of the ventricular furrow in Gyromitus, Thau�matomonas, and TM�5. The above� stated featuresshow that there are similarities in the cell structure ofthe studied thaumatomonads. A further investigationinto other representatives of Thaumatomastix willenable a more detailed comparison of their cell struc�ture with that of representatives of Thaumatomonasand make it possible to learn taxonomically importantdifferences between these two genera.

CONCLUSIONS

The cell structure of the TM–5 clone is studied.The TM�5 clone is rated among Thaumatomastixgenus because it has somatic and flagellant scales,mastigonemes on the back flagellum, two heterody�namic flagella, tubular mitochondria cristae, urticant

organelles (kinetocysts), and a ventricular furrow. Onthe basis of a similar thin�cell structure and feedingmanner, the resemblance between this flagellate andThaumatomonas and Gyromitus representatives isshown.

ACKNOWLEDGMENTS

This study was supported by the Russian Founda�tion for Basic Research, project no. 08�04�00244 and09�04�01150.

REFERENCES

1. Karpov, S.A., The Structure of the Flagellate Apparatusof the Colorless Flagellate Thaumatomonas lauterborniand Evaluation of the Concept of Evolutionary Conser�vatism of Cellular Structures, Tsitologiia, 1987, vol. 29,no. 12, pp. 1349–1354.

2. Karpov, S.A., Analysis of Orders Phalansteriida, Spon�gomonadida and Thaumatomonadida, Zool. Zh., 1990,vol. 69, no. 3, pp. 5–12.

3. Karpov, S.A., Ultrastructure of a Colorless FlagellateThaumatomonas seravini, Tsitologiia, 1993, vol. 35,no. 9, pp. 8–11.

4. Karpov, S.A. and Zhukov, B.F., Characteristics of theUltrastructure of the Colorless Flagellate Thaumatomo�nas lauterborni, Tsitologiia, 1987, vol. 29, no. 10,pp. 1168–1171.

5. Myl’nikov, A.P. and Karpov, S.A., The New Represen�tative of Colorless Flagellates Thaumatomonas seravinisp. nov. (Thaumatomonadida, Protista), Zool. Zh.,1993, vol. 72, no. 3, pp. 5–9.

6. Shirkina, N.I., Morphology and Life Cycle of Thau�matomonas lauterborni DE Saedeleer (Mastigorhoradiesing), in Fauna i biologiya presnovodnykh organizmov(Fauna and Biology of Freshwater Organisms), Lenin�grad: Nauka, 1987, pp. 87–107.

7. Moestrup, Flagellar Structure in Algae: a Review withNew Observations Particularly on the Chrysophyceae,Phaeophyceae (Fucophyceae), Euglenophyceae, andReckertia, Phycologia, 1982, vol. 21, no. 4, pp. 427–528.

8. Patterson, D.J., Vrs, N., Simpson, A.G.B., andO’Kelly, C.J., Residual Free�Living and PredatoryHeterotrophic Flagellates, in An Illustrated Guide to theProtozoa, Lawrence: Society of Protozoologists, 2000,pp. 1302–1328.

9. Swale, E.M.F. and Belcher, J.H., Gyromitus limax nov.sp.—Free�Living Colourless Amoebo�Flagellate,Arch. Protistenkd., 1975, vol. 117, nos. 1/2, pp. 20–26.

10. Thomsen, H.A., Hllfors, G., Hllfors, S., and Ikvalko,J., New Observations on the Heterotrophic ProtistGenus Thaumatomastix (Thaumatomastigaceae, Pro�tista Incertae Sedis), with Particular Emphasis onMaterial from Baltic Sea, Ann. Bot. Fennici, 1993,vol. 30, pp. 87–108.

11. Thomsen, H.A. and Ikvalko, J., Species of Thaumato�mastix (Thaumatomastigidae, Protista Incertae Sedis)from the Arctic Sea Ice Biota (North�East WaterPolynya, NE Greenland), J. Mar. Syst., 1997, vol. 10,pp. 263–277.

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