Detection and analysis of gene expression during infection by in vivo expression technology

Many limitations associated with the use of in vitro models for study of bacterial pathogenesis can be overcome by the use of technologies that detect pathogen gene expression during the course of infection within an intact animal. In vivo expression technology (IVET) accomplishes this with versatility: it has been developed with a variety of reporter systems which allow for either in vivo selection or ex vivo screening. Selectable gene fusion systems generally allow for the complementation of a bacterial metabolic defect that is lethal in vivo, or for antibiotic resistance during the course of in vivo antibiotic challenge. In contrast, the screenable gene fusion system uses a site-specific DNA recombinase that, when expressed in vivo, excises a selectable gene cassette from the bacterial chromosome. Loss of this cassette can then be either screened or selected for ex vivo. The recombinase-based IVET can be used to detect genes that are transcriptionally induced during infection, including those expressed transiently or at low levels and, in addition, can be used to monitor the spatial and temporal expression of specific genes during the course of infection.