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Measurement of bacterial burden in animal infection models is a key component for both bacterial pathogenesis studies and therapeutic agent research. The traditional quantification means for  in vivo  bacterial burden requires frequent animal sacrifice and enumerating colony forming units (CFU) recovered from infection loci. To address these issues, researchers have developed a variety of luciferase-expressing bacterial reporter strains to enable bacterial detection in living animals. To date, all such luciferase-based bacterial reporters are in cell-associated form. Production of luciferase-secreting recombinant bacteria could provide the advantage of reporting CFU from both infection loci themselves and remote sampling (eg. body fluid and plasma). Toward this end, we have genetically manipulated a pathogenic  Escherichia coli  ( E. coli ) strain, ATCC25922, to secrete the marine copepod  Gaussia princeps  luciferase (Gluc), and assessed the use of Gluc as both an  in situ  and  ex situ  reporter for bacterial burden in mouse tissue cage infections. The  E. coli  expressing Gluc demonstrates  in vivo  imaging of bacteria in a tissue cage model of infection. Furthermore, secreted Gluc activity and bacterial CFUs recovered from tissue cage fluid (TCF) are correlated along 18 days of infection. Importantly, secreted Gluc can also be detected in plasma samples and serve as an  ex situ  indicator for the established tissue cage infection, once high bacterial burdens are achieved. We have demonstrated that Gluc from marine eukaryotes can be stably expressed and secreted by pathogenic  E. coli in vivo  to enable a facile tool for longitudinal evaluation of persistent bacterial infection.

Secreted Gaussia princeps Luciferase as a Reporter of Escherichia coli Replication in a Mouse Tissue Cage Model of Infection