Comparison of PCR-based methods for typing Escherichia coli

Area of Science:

• Microbiology
• Genetics
• Infectious Diseases

Background:

• Accurate typing systems are crucial for tracking Escherichia coli (E. coli) transmission.
• Existing methods like PFGE, RAPD, and rep-PCR have limitations in discriminating E. coli strains.

Purpose of the Study:

• To establish a robust library typing system for studying E. coli cross-transmission.
• To evaluate and compare the discriminatory power of various genotyping techniques for E. coli.

Main Methods:

• Genotyping of E. coli isolates using pulsed-field gel electrophoresis (PFGE), random amplified polymorphic DNA (RAPD), repetitive (rep) PCR, and fluorescent amplified fragment length polymorphism (AFLP).
• Analysis of fingerprints using Pearson correlation and Dice coefficients, with a focus on the 'uncertain band' feature in GelCompar II software for AFLP.
• Application of the two most discriminative PCR typing methods to 112 isolates from hospitalized patients over nine months.

Main Results:

• RAPD and rep-PCR showed insufficient discriminatory power for E. coli typing.
• AFLP distinguished 18 types among 18 unrelated strains, outperforming PFGE (17 types) and IS3 rep-PCR (10 types).
• Dice coefficients, ignoring uncertain bands, demonstrated the best concordance with visual interpretation for AFLP fingerprint analysis.

Conclusions:

• Fluorescent amplified fragment length polymorphism (AFLP) exhibits the highest discriminatory capacity for E. coli isolate typing.
• Dice coefficient analysis is the most efficient method for automated retrieval of indistinguishable genotypes in AFLP fingerprint databases.
• This typing system aids in studying the cross-transmission of E. coli in healthcare settings.