Antibiofilm Activity of Chitosan Nanoparticles Against Uropathogenic Escherichia Coli

Official title

Evaluation of the Antibiofilm Activity of Chitosan Nanoparticles Against Uropathogenic Escherichia Coli Isolated From Assiut University Hospitals

Overview

1. Isolation of uropathogenic Escherichia coli (UPEC) and determination of their Antimicrobial sensitivity Patterns. 2. Evaluation of biofilm-forming capacity of UPEC isolates. 3. Assessment of the antibiofilm efficacy of chitosan nanoparticles alone and in combination with ciprofloxacin against UPEC isolates. 4. Examination of the effectiveness of chitosan nanoparticle coating in preventing biofilm formation by UPEC on urinary catheter surfaces. 5. Evaluation of the impact of chitosan nanoparticles on the expression levels of biofilm associated genes in UPEC.

Detailed description

Urinary tract infections (UTIs) remain among the most prevalent bacterial infections globally, causing substantial healthcare burden, with uropathogenic Escherichia coli (UPEC) responsible for the majority of cases in both community and hospital settings. Catheter-associated urinary tract infections (CAUTIs) represent a major proportion of healthcare-associated infections due to bacterial adhesion and biofilm formation on indwelling urinary catheters. Biofilm formation enables microorganisms to attach to abiotic surfaces and produce an extracellular polymeric matrix that enhances bacterial survival under adverse environmental conditions.

Biofilm-associated bacteria exhibit increased resistance to host immune responses and antimicrobial agents, contributing to chronic and recurrent infections. Cells embedded within biofilms may demonstrate markedly elevated antibiotic tolerance compared with planktonic bacteria, limiting therapeutic success. Although systemic antibiotics remain the mainstay of treatment, rising antimicrobial resistance among biofilm forming UPEC strains highlights the need for alternative antibiofilm strategies.

Chitosan, a naturally derived biopolymer, has attracted attention due to its biocompatibility, biodegradability, and intrinsic antimicrobial properties. Chitosan disrupts bacterial membranes and inhibits biofilm matrix formation and surface adhesion. Emerging evidence indicates that chitosan can downregulate biofilm-related gene expression involved in adhesion and extracellular polysaccharide synthesis. Furthermore, nanoparticle formulation enhances chitosan penetration into biofilms and improves antimicrobial efficiency compared with bulk polymer forms. Moreover, chitosan nanoparticles may enhance antibiotic diffusion and demonstrate synergistic antibiofilm activity when combined with ciprofloxacin.

Ciprofloxacin, a fluoroquinolone antibiotic widely used in UTIs, has demonstrated partial inhibition of biofilm formation through interference with bacterial DNA replication, initial bacterial adhesion to surfaces, reduction of expression of biofilm-related genes, quorum-sensing activity and decreases production of extracellular polymeric substances (EPS); however, its efficacy is reduced against mature biofilms. Consequently, chitosan nanoparticles may demonstrate enhanced antibiofilm activity when combined with ciprofloxacin.

Despite promising findings, limited studies have evaluated chitosan nanoparticle coatings on clinically relevant catheter surfaces against UPEC isolates. Therefore, investigating this strategy may offer an effective preventive approach for CAUTIs and support improved infection control practices.

Interventions

• Other chitosan nanoparticles
  Chitosan nanoparticles will be applied to strong biofilm-forming UPEC isolates in vitro to evaluate their antibiofilm activity. Treatments include: * Chitosan nanoparticles alone at sub-inhibitory concentrations * Ciprofloxacin alone at sub-inhibitory concentrations * Combination of chitosan nanoparticles and ciprofloxacin Biofilm formation will be quantified using crystal violet staining in 96-well microtiter plates. The intervention also includes coating of urinary catheter segments with chit

Primary outcome measures

• Percentage of biofilm inhibition of UPEC isolates by chitosan nanoparticles alone and in combination with ciprofloxacin [Time frame: 24 hours after treatment of bacterial cultures in vitro.]

Secondary outcome measures (3)

• Minimum Inhibitory Concentration (MIC) of chitosan nanoparticles and ciprofloxacin against UPEC isolates [Time frame: 18-24 hours after inoculation]
• Effectiveness of chitosan nanoparticle coating in preventing UPEC biofilm formation on urinary catheter segments [Time frame: 18-24 hours after bacterial incubation on coated catheter segments.]
• Relative gene expression of biofilm-associated genes (fimH and luxS) in UPEC isolates after chitosan nanoparticle treatment [Time frame: Immediately after 24-hour treatment of bacterial cultures in vitro.]

Eligibility criteria

Inclusion criteria

• 1\. Patients (inpatients or outpatients) clinically diagnosed with urinary tract infection (UTI).

2\. Presence of significant bacteriuria in urine culture (≥10⁵ colony-forming units per milliliter (CFU/mL) for midstream urine, or as clinically indicated).

3\. Isolation of Escherichia coli as the sole or predominant pathogen from urine culture.

4\. Patients of any age and both sexes. 5. Patients who have not received antibiotic therapy within the previous 48-72 hours.

Exclusion criteria

• 1\. Patients who received systemic antibiotic therapy within the previous 48-72 hours prior to urine sample collection.

2\. Urine cultures showing insignificant bacteriuria (<10⁵ CFU/mL for midstream urine, unless clinically justified).

3\. Polymicrobial growth in urine culture (mixed bacterial growth suggestive of contamination).

4\. Isolation of organisms other than Escherichia coli. 5. Improperly collected, leaking, or contaminated urine samples. 6. Patients unwilling to provide informed consent.

Criteria are shown verbatim from the registry (in English). Final eligibility is always assessed by the study center.

Healthy volunteers: No

Study design

Observational model

Case-only

Study locations

Egypt · 1 center

• Faculty of Medicine, Assiut University, Microbiology and Immunity department — Asyut

Publications

• Oliveira MCF, Canellas ALB, Berbert LC, Cardoso AM, Silva VA, Garutti SST, Rangel DHF, Dias RCS, Perini JA, Souza CRVM, Chagas TPG, Laport MS, Pellegrino FLPC. Assessment of Antimicrobial Resistance and Virulence of Biofilm-Forming Uropathogenic Escherichia coli from Rio de Janeiro. Antibiotics (Basel). 2025 Aug 29;14(9):869. doi: 10.3390/antibiotics14090869. PMID 41009848
• Wiegand I, Hilpert K, Hancock RE. Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nat Protoc. 2008;3(2):163-75. doi: 10.1038/nprot.2007.521. PMID 18274517
• Obaid NA, Alzahrani AM, Alaryni BA, Almegrin FF, Alsubhi RS, Alzahrani RS, et al. Effectiveness of chitosan coating catheter in preventing catheter-associated urinary tract infection (CAUTI). J Pharm Res Int. 2022;34(19A):6-19.
• Fattah RAFA, Fathy FEZY, Mohamed TAH, Elsayed MS. Effect of chitosan nanoparticles on quorum sensing-controlled virulence factors and expression of LasI and RhlI genes among Pseudomonas aeruginosa clinical isolates. AIMS Microbiol. 2021 Oct 26;7(4):415-430. doi: 10.3934/microbiol.2021025. eCollection 2021. PMID 35071940
• Yao H, Liu J, Jiang X, Chen F, Lu X, Zhang J. Analysis of the Clinical Effect of Combined Drug Susceptibility to Guide Medication for Carbapenem-Resistant Klebsiella pneumoniae Patients Based on the Kirby-Bauer Disk Diffusion Method. Infect Drug Resist. 2021 Jan 12;14:79-87. doi: 10.2147/IDR.S282386. eCollection 2021. PMID 33469322
• Hooton TM, Gupta K. Urinary tract infections and asymptomatic bacteriuria in adults. N Engl J Med. 2021;384(11):1028-37.
• Singh S, et al. Nanotechnology-based coatings for prevention of catheter-associated infections. Nanomedicine. 2024; 52:102640.
• Yan J, Bassler BL. Surviving as a Community: Antibiotic Tolerance and Persistence in Bacterial Biofilms. Cell Host Microbe. 2019 Jul 10;26(1):15-21. doi: 10.1016/j.chom.2019.06.002. PMID 31295420

Identifiers

NCT: NCT07482553 · not assigned yet

Primary sources (government registries)

• ClinicalTrials.gov (US NIH)

View this study on ClinicalTrials.gov ↗