Liquid chromatography/mass spectrometry characterization of Escherichia coli and Shigella species.
Liquid chromatography/quadrupole time of flight mass spectrometry (LC/QTOF MS) utilizing electrospray ionization was employed to monitor protein expression in Escherichia coli and Shigella organisms. Comparison with MALDI/TOF-MS revealed more proteins, particularly above 15 kDa. A combination of automated charge state deconvolution, spectral mirroring, and spectral subtraction was used to reveal subtle differences in the LC/MS data. Reproducible intact protein biomarker candidates were discovered based on their unique mass, retention time, and relative intensity. These marker candidates were implemented to differentiate closely related strain types, (e.g., two distinct isolates of E. coli O157:H7) and to correctly identify unknown pathogens. This LC/MS approach is less labor-intensive than pulsed-field gel electrophoresis, affords greater specificity than real-time PCR, and requires no primers or antibodies. Additionally, this approach would be beneficial during outbreaks of foodborne disease or bioterrorism investigations by complementing methods typically used in diagnostic microbiology laboratories.
Everley
Robert A
RA
Division of Consolidated Laboratory Services, Commonwealth of Virginia, and Department of Chemistry, Virginia Commonwealth University, Richmond, Virginia 23219, USA.
Mott
Tiffany M
TM
Wyatt
Shane A
SA
Toney
Denise M
DM
Croley
Timothy R
TR
D000818:Animals; D001426:Bacterial Proteins / Q000032:analysis; D015373:Bacterial Typing Techniques; D015415:Biomarkers / Q000378:metabolism; D002853:Chromatography, Liquid / Q000379:methods*; D004926:Escherichia coli / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification; D008970:Molecular Weight; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012760:Shigella / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013997:Time Factors
D001426:Bacterial Proteins; D015415:Biomarkers