Immune defence mechanisms of barramundi (Lates calcarifer) peripheral blood against Streptococci

Abstract

IMMUNE DEFENCE MECHANISMS OF BARRAMUNDI (Lates calcarifer) PERIPHERAL BLOOD AGAINST STREPTOCOCCIAquaculture now supplies more than half of the fish used for human consumption and is a major contributor to global food security. As aquaculture growth must be sustained to continue to supply increasing population-driven demand for food fish, efficiency must be increased. In spite of major inroads in disease control through mass vaccination of farm fish, bacterial diseases continue to cause substantial losses, particularly in tropical and warmtemperate regions where aquaculture is expanding fastest. To develop new vaccines, and improve existing vaccines for effective disease control in these regions, an improved understanding of the mechanisms of pathogen immune evasion and dissemination within the host may identify new vaccine targets. Streptococcus agalactiae and Streptococcus iniae are significant pathogens of warm and temperate farmed and wild fish. Both have a broad host range, express a polysaccharide capsule as a major virulence determinant and cause similar pathologies characterized by rapid sepsis, followed by meningitis and death. S. iniae causes significant mortality in barramundi, Lates calcarifer. Interestingly, S. agalactiae ST261, although causing mortality in wild fish along the Queensland coast, does not appear to cause mortality in barramundi farmed in close proximity. This difference provides a basis for comparison, in order to explore the requirements for blood colonization and sepsis by S. iniae in barramundi, and to increase our understanding of sepsis in fish.S. iniae grew rapidly in barramundi blood, doubling in less than 30 minutes in a whole blood bactericidal assay. In contrast, S. agalactiae was unable to multiply. Moreover, E. coli DH5 was completely killed in barramundi blood during the same incubation period, suggesting 3 that antibacterial humoral and cellular immune defences were functional within the blood bactericidal model. A capsular defective strain of S. iniae also survived in the bloodbactericidal assay, however the rapid proliferation was reduced, suggesting that while the capsule is important for bacterial colonisation it is not the only means used to evade bloodborne defences. As gram-positive pathogens are particularly susceptible to lysozyme due to the dominance of muramic acids in the cell wall, the role of lysozyme was investigated.Lysozyme levels in barramundi serum and plasma were undetectable in a lysis assay using Micrococcus lysodeitikus, indicating very low levels of circulating lysozyme in healthy barramundi.The first responder immune cells in blood are neutrophils and these are critical in the prevention of sepsis; for example, granulocytopenia (circulating granulocyte deficiency) is associated with increased susceptibility to Escherichia coli K1 and Klebsiella pneumoniae sepsis in neonates, and this can be partially repaired by injection of granulocyte colony stimulating factor (G-CSF) to increase neutrophil numbers (Deshmukh et al., 2014). This critic...