The rickettsial agent of Q fever, Coxiella burnetii is an obligate intracellular parasite which replicates within the cellular phagolysosome. Classical genetic methods to define functional characteristics are not available and as a result, the physiology and genetic capabilities of C. burnetii are poorly understood. Although the mechanisms of virulence of C. burnetii is not known, a distinct antigenic phase variation has been associated with a reduction in virulence. The antigenic shift occurs when organisms isolated from infected animals (phase I) are repeatedly passed in eggs (phase II). As the organisms undergo an antigenic change to phase II, there is a concomitant decrease in virulence. In order to develop an understanding of the nature of the genes involved in virulence, we will examine a recently isolated plasmid and chromosomal DNA for genes relating to phase variation and virulence. We will first use the plasmid isolated from the Nine Mile, phase I strain (labeled by nick translation) to determine how widely plasmid sequences are distributed in other strains of C. burnetii (Southern blotting, hybridization). We will also attempt to cure C. burnetii of its plasmid. These studies will allow us to see if a correlation exists between virulence and the plasmid. Next recombinant DNA techniques will be used to place plasmid and chromosomal genes in E. coli. Those genes coding for phase specific and other surface antigens will be identified by specific antisera. Other bacteria which share properties with C. burnetii will be examined for the presence of DNA sequences related to those of the plasmid. In addition, possible metabolic functions will be examined using recombinant DNA techniques to place plasmid sequences into defined E. coli auxotrophs and look for growth due to complementation. These studies will provide the first analysis of the genetic capabilities of a rickettsial agent, and are part of the long range objectives of our laboratory in understanding the biochemical basis of the hostparasite relationship.
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