Lumpfish (Cyclopterus lumpus) has become the predominant cleaner fish species used in North American salmon aquaculture for sea lice (e.g., Lepeophtheirus salmonis) biocontrol. Lumpfish utilization has contributed significantly towards eliminating the utilization of chemotherapeutants by effectively controlling the abundance of this damaging pest of Atlantic salmon (Salmo salar) aquaculture. Vibrio anguillarum is a frequent pathogen of lumpfish in Atlantic Canada. Here, several vaccine trials against V. anguillarum were conducted. We determined that the V. anguillarum growth conditions are essential for expressing protective antigens in the vaccine formulation. Generic commercial vaccines provide between 2-55% protection against V. anguillarum, not adequately protecting the lumpfish. Generic or autogenous vaccines delivered by mucosal routes (e.g., dip or bath) stimulate the naïve fish. However, only systemic immunization regimes (e.g., intraperitoneal injection) induced significant protective immunity against the lethal V. anguillarum systemic or mucosal challenge. Mucosal immunization conferred an evident immune stimulation but not immune protection. Transcriptome analysis of immunized fish revealed an evident difference in the transcriptome profile of the spleen and the head kidney. A modest number of DEG was detected in the head kidney, spleen from single immunized animals, and head kidney from boosted fish. A more significant number of DEGs was detected in the spleen of boosted animals. Common DEGs were observed between the boosted organs but not in single immunized organs. Gene ontology analysis in the boosted spleen revealed enrichment for humoral immune response, plasma lipoprotein particle, vitamin transport, metallocarboxypeptidase activity, among other immune-related gene groups. Hepcidin and c6 complement encoding genes were consistently upregulated in all organs studied and might be valuable biomarkers of effective vaccination.