Can the fliC PCR-Restriction Fragment Length Polymorphism Technique Replace Classic Serotyping Methods for Characterizing the H Antigen of Enterotoxigenic Escherichia coli Strains?

Abstract

In this study, we performed fliC PCR-restriction fragment length polymorphism (RFLP) to investigate whether this technique would be better than classic serotyping for the characterization of the H antigen in enterotoxigenic Escherichia coli (ETEC) strains. We showed that the fliC genes from ETEC strains can be characterized by restriction analysis of their polymorphism. Only one allele of the fliC gene from ETEC strains was found for each flagellar antigen, with the exception of H21. Nonmotile strains could also be characterized using this molecular technique. Moreover, determination of the somatic antigen was guided by the identification of the flagellar antigen from previously unknown serotypes of ETEC strains by PCR-RFLP, thus reducing the number of anti-antigen O sera used. The PCR-RFLP technique proved to be faster than classic serotyping for the characterization of the E. coli H antigen, taking 2 days to complete instead of the 7 or more days using classic serotyping. In conclusion, the H molecular typing for Enterobacteriaceae members may become an important epidemiological tool for the characterization of the H antigen of E. coli pathotypes. The PCR-RFLP technique is capable of guiding the determination of the H antigen and could partially replace seroagglutination. With the determination of the molecular profiles of alleles from strains obtained in epidemiological studies, new patterns will be described for ETEC strains or other E. coli pathotypes, thus permitting widespread use of this technique to characterize fliC genes and determine the H antigen of E. coli strains.

Phenotypic H-Antigen Typing by Mass Spectrometry Combined with Genetic Typing of H Antigens, O Antigens, and Toxins by Whole-Genome Sequencing Enhances Identification of Escherichia coli Isolates

“…Similar to reference strain E32511, H types were still obtained by WGS for the remaining six NM isolates that neither serotyping nor MS-H could identify. This further confirmed that H antigens characterized by WGS do not necessarily represent phenotypic H types, as genes responsible for flagellum production are not always expressed ( 27 ).…”

Molecular Characterization Reveals Three Distinct Clonal Groups among Clinical Shiga Toxin-Producing Escherichia coli Strains of Serogroup O103

“…Although the H type can be a useful phenotypic marker for classifying strains, we could not determine the H type of some O103 isolates, because of unclear agglutination or lack of bacterial motility. As many researchers have shown before (8,18,29), sequence variation in the fliC gene could be a proxy for these agglutination tests. In the present study, on the basis of sequence variation in fliC genes, we developed a multiplex PCR method for such classification of STEC O103 strains.…”

New Approaches for Escherichia coli Genotyping

“…Herein we propose two new, original tools that can be useful for genotyping of E. coli isolates. The first of the developed method, a PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) test uses a highly variable fliC gene [8][9][10], encoding the H antigen as a molecular target. The second approach is based on the PCR amplification of seven selected genes that characterize with high sequence and size variability within the sequenced genomes of E. coli [11].…”