title: Utilization of Whole-Cell MALDI-TOF Mass Spectrometry to Differentiate Burkholderia pseudomallei Wild-Type and Constructed Mutants.
abstract: Whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (whole-cell MALDI-TOF MS) has been widely adopted as a useful technology in the identification and typing of microorganisms. This study employed the whole-cell MALDI-TOF MS to identify and differentiate wild-type and mutants containing constructed single gene mutations of Burkholderia pseudomallei, a pathogenic bacterium causing melioidosis disease in both humans and animals. Candidate biomarkers for the B. pseudomallei mutants, including rpoS, ppk, and bpsI isolates, were determined. Taxon-specific and clinical isolate-specific biomarkers of B. pseudomallei were consistently found and conserved across all average mass spectra. Cluster analysis of MALDI spectra of all isolates exhibited separate distribution. A total of twelve potential mass peaks discriminating between wild-type and mutant isolates were identified using ClinProTools analysis. Two peaks (m/z 2721 and 2748 Da) were specific for the rpoS isolate, three (m/z 3150, 3378, and 7994 Da) for ppk, and seven (m/z 3420, 3520, 3587, 3688, 4623, 4708, and 5450 Da) for bpsI. Our findings demonstrated that the rapid, accurate, and reproducible mass profiling technology could have new implications in laboratory-based rapid differentiation of extensive libraries of genetically altered bacteria.
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Suthamat
authors.initials: S
authors.lastname: Niyompanich
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Kitima
authors.initials: K
authors.lastname: Srisanga
authors.affiliation: National Center for Genetic Engineering and Biotechnology (BIOTEC), Thailand Science Park, Pathum Thani, Thailand.
authors.forename: Janthima
authors.initials: J
authors.lastname: Jaresitthikunchai
authors.affiliation: National Center for Genetic Engineering and Biotechnology (BIOTEC), Thailand Science Park, Pathum Thani, Thailand.
authors.forename: Sittiruk
authors.initials: S
authors.lastname: Roytrakul
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Sumalee
authors.initials: S
authors.lastname: Tungpradabkul
mesh_terms: D000942:Antigens, Bacterial / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D016957:Burkholderia pseudomallei / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D005819:Genetic Markers / Q000235:genetics*; D005838:Genotype; D006801:Humans; D008554:Melioidosis / Q000382:microbiology; D009154:Mutation / Q000235:genetics*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D012808:Sigma Factor / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C444341:Bps-1 recombinant protein antigen; D005819:Genetic Markers; D011994:Recombinant Proteins; D012808:Sigma Factor; C069567:sigma factor KatF protein, Bacteria; D017853:Phosphotransferases (Alcohol Group Acceptor)