Visualizing pneumococcal infections in the lungs of live mice using bioluminescent Streptococcus pneumoniae transformed with a novel gram-positive lux transposon

Animal studies with Streptococcus pneumoniae have provided valuable models for drug development, In order to monitor long-term pneumococcal infections noninvasively in living mice, a novel gram-positive lux transposon cassett e, Tn4001 IuxABCDE Km(r), that allows random integration of lux genes onto the. bacterial chromosome was constructed. The cassette was designed so tha t the luxABCDE and kanamycin resistance genes were linked to form a single promoterless operon. Bioluminescence and kanamycin resistance only occur in a bacterial cell if this operon has transposed downstream of a promoter on the bacterium's chromosome, S. pneumoniae D39 was transformed with plasmid pAUL-A Tn4001 luxABCDE Km(r), and a number of highly bioluminescent coloni es were recovered, Genomic DNA from the brightest D39 strain was used to tr ansform a number of clinical S, pneumoniae isolates, and several of these s trains were tested in animal models, including a pneumococcal lung infectio n model. Strong bioluminescent signals were seen in the lungs of the animal s containing these pneumococci, allowing the course and antibiotic treatmen t of the infections to be readily monitored in real time in the living anim als. Recovery of the bacteria from the animals showed that the bioluminesce nt signal corresponded to the number of CFU and that the lux construct was highly stable even after several days in vivo. We believe that this lux tra nsposon mill greatly expand the ability to evaluate drug efficacy against g ram-positive bacteria in living animals using bioluminescence.