Biofilm Formation In Uropathogenic Escherichia Coli Strains; Relationship With Virulence Factors And Antimicrobial Resistance In Tertiary Care Hospital In North Karnataka Region

Abstract

Background: Urinary tract infection (UTI) is a clinical condition, with characteristic symptoms, colonization and multiplication of bacteria in significant numbers i.e., 105 cfu/ml within the urinary tract. At this juncture, antimicrobial resistance (AMR) in uropathogens has become one of the major concerns globally. Aim & Objectives: The present study aimed to demonstrate the biofilm formation in Uropathogenic Escherichia coli strains; characterize the phenotypic & genotypic virulence factors and their relationship with antimicrobial resistance. Methods: In the present study, 1000 suspected UTI cases were included. Urine samples were processed for culture and antimicrobial drug susceptibility testing. Only Escherichia coli (E. coli) isolates were further studied for production of biofilm, ESBL and haemolysin enzymes by phenotypic methods. Virulence genes, papEF, traT & PAI were detected by multiplex PCR. Molecular confirmation of virulence genes was done by Sanger sequencing. Sequences were studied for characterization and mutations. Results: From the 1000 urine samples, 395 E. coli were isolated. Higher resistance was observed with antibiotics - ampicillin (82.53%), cefuroxime (72.41%), amoxicillinclavulanic acid (71.90%), ceftriaxone (66.58%), ciprofloxacin (65.82%) and cefepime (57.47%). Further, in-vitro biofilm assay confirmed formation of biofilms by 71.39% isolates. Biofilm-forming E. coli strains developed higher degree of resistance towards antibiotics ampicillin (87.36%) followed by cefuroxime (81.58%), amoxicillin-clavulanic acid (77.61%), ciprofloxacin (71.48%), cefepime (64.98%) and ceftriaxone (54.6%). Phenotypic methods detected 62.3% isolates as ESBL producers and 40.2% were β-haemolytic. Virulence gene characterization revealed presence of gene traT in 73.2% isolates, PAI in 62.9% isolates and papEF among 33.5% isolates. ESBL & Hemolysin producing UPEC exhibited significantly higher resistant to ampicillin, amoxicillin-clavulanic acid, aztreonam, cefriaxone, cefuroxime, cefepime, ciprofloxacin, chloramphenicol, gentamicin and norfloxacin. Association between the expression of virulence genes and antimicrobial resistance was observed. Statistically proved association was seen with expression of genes RPAI and traT with antibioticsnitrofurantoin and amikacin respectively. All the isolates were sensitive to antibiotics nitrofurantoin, piperacillin-tazobactam, and imipenem. Strains showed 99% sequence identity in Sanger sequencing and no mutations were detected among the study strains.

Conclusion: Increase in trend of resistance was observed with antibiotics which were routinely used to treat UTI. Predominance of the traT gene and PAI markers was seen among UPEC strains. Results indicated significant correlation between phenotypic and genotypic virulence factors and antibiotic resistance. Antibiotics nitrofurantoin, piperacillin - tazobactam, and imipenem can be effective for severe UTIs. This study concludes that expression of virulence factors by UPEC strains is responsible for increased antibiotic resistance. Hence, characterizing the UPEC strains help clinicians and microbiologists to reach a better therapeutic outcomes and treatment regimens in this region.