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Protistology 6 (4), 258-263 (2010/11)
Protistology
Morphological, ecological and histopathological studies of Trichodina gobii Raabe, 1959 (Ciliophora: Peritrichida) infecting the gills of Solea aegyptiaca
Chiraz Yemmen1, Yann Quilichini2, Mohamed Hédi Ktari1, Bernard Marchand2 and Sihem Bahri1
Specimens of Solea aegyptiaca caught from Ghar El Melh lagoon in Tunisia were investigated for trichodinid ectoparasites. One species of the genus Trichodina was recorded on the gills of S. aegyptiaca. This species is identified as Trichodina gobii Raabe, 1959 which is reported for the first time on the soleidae fishes from Tunisian waters. The prevalence and the mean intensity level of these ciliophorans varied throughout the year with maximum rate of infection during the winter. Taxonomic descriptions of this species based on silver nitrate method and scanning electron microscopic observations are provided in this paper. Histopathological examinations of gills infested with T. gobii, revealed several lesions such as hyperplasia and lamellar epithelium lifting of the secondary lamellae.
Introduction
Trichodinids are one of the largest and most widely dispersed groups of ectoparasites in freshwater, marine and euryhaline environments. Today more than 260 species representing ten genera were described within the family of Trichodinidae from the skin, gills, urinary bladder of fishes, amphibians and from the integument and the genital tract of
invertebrates (Lom and Dykova, 1992). Some species inhabiting inhabit also the digestive tract of marine fishes (Da Cunha and Pinto, 1928; Basson et al., 1990). The genus Trichodina is the largest of this family; about 200 species of Trichodina have been described from fishes by silver impregnation technique (Asmat et al., 2005). In the case ofmassive infestation, some species of Trichodinids become highly pathogenic and may cause severe damage
© 2010 by Russia, Protistology
and even the death of their host (MacArdle, 1984; Hassan, 1999). In Tunisia, there are a few studies mentioned the presence of Trichodinids on marine fishes, all of them without presenting any taxonomic study. The aim of the present work is to identify the Trichodina species that occur on the gills of S. aegyptiaca - one of the important economical fish in Tunisian waters, to follow the variations of the prevalence and the mean intensity of this parasite during the investigation period, and to evaluate its pathogenicity.
Material and methods
From September 2009 to August 2010, a total number of130 specimens of S. aegyptiaca measuring 16.2 to 28.4 cm were caught by gill net from Ghar El Melh lagoon which is situated in Northeast of Tunisia (37°10& N and 10°09& E). During the investigation period, host fishes were transported alive in local water directly to the laboratory. Wet smears ofgills were prepared and examined in order to detect the presence of Trichodinids. Smears from infested infected fishes were air-dried and impregnated for 10 min in 2 % aqueous AgNo3 solution (Klein’s silver impregnation technique) (Klein, 1958), washed in distilled water, and exposed to ultraviolet light for 20 to 25 min, in order to study details of the adhesive disc and the aboral ciliary spiral. Preparations and photographs were made under a Nikon E 600 microscope. All measurements are in micrometers and based on 30 trichodinid specimens and follow the uniform specific characteristics given by Lom (1958). The description of denticule elements follows the format recommended by Van As and Basson (1989). The intensity of infestation and the seasonal prevalence of parasites were calculated according to Bush et al. (1997).
For histological study, gills were fixed in 4% formaldehyde and processed using standard technique with haemotoxylin-eosin staining and examined microscopically.
For scanning electron microscopic study, gills were fixed for 1 hour in 2.5% glutaraldehyde buffered with 0.1 M sodium cacodylate at pH 7.2, then dehydrated in a graded acetone series and dried using CO2 in an Emitech K850 critical point dryer. After mounting, gills were coated with gold/ palladium in a Quorum Technologies SC7640 sputter coater and examined with a Hitachi S-3400N scanning electron microscope at an acceleration voltage of 10 kV.
Morphological data of Trichodina gobii Raabe, 1959
All collected specimens from the gills of Solea aegyptiaca are assigned to T. gobii. Parasite presents a medium size with 36.8-40.8 (38.5 ± 1.4) ^m in diameter. Adhesive disc concave, 27.2-32.8 (29.8 ± 1.7) ^m in diameter, with a well defined central circle 7.2-9.6 (8.3 ± 0.9) ^m and surrounded by a border membrane of 3.2-4.5 (3.6 ± 0.5 ) ^m in width (Fig. 1, Table 1). Diameter of denticulate ring 20.229.6 (24.6 ± 2) ^m, consists of20-23 denticles, with length of 6.2-7 (6.4 ± 0.6) ^m. Blade 4-5.6 (4.6 ± 0.4) ^m less curved and more or less rectangular, slightly narrowing to rounded distal ends and filling large portion of sectors between y and y-1 axes; tangent point almost rounded, situated slightly lower than distal margin. Posterior margin curve forms small narrow arch with y axes; deepest point on same level as apex. Section connection blade is relatively thick. Blade apophysis and posterior projection are absent. Sections of central part above and below x axis are similar in shape. Central part is thick with rounded point extending more than half way to y axis. Thorns 2.4-4 (3.24 ± 0.2) are slender and smooth with rounded end. Apophysis of thorn is not visible. Section connection of thorn is thinner than those connecting blade. Number of radial pins per denticles is 7-8.
SEM observations ofthe ciliate show a spherical shape in oral view. The surface topography was smooth (Fig. 2). There are two ciliary bands well developed, one adoral and the other around the aboral adhesive disk (Fig. 3). The adoral ciliary band is composed of two rows of closely-set cilia. The buccal region is therefore flanked by two bands of cilia (Fig. 2). The adhesive disk is surrounded by a well developed border membrane, and contains fine vertical stria over its entire surface (Figs 3, 4). These strias on the internal surface of the border membrane are the radial pins.
Prevalence and Mean intensity
The prevalence and the mean intensity of T. gobii fluctuate during the sampling period. Indeed, the prevalence ranged from 12.03 % to 84.4 %. The prevalence reached its maximum in winter, when water temperature was low (12 °C), then decreased along with increased temperature and reached its minimum in summer when the temperature was at maximum (29.33 °C) (Fig. 5).
Figs 1-4. Photomicrographs of Trichodina gobii from the gills of Solea aegyptiaca. 1 — Silver impregnated adhesive disc of T. gobii; 2-4 — SEM micrographs of T. gobii: 2 — oral view showing adoral spiral entering infundibulum; 3 — aboral view showing border membrane (b) and ciliary band (c); 4 — aboral view showing denticulate ring. Scale bars = 12^m.
The mean intensity level of T. gobii has increased from 29.27 ± 3.08 in autumn and reached a peak value of 117.66 ± 29.5 in winter, when the water temperature was low, then mean intensity decreased gradually with the increasing of temperature and reached its minimum in summer (11.63 ± 1.04) (Fig. 6).
Histopathology
Histopathological examination of infested fishes infested by with the present species, revealed significant gill changes. In most cases fusion of secondary lamellae and hyperplasia were observed (Fig. 7). We noted also, an intense lamellar epithelium lifting (epithelium detachment) in secondary lamellae (Fig. 8). In addition to these lesions, an excessive accumulation of mucus on gills was also seen on gills of in infested fishes.
Discussion
The present species of Trichodina collected from the gills of S. aegyptiaca, was compared with previously described Trichodinid species with similar morphological characters. T. jadranica Raabe, 1958 is one of the most frequently reported trichodinids from various marine fishes, especially flatfishes.
Based on the silver impregnated specimens, the species of Trichodina collected from S. aegyptiaca are very similar to T. jadranica by the well defined central circle, a low number of denticles and a relatively smaller body. However, our species is different by less curved and more or less rectangular blades slightly narrowing to rounded distal ends. The denticle blades of T. jadranica are distinctly curved, almost sickle-shaped with a sharp tangent point distally. The present species shows similarity in the denticles shape which is almost rectangle-like blades with T. gobii Raabe, 1959 originally described from Gobius minutes from the Baltic Sea. The similarity in the denticle shape and dimensions of the present species is also shared with the others populations of T. gobii already described from various hosts (Table 1). The SEM observations of T. gobii are reported for the first time in this study. The ciliate was spherical in shape and the surface topography was smooth, typically for the most Trichodinids (Qi et al., 1995).
It seems that T. gobii has low host specificity
Fig. 5. Seasonal variations of temperature (°C) and prevalence of infection of T. gobii.
Fig. 6. Seasonal variations of temperature (°C) and mean intensity level of T. gobii.
and a widely geographical distribution, it can occur It seems that the high temperature ofwater, prevent
in both marine and freshwater environments. This the proliferation of the ciliate. The present finding
study reports for the first time the presence ofT. gobii agree with those of McArdle (1984), Abu El-Wafa
on soleidae fishes from Tunisian coast. (1988), El-Khatib (1989) and Hassan (1999), who
T. gobii was present throughout the year with mentioned that trichodinids were prevalent all over
higher rate of infestation during the coldest season. the year with maximum rate of infestation during
Table 1. Morphometric data (in micrometers) of Trichodina jadranica and Trichodina gobii from different hosts.
Raabe (1958) Raabe (1959) Lom (1970) Grupcheva et al. (1989) Su and White (1995) Madsen et al. (2000) This paper
Species T. jadranica T. gobii T. gobii T. gobii T. gobii T. gobii T. gobii
Host Mullus barbatus Gobius minutus Callionymus lyra Crenilabrus ocellatus, C. griseus, Neogobius malanostomus Nesogobius sp. Anguilla anguilla Solea aegyptiaca
Site Gills Gills Gills Gills Gills Gills Gills
Locality Adriatic Sea Baltic Sea Brittany coast of France Black Sea, Blalchik Tasmania, Australia Eel farm, Denmark Lagoon of Ghar El Melh, Tunisia
Body diameter 34-43 25-35 28-41 (36) 35-50 40-55(45.1) - 36.8-40.8 (38.5 ± 1.4)
Adhesive disc 28-38 25-35 (30) 22-31 (27) 20-27 24-37(32.3) 31-41(37±4.1) 27.2-32.8 (29.8 ± 1.7)
Border membrane width - -- 3 2,9-3,6 3.1 - 3.2-4.5 (3.6 ± 0.5 )
Denticle ring diameter 16-22 13-18 (15,5) 14-18 (16) 12,5-16 13-21(16 .6) 19-29(23±2.8) 20.2-29.6 (24.6 ± 2)
Denticle number Radial pins per denticle 22-25 8 19-23 (20-21) 6 18-23 (22) 6-7 19-21 6 20-24(23) 6-9(7) 24-27(26±0.9) 6-8 20-23 7-8
Denticle Span - - - 7.3-7.8 - - Denticle length - - 4,5-5 2.9-4.3 4-5(4.5) - 6.2-7 (6.4 ± 0.6)
Blade length 3-4 3.5-4.5 (4) 2.2 - 3 2.9-3.4 4-5(4.7) - 4-5.6 (4.6 ± 0.4)
Central part width - - 1.5 1.5 1-1.5 (1.1) - 1-1.5
Ray length 2.5-3 3-4 (3.5) 2-2.8 2.9 3-4 (3.7) - 2.4-4 (3.24 ± 0.2)
Central circle diameter - - - - 7-8 (7.5) - 7.2-9.6 (8.3 ± 0.9)
Figs 7, 8. Histopathology of infested fishes. 7 — Histological section of infested gills showing pronounced hyperplasia in secondary lamellae; 8 — intense lamellar epithelium lifting in secondary lamellae (white arrow) (T. gobii indicated by black arrow).
winter. Fluctuations ofprevalence and mean intensity of T. gobii observed in this study are connected in relation with the ecological conditions such as temperature, oxygen and salinity. This result has been mentioned by Maslin-Leny (1988), Hassan (1999), Ogut and Palm (2005) and Kristmundsson et al. (2006).
In the present study it is shown, that the infestation with T. gobii caused serious pathological lesions in gills, such as hyperplasia of the epithelial cells, clubbing and fusion of the gill filaments, which have been previously reported by many authors (Padnos and Nigrelli, 1942; Davis, 1947; Sarig, 1971; Hassan, 1999). In addition to these lesions, massive production of mucus was observed on infested fishes as a defence mechanism produced by the host to eliminate the parasite.
Acknowledgements
The authors are grateful to Dr. Kuidong Xu, Institute of Oceanology, Chinese Academy of Sciences, 266071 Qingdao, China, for his help in the identification of Trichodina species found in this study.
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Address for correspondence: Chiraz Yemmen. Unité de Biodiversité et Biologie des Populations. Département de Biologie, Faculté des Sciences de Tunis, Université Tunis-El Manar; e-mail: chiraz._yemmen@yahoo.fr
