Haemophilus ducreyi is the etiologic agent of chancroid, a sexually transmitted genital ulcer disease. Very little Is known about how this unencapsulated. Gram-negative bacterium evades host defenses and causes demrial lesion development. However, it has been shown that H. ducreyi has the ability to prevent phagocytosis of both itself and secondary targets by macrophages in vitro. We identified two extremely large H. ducreyi proteins, designated LspA1 and LspA2, that are released into H. ducreyi culture supernatant fluid and which are necessary for the observed inhibition of phagocytic activity. That these proteins are relevant to disease production was proven by the finding that a H. ducreyi mutant unable to express either LspA1 or LspA2 had drastically reduced virulence in both animal and human models of experimental chancroid. Little is known, however, about how the LspA proteins inhibit phagocytic activity. The proposed research project in this MERIT extension application will investigate the stoicture, function, and expression of the LspA proteins. In the first Specific Aim, we will capitalize on our recent successful cloning of the H. ducreyi IspAl gene to purify a functional recombinant LspAl protein and then determine the composition of the active form of this protein. In the second Specific Aim, we will elucidate the mechanism of action involved in the inhibition of phagocytic activity by the LspA proteins. We already have data which indicate that the LspA proteins can affect signaling pathways that control phagocytosis in macrophages, and our research efforts will be focused on the most proximal elements, and especially the Src family protein tyrosine kinases, in the phagocytic signaling cascade. In the third Specific Aim, we will characterize the H. ducreyi CpxRA two-component signal transduction system that is responsible for control of expression of both the LspA proteins as well as other co-regulated gene products of this bacterium.
The relevance of this research to public health involves the new infomnation that will be gained about H. ducreyi inhibits phagocytosis, one of the primary defense mechanisms of the human body. The ability of phagocytes to engulf and kill bacteria is essential to preventing or curing infectious diseases. Infomnation gained from this study will help us understand how phagocytes control this protective activity.
