Noninvasive biophotonic imaging for monitoring of catheter-associated urinary tract infections and therapy in mice

Abstract

Urinary tract infections (UTIs) are among the most common bacterial infections acquired by humans, particularly in catheterized patients. A major problem with catheterization is the formation of bacterial biofilms on catheter material and the risk of developing persistent UTIs that are difficult to monitor and eradicate. To better understand the course of UTIs and allow more accurate studies of in vivo antibiotic efficacy, we developed a catheter-based biofilm infection model with mice, using bioluminescently engineered bacteria. Two important urinary tract pathogens, Pseudomonas aeruginosa and Proteus mirabilis, were made bioluminescent by stable insertion of a complete lux operon. Segments of catheter material (precolonized or postimplant infected) with either pathogen were placed transurethrally in the lumen of the bladder by using a metal stylet without surgical manipulation. The bioluminescent strains were sufficiently bright to be readily monitored from the outside of infected animals, using a low-light optical imaging system, including the ability to trace the ascending pattern of light-emitting bacteria through ureters to the kidneys. Placement of the catheter in the bladder not only resulted in the development of strong cystitis that persisted significantly longer than in mice challenged with bacterial suspensions alone but also required prolonged antibiotic treatment to reduce the level of infection. Treatment of infected mice for 4 days with ciprofloxacin at 30 mg/kg of body weight twice a day cured cystitis and renal infection in noncatheterized mice. Similarly, ciprofloxacin reduced the bacterial burden to undetectable levels in catheterized mice but did not inhibit rebound of the infection upon cessation of antibiotic therapy. This methodology easily allows spatial information to be monitored sequentially throughout the entire disease process, including ascending UTI, treatment efficacy, and relapse, all without exogenous sampling, which is not possible with conventional methods.

FIG. 1.

(A and B) Growth and bioluminescence curves of (A) P. aeruginosa Xen 5 and (B) P. mirabilis Xen 44 UTI in mice infected with precolonized catheters or bacterial suspensions. UTI was induced by implanting a precolonized catheter in the bladder or by transurethral inoculation of the bladder with a bacterial suspension. The total photon emissions from the infected sites were quantified using Living Image software (cumulative results are shown). Each data point is the mean ± standard error for 13 to 19 mice. Results for postimplant infection were similar to those for precolonized infection except that the bioluminescence signal peaked 2 days after infection.

FIG. 2.

Monitoring the spread of P. aeruginosa Xen 5 infection from the bladder to the kidney by bioluminescence. Mice were infected by placement of a precolonized catheter in the bladder through the urethral meatus, and the infection was monitored by recording photon emission over time. Bacteria ascending from the bladder up the ureters and to the kidney can be clearly visualized (dorsal view) in live animals, using a luciferase tag pathogen and bioluminescence imaging.

FIG. 3.

In vivo bioluminescence monitoring of P. mirabilis Xen 44 in the mouse model of UTI during treatment with ciprofloxacin. Effect of ciprofloxacin treatment 2 days after infection and readministration of ciprofloxacin for four additional days after the infection was reestablished. The total number of photons detected per second over the infected bladder was quantified using Living Image software and plotted with respect to time. Each data point is the mean ± standard deviation for 6 to 13 mice. Arrows indicates the days of antibiotic administration. Data are averages of results from three experiments. The viable counts in each catheter were determined immediately after removal from the bladder and are shown in the upper quadrants of the plot.

FIG. 4.

Real-time monitoring of the effects of ciprofloxacin on P. mirabilis Xen 44 UTI. A representative animal from the group receiving antibiotic is shown. Note that the response to treatment, including relapse and renal infection, can be monitored noninvasively within the same animal throughout the study period.

FIG. 5.

Scanning electron micrograph of a longitudinal section of a catheter from the bladder of a mouse 2 days after infection. Cross-section of catheter at low magnification (left) shows the lumen of the catheter filled with a thick biofilm. Also note a thin film outside the surface of the catheter. Numerous rod-shaped bacterial cells mostly embedded within the intracellular material are clearly seen at higher magnification (right).