Studies on the monogenean, Entobdella hippoglossi Muller, 1776, parasitising a commercially important cultured fish, the Atlantic halibut, Hippoglossus hippoglossus Linnaeus, 1758.

Abstract

The skin monogenean parasite, Entobdella hippoglossi Muller 1776 (family Capsalidae) has been found to commonly occur on broodstock halibut during the development of the culture of Atlantic halibut, Hippoglossus hippoglossus L., 1758. Due to the lack of information relating to its host’s ecology and recent establishment of the halibut farming industry, research on E. hippoglossi is lacking. This study, therefore, was carried out to investigate the biological and pathological aspects of E. hippoglossi, on the skin of Atlantic halibut under culture conditions. A comparison of the parasite populations from two different sites, Machrihanish and Ardtoe, showed that the parasite burden from Machrihanish was twice that of the parasite population on Ardtoe halibut (641 ± 233.3 and 307 ± 276.6, respectively). The mean length of parasites, however, from the Ardtoe halibut was longer than those collected from the Machrihanish halibut (10.6 ± 3.3 mm and 6.0 ± 2.4 mm, respectively) (P< 0.05). The mean intensity of the parasite from female halibut was 605 ± 244.7, while from male hosts it was 231 ± 226.6 (P< 0.05). Also the mean length of parasites from female fish (8.4 ± 3.6 mm) was significantly longer than from male hosts (7.9 ± 3.8 mm) (P<0.05). While the parasite burden from the dorsal and ventral surfaces was almost the same, there was a significant difference in the mean length of parasites. The mean length of the parasites from the ventral surface was 9.6 ±3.7 mm while from the dorsal surface it was 6.9 ±3.1 mm (P<0.05). A comparison of the mean parasite length in four principal zones (front dorsal, middle dorsal, rear dorsal and ventral) on the fish revealed that the parasites from the ventral surface were significantly longer than those from the other zones (ventral: 9.7 ± 3.7 mm; front dorsal: 8.1 ± 3.3 mm; middle dorsal: 5.7 ± 1.9 mm and rear dorsal: 4.9 ±1.8 mm) (P<0.05). On the dorsal surface, the mean parasite length from the front zone was significantly bigger than those from the middle and the rear zones (P<0.05). The dominant length class of E. hippoglossi found from Atlantic halibut maintained at Machrihanish was 3-5 mm (30.9 %), whilst only 6.7 % of the parasites in the same size class were found on the fish at Ardtoe. In comparison, the majority of parasites in the Ardtoe population (26.1 %) measured from 11-13 mm. The same size class in the Machrihanish population was 3.6 %. The smallest size class (less than 3 mm) composed 0.98 % of the Ardtoe population whilst 6.6 % of the population were in this size class for Machrihanish halibut. The 5-7 mm group represented the dominant size class of parasites collected from the dorsal surface (27.7 %) whilst the majority of the parasites from the ventral surface belonged to the size class, 11-13 mm (20.9 %). The eggs of E. hippoglossi are discharged from the uterus by a very powerful contraction of the anterior part of the parasite. Three kinds of egg laying methods were observed. In the first type the eggs were laid in a chain-like manner which grouped to form an egg bundle. The second style was that eggs were laid attached to an egg ball. From one to hundreds of eggs attached to the egg ball which was then anchored on the bottom of the experiment vessel. The third method found was that some parasites laid eggs singly. The singly laid eggs were found to be sterile and did not develop any further. The egg laying rates of parasites on male and female hosts were compared. There was no significant difference between the mean number of eggs produced by parasites from male halibut (26.9 ± 10.4 eggs) and eggs produced by parasites from female halibut (35.2 ± 9.9 eggs). There was however, a significant difference between the mean number of eggs produced by parasites collected from the dorsal (8.9 ± 2.8 eggs) and the ventral surface (53.2 ±13.2 eggs). Newly laid eggs of E. hippoglossi were yellowish in colouration and tetrahedral in shape. The egg surface was pitted. Each side of the tetrahedron measured about 200 pm in length. An egg filament is attached to the proximal apex of the egg and is entwined at its free end within the egg bundle. The egg filament possesses buoy-like structures. These may contribute to eggs floating from the sea bed. The shape and size of these structures is totally different to the sticky droplets found from eggs of E. soleae. The operculum at one apex of the egg operates as a hatching gate for the emergent oncomiracidium. The larvae began to hatch naturally, without any hatching stimuli during illuminated or dark conditions, after 27-30 days incubation at 12°C. The anterior region emerged first through the operculum of the egg, while the posterior region was captured by the gap between the egg and operculum. Whether the oncomiracidia were trapped or they simply paused there for a certain purpose is not clear. Once they were released from the gap, they actively and freely swam. The larvae had 3 zones of cilia corresponding to the anterior, the middle and the posterior regions of the body. The oncomiracidium had 4 eye-spots with pigmented cups and lenses. The haptor bore 3 pairs of medianly situated sclerites, the anterior hamuli, the posterior hamuli and the accessory sclerites, and fourteen marginally situated sclerites, the marginal hooklets. The oncomiracidium of E. hippoglossi has a total of 64 epidermal ciliary plates, comprising 27 cells on the anterior region, 20 cells on the middle region and 17 cells on the posterior region. In comparison with those from the oncomiracidium of E. soleae, the anterior and the middle regions were the same in number but E. soleae had only 13 ciliary cells on the posterior region. When the oncomiracidia were placed on the top of a 150 cm glass tube filled with sea water, they actively swam downwards. Of 20 parasites, 11 (52.3 %) reached their final destination within 8 minutes, four parasites (19 %) arrived around 11 minutes. Five parasites (23.8 %) swam actively but they changed their direction often before they reached the bottom (13-27 minutes), while one parasite did not move at all in a 30 minute period. The average swimming speed of the oncomiracidium was 0.32 ±0.1 cm/second (3.1 ± 1.8 sec/cm) throughout the experiments. The average swimming speed of parasites in the first 3 minutes as they headed downwards was 0.30 ± 0.12 cm/sec (3.3 ± 1.88 sec/cm) and that of the parasites after 3 to 6 minutes was 0.32 ± 0.12 cm/sec (3.1 ± 1.92 sec/cm). The average swimming speed of the parasites after 6 minutes was 0.37 ± 0.20 cm/sec (2.7 ± 2.2 sec/cm). The oncomiracidium showed an upward and downward movement continuously within 10 cm of the bottom of the experimental vessel, continuing until they became moribund. This behaviour seems to have a very important role in searching for its flat fish host. A positive photo - response was observed in oncomiracidia of E. hippoglossi. The responses of 50 larvae were recorded, 43 of them (86 %) were photo-positive (P<0.05). Four larvae were found to be photo-negative and 5 others showed no clear cut directional responses. In a second test, using halibut mucus taken from a mature halibut as a stimulus, the responses of 50 freshly hatched larvae were recorded. Twenty four out of a total of 50 oncomiracidia (48 %) responded positively to halibut mucus (P<0.05) while 21 of the larvae (42 %) did not respond clearly and 10 % of these showed a negative selection for halibut mucus. When a test was carried out dividing mucus and light as different stimuli, eighty percent of the parasites moved towards the light (P<0.05) and 14 % of the parasites went towards the mucus side arm, while 6 % of the parasites did not show a clear response to either mucus or light. Experimental infection of juvenile halibut with adult parasites was carried out. Almost all of the parasites were found to move to the head region of the ventral surface, whether they were placed on the dorsal surface or on the ventral surface. Some parasites settled there for at least 8 weeks whilst other parasites seemed to detach from the juvenile halibut. When the experiment was terminated, no oncomiracidia or juveniles were found on the juvenile halibut, although 8 weeks at 12°C was enough time to for this parasite to hatch. Skin haemorrhages were found on experimentally infected juvenile halibut. A mass of sloughed necrotic epithelial cells mixed with mucus and debris was found on the surface of infected tissue. In the infected tissue, the mucous cells were irregularly distributed in the epidermis. An SEM study revealed the sucking action of the opisthaptor, indentation by papillae of the parasite and lesions caused by the accessory sclerites on the juvenile halibut skin. It seems that the papillae might act as grips in attachment of the parasite. The arrangement of papillae showed quite a regular distribution on the ventral part of the haptor. The accessory sclerites of E. hippoglossi are quite different from those of E. soleae. The accessory sclerites of E. hippoglossi have a very sharp end compared to the smooth curved ending in E. soleae. Because of their deep penetration they may act as hooks rather than just acting as props as has been suggested for E. soleae. Generally, the mucous cell size, epidermal thickness and mucous cell number of the front region of mature halibut skin, whether on the dorsal or ventral surface, were all greater than those of the other regions. On juvenile halibut, however, the mucous cell sizes from the rear and the middle regions were significantly bigger than those from the front region of the dorsal surface. The distribution and concentration of mucous cells on the halibut skin were clearly positively correlated with the parasite population on the halibut host.