Immune response of Barramundi (Lates calcarifer) to Vibrio harveyi bacterin

Commercial farming of barramundi or Asian seabass (Lates calcanfer) is a burgeoning industry in Australia and South East Asia. At present there are few serious bacterial infections, however disease caused by Vibrio harveyi infection is an on-going problem with the potential to become chronic as the industry expands. Therefore, the development of an effective vaccine against vibriosis was seen as a prudent strategy for the future success of barramundi production. This thesis reports on the development of appropriate reagents for in vitro monitoring of immune responses to a bacterin, and subsequent validation in laboratory trials where fish were immunised with a bacterin via various routes or, with lipopolysaccharide (LPS) via intraperitoneal (IP) injection. Barramundi immunoglobulin was isolated by affinity chromatography using either mannanbinding protein (MBP) or Staphylococcal protein A (SpA) as capture ligands. Both isolation products were used to produce polyclonal antisera in rabbits, which were subsequently compared for specificity and titer. Two antisera were used to monitor anti-V. harveyi antibody activity in barramundi serum after primary and secondary immunisations with bacterin via the IP route. Elevated antibody levels after secondary immunisation indicated a memory response. The routes of bacterin administration were investigated in terms of antibody production and some non-specific immune parameters. It was found that barramundi respond systemically to bacterin delivered by IP injection, immersion (IMIM) and anal intubation (Al). The highest and most consistent response was in fish treated via IP followed by AT then IMM. This consistency of response from IP-treated fish carried over to bacterial inhibitory activity of serum where all demonstrated >50% inhibitory activity relative to non-immune controls. There was no significant enhancement of serum lysozyme activity nor head kidney macrophage phagocytic activity as a result of any immunisation procedure. Finally, the response of barramundi to IP immunisation with bacterin or lipopolysaccharide (LPS) extracted from the bacterium was compared to elucidate the role of the latter. There were systemic antibody responses to both preparations with little difference noted in the magnitude of the response. However, bacterial inhibitory activity of serum was highest in the bacterin treament group. This may indicate that anti-LPS antibodies do not agglutinate to the same extent as anti-whole bacterial cell antibodies. In conclusion, systemic antibody and bacterial inhibitory activity after immunisations with the bacterin suggests that it may be an effective vaccine and oral administration is worth investigation. The protective role of LPS is unclear, though relatively lower bacterial inhibitory activity of anti-LPS antibodies suggests it may be less protective than anti-whole bacterial cell antibodies.