title: Construction of Aeromonas salmonicida subsp. achromogenes AsaP1-toxoid strains and study of their ability to induce immunity in Arctic char, Salvelinus alpinus L.
abstract: The metalloendopeptidase AsaP1 is one of the major extracellular virulence factors of A. salmonicida subsp. achromogenes, expressed as a 37-kDa pre-pro-peptide and processed to a 19-kDa active peptide. The aim of this study was to construct mutant strains secreting an AsaP1-toxoid instead of AsaP1-wt, to study virulence of these strains and to test the potency of the AsaP1-toxoid bacterin and the recombinant AsaP1-toxoids to induce protective immunity in Arctic char. Two A. salmonicida mutants were constructed that secrete either AsaP1E294Aor AsaP1Y309F. The secreted AsaP1Y309F-toxoid had weak caseinolytic activity and was processed to the 19-kDa peptide, whereas the AsaP1E294A-toxoid was found as a 37-kDa pre-pro-peptide suggesting that AsaP1 is auto-catalytically processed. The LD50of the AsaP1Y309F-toxoid mutant in Arctic char was significantly higher than that of the corresponding wt strain, and LD50of the AsaP1E294A-toxoid mutant was comparable with that of an AsaP1-deficient strain. Bacterin based on AsaP1Y309F-toxoid mutant provided significant protection, comparable with that induced by a commercial polyvalent furunculosis vaccine. Detoxification of AsaP1 is very hard, expensive and time consuming. Therefore, an AsaP1-toxoid-secreting mutant is more suitable than the respective wt strain for production of fish bacterins aimed to protect against atypical furunculosis.
authors.affiliation: Institute for Experimental Pathology, University of Iceland, Keldur, Reykjavík, Iceland.|Southern Research Institute, Birmingham, AL, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Schwenteit
authors.affiliation: Department of Molecular Biology, Umeå Centre for Microbial Research (UCMR), Umeå University, Umeå, Sweden.
authors.forename: B
authors.initials: B
authors.lastname: Weber
authors.affiliation: Southern Research Institute, Birmingham, AL, USA.|Department of Molecular Biology, Umeå Centre for Microbial Research (UCMR), Umeå University, Umeå, Sweden.
authors.forename: D L
authors.initials: DL
authors.lastname: Milton
authors.affiliation: Department of Biotechnology and Enzyme Catalysis, Institute of Biochemistry, Greifswald University, Greifswald, Germany.
authors.forename: U T
authors.initials: UT
authors.lastname: Bornscheuer
authors.affiliation: Institute for Experimental Pathology, University of Iceland, Keldur, Reykjavík, Iceland.|Faculty of Medicine, University of Iceland, Reykjavík, Iceland.
authors.forename: B K
authors.initials: BK
authors.lastname: Gudmundsdottir
keywords: Aeromonas salmonicida subsp. achromogenes; Arctic char; AsaP1-toxoid; Salvelinus alpinus L; bacterin; extracellular protease AsaP1