 |
 |
|
|||
 |
 |
 |
 |
 |
 |
|
MEASURING SPATIAL DIFFERENCES IN PSEUDOMONAS AERUGINOSA DISTRIBUTION AND VIRULENCE ACTIVATION RELATED TO INTESTINAL ISCHEMIA REPERFUSION INJURY. David Fink, MD, Kathleen Romanowski, MD, Trissa Babrowski, MD, Vesta Valuckaite, MD, Alex Zaborin, PhD, Valeriy Poroyko, PhD, Irina Morozova, MS, Olga Zaborina, PhD, John Alverdy, MD, University of Chicago
Background: The precise mechanism by which intestinal microbes influence the development of sepsis leading to mortality remains to be elucidated. Although altered barrier function and systemic absorption of microbes and their products is hypothesized to be an important element of this process, equally important may be the degree to which intestinal microbes express enhanced virulence both in the gut and remote organs. In the current study, we have developed a comprehensive approach to this problem using photon-camera based detection of propagation (growth), dissemination, and virulence expression of intestinal bacteria using two strains of bioluminescent bacteria. Methods: The Xenogen system was used to capture images of luminescent strains of Pseudomonas aeruginosa. Mice (n=3 per group) underwent either sham laparotomy or 15 minutes of intestinal ischemia reperfusion (IIR) via superior mesenteric artery occlusion. At surgery, either XEN41 (constitutively luminescent strain of P. aeruginosa) or PAO1/lecA::lux (inducible expression of barrier disrupting lecA protein) were injected into the terminal ileum. 12 hours post-operatively animals were sacrificed and their organs immediately imaged ex-vivo using the Xenogen system. Spatial regions of interest were defined around each organ or portion of the gastrointestinal tract and the total photon count from each area was quantified. In reiterative experiments, mouse mortality was followed. Results: Xenogen analysis of propagation and distribution of constitutively luminescent P. aeruginosa XEN41 demonstrated that intestinal IIR had a significant effect on the regional distribution of bacteria whereby P. aeruginosa was found to migrate from the site of injection (distal ileum) toward the site of injury (jejunum) (3.82 fold increase IIR versus Sham). Bacterial growth was not affected by IIR. Intriguingly, we found a lower dissemination rate of P. aeruginosa following IIR versus Sham where mesenteric lymph node (MLN) photon counts were 49% lower in IIR compared to Sham. Virulence expression using inducible PAO1/lecA::lux demonstrated that despite lower total number of bacteria in organs from IIR mice, the “virulence signal” was 10.56 times higher in the IIR mice compared to sham. Increased virulence of P. aeruginosa was associated with enhanced lethality of 15 min of IIR mice (data not shown). Conclusion: Approaches assessing both bacterial counts, dissemination, and site-specific virulence expression using constitutively and inducible bioluminescent bacterial strains may shed new light on how intestinal microbes contribute to sepsis and mortality. AVERAGE PHOTON COUNTS PER ORGAN
Back to Program |
 |
 |
 |
 |
 |
 |
Privacy Policy