MP02-10: Genus-Level Identification of Bladder-Resident Bacteria Associated With Recurrent Urinary Tract Infection in Post-Menopausal Women by Fluorescence In-Situ Hybridization

Parker R.M. Kenee*, Dallas, TX, Jashkaran Gadhvi, Fatima Khan, Richardson, TX, Alana L. Christie, Dallas, TX, Nicole J. De Nisco, Richardson, TX, Philippe E. Zimmern, Dallas, TX

Introduction: Many women with antibiotic-recalcitrant recurrent urinary tract infection (rUTI) elect to undergo electrofulguration (EF) of areas of chronic cystitis when standard antibiotic therapies fail. One potential benefit of this procedure is the removal of tissue-resident bacterial communities.1 Although tissue-resident bacteria have been observed in the bladder walls of postmenopausal women with rUTI by fluorescence in situ hybridization (FISH), genus and species level identification of these bacteria by FISH has not yet been reported.2 We used genus-specific probes to quantify Escherichia spp. in the bladder biopsies of postmenopausal women with rUTI undergoing EF and investigate the relationship between detected bacterial community sizes and stage of cystitis.

Methods: Following IRB approval, bladder biopsies were obtained from consenting postmenopausal women who elected EF for the advanced management of rUTI. Biopsies were immediately fixed in paraformaldehyde and then paraffin-embedded and sectioned (5 µm). FISH was performed as described previously2 using the following probe sets: 1.  Scramble-AlexaFluor488/647 (control), 2. Universal 16s rRNA-AlexaFluor647 (all bacteria), and 3. Escherichia 16s rRNA-AlexaFluor488. Slides were imaged using a Zeiss LSM880 with a 63x objective. 10 randomly sampled images were collected for each biopsy section. Images analysis was performed blinded to quantify bacterial community sizes. Patients were classified by cystitis stage (Stage 1 (trigone alone) to 4 (pancystitis)) determined by cystoscopy at time of the biopsy. Least mean squares statistical analysis was used to generate an average number of bacterial organisms per 10x1 µm2 for each cystitis stage.

Results: Genera-specific FISH was performed on bladder biopsies from 23 women. The universal 16s rRNA probe detected tissue-resident bacteria in the biopsies of 95.7% (22/23) women. Tissue-resident Escherichia spp. were detected in the bladder biopsies of 52.1% (12/23) women. The highest average bacterial community sizes were observed in Stage 1 cystitis bladder biopsies (8.4 per 10x1 µm2, 95% C.I. 6.6 – 10.1).

Conclusions: For the first time, 16s rRNA FISH detected Escherichia spp. in the bladders of postmenopausal women electing to undergo EF for the advanced management of rUTI. Interestingly, bladder-resident bacterial community sizes were highest in bladder biopsies from women with Stage 1 cystitis.

References 1. Mysorekar et al. PNAS 2006  2. DeNisco et al. JMB 2019

Source of Funding: N/A