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

Abstract

Mass spectrometry-based phenotypic H-antigen typing (MS-H) combined with whole-genome-sequencing-based genetic identification of H antigens, O antigens, and toxins (WGS-HOT) was used to type 60 clinical Escherichia coli isolates, 43 of which were previously identified as nonmotile, H type undetermined, or O rough by serotyping or having shown discordant MS-H and serotyping results. Whole-genome sequencing confirmed that MS-H was able to provide more accurate data regarding H antigen expression than serotyping. Further, enhanced and more confident O antigen identification resulted from gene cluster based typing in combination with conventional typing based on the gene pair comprising wzx and wzy and that comprising wzm and wzt. The O antigen was identified in 94.6% of the isolates when the two genetic O typing approaches (gene pair and gene cluster) were used in conjunction, in comparison to 78.6% when the gene pair database was used alone. In addition, 98.2% of the isolates showed the existence of genes for various toxins and/or virulence factors, among which verotoxins (Shiga toxin 1 and/or Shiga toxin 2) were 100% concordant with conventional PCR based testing results. With more applications of mass spectrometry and whole-genome sequencing in clinical microbiology laboratories, this combined phenotypic and genetic typing platform (MS-H plus WGS-HOT) should be ideal for pathogenic E. coli typing.

A null model for microbial diversification

“…Regardless of the underlying evolutionary forces, D + gene families promise to be useful phylogenetic markers for delineating microdiversity within a group of closely related bacteria. Some of the identified D + gene families have already been either shown to be efficacious markers for typing of pathogenic isolates [e.g., porB (74) and lip (75) for Neisseria, and the above-discussed fliC (76, 77) for E. coli] or identified to have a potential to be such markers [e.g., E. coli housekeeping genes ugd and galF that often flank frequently horizontally exchanged genes involved in the O antigen biosynthesis (78,79)]. Therefore, the presented framework could help identify loci promising for describing bacterial genotypic variation that is driven by ecological forces.…”