Intrarenal Injection of Escherichia coli in a Rat Model of Pyelonephritis

The overall goal of this surgical rat model of Peylonephritis is to provide a method that does not require renal reflux, which is used in other models. This method can help answer key questions in the the medical field such as how best to administer antibiotic therapy, how we can monitor the effects of recurrent infections or residual infections after therapy that can lead to recurrence. The main advantage of this technique is that this model directly applies bacteria to the kidney in a reasonably reliable quantity without the requirement of renal reflux, which can add some variability in terms of infection.

Prior to starting this procedure, prepare the bacterial injection bolus containing between one million and 10 million cells. After anesthetizing the rat and preparing its skin for surgery, apply vet eye ointment to the eyes to prevent dryness. Position the rat on a warmed surgical platform in a left decubitus position with its right flank facing up.

Scrub the skin with a disinfectant. Repeat three times and then scrub the surgical site with 70%alcohol swap until the skin is clear. To begin the surgery, using a Number 10 scalpel, make a small, two to three centimeter right dorsal retroperitoneal incision beginning at the bottom of the rib cage.

Continue by dissecting away the subcutaneous tissue, fat and muscles to ultimately visualize and access the abdominal cavity. Once the liver is clearly visible and accessible, use blunted forceps to retract it upwards. Then, using a second pair of blunted forceps, expose the right kidney so it sits just outside of the abdominal cavity.

Now, position the kidney for injection in the nondominant hand. Using the dominant hand, precisely target the injection needle on the renal pelvis. Then, slowly and steadily inject 0.1 milliliters of bacterial or saline solution.

Here, saline is being injected. The bolus will look like a white bubble in the tissue. Before retracting the needle, place a strip of absorbable hemostat over the injection site to prevent an outflow of the inoculum into the peritoneum.

Then, slowly pull the needle out. Now, using a large syringe, thoroughly rinse the kidney with normal saline. Then, return the kidney to the abdominal cavity.

For the surgical site closure, use absorbable or monofilament sutures in body tissue and 4-0 nonabsorbable braided sutures on body surfaces. To place sutures, start at the site furthest from the operator, using fine toothed forceps to grasp the skin and avert it slightly. Then, rotate the needle holder into a pronated position and supinate the wrist to rotate the needle and drive it through the full thickness of the skin.

Tighten the suture just enough to expose the tissue edges. Any tighter will obstruct the blood supply and thus slow wound healing and possibly result in dehiscence. Then, tie off the suture using a square knot, as if it were a simple interrupted stitch, except that only the short strand is cut, leaving a three to four millimeter tail.

Now, placing each subsequent stitch about three millimeters from the previous, continue the running suture. Once the end of the incision is reached, do not pull the last stitch completely through. Instead, use the loop being held by the needle holder as the short strand and tie off the distal end of the suture closure.

Now, using instrument ties, tie off the suture using square knots. The result will be three strands sticking up from a completed knot. After providing analgesics and a 0.6 milliliter peritoneal injection of normal saline, observe the animal while it recovers from the anesthesia on a heating pad with absorbent soft bedding.

Over the first 24 hours of recovery, rehydrate the animal with oral or parenteral fluids until it can return to normal feeding. Thereafter, regularly monitor the incision for swelling, exudate or dehiscence and provide analgesics for pain as needed. An acute peylonephritis infection was induced by injection of one million and 10 million UT189 E.coli cells into the rodent model.

MRI of the kidney showed a progressive increase in the region of infection from one to four days after injection. Once mastered, this technique can be done in 30 minutes if performed properly. While attempting this procedure, it's important to remember to check on the animals following surgery to watch for signs of E.Coli infection outside of the kidney, especially at the incision points, to make sure that the animals do not become overly septic.

Following this procedure, other methods, like MRI scanning, can be performed in order to answer additional questions like, is the infection viable, how fast is the infection growing, and is this therapy effective?