| dc.description.abstract | Pasteurellosis caused by Pasteurella atlantica genomvar salmonicida has been an emerging disease in Norway since 2018, causing poor fish welfare and mortalities in the fish farming industry. The development of a commercial vaccine is challenging, and knowledge of the bacteria’s pathogenesis, virulence factors, and immunogenic components is still restricted.
The aim of this study was to predict immunogenic proteins of P. atlantica gv. salmonicida (PaL-1 isolate) by in silico analysis, express two proteins recombinantly using E. coli and test the antigenicity of the proteins by western blot analysis (WB). Furthermore, the growth of PaL-1 and a Pasteurella atlantica gv. cyclopteri isolate in TSB supplied with 1.5% NaCl and 10% FCS (TSB+) at different temperatures (10°C, 20°C and 30°C) was investigated. Lastly, the growth of PaL-1 in TSB+ was compared to growth in TSB (2% NaCl) supplemented with reduced and oxidized glutathione (GSH and GSSG, respectively), and the ligand binding of one of the recombinant proteins with GSH and GSSG was assessed.
PaL-1 grew well in TSB+ at 20°C, and the exponential growth phase started after approximately 8 hours. The P. atlantica isolates incubated at 20 °C and 30 °C gave a higher yield in cell number compared to when cultured at 10 °C. Several immunogenic proteins were identified by in silico analysis of the PaL-1 proteome. The most prominent were a Heme binding protein A-like protein (HbpA) and a BamA-like protein, which were chosen for recombinant protein expression. The HbpA-like protein was successfully expressed recombinantly but showed no reactivity when blotted against sera from PaL-1 vaccinated fish.
The growth of PaL-1 in TSB supplemented with GSH/GSSG gave a higher yield than the growth in TSB+. By further testing, this may have the potential to be a serum-free growth medium for P. atlantica. The ligand binding assessment of the recombinant HbpA-like protein and GSH/GSSG did not show any reactivity. However, this may be due to methodological issues. | |
