MORE THAN JUST ANTIBODIES: PROTECTIVE MECHANISMS OF A MUCOSAL VACCINE AGAINST FISH PATHOGEN FLAVOBACTERIUM COLUMNARE  

A recently developed attenuated vaccine (17-23) for Flavobacterium columnare has been demonstrated to provide superior protection for channel catfish, Ictalurus punctatus, against genetically diverse columnaris isolates. First, we were interested in elucidating the host responses generated by a virulent (BGSF-27) or attenuated (17-23) isolate utilizing RNA-seq transcriptome analyses. We hypothesized that a comparison of signatures between isolates would reveal mechanisms of pathogen attachment and invasion as well as patterns of immune evasion or host manipulation linked to virulence. Thirty day old fry were accordingly challenged with either virulent or vaccine isolates and gill tissues collected at 1 h and 2 h post infection, time points previously demonstrated to be critical in early host-pathogen interactions. Gene pathway analyses suggested potent suppression of early host immune responses by the virulent isolate through higher expression of nuclear receptor co-repressors (NCOR) responsible for antagonizing macrophage and T cell signaling.  Conversely, in vaccinated fry, we observed induction of Ca²⁺/calmodulin-dependent protein kinase II (CAMKII), responsible for clearing NCOR, and commensurate up-regulation of transcription factor AP-1 subunits, c-Fos and c-Jun.  In a secondary challenge, 58 day old fingerling catfish (28 days post-vaccination or unvaccinated control) were challenged with a virulent F. columnare isolate (BGSF-27) and gill tissues collected pre-challenge (0 h), and 1 h and 2 h post infection. In this study, differential expression analysis within and between treatments revealed several patterns and pathways potentially underlying improved survival of vaccinated fish.  Most striking was a pattern of dramatically higher basal expression of an array of neuropeptides (e.g. somatostatin), hormones, complement factors, and proteases at 0 h in vaccinated fish. Previous studies indicate these are likely the preformed mediators of neuroendocrine cells and/or eosinophilic granular (mast-like) cells within the fish gill. Following challenge, these elements fell to almost undetectable levels (>100-fold downregulated) by 1 h in vaccinated fish, suggesting their rapid release and/or cessation of synthesis following degranulation.  Concomitantly, levels of pro-inflammatory cytokines (IL-1b, IL-8, IL-17) were induced in unvaccinated fish.  In contrast, in vaccinated catfish, we observed up-regulated MHC class II responses and widespread induction of genes needed for collagen deposition and tissue remodeling.  These results and their implications for potential approaches to combatting columnaris disease will be discussed.