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Flavobacterium columnare  causes substantial losses among cultured finfish species. The Gram-negative bacterium is an opportunistic pathogen that manifests as biofilms on the host's mucosal surfaces as the disease progresses. We previously demonstrated that the dominant mucosal IgM antibody response to  F. columnare  is to the chaperone protein DnaK that is found in the extracellular fraction. To establish the efficacy of using recombinant protein technology to develop a new vaccine against columnaris disease, we are reporting on two consecutive years of vaccine trials using a recombinant  F. columnare  DnaK protein (rDnaK). In year one, three groups of channel catfish ( n  = 300) were immunized by bath immersion with a live attenuated  F. columnare  isolate, rDnaK or sham immunized. After 6 weeks, an  F. columnare  laboratory challenge showed a significant increase in survival (>30%) in both the live attenuated and rDnaK vaccines when compared to the non-immunized control. A rDnaK-specific ELISA revealed significant levels of mucosal IgM antibodies in the skin of catfish immunized with rDnaK at 4- and 6-weeks post immunization. In the second year, three groups of channel catfish ( n  = 300) were bath immunized with rDnaK alone or with rDnaK after a brief osmotic shock or sham immunized. After 6 weeks a laboratory challenge with  F. columnare  was conducted and showed a significant increase in survival in the rDnaK (> 25%) and in rDnaK with osmotic shock (>35%) groups when compared to the non-immunized control. The rDnaK-specific ELISA demonstrated significant levels of mucosal IgM antibodies in the skin of catfish groups immunized with rDnaK at 4- and 6-weeks post immunization. To further understand the processes which have conferred immune protection in the rDnaK group, we conducted RNA sequencing of skin samples from the non-immunized ( n  = 6) and rDnaK treated channel catfish at 1-week ( n  = 6) and 6 weeks ( n  = 6) post immunization. Significantly altered gene expression was identified and results will be discussed. Work to further enhance the catfish immune response to  F. columnare  rDnaK is underway as this protein remains a promising candidate for additional optimization and experimental trials in a production setting.

frontiers in immunology

Evaluation of a Recombinant Flavobacterium columnare DnaK Protein Vaccine as a Means of Protection Against Columnaris Disease in Channel Catfish (Ictalurus punctatus)