Luciferase imaging of chylamdia muridarum ascending infection in mice

Abstract

To understand Chlamydia pathogenic mechanisms by visualizing chlamydial ascending infection in the genital tract. Constructing luciferase expressing- Chlamydia muridarum (C. muridarum) organisms for monitoring C. muridarum trafficking in the female mouse genital tract using bioluminescence technology. A shuttle vector containing the luciferase gene was used to transform C. muridarum organisms. A stable transformant was characterized in Hela cells by monitoring both luciferase mRNA levels using qRT-PCR and luciferase activity using the substrate D-luciferin. The in vitro characterized transformant was then used to infect 10 Balb/cJ mice intravaginally, and the infection was monitored using the Xenogen IVIS imaging system at the whole animal scale. Luciferase activity was detectable at 12 hours (h), peaked by 15h, and was significantly reduced by 30h after infection in cell cultures infected with luciferase-expressing C. muridarum transformant. Mice intravaginally infected with the same transformant were monitored for luciferase activity on days 3, 7, 10, 14, 21 & 28 after infection. Six of the 10 inoculated mice displayed luciferase signal in lower with two also in upper genital tracts on day 3 after infection. By day 7, all 10 mice developed luciferase signal with 1 only in lower and the remaining 9 in upper genital tracts. The luciferase signal was maintained in upper genital tract in 6 and 2 mice by days 14 and 21, respectively. No luciferase activity was detected in 9 of 10 by day 28. Whole body images revealed a transient airway co-infection among 5 mice housed in the same cage with 5 mice positive on day 3 and 1 remaining positive on day 7. Its presence did not significantly alter the genital tract infection courses. We have successfully visualized Chlamydia muridarum ascension in mouse genital tract. Our observations suggest that C. muridarum organisms are most active in ascending to the upper genital tract within the first 7 days after intravaginal inoculation. Mice of the same inbred strain display variation in the time course of ascending infection. Unexpectedly, co-infection in the airway is common in some cages, but it does not seem to affect the time-course of ascending infection. The luciferase detection-based in vivo imaging (without the need for sacrificing mice) will greatly facilitate our understanding of chlamydial pathogenic mechanisms.