Intracellular pathogens are responsible for many globally important diseases in human beings. The ability to evade the bactericidal activity of the macrophage is key to their survival and multiplication in the mammalian host. Rhodococcus equi is a facultative intracellular pathogen which causes severe granulomatous pneumonia in foals and immunocompromised human beings. R. equi is an important opportunistic pathogen in human patients suffering from the acquired immunodeficiency syndrome (AIDS) as evidenced by the increasing number of reported cases in the last 5 years. Diagnosis of rhodococcal pneumonia can be difficult and may be delayed until the development of advanced disease when treatment is often unsuccessful. Entry and survival within the pulmonary macrophage leads to persistence of this organism in the lung of infected human and equine patients by evasion of host immune responses. Virulent and avirulent R. equi strains have been identified based on their ability to persist within the lung of severe combined immunodeficient (SCID) mice following intratracheal inoculation or by lethal dose 50 determination using immunocompetent mice. Similarly, virulent strains are capable of persisting and multiplying within macrophages in vitro. A 15-17 kDa surface protein (VAP A) is consistently expressed by virulent R. equi but not by avirulent strains leading to its designation as a virulence- associated protein. The close correlation between the virulent phenotype and ability to express this protein indicates that this protein may have an essential role in persistence of R. equi within the lung and macrophages. The goal of this proposal is to determine if VAP A is essential for persistence within the lung of an immunodeficient host and within macrophages in vitro. These hypotheses will be tested by elimination of expression of VAP A by mutagenesis and testing of virulence 1) in vivo and 2) within macrophages in vitro.
