Aluminum adjuvant potentiates gilthead seabream immune responses but induces toxicity in splenic melanomacrophage centers

A key goal of a successful vaccine formulation is the strong induction of persistent protective immune responses without producing side-effects. Adjuvants have been proved to be successful in several species at inducing increased immune responses against poorly immunogenic antigens. Fish are not the exception and promising results of adjuvanted vaccine formulations in many species are needed. In this study, over a period of 300 days, we characterized the apparent damage and immune response is gilthead seabream immunized by intraperitoneal injection with the model antigen keyhole limpet hemocyanin (KLH) alone or formulated with Montanide ISA water-in-oil (761 or 763), or Imject (TM) aluminum hydroxide (aluminium), as adjuvants. Throughout the trial, external tissue damage was examined visually, but no change was observed. Internally, severe adhesions, increased fat tissue, and hepatomegaly were recorded, but, without impairing animal health. At 120 days post priming (dpp), histopathological evaluations of head-kidney, spleen and liver revealed the presence of altered melanomacrophage centers (MMC) in HK and spleen, but not in liver. Surprisingly, in all aluminium treated fish, classical stains unmasked a toxic effect on splenic-MMC, unequivocally characterized by a strong cell depletion. Furthermore, at 170 dpp transmission electron microscopy confirmed this data. Paradoxically, at the same time powerful immune responses were recorded in most vaccinated groups, including the aluminium treatment. Whatever the case, despite the observed adhesions and MMC depletion, fish physiology was not affected, and most side-effects were resolved after 300 dpp. Therefore, our data support adjuvant inclusion, but strongly suggest that use of aluminium must be further explored in detail before it might benefit the rational design of new vaccination strategies in aquaculture.