Genomics is the study of the structure, function, and evolution of genomes (2). Genomics is differentiated from traditional genetics by the application of computational biologic and robotics technologies that permits the simultaneous analysis of 1000 or more genes. This approach allows the study of large numbers of genes expressed during a variety of fundamental biologic processes such as meiosis, development, infection, or virulence. These data are able to be disseminated to the scientific community and immediately utilized in research strategies encompassing purely computer generated searches for gene homologies or genetic comparisons, or for unique in vivo approaches to identify genes involved in pathogenesis or virulence. The group of organisms known as Pneumocystis carinii is an especially attractive target for a genome project. There is a dearth of basic biologic information about these opportunistic pathogens that are a major cause of pneumonia in patients with AIDS, as well as pathogens of concern in commercial animal colonies. The lack of a continuous in vitro cultivation system for any member of this family has largely contributed to the scientific ignorance concerning its life cycle, mode of transmission, infective form and poor understanding of its metabolic processes. Pneumocystis has only recently been shown to be a member of the fungi by genetic sequence analysis of the nuclear 16S-like gene and other genes. It is a unique member of the fungal kingdom distinguished by its phylogenetic isolation; lack of ergosterol and insensitivity to standard anti-fungal therapies; paucity of nuclear ribosomal genes (1-2 copies); presence of a highly repeated gene family encoding surface antigens; and relatively delicate cell wall. Because of its compact genome size, 7.7 Mbp, and the availability of clearly resolved chromosomes by pulsed field gel electrophoresis, it is now possible to address basic biological questions by application of genomic technologies. The goals of the present proposal are to produce physical maps and gene inventories for Pneumocystis populations form the rat and human being. This project represents the first time a non- culturable microbe will be the subject of a genomics project. It is a unique opportunity to make a dramatic difference in the progress of Pneumocystis research and to concurrently broaden our understanding of that group of organisms known as the Fungi. Benefits from such a project include construction of a physical map that will provide structural information of the Pneumocystis genome; identification of gene networks involved in metabolism and virulence; determination of the genetic relatedness of Pneumocystis with extant fungi; insights into the mechanisms for host specificity demonstrated by different populations of Pneumocystis; identification of potential drug targets; and possible strategies leading to establishment of a continuous culture system.
