Funding for this work was provided by ORWH SCOR P50 DK064540, RO1 DK 051406, RO1 AI 108749-01, F32 DK 101171, and F32 DK 104516-01.

Ethics statement: The Washington University Animal Studies Committee approved all mouse infections and procedures as part of protocol number 20150226, which expires 12/10/2018. Overall care of the animals was consistent with The Guide for the Care and Use of Laboratory Animals from the National Research Council and the USDA Animal Care Resource Guide. Euthanasia procedures are consistent with the &#8220;AVMA guidelines for the Euthanasia of Animals 2013 edition.&#8221;
1. UPEC UTI Protocol, Inoculation Needle Preparation (Figure S1)
<ol>
	<li>Remove the cap of the 30 G needle. Thread approximately 1 inch of PE10 tubing onto the shaft of the needle. UV sterilize the needle assemblies overnight. Catheterized needles can be stored indefinitely in sterile petri plates.</li>
</ol>
2. UPEC Bacterial Inoculum Preparation
<ol>
	<li>Prepare UTI89 inoculum (start 72 hr prior to the inoculation day)
	<ol>
		<li>Streak UTI89 onto an LB (Luria-Bertani) agar plate from a frozen stock. Incubate overnight at 37 &#176;C. Pick a colony and inoculate it into 10 ml of LB in a 125 ml flask. Incubate at 37 &#176;C for 24 hr, statically.</li>
		<li>Inoculate 10 &#181;l of overnight culture into 10 ml of LB in a new 125 ml flask for an additional 24 hr at 37 &#176;C, statically.</li>
		<li>Transfer 3 ml (will vary based on strain and desired inoculum size) to sterile 1.5 ml tubes and centrifuge at 7,000 x g for 3 min and resuspend the pellet in 1x PBS and centrifuge a second time. Resuspend the bacterial pellet in 1 ml 1x PBS.</li>
	</ol>
	</li>
	<li>Measure bacterial optical density and adjust concentration to the desired inoculum density. The standard concentration of the inoculum used is 107 bacteria; however, this may vary due to the design of the study.</li>
</ol>
3. Bacterial Inoculation
<ol>
	<li>Clean the workstation with 70% ethanol and cover area with absorbent paper (or use sterile flow hood).</li>
	<li>Draw up to 0.9 ml of the prepared bacterial inoculum into a 1 ml (TB) syringe (remove air bubbles). Attach a prepared sterile inoculation needle with PE10 tubing, onto the syringe containing the inoculum, then sterilely trim the polyethylene tubing. 
	NOTE: Leave 1 mm of tubing above the tip of the needle to avoid puncturing the bladder.</li>
	<li>Cut a 1 inch square piece of parafilm and put a dab of surgical lubricant (approximately the size of a dime) on top.</li>
	<li>Anesthetize female C3H/HeN mice by putting them in a vaporizer chamber (following manufactures protocol) until unconscious but still breathing normally (1 breath/sec). 
	NOTE: Some IACUC committees do not approve the use of a vaporizer. Please follow the indication of the IACUC committee of your institution. 
	CAUTION: Isoflurane is an inhalation anesthetic. Use in a well-ventilated area and minimize inhalation.</li>
	<li>Remove the mouse from the vaporizer and place it on its back on a paper towel and spread the legs.</li>
	<li>Cover the nose of the mouse with a nose cone (a tube connected to the vaporizer that provides a controlled isoflurance dose that comes equipped on some vaporizer units) to maintain anesthetization.</li>
	<li>Gently palpitate the bladder to induce urination and ensure a voided bladder. Wipe the periurethral area with 100% ethanol wipe. Dab the inoculation needle/syringe, point first, into the surgical lubricant.</li>
	<li>Inoculate each mouse with 50 &#181;l of the bacteria solution by inserting the inoculation needle transurethrally, approximately 12 mm, and pressing down on the syringe plunger gently to dispense the inoculum into the bladder gently (10 &#181;l/sec). Remove the inoculation needle from the mouse. 
	NOTE: Immediate return of inoculum at the urethral opening when beginning to inoculate indicates improper or incomplete insertion of the needle.</li>
	<li>Remove the mouse from nose cone and return it to its cage. Repeat steps 3.3-3.8 for each mouse. When/if switching inoculum conditions/strains, dispose of the syringe and inoculation needle in an approved sharps container and start again step 3.2.&#160;&#160; 
	NOTE: Inoculation with uropathogenic organisms generally does not cause severe pain symptoms. However, in rare instances, administering large doses of pathogens may cause fever, reduction in food and water intake and abnormal behavior. Animal health should be monitored throughout the experiment. If overt pain symptoms are notice, an analgesic, such as Buprenorphine (0.05&#8722;0.1 mg/kg given subcutaneously), can be applied. Procedures should be in accordance with each institution&#8217;s IACUC.</li>
</ol>
4. Determination of Bacterial Burdens
<ol>
	<li>Prepare separate 5 ml tubes for each bladder and kidneys pair to be harvested. Add 1ml of 1x PBS/tube/mouse bladder to be harvested. Add 0.8 ml of 1x PBS/tube/pair of mouse kidneys to be harvested. Label each tube to correspond to a given mouse. Prepare a 96-well plate containing 180 &#181;l of 1x PBS in each well for 1:10 serial dilutions.</li>
	<li>Clean the work area with 70% ethanol and cover the area with an absorbent cover or paper towel.</li>
	<li>Anesthetize the mouse with isoflurane (see step&#160;3.4) until the mouse stops breathing for approximately 1 min, then place it on the absorbent cover or paper towel. Other methods of euthanasia are also accepted by IACUC committees, such as carbon dioxide, and can be substituted for isoflurane overdose.</li>
	<li>Sacrifice the mouse by rapid cervical dislocation that consists of restraining the neck at the base of the head and pulling the tail horizontally away from the body until dislocation occurs. 
	NOTE: Most IACUC committees require a secondary means of euthanasia and cervical dislocation is a common method. However, other methods can be used if approved by the IACUC committee.</li>
	<li>Place the dead mouse on its back and spray 70% ethanol on the abdomen. Dissect the mouse, harvest the bladder and kidneys, in that order to minimize potential contamination from the blood following the kidney dissection. Place the organs independently into the prepared 5 ml tubes containing 1x PBS (see step 4.1). 
	NOTE: Use different scissors for the external dissection and for organ harvesting to minimize contamination.</li>
	<li>Tap the tube to ensure the organs are in the 1x PBS. Repeat step 4.3-4.5 for each mouse. Clean scissors and forceps in 70% ethanol after each mouse.</li>
</ol>
5. Bacterial Recovery
<ol>
	<li>Homogenize harvested bladder and kidneys in the 5 ml tubes with a tissue homogenizer, 15 sec per kidney and 30 sec per bladder. Rinse homogenizer sequentially in 1x PBS, 70% ethanol and 1x PBS between samples.</li>
	<li>Make serial 1:10 dilutions out to 10-8 and plate each dilution for CFU (colony forming units) on LB agar plates. Incubate the plates at 37 &#176;C overnight and count colonies the next day.</li>
</ol>
6.&#160;IBC Enumeration
<ol>
	<li>Aseptically remove the bladder and place it in 1x PBS in a 6-well plate with a silicone bottom. Plates can be prepared using a silicone elastomer kit and pouring approximately 1 cm of silicone in each well, see manufactures instructions. Cut the bladder in half using scissors.</li>
	<li>Using small metal pins gently splay the bladder so that the bladder lumen is facing upward. Be sure to place the pins at the outermost edges of the bladder sections to ensure the maximal amount of urothelium is exposed.</li>
	<li>After splaying, gently wash the bladder once with 1x PBS. Remove the 1x PBS by pipet. Fix the bladder by adding enough 3% paraformaldehyde to cover the entire splayed bladder. CAUTION: Paraformaldehyde is carcinogenic. Proper protective equipment, including gloves, should be worn at all times. Disposal should follow institutional guidelines.</li>
	<li>Incubate at room temperature for 1 hr. Remove paraformaldehyde solution. Wash once with 1x PBS.</li>
	<li>Wash with LacZ wash solution (2 mM MgCl2, 0.01% sodium deoxycholate and 0.02% nonidet-p40in 1x PBS) three times for 5 min per wash.</li>
	<li>Remove wash solution and add enough LacZ stain (9.5 ml LacZ wash solution, 0.4 ml 25 mg/ml X-gal and 0.1 ml of 100 mM K-ferrOcyanide/K-ferrIcyanide) to cover the bladders. Incubate at 30 &#176;C overnight protected from light.</li>
	<li>Remove splayed bladders from incubator and observe IBCs, which appear as blue puncta as visualized with a dissecting scope, 40-60X magnification. 
	NOTE: Sometimes a mouse may urinate prior to placement of the tube under the urethra. In this case, wait 15-20 min before attempting to collect urine again.</li>
</ol>
7. Urine Collection for Bacteriuria CFU Enumeration (Not Applicable for CAUTI)
<ol>
	<li>To collect urine pre or post infection gently restrain the mouse by holding the tail and placing it on a flat elevated surface (top of mouse cage).</li>
	<li>Hold a sterile 1.5 ml tube under the mouse urethra. Gently press down on the back of the mouse near the tail to apply pressure to the bladder. Catch the urine in the 1.5 ml tube.</li>
	<li>Make serial 1:10 dilutions out to 10-7 and plate each dilution on appropriate LB. Incubate the plates at 37 &#176;C overnight and count colonies the next day.</li>
</ol>
8. CAUTI Model Protocol, Catheter Needle Preparation for CAUTI Model (Figure S2)
<ol>
	<li>Remove the cap of the 30 G needle. Cut a 7 mm piece of PE10 tubing and 5 mm piece of silicone tubing such as RenaSIL. Thread the needle with PE10 tubing until the tubing touches the base of the needle.</li>
	<li>Then feed the 5 mm piece onto the PE10-containing needle. UV sterilize the needles assemblies overnight. 
	NOTE: When loading the silicone tubing, leave approximately 1 mm of the tubing extending past the needle tip.</li>
</ol>
9. E. faecalis OG1RF Bacterial Inoculum Preparation
<ol>
	<li>Prepare OG1RF inoculum starting 48 hr prior to the inoculation day. Streak OG1RF onto a BHI (brain heart infusion) plate from a frozen stock.</li>
	<li>Pick a colony and inoculate it into 10 ml of BHI and grow it statically for 18 hr at 37 &#176;C. Centrifuge the culture and wash the pellet (3 times) in 1 ml volumes of sterile 1x PBS.</li>
	<li>Resuspend the bacterial pellet into 1x PBS. Measure bacterial optical density and adjust the concentration to the desired inoculum size. The standard concentration of the inoculum used is 107; however, this may vary due to the design of the study</li>
</ol>
10. Catheter Implantation
<ol>
	<li>Clean the workstation with 70% ethanol and cover the area with an absorbent cover (or use sterile flow hood).</li>
	<li>Attach catheter-needle to an empty 1 ml (TB) syringe. Cut a 1 inch square piece of parafilm and put a dab of surgical lubricant (approximately the size of a dime) on top. 
	NOTE: Two different needles are used for deposition of the catheter and for the inoculum to minimized accidental inoculation due to the mechanical manipulation necessary for catheter implantation. This also allows for the convenience of preparing fewer inoculation needles.</li>
	<li>Anesthetize C57BL/6 mice by putting them in a 32 ounce glass jar containing a tea-infuser ball with cotton balls soaked in 3 ml of isoflurane or vaporizer chamber (following manufactures protocol) until unconscious but still breathing normally (1 breath/sec).</li>
	<li>Then put the mouse on its back on a paper towel and spread the legs. Cover the nose of the mouse with a nose cone or 50 ml conical tube with cotton balls containing a small amount (approximately 1-2 ml) of isoflurane. 
	CAUTION: Isoflurane is an inhalation anesthetic. Use in a well-ventilated area and minimize inhalation by researcher.</li>
	<li>Gently palpitate the bladder to induce urination and ensure a voided bladder. Wipe periurethral area with a 100% ethanol wipe.</li>
	<li>Disinfect the periurethral area with 10% povidone-iodine solution using a cotton-tip applicator. Dab the inoculation needle/syringe, point first, into the surgical lubricant. 
	NOTE: Povidine-iodine is used for catheter implantation, which is a stronger aseptic solution than alcohol. Catheter implantation requires more mechanical manipulation to insert the catheter and thus a greater potential for contamination.</li>
	<li>Insert the catheter into the urethral opening. Once in, use tweezers to push the (7 mm PE10) long piece toward the bladder, consequently pushing the (5 mm silicone) small catheter and depositing it into the bladder. Remove the needle, with the 7 mm tubing still attached, immediately.</li>
</ol>
11. Bacterial Inoculation
<ol>
	<li>Follow the bacterial inoculation protocol in section 3, using the prepared inoculum from section 9. Remove the mouse from the nose cone and return it to its cage</li>
</ol>
12.&#160;At Each Time Point of Sacrifice
<ol>
	<li>Prepare 5 ml tubes for bladder and kidney (For organs follow step 4.1) and 1.5 ml Eppendorf tube for catheters. Add 1 ml of 1x PBS into 1.5 ml Eppendorf tube for catheter samples</li>
	<li>Follow steps 4.3 to 4.5. Dissect the mouse, harvesting the bladder, kidneys and catheter, placing the tissues independently into 5 ml tubes containing 1x PBS and the catheter into the 1.5 ml Eppendorf (see step 12.1). 
	NOTE: Use different scissors for the external dissection and for organ harvesting and catheter retrieval to minimize contamination</li>
	<li>Then follow step 4.6.</li>
</ol>
13.&#160;Bacterial Recovery
<ol>
	<li>For organs, follow steps 5.1 and 5.2. For bacterial recovery from catheters, vortex at maximum speed for 30 sec, sonicate the catheter samples for 5 min using a bath sonicator, and then vortex at maximum speed for an additional 30 sec.</li>
	<li>Make serial 1:10 dilutions out to 10-8 and plate each dilution for CFU enumeration on BHI plates. Incubate the plates at 37 &#186;C overnight and count colonies the next day.</li>
</ol>

The authors declare that they have no competing financial interests.

Uncomplicated community acquired UTI is a common and costly infection accounting for several million primary care visits every year 46. In addition, CAUTIs are a common healthcare acquired infection that has become extremely costly to healthcare providers as the Centers of Medicare and Medicaid Services no longer reimburses providers for the added cost of treatment resulting from hospital acquired CAUTI 45. The mouse models of UTI, both uncomplicated cystitis and CAUTI, described in these protocols ...

An erratum was issued for: <a href="https://www.jove.com/video/52892/establishment-and-characterization-of-uti-and-cauti-in-a-mouse-model">Establishment and Characterization of UTI and CAUTI in a Mouse Model</a>. The Protocol section has been updated.
The ethics statement has been updated from:
Ethics statement: The Washington University Animal Studies Committee approved all mouse infections and procedures as part of protocol number 20120216, which was approved 01/11/2013 and expires 01/11/2016. Overall care of the animals was consistent with The guide for the Care and Use of Laboratory Animals from the National Research Council and the USDA Animal Care Resource Guide. Euthanasia procedures are consistent with the &#8220;AVMA guidelines for the Euthanasia of Animals 2013 edition.&#8221;
to:
Ethics statement: The Washington University Animal Studies Committee approved all mouse infections and procedures as part of protocol number 20150226, which expires 12/10/2018. Overall care of the animals was consistent with The Guide for the Care and Use of Laboratory Animals from the National Research Council and the USDA Animal Care Resource Guide. Euthanasia procedures are consistent with the &#8220;AVMA guidelines for the Euthanasia of Animals 2013 edition.&#8221;
Step 3 has been updated from:
3. Bacterial Inoculation
<ol>
	<li>Clean the workstation with 70% ethanol and cover area with absorbent paper (or use sterile flow hood).</li>
	<li>Draw up to 0.9 ml of the prepared bacterial inoculum into a 1 ml (TB) syringe (remove air bubbles). Attach a prepared sterile inoculation needle with PE10 tubing, onto the syringe containing the inoculum, then sterilely trim the polyethylene tubing. 
	NOTE: Leave 1 mm of tubing above the tip of the needle to avoid puncturing the bladder.</li>
	<li>Cut a 1 inch square piece of parafilm and put a dab of surgical lubricant (approximately the size of a dime) on top.</li>
	<li>Anesthetize female C3H/HeN mice by putting them in a 32 ounce glass jar containing a tea-infuser ball with cotton balls soaked with 3 ml of isoflurane or vaporizer chamber (following manufactures protocol) until unconscious but still breathing normally (1 breath/sec). 
	NOTE: Some IACUC committees do not approve the use of a jar or vaporizer. Please follow the indication of the IACUC committee of your institution. 
	NOTE: If glass jar with tea-ball and/or nose cone with isoflurane-soakd cotton balls are used, the animal must be carefully observed to avoid anesthetic overdose and death. The advantage of using a vaporizer and anesthesia chamber is that it provides controlled isoflurane administration that avoids accidental overdoses. CAUTION: Isoflurane is an inhalation anesthetic. Use in a well-ventilated area and minimize inhalation.</li>
	<li>Remove the mouse from the jar/vaporizer and place it on its back on a paper towel and spread the legs.</li>
	<li>Cover the nose of the mouse with a nose cone (a tube connected to the vaporizer that provides a controlled isoflurance dose that comes equipped on some vaporizer units) or 50 ml conical tube with a cotton ball containing a small amount (approximately 1-2 ml) of isoflurane.</li>
	<li>Gently palpitate the bladder to induce urination and ensure a voided bladder. Wipe the periurethral area with 100% ethanol wipe. Dab the inoculation needle/syringe, point first, into the surgical lubricant.</li>
	<li>Inoculate each mouse with 50 &#181;l of the bacteria solution by inserting the inoculation needle transurethrally, approximately 12 mm, and pressing down on the syringe plunger gently to dispense the inoculum into the bladder gently (10 &#181;l/sec). Remove the inoculation needle from the mouse. 
	NOTE: Immediate return of inoculum at the urethral opening when beginning to inoculate indicates improper or incomplete insertion of the needle.</li>
	<li>Remove the mouse from nose cone and return it to its cage. Repeat steps 3.3-3.8 for each mouse. When/if switching inoculum conditions/strains, dispose of the syringe and inoculation needle in an approved sharps container and start again step 3.2.&#160;&#160; 
	NOTE: Inoculation with uropathogenic organisms generally does not cause severe pain symptoms. However, in rare instances, administering large doses of pathogens may cause fever, reduction in food and water intake and abnormal behavior. Animal health should be monitored throughout the experiment. If overt pain symptoms are notice, an analgesic, such as Buprenorphine (0.05&#8722;0.1 mg/kg given subcutaneously), can be applied. Procedures should be in accordance with each institution&#8217;s IACUC.</li>
</ol>
to:
3. Bacterial Inoculation
<ol>
	<li>Clean the workstation with 70% ethanol and cover area with absorbent paper (or use sterile flow hood).</li>
	<li>Draw up to 0.9 ml of the prepared bacterial inoculum into a 1 ml (TB) syringe (remove air bubbles). Attach a prepared sterile inoculation needle with PE10 tubing, onto the syringe containing the inoculum, then sterilely trim the polyethylene tubing. 
	NOTE: Leave 1 mm of tubing above the tip of the needle to avoid puncturing the bladder.</li>
	<li>Cut a 1 inch square piece of parafilm and put a dab of surgical lubricant (approximately the size of a dime) on top.</li>
	<li>Anesthetize female C3H/HeN mice by putting them in a vaporizer chamber (following manufactures protocol) until unconscious but still breathing normally (1 breath/sec). 
	NOTE: Some IACUC committees do not approve the use of a vaporizer. Please follow the indication of the IACUC committee of your institution. 
	CAUTION: Isoflurane is an inhalation anesthetic. Use in a well-ventilated area and minimize inhalation.</li>
	<li>Remove the mouse from the vaporizer and place it on its back on a paper towel and spread the legs.</li>
	<li>Cover the nose of the mouse with a nose cone (a tube connected to the vaporizer that provides a controlled isoflurance dose that comes equipped on some vaporizer units) to maintain anesthetization.</li>
	<li>Gently palpitate the bladder to induce urination and ensure a voided bladder. Wipe the periurethral area with 100% ethanol wipe. Dab the inoculation needle/syringe, point first, into the surgical lubricant.</li>
	<li>Inoculate each mouse with 50 &#181;l of the bacteria solution by inserting the inoculation needle transurethrally, approximately 12 mm, and pressing down on the syringe plunger gently to dispense the inoculum into the bladder gently (10 &#181;l/sec). Remove the inoculation needle from the mouse. 
	NOTE: Immediate return of inoculum at the urethral opening when beginning to inoculate indicates improper or incomplete insertion of the needle.</li>
	<li>Remove the mouse from nose cone and return it to its cage. Repeat steps 3.3-3.8 for each mouse. When/if switching inoculum conditions/strains, dispose of the syringe and inoculation needle in an approved sharps container and start again step 3.2.&#160;&#160; 
	NOTE: Inoculation with uropathogenic organisms generally does not cause severe pain symptoms. However, in rare instances, administering large doses of pathogens may cause fever, reduction in food and water intake and abnormal behavior. Animal health should be monitored throughout the experiment. If overt pain symptoms are notice, an analgesic, such as Buprenorphine (0.05&#8722;0.1 mg/kg given subcutaneously), can be applied. Procedures should be in accordance with each institution&#8217;s IACUC.</li>
</ol>

An erratum was issued for: <a href="https://www.jove.com/video/52892/establishment-and-characterization-of-uti-and-cauti-in-a-mouse-model">Establishment and Characterization of UTI and CAUTI in a Mouse Model</a>. The Protocol section has been updated.

Erratum: Establishment and Characterization of UTI and CAUTI in a Mouse Model

Urinary tract infections (UTIs) are one of the most common bacterial infections and can be divided into two categories based on the mechanism of acquisition, community and nosocomial acquired UTI. Community-acquired UTIs often occur in otherwise healthy women and studies have shown that approximately 50% of women will have at least one UTI in their lifetime 1. Additionally, recurrence is a major problem. A woman who has an initial acute infection has a 25-40% chance of having a second infection within six months despite appropriate antibiotic treatment and many women continue to have frequent recurrences 2. The bacteria that cause these infections are also becoming increasingly antibiotic resistant further confounding treatment protocols 3-6. UTI affect millions of individuals every year costing approximately 2.5 billion dollars in health care related expenses in the US, underscoring the impact and prevalence of the disease 1,7.Nosocomial acquired UTIs are mainly associated with the presence of foreign bodies such as indwelling catheters. Catheter-associated UTIs (CAUTI) remain the most common nosocomial acquired UTI, accounting for ~70-80% of such infections 8. Furthermore, CAUTI is associated with increased morbidity and mortality and it is the most common cause of secondary bloodstream infections 9.
UPEC associated community acquired UTIs are thought to be caused by the introduction of bacteria into the bladder from reservoirs in the gastrointestinal tract via mechanical manipulation during sexual intercourse, poor hygiene or other microbial population dynamics between different host niches 10. Once inside the bladder, UPEC employ numerous virulence factors, including capsule, iron acquisition systems, toxins, a virulence plasmid, tRNAs, pathogenicity islands and colonization factors that have been shown to play a role in pathogenesis 11-14. Critical to the establishment of UPEC colonization, UPEC also encode multiple types of adhesive chaperone usher pathway (CUP) pili that recognize receptors with stereochemical specificity 15. Type 1 pili, tipped with the FimH adhesin, are expressed by UPEC and bind mannosylated uroplakins 16 and &#945;-1, &#946;-3 integrins 17, which are expressed on the luminal surface of both human and mouse bladders 18. These FimH-mediated interactions facilitate bacterial colonization and invasion of the superficial epithelial cells19,20. Once inside the cell, UPEC can escape into the cytoplasm where a single bacterium can rapidly divide to form an intracellular bacterial community (IBC), which upon maturation, can contain ~104 bacteria 21. IBC formation has been demonstrated in at least six different mouse strains, C3H/HeN, C3H/HeJ, C57Bl/6, CBA, FVB/NJ and BALB/c, and with a wide variety of different UPEC strains and other Enterobacteriaceae 22-24. However temporal and spatial differences of IBC formation can vary depending upon the mouse background and the infecting UPEC strain. In C3H/HeN mice infected with the prototypical UPEC strains UTI89 or CFT073, IBC formation can be visualized as small biomasses of bacteria as early as 3 hpi (hr post infection). This community continues to expand and reaches a &#8220;midpoint&#8221; of development approximately 6 hpi when the rod shaped bacteria occupy a large percentage of the cytoplasmic space of terminally differentiated superficial umbrella cells These early IBCs form in a relatively synchronous manner with the majority displaying similar dimensions and morphologies. ~8 hpi the bacteria in the IBC change from a bacilli to cocci morphology. IBCs are transient in nature. Thus, IBC maturation from 12-18 hpi results in continued expansion of the bacterial population, followed by their filamentation and dispersal out of the cell with subsequent spread to neighboring cells 23. Thus, the IBC niche allows for rapid bacterial growth in an environment protected from host immune responses and antibiotics 25. The distinct stages of UPEC infection that are seen in mice are also observed in humans, such as IBCs and filamentation, supporting the mouse model of UTI as a beneficial tool that can be used to model UTI in humans 22,26-28.
While a majority of women experience a UTI in their lifetime, the outcome of these infections can range from acute self-limiting infection with no recurrence, to frequent recurrent cystitis. Further, studies have shown a strong familial occurrence of UTI, suggesting a genetic component contributes to UTI susceptibility 29. We have found that the differing UTI outcomes seen in clinics can be mirrored by the differing outcomes of experimental UPEC infection among inbred mouse strains 30. For example, C3H/HeN, CBA, DBA, and C3H/HeOuJ mice are susceptible, in an infectious dose-dependent manner, to long-lasting, chronic cystitis characterized by persistent, high titer bacteria (&#62;104 colony forming units (CFU)/ml), high titer bacterial bladder burdens at sacrifice &#62;4 weeks post-infection (wpi), chronic inflammation, and urothelial necrosis . These mice also display elevated serum levels of IL-6, G-CSF, KC, and IL-5 within the first 24 hpi that serve as biomarkers for the development of chronic cystitis. This may accurately represent the natural course of UTI in some women, as placebo studies have shown that a large percentage of women experiencing UTI will retain high levels of bacteria in their urine for several weeks after the first symptoms of cystitis if not given antibiotic treatment 31,32. Further, using C3H/HeN mice, we found that a history of chronic cystitis is a significant risk factor for subsequent severe recurrent infections. Recurrent UTI is the most significant clinical manifestation of UTI and the C3H/HeN mouse is currently the only studied model that recapitulates an increased predisposition after previous exposure. A second UTI outcome is recapitulated in C57Bl/6 mice where acute UPEC infection is self-limiting, with resolution of bladder inflammation and bacteriuria within approximately a week. Interestingly, in this model, UPEC readily form quiescent intracellular reservoirs within the bladder tissue from which UPEC are capable of emerging from a dormant state to reinitiate an active UTI, potentially explaining one mechanism for same strain recurrent UTI in humans 33,34.
In addition to genetic influences on UTI susceptibility, introduction of a catheter into the bladder greatly increases the likelihood of having an infection as well as increasing the range of bacteria able to cause an infection. It has been demonstrated that human urinary catheterization causes histological and immunological changes in the bladder due to mechanical stress that results in a robust inflammatory response, exfoliation, edema of the lamina propria and submucusa, urothelial thinning, and mucosal lesion of the urothelium and kidney 35,36. Additionally, the catheter provides a surface for bacterial attachment thereby creating an environment utilized by several species to cause CAUTI. While UPEC are still a major contributor, Enterococcus faecalis accounts for 15% of these CAUTI 37. E. faecalis is becoming increasingly resistant to antibiotics with vancomycin resistance emergence, posing a serious health concern 38. E. faecalis possess numerous virulence factors including toxins and adhesins necessary for attachment to both the catheter and epithelium 38. During urinary catheterization, the host is vulnerable to microbial adhesion, multiplication and dissemination in the urinary tract 39,40. E. faecalis forms a biofilm on the catheter as part of a mechanism to persist in the bladder and disseminate to the kidneys, which is reproduced in a mouse CAUTI model41. Recently, it has been shown during urinary catheterization, fibrinogen (Fg) is released into the bladder as part of the inflammatory response. Fg accumulates in the bladder, coats the catheter and is essential for E. faecalis biofilm formation, functioning as an attachment scaffold. In a C57BL/6 mouse model of CAUTI, we discovered that E. faecalis biofilm formation on the catheter, and thus persistence in the bladder, was dependent on the Ebp pilus, specifically its tip adhesin EbpA. We found that the N-terminal domain of EbpA specifically binds to Fg coating the catheter. Additionally, it was found that E. faecalis utilizes Fg as metabolite source during infection, thus enhancing biofilm formation 42.
Mouse models have proven critical to understanding as well as predicting clinical manifestations of UTI and CAUTI 41. In this article we demonstrate inoculum preparation of the cystitis UPEC isolate UTI89 and transurethral inoculation of C3H/HeN mice. Additionally, we demonstrate a protocol for catheter insertion in C57BL/6 mice and inoculation of the E. faecalis OG1RF strain. Both of these techniques lead to consistent and reliable UTI or CAUTI in mice. We also display techniques used to observe IBC formation during acute cystitis and urine collection for the analysis of chronic or recurrent cystitis. C3H/HeN mice have been used to study aspects of UPEC pathogenesis including initial bacterial invasion, IBC formation, filamentation and the development of chronic cystitis 23,33,43. These virulence parameters have also been studied in a variety of other mouse backgrounds 22,33. For CAUTI, the C57BL/6 model allows for foreign body implantation into the bladder with subsequent bacterial colonization, which can be maintained for 7 days post infection 41. These models have been useful for assessing bacterial virulence mechanisms, host responses to UTI and mechanisms to subvert host responses, much of which has been subsequently recapitulated or observed in clinical human populations.

<table><tbody><tr><td>Material for catheter and needle preparation:</td><td></td><td></td><td></td></tr><tr><td>30 G needles</td><td>BD Precision Glide</td><td>305106</td><td>30 G x &amp;frac12; (0.3 mm x 13 mm)</td></tr><tr><td>PE10 polyethylene tubing</td><td>BD</td><td>427400</td><td>Inside diameter -0.011 in (0.28 mm); outside diameter &amp;ndash; 0.024 in (0.61 mm)</td></tr><tr><td>RenaSIL 025 platinum cured silicon tubing</td><td>Braintree Scientific, Inc</td><td>SIL 025</td><td>inside diameter-0.012 x outside diameter 0.025, 25 ft coil</td></tr><tr><td colspan="4">Material for infections:</td></tr><tr><td>Isoflourane &amp;ndash; Isothesia</td><td>Butler Schein</td><td>29405</td><td>250 ml</td></tr><tr><td>Clear Glass Straight-Sided Jar</td><td>Kimble Chase</td><td>5413289V 21</td><td></td></tr><tr><td>Stainless Steel Tea Infuser</td><td>Schefs-Amazon</td><td></td><td>Premium Loose Leaf Tea Infuser By Schefs - Stainless Steel - Large Capacity -</td></tr><tr><td>Non-sterile cotton balls</td><td>Fisherbrand</td><td>22-456-880</td><td></td></tr><tr><td>50 ml Falcon tubes</td><td>VWR</td><td>89039-660</td><td></td></tr><tr><td>Isotec 3 -vaporizer</td><td>Ohmeda</td><td>1224478</td><td></td></tr><tr><td>Ear punch</td><td>Fisher Scientific</td><td>13-812-201</td><td>(when necessary)</td></tr><tr><td>Betadine solution</td><td>Betadine solution</td><td></td><td>10% Povidie-iodine topical solution</td></tr><tr><td>Q-tips</td><td>Fisher Scientific</td><td>22-037-924</td><td>6 inches</td></tr><tr><td>Diapers for bench</td><td>Fisherbrand</td><td>14206 63</td><td>Absorbent Underpads (20&amp;rdquo;X36&amp;rdquo;mats)</td></tr><tr><td>Surgical lubricant</td><td>Surgilube</td><td>0281-0205-36</td><td></td></tr><tr><td>Dissecting scissor</td><td>Fine Science tools, INC</td><td>14084-08</td><td></td></tr><tr><td>Micro-Adson Forceps</td><td>Fine Science tools, INC</td><td>11018-12</td><td></td></tr><tr><td>1 ml syringe</td><td>BD</td><td>309659</td><td>Tuberculin slip tip</td></tr><tr><td>Parafilm</td><td>Bemis</td><td>PM996</td><td>4 inches x 125 FT</td></tr><tr><td>Eppendorf rack</td><td>Fisherbrand</td><td>05-541-1</td><td></td></tr><tr><td>Eppendorf tubes</td><td>MIDSCI</td><td>AVX-T-17-C</td><td></td></tr><tr><td colspan="4">Harvesting catheters, bladders and kidneys:</td></tr><tr><td>Homogenizer</td><td>PRO Scientific INC</td><td>Bio-Gen Pro 200</td><td></td></tr><tr><td>5 ml polypropylene round-bottom tube</td><td>BD</td><td>352063</td><td>for organ homogenization</td></tr><tr><td>Paper towel</td><td>Georgia-Pacific</td><td></td><td></td></tr><tr><td>Ethanol</td><td>Pharmco-AAPER</td><td>11100020S</td><td>200 proof</td></tr><tr><td>Costar&amp;trade; Clear Polystyrene 96-Well Plates</td><td>Corning</td><td>3788</td><td></td></tr><tr><td>1x Phosphate sodium saline</td><td>Sigma-Aldrich</td><td>P3813</td><td></td></tr><tr><td>BRANSONIC Ultrasonic cleaner 1210</td><td>Branson Ultrasonics Corporation</td><td>1210</td><td></td></tr><tr><td colspan="4">IBC materials:</td></tr><tr><td>6-well tissue culture test plate</td><td>Techno Plastic Products</td><td>92006</td><td></td></tr><tr><td>Pins</td><td>Fine Science Tools</td><td>26002-20</td><td></td></tr><tr><td>Sylgard 184</td><td>Dow Corning</td><td>3097358-1004</td><td>Silicone Elastomer Kit</td></tr><tr><td>X-gal (5-bromo-4-chloro-3-indolyl-b-D-galactoside)</td><td>Invitrogen</td><td>15520-034</td><td>Ultrapure</td></tr><tr><td>N, N-Dimethylformamide</td><td>Sigma Aldrich</td><td>D4551</td><td></td></tr><tr><td>MgCl&lt;sub&gt;2&lt;/sub&gt; (Magnesium chloride)</td><td>Sigma Aldrich</td><td>M8266</td><td></td></tr><tr><td>Sodium deoxycholate</td><td>Sigma Aldrich</td><td>D6750</td><td></td></tr><tr><td>Nonidet-P40</td><td>Roche</td><td>11754599001</td><td>Octylphenolpoly(ethyleneglycolether)n</td></tr><tr><td>Potassium hexacyanoferrate(II) trihydrate (K-ferrOcyanide)</td><td>Sigma Aldrich</td><td>P3289</td><td></td></tr><tr><td>Potassium hexacyanoferrate(III) (K-ferrIcyanide)</td><td>Sigma Aldrich</td><td>60299</td><td></td></tr></tbody></table>

establishment and characterization of uti and cauti in a mouse model

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. Females are disproportionately afflicted by UTI: 50% of all women will have a UTI in their lifetime. Additionally, 20-40% of these women who have an initial UTI will suffer a recurrence with some suffering frequent recurrences with serious deterioration in the quality of life, pain and discomfort, disruption of daily activities, increased healthcare costs, and few treatment options other than long-term antibiotic prophylaxis. Uropathogenic Escherichia coli (UPEC) is the primary causative agent of community acquired UTI. Catheter-associated UTI (CAUTI) is the most common hospital acquired infection accounting for a million occurrences in the US annually and dramatic healthcare costs. While UPEC is also the primary cause of CAUTI, other causative agents are of increased significance including Enterococcus faecalis. Here we utilize two well-established mouse models that recapitulate many of the clinical characteristics of these human diseases. For UTI, a C3H/HeN model recapitulates many of the features of UPEC virulence observed in humans including host responses, IBC formation and filamentation. For CAUTI, a model using C57BL/6 mice, which retain catheter bladder implants, has been shown to be susceptible to E. faecalis bladder infection. These representative models are being used to gain striking new insights into the pathogenesis of UTI disease, which is leading to the development of novel therapeutics and management or prevention strategies.

The intravesical models of uncomplicated and catheter associated UTI provide flexible platforms for elucidating the molecular mechanisms of bacterial pathogenesis, the impact of these diseases on host tissue, and the development and testing of novel approaches to manage these common and costly infections. Depending on the mouse strain and pathogen, intravesical inoculation can be used to study host-pathogen interactions to elucidate factors necessary for initiating or modulating acute (Figure 1 and <stro...

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Establishment and Characterization of UTI and CAUTI in a Mouse Model

The ability to model urinary tract infections (UTI) is crucial in order to be able to understand bacterial pathogenesis and spawn the development of novel therapeutics. This work&#8217;s goal is to demonstrate mouse models of experimental UTI and catheter associated UTI that recapitulate and predict findings seen in humans.

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. ...

establishment-and-characterization-of-uti-and-cauti-in-a-mouse-model

Research

JoVE Journal

Medicine

20.0K Views.  Washington University School of Medicine. The ability to model urinary tract infections (UTI) is crucial in order to be able to understand bacterial pathogenesis and spawn the development of novel therapeutics. This work’s goal is to demonstrate mouse models of experimental UTI and catheter associated UTI that recapitulate and predict findings seen in humans.

Video: Establishment and Characterization of UTI and CAUTI in a Mouse Model

Transurethral Induction of Mouse Urinary Tract Infection

Uropathogenic bacterial strains of interest are grown on agar. Generally, uropathogenic E. coli (UPEC) and other strains can be grown overnight on Luria-Bertani (LB) agar at 37&deg;C in ambient air. UPEC strains grow as yellowish-white translucent colonies on LB agar. Following confirmation of appropriate colony morphology, single colonies are then picked to be cultured in broth. LB broth can be used for most uropathogenic bacterial strains. Two serial, overnight LB broth cultures can be employed to enhance expression of type I pili, a well-defined virulence factor for uropathogenic bacteria. Broth cultures are diluted to the desired concentration in phosphate buffered saline (PBS). Eight to 12 week old female mice are placed under isoflurane anesthesia and transurethrally inoculated with bacteria using polyethylene tubing-covered 30 gauge syringes. Typical inocula, which must be empirically determined for each bacterial/mouse strain combination, are 106 to 108 cfu per mouse in 10 to 50 microliters of PBS. After the desired infection period (one day to several weeks), urine samples and the bladder and both kidneys are harvested. Each organ is minced, placed in PBS, and homogenized in a Blue Bullet homogenizer. Urine and tissue homogenates are serially diluted in PBS and cultured on appropriate agar. The following day, colony forming units are counted.

This video will demonstrate methods to transurethrally induce mouse urinary tract infections and quantify the extent of resulting infections.

Uropathogenic bacterial strains of interest are grown on agar. Generally, uropathogenic E. coli (UPEC) and other strains can be grown overnight on ...

Immunology and Infection

A Mouse Model of in Utero Transplantation

The transplantation of stem cells and viruses in utero has tremendous potential for treating congenital disorders in the human fetus. For example, in utero transplantation (IUT) of hematopoietic stem cells has been used to successfully treat patients with severe combined immunodeficiency.1,2 In several other conditions, however, IUT has been attempted without success.3 Given these mixed results, the availability of an efficient non-human model to study the biological sequelae of stem cell transplantation and gene therapy is critical to advance this field. We and others have used the mouse model of IUT to study factors affecting successful engraftment of in utero transplanted hematopoietic stem cells in both wild-type mice4-7 and those with genetic diseases.8,9 The fetal environment also offers considerable advantages for the success of in utero gene therapy. For example, the delivery of adenoviral10, adeno-associated viral10, retroviral11, and lentiviral vectors12,13 into the fetus has resulted in the transduction of multiple organs distant from the site of injection with long-term gene expression. in utero gene therapy may therefore be considered as a possible treatment strategy for single gene disorders such as muscular dystrophy or cystic fibrosis. Another potential advantage of IUT is the ability to induce immune tolerance to a specific antigen. As seen in mice with hemophilia, the introduction of Factor IX early in development results in tolerance to this protein.14 
In addition to its use in investigating potential human therapies, the mouse model of IUT can be a powerful tool to study basic questions in developmental and stem cell biology. For example, one can deliver various small molecules to induce or inhibit specific gene expression at defined gestational stages and manipulate developmental pathways. The impact of these alterations can be assessed at various timepoints after the initial transplantation. Furthermore, one can transplant pluripotent or lineage specific progenitor cells into the fetal environment to study stem cell differentiation in a non-irradiated and unperturbed host environment.
The mouse model of IUT has already provided numerous insights within the fields of immunology, and developmental and stem cell biology. In this video-based protocol, we describe a step-by-step approach to performing IUT in mouse fetuses and outline the critical steps and potential pitfalls of this technique.

A Mouse Model of in Utero Transplantation

The mouse model of in utero transplantation is a versatile tool that can be used to study the potential clinical applications of stem cell transplantation and gene therapy in the fetus. In this protocol, we present a general approach to performing this technique

The transplantation of stem cells and viruses in utero has tremendous potential for treating congenital disorders in the human fetus.  For example, in ...

Multi-photon Imaging of Tumor Cell Invasion in an Orthotopic Mouse Model of Oral Squamous Cell Carcinoma

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient management strategies. Orthotopic mouse models of oral cancer have been developed to facilitate the study of factors that impact invasion and serve as model system for evaluating anti-tumor therapeutics. In these systems, visualization of disseminated tumor cells within oral cavity tissues has typically been conducted by either conventional histology or with in vivo bioluminescent methods. A primary drawback of these techniques is the inherent inability to accurately visualize and quantify early tumor cell invasion arising from the primary site in three dimensions. Here we describe a protocol that combines an established model for squamous cell carcinoma of the tongue (SCOT) with two-photon imaging to allow multi-vectorial visualization of lingual tumor spread. The OSC-19 head and neck tumor cell line was stably engineered to express the F-actin binding peptide LifeAct fused to the mCherry fluorescent protein (LifeAct-mCherry). Fox1nu/nu mice injected with these cells reliably form tumors that allow the tongue to be visualized by ex-vivo application of two-photon microscopy. This technique allows for the orthotopic visualization of the tumor mass and locally invading cells in excised tongues without disruption of the regional tumor microenvironment. In addition, this system allows for the quantification of tumor cell invasion by calculating distances that invaded cells move from the primary tumor site. Overall this procedure provides an enhanced model system for analyzing factors that contribute to SCOT invasion and therapeutic treatments tailored to prevent local invasion and distant metastatic spread. This method also has the potential to be ultimately combined with other imaging modalities in an in vivo setting.

A comprehensive overview of the techniques involved in generating a mouse model of oral cancer and quantitative monitoring of tumor invasion within the tongue through multi-photon microscopy of labeled cells is presented. This system can serve as a useful platform for the molecular assessment and drug efficacy of anti-invasive compounds.

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient ...

Establishment of Human Epithelial Enteroids and Colonoids from Whole Tissue and Biopsy

The epithelium of the gastrointestinal tract is constantly renewed as it turns over. This process is triggered by the proliferation of intestinal stem cells (ISCs) and progeny that progressively migrate and differentiate toward the tip of the villi. These processes, essential for gastrointestinal homeostasis, have been extensively studied using multiple approaches. Ex vivo technologies, especially primary cell cultures have proven to be promising for understanding intestinal epithelial functions. A long-term primary culture system for mouse intestinal crypts has been established to generate 3-dimensional epithelial organoids. These epithelial structures contain crypt- and villus-like domains reminiscent of normal gut epithelium. Commonly, termed &#8220;enteroids&#8221; when derived from small intestine and &#8220;colonoids&#8221; when derived from colon, they are different from organoids that also contain mesenchyme tissue. Additionally, these enteroids/colonoids continuously produce all cell types found normally within the intestinal epithelium. This in vitro organ-like culture system is rapidly becoming the new gold standard for investigation of intestinal stem cell biology and epithelial cell physiology. This technology has been recently transferred to the study of human gut. The establishment of human derived epithelial enteroids and colonoids from small intestine and colon has been possible through the utilization of specific culture media that allow their growth and maintenance over time. Here, we describe a method to establish a small intestinal and colon crypt-derived system from human whole tissue or biopsies. We emphasize the culture modalities that are essential for the successful growth and maintenance of human enteroids and colonoids.

We describe a method to establish human enteroids from small intestinal crypts and colonoids from colon crypts collected from both surgical tissue and biopsies. In this methodological article, we present the culture modalities that are essential for the successful growth and maintenance of human enteroids and colonoids.

The epithelium of the gastrointestinal tract is constantly renewed as it turns over. This process is triggered by the proliferation of intestinal stem ...

Urinary Tract Infection in a Small Animal Model: Transurethral Catheterization of Male and Female Mice

Urinary tract infections (UTI) are extremely common worldwide, incurring significant morbidity and healthcare-associated expenses. Small animal models, which accurately reflect disease establishment and progression, permit dissection of host-pathogen interactions and generation of immunity to infection. In mice, intravesical instillation of uropathogenic E. coli, the causative agent in more than 85% of community acquired UTI, recapitulates many of the stages of infection observed in humans. Until recently, however, UTI could only be modeled in female animals. This limitation has hindered the study of sex-related differences in UTI, as well as other bladder pathologies, such as cancer. Here, we describe a method to instill male mice that allows direct comparison between female and male animals and provide a detailed protocol to assess bladder tissue by flow cytometry as a means to better understand host responses to infection. Together, these approaches will aid in the identification of host factors that contribute to sex biases observed in UTI and other bladder-associated diseases.

The established model of transurethral catheterization of mice allows the study of bladder pathologies, including urinary tract infection, but can only be performed in females. A new model of male transurethral instillation, presented here, will enable research in an area marked by strong clinical and epidemiological differences between the sexes.

Urinary tract infections (UTI) are extremely common worldwide, incurring significant morbidity and healthcare-associated expenses. Small animal models, ...

Recurrent Escherichia coli Urinary Tract Infection Triggered by Gardnerella vaginalis Bladder Exposure in Mice

Recurrent urinary tract infections (rUTI) caused by uropathogenic Escherichia coli (UPEC) are common and costly. Previous articles describing models of UTI in male and female mice have illustrated the procedures for bacterial inoculation and enumeration in urine and tissues. During an initial bladder infection in C57BL/6 mice, UPEC establish latent reservoirs inside bladder epithelial cells that persist following clearance of UPEC bacteriuria. This model builds on these studies to examine rUTI caused by the emergence of UPEC from within latent bladder reservoirs. The urogenital bacterium Gardnerella vaginalis is used as the trigger of rUTI in this model because it is frequently present in the urogenital tracts of women, especially in the context of vaginal dysbiosis that has been associated with UTI. In addition, a method for in situ bladder fixation followed by scanning electron microscopy (SEM) analysis of bladder tissue is also described, with potential application to other studies involving the bladder.

Recurrent Escherichia coli Urinary Tract Infection Triggered by Gardnerella vaginalis Bladder Exposure in Mice

A mouse model of uropathogenic E. coli (UPEC) transurethral inoculation to establish latent intracellular bladder reservoirs and subsequent bladder exposure to G. vaginalis to induce recurrent UPEC UTI is demonstrated. Also demonstrated are the enumeration of bacteria, urine cytology, and in situ bladder fixation and processing for scanning electron microscopy.

Recurrent urinary tract infections (rUTI) caused by uropathogenic Escherichia coli (UPEC) are common and costly. Previous articles describing models of ...

Longitudinal Follow-Up of Urinary Tract Infections and Their Treatment in Mice using Bioluminescence Imaging

Urinary tract infections (UTI) rank among the most common bacterial infections in humans and are routinely treated with empirical antibiotics. However, due to increasing microbial resistance, the efficacy of the most used antibiotics has declined. To find alternative treatment options, there is a great need for a better understanding of the UTI pathogenesis and the mechanisms that determine UTI susceptibility. In order to investigate this in an animal model, a reproducible, non-invasive assay to study the course of UTI is indispensable.
For years, the gold standard for the enumeration of bacterial load has been the determination of Colony Forming Units (CFU) for a particular sample volume. This technique requires post-mortem organ homogenates and serial dilutions, limiting data output and reproducibility. As an alternative, bioluminescence imaging (BLI) is gaining popularity to determine the bacterial load. Labeling pathogens with a lux operon allow for the sensitive detection and quantification in a non-invasive manner, thereby enabling longitudinal follow-up. So far, the adoption of BLI in UTI research remains limited.
This manuscript describes the practical implementation of BLI in a mouse urinary tract infection model. Here, a step-by-step guide for culturing bacteria, intravesical instillation and imaging is provided. The in vivo correlation with CFU is examined and a proof-of-concept is provided by comparing the bacterial load of untreated infected animals with antibiotic-treated animals. Furthermore, the advantages, limitations, and considerations specific to the implementation of BLI in an in vivo UTI model are discussed. The implementation of BLI in the UTI research field will greatly facilitate research on the pathogenesis of UTI and the discovery of new ways to prevent and treat UTI.

This manuscript describes the intravesical administration of uropathogenic bacteria with a lux operon to induce a urinary tract infection in mice and subsequent longitudinal in vivo analysis of the bacterial load using bioluminescence imaging.

Urinary tract infections (UTI) rank among the most common bacterial infections in humans and are routinely treated with empirical antibiotics. However, ...

Transurethral Catheterization and Bacterial Inoculation to Generate a Urinary Tract Infection Model in Male Mice

Source: Zychlinsky Scharff, A., et al. <a href="https://app.jove.com/v/54432">Urinary Tract Infection in a Small Animal Model: Transurethral Catheterization of Male and Female Mice.</a> J. Vis. Exp. (2017).
This video demonstrates the establishment of uropathogenic Escherichia coli infection in a female mouse bladder through transurethral catheter instillation. The injected bacteria successfully establish a urinary tract infection within the bladder, providing a valuable platform for studying healthcare-associated diseases related to the urinary tract.

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

JoVE Encyclopedia of Experiments

Immunology

Immunopathology

Host-Pathogen Interaction Models

Generating Urinary Tract Infection Model: A Technique to Administer Uropathogenic Bacteria in Mouse to Develop Urinary Tract Infection Model

Source: Luyts, N. et al., <a href="https://www.jove.com/v/62614">Longitudinal Follow-Up of Urinary Tract Infections and Their Treatment in Mice using Bioluminescence Imaging.</a> J. Vis. Exp. (2021).
In this video, we describe the technique for the intravesical administration of uropathogenic bacteria to generate a urinary tract infection model in mice. This model can be used to understand the spread of uropathogenic bacterial infection within the urinary system and the mechanisms that determine UTI susceptibility.

Source: Luyts, N. et al., Longitudinal Follow-Up of Urinary Tract Infections and Their Treatment in Mice using Bioluminescence Imaging. J. Vis. Exp. ...

Biology

Rodent Models

Adult

Establishment of a Urinary Tract Infection Model in Female Mice

After anesthetizing a female mouse, according to an approved protocol, place the animal supine, and apply medium pressure to the lower abdomen to empty the bladder of urine. Full bladders feel like a pea under the skin between the iliac crests.
When the bladder is empty, using the non-dominant hand, place the thumb on the tail and a finger of the same hand on the abdomen of the mouse, and apply gentle pressure in opposing directions to hold the mouse firmly in place. Next, place the tip of the catheter perpendicular to the mouse at the urethral orifice. Then, with gentle pressure, slide the catheter into the urethra until the hub meets the urethral orifice while simultaneously lowering the syringe so that it is parallel to the working surface.
Once the catheter is in place, use a finger from the non-dominant hand that&#39;s resting on the mouse&#39;s abdomen to very gently pull the abdominal skin towards the head of the mouse. Note that the urethral orifice will not move. However, if the catheter is in the vagina, the tissue will move up and away from the catheter. With the hub of the catheter against the urethral orifice, slowly dispense 50 microliters of the bacterial inoculum.
A slow installation rate minimizes vesicoureteral reflux into the kidney. Slowly remove the catheter to prevent leakage, then, place the animal in its cage in a supine position.

Establishment and Characterization of UTI and CAUTI in a Mouse Model

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