Role of scuticociliate proteinases in infection success in turbot, Psetta maxima (L.).

Scuticociliatosis caused by Philasterides dicentrarchi is one of the most severe diseases of farmed turbot, Psetta maxima (L.). Immunized fish showed elevated levels of specific antibodies (Ab), which caused the destruction of parasites through the activation of complement by the alternative and classical pathways. By using affinity chromatography on bacitracin-sepharose columns, we demonstrated the existence of high levels of parasite proteinases in the serum and, to a lesser extent, in the ascitic fluid of experimentally infected fish, and the absence of such proteinases in the serum of uninfected fish. Serum from uninfected fish displayed haemolytic activity against sheep red blood cells. However, incubation of this serum with parasite proteinases led to a decrease in serum haemolytic activity, suggesting that proteinases are able to destroy fish complement. Proteinases isolated from serum or ascitic fluid of infected fish were also able to degrade turbot Ab. Preincubation of turbot serum containing specific Ab for P. dicentrarchi with the proteinases led to a significant decrease in the killing activity of the serum. The results confirm that P. dicentrarchi proteinases in serum from infected fish may provide a mechanism for circumventing normal host immunity by inactivating the Ab and complement factors required for complement activation.

Antigenic and cross-protection studies on two turbot scuticociliate isolates.

The protection induced in turbot by inactivated vaccines containing either of two isolates (I(1) and C(1)) of the scuticociliate parasite Philasterides dicentrarchi, which causes important mortalities in turbot cultures, was evaluated in the present study. The results obtained after challenging the fish with the two isolates show that vaccination protected fish only against the homologous isolate, but did not confer cross-protection. The two isolates constitute two serotypes, as shown in the immobilization tests with mouse and turbot anti-I(1) and anti-C(1) antisera, in which only the homologous antisera immobilized the ciliates. ELISA assays, using total antigen free of proteases (TAWP), cytosolic antigens (CYA), ciliar antigens (CA) or membrane protein fraction (MPF), were also carried out. Differences in the levels of antibodies produced in mouse against the homologous and heterologous antigens were observed; these differences were significantly different when the antigen preparations used in the ELISA were TAWP, CYA or CA. Nevertheless, ELISA assays using turbot sera against TAWP did not show significant differences in the levels of antibodies against the homologous and heterologous antigens. Antigenic cross-reactivity was also detected in the Western blot assays, as well as significant differences in the patterns of antigenic recognition in the two isolates - in both reduced and non-reduced TAWP antigens, but which was noteworthy when mouse antisera were used. The results obtained in the present study demonstrate for the first time the existence of serotypes of the ciliate parasite of turbot Philasterides dicentrarchi that display clear antigenic differences, which must be taken into consideration in the future development of a vaccine against scuticociliatosis.