Antigens associated with pathogenesis of Aeromonas salmonicida and related protective mechanisms in fish
This project investigated several factors that influence the virulence of the fish pathogen Aeromonas salmonicida subspecies salmonicida. In this study the effect of iron supplementation was investigated on whole cell bacterin used as a vaccine. It was shown that A-layer negative strains grown under iron supplementation do not confer high protection. A-layer positive strains though, do provide protection at levels similar to the A-layer negative iron restricted bacterin. Use of a combined vaccine appeared to compound the protection. Cells grown under iron normal and iron restricted conditions have previously been shown to produce, respectively, ferric superoxide dismutase (SOD) and manganese SOD to protect against superoxide anions. Analysis here followed the transfer from iron restricted culture to iron supplemented culture to investigate the change in superoxide dismutase (SOD) induction. It appeared that both ferric and manganese SOD were expressed even though only one form was active in either iron condition. This implies post-translational control of activity. Catalase in another enzyme with a protective role and is induced by hydrogen peroxide. Cells grown without the addition of hydrogen peroxide do not appear to express a functional catalase. Analysis of catalase induction has shown that iron normal and iron supplemented growth using hydrogen peroxide induces expression of a catalase enzyme. Iron restricted bacteria show sensitivity to normal induction doses of hydrogen peroxide. Lowering the induction concentration allows expression of the catalase. This suggests that the catalase is heme co-factored. The relationship between iron levels and catalase induction suggests that catalase has a high priority for any available iron. The in vivo expression of the serine proteinase and iron regulated outer membrane proteins (IROMP's) was investigated. Whilst the serine proteinase was induced in vivo in a normal virulent strain, a proteinase-negative strain did not appear to produce the proteinase. This suggests that the serine proteinase is not essential for virulence. The expression of IROMP's in vivo indicates an iron restricted environment. A. salmonicada secretes a glycerophospholipid: cholesterol acyl transferase (GCAT) that is often complexed with LPS and believed to be a major toxin of the ECP. Analysis of culture supernatants has identified a secreted extracellular polysaccharide that may also complex with GCAT.