Formulation of fish vaccine based on lipidic nanovesicles, in particular, a proteoliposome or cochleate, with activity against the salmonid rickettsial syndrome (srs)

Method to prepare a formulation of fish vaccine based on lipidic nanovesicles, especially, a proteoliposome and cochleates, with activity against the Salmonid Rickettsial Syndrome (SRS) CHARACTERIZED THAT it includes:

• a) to culture P. salmonis, harvest it by centrifugation and store it frozen;
• b) purify membranes and fragments of cell wall from the frozen sediment of the stage a) by resuspending the frozen sediment in a buffer solution adding pearls of zirconia/silica, to then freeze, thaw by sonication and freeze until complete 7 repetitions of the cycle freeze-thaw-freeze, then to centrifugate reserving supernatant and discarding sediment, and then centrifugate reserving sediment and discarding the supernatant,
• c) prepare membrane proteoliposomes from the frozen sediment of step b) resuspend the sediment in a membranes solubilisation solution, incubate with agitation, centrifugate and preserve the supernatant, then add non-polar resins able to capture detergent previously resuspended in solution and sterilised, to subsequently incubate with agitation and to add those non-polar resins again to incubate with agitation, leave to settle, preserving the supernatant,
• d) prepare membranes from the frozen sediment of step b) resuspend the sediment in a physiological solution or physiological saline, incubate with agitation, centrifugate preserving the supernatant,
• e) prepare membrane cochleates from the frozen sediment of stage b) resuspend the sediment in a solubilisation solution (TLis), incubate with agitation, centrifugate and preserve the supernatant. This supernatant is added by dropping at equal volume of formation solution, then 4 volumes of washing solution are added. Finally, the cochleates are centrifuged, discarding the supernatant and the sediment resuspended in washing solution, and
• f) the membranes, fragments, membranes proteoliposomes and membrane cochleates obtained in steps c) to e) are mixed with physiological saline, under agitation to an adjuvant and homogenise the mixture, to then optionally store in sealed containers.