Hemolysin EthA in Edwardsiella tarda is essential for fish invasion in vivo and in vitro and regulated by two-component system EsrA-EsrB and nucleoid protein HhaEt

Edwardsiella tarda is a Gram-negative pathogen for hemorrhagic septicemia in fish. Recently, two-component system (TCS) EsrA-EsrB in E. tarda has been found to play key roles in regulating type III secretion system (TTSS) and type VI secretion system (T6SS). In this study, a markedly attenuated Delta esrB mutant was investigated to exhibit enhanced cell-invasion capability, as well as the increased cytotoxicity of its extracellular products (ECPs). Compared with the parental strain, the Delta esrB mutant unexpectedly displayed the significantly increased hemolytic activity, and the restoration of hemolysin production was observed in the complemented strain esrB(+). A hemolysis-associated 147 kDa protein, EthA, was found to be up-regulated in the ECPs of Delta esrB. The deletion of ethA gene in E. tarda wild type and Delta esrB strains drastically decreased their capacities in internalization of epithelial papilloma of carp (EPC) cells. These results indicated that the increased production of EthA was responsible for the enhanced cell-invasion related capabilities in Delta esrB. Furthermore, the expression of EthA in Delta esrB exhibited a temperature-induced manner, and a nucleoid protein Hha(Et) was identified to mediate ethA expression by directly binding to its promoter. These results demonstrated that the virulence determinant EthA was fully required for invasion abilities of E. tarda and was subjected to the control of a complicated and precisely regulated network primed for its invasion, colonization and infection process in fish. (C) 2010 Elsevier Ltd. All rights reserved.