Chemical Attenuation of Miamiensis avidus Using Oxytetracycline Reduces Virulence and Confers Partial Protection in Olive Flounder ( Paralichthys olivaceus )

Scuticociliatosis caused by the ciliate Miamiensis avidus remains a serious constraint in the intensive culture of olive flounder ( Paralichthys olivaceus ), yet effective vaccines are still unavailable. In this study, we evaluated chemical attenuation as a novel approach to generate a safe whole‐cell vaccine candidate against M. avidus . Ciliates were attenuated by oxytetracycline (OTC) treatment, and the safety, residual virulence and protective potential of the chemically attenuated preparation were assessed in vitro and in vivo. Treatment with 5 mM OTC for 3 h suppressed ciliate proliferation while preserving motility and morphology. The attenuated ciliates exhibited markedly reduced cytopathic effects on EPC feeder cells and caused no mortality in olive flounder, confirming successful attenuation. Fish immunised with the chemically attenuated whole‐cell vaccine candidate of M. avidus showed reduced cumulative mortality following experimental challenge compared with PBS‐injected controls and fish immunised with non‐adjuvanted formalin‐killed cells, particularly when a booster immunisation was administered. Serum agglutination titre was significantly different between the PBS control and vaccinated groups, but not among the vaccinated groups. Overall, these results suggest, for the first time, that chemical attenuation can be applied to a scuticociliate parasite to generate a safe whole‐cell vaccine candidate that provides partial protection. This proof‐of‐concept study highlights the potential of chemical attenuation as a foundation for further optimisation of vaccine strategies against scuticociliatosis.