Coexistence of several Philasterides dicentrarchi strains on a turbot fish farm

Philasterides dicentrarchi is a histophagous scuticociliate parasite that causes one of the most severe diseases in the turbot, Scophthalmus maximus (L.), farmed in open circuit systems or in recirculation aquatic systems (RAS). In this study, we investigated the existence of intraspecific variation in P. dicentrarchi isolates obtained from outbreaks of turbot scuticociliatosis on a fish farm with a RAS. Ten isolates, obtained during two years of sampling, were morphologically characterized by examination of thirteen biometric parameters related to oral ciliature and three biometric parameters related to ciliate size. The data were analyzed by canonical discriminant analysis (CDA), which separated the P. dicentrarchi isolates into four morphotypes. Serological analysis of the isolates by western blot revealed the presence of three serotypes: “serotype D2/D6”, which includes isolates D2 and D6, D7 and D8, characterized by a 37kD band (non-reducing conditions) or a 50kD band (reducing conditions); “serotype D3”, including isolates D3, D4, D9 and D10, characterized by a broad band at 32–36kD (non-reducing conditions) or a band at 38–45kD (reducing conditions), and serotype D11, including isolates D11 and D13, characterized by the presence of several bands between 39 and 48kD (non-reducing conditions) and 30–32kD (reducing conditions). On the basis of the nucleotide sequences of amplicons of the β-tubulin gene, the isolates were grouped into three genotypes corresponding exactly to the D2/D6, D3 and D11 serotypes. We also verified the coexistence of two serotypes involved in the same outbreak of scuticociliatosis in the same tank. Competition tests between isolates indicated that none of the serotypes was capable of eliminating the others in vitro, while cross-protection assays confirmed the lack of heterologous protection between D2 and D3 isolates. This is the first report of the coexistence of several serotypes of P. dicentrarchi on a fish farm, and shows the need for detailed characterization of the intraspecific variability among these scuticociliates to enable production of an effective vaccine.