First record of Trichodina heterodentata ( Ciliophora : Trichodinidae ) from channel catfish , Ictalurus punctatus cultivated in Brazil

This study characterises morphologically Trichodina heterodentata Duncan, 1977 from channel catfish, Ictalurus punctatus (Rafinesque, 1818) in the State of Santa Catarina, Brazil. Body and gill smears were air-dried at room temperature, impregnated with silver nitrate and/or stained with Giemsa. Ten characteristics were selected to compare the present material with other morphological characterisations of T. heterodentata. Prevalence rate was 100%, mean intensity 89,333.70 (3,125 to 299,100 parasites per host). Trichodina heterodentata was considered medium-sized trichodinid with mean body diameter 59.4 ± 8.5 μm, denticulate ring 38.5 ± 4.5 μm, adhesive disc 60.2 ± 6.7 μm diameter and 24.4 ± 1.6 denticles. In relation to previous reports of T. heterodentata this material resembles in 90% of the analysed characters. This work confirms the biometrical variation that exists in the different populations of T. heterodentata. A list of hosts and comparative measurements of T. heterodentata are presented and the channel catfish is considered a new host.


Introduction
Trichodinids are among one of the most common ectoparasites in wild and cultivated fish (Basson andVan As, 1994, Martins andGhiraldelli, 2008).This parasite occurs normally in a few numbers on mucous surface and gills.When the relationship host/parasite/environment is broken by nutritional deficiency, poor water quality, infectious and/or parasitic diseases trichodinids may proliferate being responsible for severe epidermal lesions and disease outbreaks, as reported by Madsen et al. (2000), Martins et al. (2002), Khan (2004) and Huh et al. (2005).

Material and Methods
Four specimens of channel catfish of 29.5 ± 2.3 cm total length were collected in July 2007 in a farm situated in the municipality of Porto União (26° 14' 17'' S and 51° 04' 42'' W), Santa Catarina, Brazil.Fish were maintained at 1.4 fish.m -2 stocking density in ponds of 3,500 m 2 with constant flow of water.In the day of sampling the pH was 5.4 to 6.9; alkalinity 24 to 44 mg.L -1 ; total ammonia 0.5 to 3.0 mg.L -1 ; dissolved oxygen 6.2 to 10.8 mg.L -1 .Water temperature three days before fish sampling was 13.3 to 15.2 °C and on the day of parasitological examination, it was 9.2 to 10.8 °C.
Wet smears of skin and gills were prepared in the field and examined under the microscope.When parasites were present the smears were air-dried and impregnated with Klein's dry silver method for observation of the adhesive disc as suggested by Lom (1958).Other smears were stained with Giemsa's solution to reveal the nuclear apparatus.The span of the denticle was measured from the tip of blade to the tip of ray as described by Arthur and Lom (1984).The body diameter is the dimension of the adhesive disc plus the border membrane.Wet mounts from the specimens preserved in 5% formalin solution were studied for the observation of adoral ciliature.All measurements are in micrometres and follow the recommendations of Lom (1958) and Van As and Basson (1989).Arithmetic means ± standard deviation is followed, in parentheses, by the minimum and maximum values and number of specimens or structures measured.To compare biometrically our specimens of T. heterodentata with others from previous records, 10 characteristics were selected as the most important in a primary observation before morphological comparison, which are: body size, adhesive disc, denticle ring, number of denticles, denticle length, blade length, ray length, central part width and span.
Characterised as a medium-sized trichodinid with disc-shaped body; convex adoral surface with ciliature of 307.0° ± 12.6 (292-325, 11); aboral side with slightly concave adhesive disc; centre of adhesive disc did not show granules in silver-impregnated specimens.From the Giemsa-stained specimens the macronucleus also presents a horseshoe-shaped 43.2 ± 4.7 (32-49, 26) of external diameter and 11.0 ± 2.2 (7-17, 26) thickness.The distance between the terminations of macronucleous is 17.6 ± 8.4 (11-27, 26); micronucleus was not observed.The denticles are characterised by wide blade and sickleshaped provided by apophysis on blade connection with the central part (Figure 1a).The blade fills the space between y and y -1 axes.Central part robust ending rounded fitting on the next denticle filling the space between y and y -1 axes.Ray long, robust, slightly directed anteriorly situated between the axes y and y + 1, tapering rounded provided by a relatively short apophysis (Figure 1b).Several specimens studied showed inconstant shape and length of ray.

Remarks
Trichodinids reproduces by binary fission and it has been the subject of study since the previous century (Kruger et al., 1995).Figure 2 shows one of the last steps of T. heterodentata`s reproduction, showing blades in the process of development whilst the old ring is being resorbed.According to Kruger et al. (1995), the original denticle ring maintains its position but loses its shape.The altering of the original shape of the denticle ring is the result of the resorption process that appears to erode the denticles from the aboral side.
Comparing the measurements of the specimens herein described, 50% out of 10 characters were similar to  (Duncan, 1977).Later, it was also recorded from a large number of fish species, including several Families (Table 1-4).In siluriform fish, T. siluri Lom, 1970 was reported by Bondad-Reantaso and Arthur (1989) but almost all measurements except for blade length were different.
The present specimens showed similar measurements of adhesive disc, denticulate ring and the dimensions of denticles when compared to population B and C of Duncan (1977) and those studied by Van As and Basson (1989).A lower number of denticles was reported in specimens described by Van As andBasson (1989, 1992), but a higher number was noted in two populations originally described by Duncan (1977) (Table 1).In fact, comparing the present material from channel catfish with T. heterodentata, a great number of characters measured coincided.Another point that must be commented is the position of ray and blade in relation to y axes.As demonstrated by illustrations of Van As and Basson (1989Basson ( , 1992) ) the blade fills the space between y and y -1 axes.The central part is robust filling the space between y and y -1 axes.Finally, the length of ray is longer than the length of blade slightly directed anteriorly situated between the axes y and y + 1.
This species resembles the size of Trichodina vallata Wellborn, 1967 described from channel catfish (Wellborn, 1967).On the other hand, the adhesive disc and the denticulate ring diameters, as well as the number of denticles were lower than that observed in this work.In relation to Trichodina discoidea also described by Wellborn (1967) in channel catfish the number of denticles and the length of blade and ray were similar, but significant lower diameters of body, adhesive disc and denticulate ring were found.Despite the fact that it occurs in wild and cultured channel catfish, the shape of the denticle of T. discoidea and T. vallata was significantly different when compared to T. heterodentata herein characterised.
Due to the fact that the denticles may present some variability, as supported by Van As and Basson (1989) and Basson and Van As (1994), the specimens herein studied correspond to another report of T. heterodentata in Brazil and channel catfish is reported as a new host.As to confirming the variation that exists in T. heterodentata populations, the present material showed slightly higher measurements of the adhesive disc, denticulate ring, central part width and span than those found in T. heterodentata reported by Al-Rasheid et al. (2000) and Asmat (2004), and higher denticulate ring diameter when compared to Dove and O'Donoghue (2005) reports.The characterisation of T. heterodentata from tadpoles (Dias et al., 2009) showed lower measurements of denticulate ring and denticle length.

Figure 1 .
Figure 1.a) Photomicrographs of Trichodina heterodentata from Ictalurus punctatus cultured in the State of Santa Catarina; and b) Schematic drawing of the denticles of T. heterodentata.
Width; L Length; *the number of structures measured.

Table 3 .
Measurements of the most similar species of Trichodina with the present material.The means are followed by standard deviation, and in parenthesis the minimum and maximum values.

Table 4 .
Measurements of the most similar species of Trichodina with the present material.The means are followed by standard deviation, and in parenthesis the minimum, maximum values and the number of structures measured.