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MS-H: a novel proteomic approach to isolate and type the E. coli H antigen using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS).


ABSTRACT: Serotyping is the long-standing gold standard method to determine E. coli H antigens; however, this method requires a panel of H-antigen specific antibodies and often culture-based induction of the H-antigen flagellar motility. In this study, a rapid and accurate method to isolate and identify the Escherichia coli (E. coli) H flagellar antigen was developed using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Flagella were isolated from pure culture, digested with trypsin, and then subjected to LC-MS/MS using one of two systems (Agilent-nano-LC-QSTAR XL or Proxeon-nano-LC-LTQ-Orbitrap XL). The resulting peptide sequence data were searched against a custom E. coli flagella/H antigen database. This approach was evaluated using flagella isolated from reference E. coli strains representing all 53 known H antigen types and 41 clinical E. coli strains. The resulting LC-MS/MS classifications of H antigen types (MS-H) were concordant with the known H serogroup for all 53 reference types, and of 41 clinical isolates tested, 38 (92.7%) were concordant with the known H serogroup. MS-H clearly also identified two clinical isolates (4.9%) that were untypeable by serotyping. Notably, successful detection and classification of flagellar antigens with MS-H did not generally require induction of motility, establishing this proteomic approach as more rapid and cost-effective than traditional methods, while providing equitable specificity for typing E. coli H antigens.

SUBMITTER: Cheng K 

PROVIDER: S-EPMC3578835 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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MS-H: a novel proteomic approach to isolate and type the E. coli H antigen using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Cheng Keding K   Drebot Mike M   McCrea Joanne J   Peterson Lorea L   Lee David D   McCorrister Stuart S   Nickel Richard R   Gerbasi Alyssia A   Sloan Angela A   Janella Debra D   Van Domselaar Gary G   Beniac Daniel D   Booth Tim T   Chui Linda L   Tabor Helen H   Westmacott Garrett G   Gilmour Matthew M   Wang Gehua G  

PloS one 20130221 2


Serotyping is the long-standing gold standard method to determine E. coli H antigens; however, this method requires a panel of H-antigen specific antibodies and often culture-based induction of the H-antigen flagellar motility. In this study, a rapid and accurate method to isolate and identify the Escherichia coli (E. coli) H flagellar antigen was developed using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Flagella were isolated from pure culture, digested  ...[more]

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