Characterization of UTI causing E.coli and K.pneumoniae clinical isolates and assessing their effect on urinary tract cells

Abstract

Title: Characterization of UTI Causing E.coli and K.pneumoniae Clinical Isolates and Assessing their Effect on Urinary Tract Cells

Background: Urinary tract infections are among the most common bacterial infections in the world. They are highly recurrent affecting more than 150 million patients every year. UTIs cause a considerable burden on both the economy and health care system. They are caused by a group of uropathogenic organisms, where Escherichia coli (E.coli) is the major player in addition to Klebsiella pneumonia (K.pneumoniae) and other gram negative and gram positive bacteria. With the alarming rise of antimicrobial resistance among uropathogens, the treatment of urinary tract infections becomes difficult and thus the need for understanding bacterial pathogenicity in the urinary tract becomes essential.

Aim: The purpose of this study is to characterize the genetic and molecular factors of the E.coli and K.pneumoniae clinical isolates isolated from UTI and non-UTI patients and study their effect on uroepithelium cell line.

Methods: Twelve E.coli and fifteen K.pneumoniae clinical isolates were collected from UTI and non-UTI patients. The antimicrobial susceptibility testing was performed on the clinical isolates by disk diffusion and broth microdilution assays using 17 antimicrobials from different antimicrobial families. To study the ability of the E.coli clinical isolates to form biofilm in vitro, biofilm formation assay was done. The effect of planktonic and biofilm extracts of the E.coli clinical isolates on the SV-HUC urothelium cell line was examined using MTT assay. Eight E.coli and seven K.pneumoniae clinical isolates were subjected to whole genome sequencing in order to search for differences at the genomic level including antimicrobial resistance genes and virulence factors.

Results: Disk diffusion and BMDs results showed that most of the E.coli and K.pneumoniae clinical isolates belonged to the MDR, XDR and PDR resistance phenotypes. Ten out of the twelve E.coli clinical isolates were able to form biofilm in vitro. The biofilm extract of 7 out of the 10 E.coli biofilm producers had an anti-proliferative effect on the SV-HUC urothelium cell line. The biofilm extract of E.coli 49-448 had the most noticeable suppressive effect on the proliferation of SV-HUC urothelium cell line. The planktonic extract of 7 out of the 12 E.coli clinical isolates showed an anti-proliferative effect on the SV-HUC uroepithelium cell line. The planktonic extract of E.coli 50-452 had an important anti-proliferative effect on the SV-HUC urothelium cell line. WGS results showed that several antimicrobial resistance genes and chromosomal point mutations conferring resistance for all antimicrobial families were detected in both E.coli and K.pneumoniae clinical isolates. Thirteen different plasmids were detected in the 8 sequenced E.coli isolates with 8 different sequence types. WGS results of the E.coli isolates showed 33 different virulence factors ranging between fimbrial proteins, iron siderophore receptors and toxins. Twelve different plasmids were detected in the 7 sequenced K.pneumoniae clinical isolates with only 1 known detected sequence type. Conclusion: Antimicrobial resistance is severely arising among uropathogens, E.coli and K.pneumoniae, thus resulting in a significant decrease in the efficiency of antimicrobials used for treatment. Biofilm production plays an essential role in increasing the chance of recurrent and persistent infections in UTI patients. Several virulence factors expressed in the E.coli isolates may be behind the anti-proliferative inhibitory effect of biofilm and planktonic extracts on the SV-HUC urothelium cell line.