MP02-04: Invasion and Assessment of Intracellular Bacterial Colony (IBC) Formation in Uroepithelial Cells by Uropathogenic E. Coli (UPEC) of Different Phylogenetic Groups Suggest Alternate Entry Mechanisms

Jacob Hogins*, Richardson, TX, Philippe Zimmern, Dallas, TX, Larry Reitzer, Richardson, TX

Introduction: UPEC strains cause most urinary tract infections (UTIs), and phylogenetic group B2 strains cause ~75% of these UTIs, while strains from 3 other groups cause ~25% UTIs. UTIs require bacterial attachment, entry, and replication within epithelial cells to form IBCs. The attachment mechanism has been studied in two strains: one strain required pili, while a second strain used flagella. Several transcriptomic analyses showed significant differences in pili and flagella gene expression, which suggested alternate infection mechanisms. We examined invasion and IBC formation from one UPEC and one non-pathogenic E. coli (NPEC) strain from groups A, B2, and D in two different urothelial cell lines.

Methods: Two urothelial cancer cell lines, CRL2169 (female grade 1 transitional carcinoma) and HTB-9 (male grade 2 carcinoma), and six bacterial strains (W3110, ECOR14, ECOR51, UTI89, ECOR48, ECOR44) were assessed for bacterial invasion. Cancer cells at 80% confluence were incubated with actively growing bacteria for 2 hours and then the culture media was replaced with fresh media with antibiotics to kill external bacteria. The cancer cells were incubated overnight, lysed, and the surviving bacteria counted after plating on LB agar plates.

Results: Group B2 bacteria were significantly better invaders than group A or D strains, and, unexpectedly, UPEC strains did not invade better than the NPEC strain within each group. Transcriptomic analysis showed expression of flagellar genes, but not pili genes, in most strains, but the converse was shown for UTI89. These results suggested different mechanisms of entry. Examination of TLR receptor gene expression in the cancer cell lines showed that CRL-2169 had greater TLR5 (bacterial flagella recognizing TLR) expression which correlated to the higher invasiveness by flagella overexpressing strains. The NPEC group A strain W3110 was a poor invader, but the few W3110 cells that entered the epithelial cells had increased flagella gene expression. We also note that one group D UPEC strain does not express either pili or flagella.

Conclusions: Bacteria can attach and invade by different mechanisms, which suggests a synergy between the bacteria and the patient which is being infected. For example, a patient with high TLR5 expression is likely to be susceptible to strains with flagella.

Source of Funding: N/A