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Title: Isolation of rifampicin resistant Flavobacterium psychrophilum strains and their potential as live attenuated vaccine candidates

Author
item Lafrentz, Benjamin
item LAPATRA, SCOTT - CLEAR SPRINGS FOODS, INC.
item CALL, DOUGLAS - WASHINGTON STATE UNIV.
item CAIN, KENNETH - UNIV. OF IDAHO

Submitted to: Vaccine
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/29/2008
Publication Date: 10/16/2008
Citation: Lafrentz, B.R., Lapatra, S.E., Call, D.R., Cain, K.D. 2008. Isolation of rifampicin resistant Flavobacterium psychrophilum strains and their potential as live attenuated vaccine candidates. Vaccine 26:5582-5589.

Interpretive Summary: Flavobacterium psychrophilum is a Gram-negative bacterial fish pathogen that causes bacterial coldwater disease (CWD) and is considered to be one of the most important pathogens impacting salmonid aquaculture due to its wide distribution and economic impact. Vaccination holds the greatest potential for the prevention of CWD; however, there are no commercial vaccines currently available. A highly attenuated strain of F. psychrophilum was developed by exposing the bacterium to the antibiotic rifampicin. Immunization of rainbow trout with live cells of this strain by injection and immersion delivery stimulated protective immune responses, which demonstrates the potential for the use of this strain as a live vaccine candidate. This research lays a foundation for the development of a live attenuated vaccine for the prevention of CWD. Such a vaccine will greatly benefit salmonid aquaculture worldwide.

Technical Abstract: Previous studies have demonstrated that passage of pathogenic bacteria on increasing concentrations of the antibiotic rifampicin leads to the attenuation of virulence and these resistant strains may serve as live attenuated vaccines. Two rifampicin resistant strains of Flavobacterium psychrophilum, 259-93A.16 and 259-93B.17, were generated by passage on TYES plates containing increasing concentrations of rifampicin. Electrophoretic analysis of whole-cell lysates prepared from the parent and resistant strains identified five differentially expressed proteins between the 259-93B.17 strain and parent strain, while there were no differences identified between the 259-93A.16 and parent strain. The LPS banding patterns were identical between all three strains. Bacterial challenges of rainbow trout (Oncorhynchus mykiss Walbaum) with the resistant strains demonstrated that the 259-93B.17 strain was highly attenuated and the 259-93A.16 strain was modestly attenuated at the challenge doses tested. Immunization of rainbow trout with the live attenuated 259-93B.17 strain by intraperitoneal injection resulted in significant protection against challenge with the virulent parent F. psychrophilum strain at 8 and 15 weeks post-immunization and fish exhibited elevated specific antibody titers. Importantly, immersion delivery of the 259-93B.17 strain stimulated protective immune responses in fish at 10 weeks post-immunization. The results demonstrate that the rifampicin resistant 259-93B.17 strain may serve as an effective live attenuated vaccine for the prevention of F. psychrophilum infections.