title: Detection of biomarkers of pathogenic Naegleria fowleri through mass spectrometry and proteomics.
abstract: Emerging methods based on mass spectrometry (MS) can be used in the rapid identification of microorganisms. Thus far, these practical and rapidly evolving methods have mainly been applied to characterize prokaryotes. We applied matrix-assisted laser-desorption-ionization-time-of-flight mass spectrometry MALDI-TOF MS in the analysis of whole cells of 18 N. fowleri isolates belonging to three genotypes. Fourteen originated from the cerebrospinal fluid or brain tissue of primary amoebic meningoencephalitis patients and four originated from water samples of hot springs, rivers, lakes or municipal water supplies. Whole Naegleria trophozoites grown in axenic cultures were washed and mixed with MALDI matrix. Mass spectra were acquired with a 4700 TOF-TOF instrument. MALDI-TOF MS yielded consistent patterns for all isolates examined. Using a combination of novel data processing methods for visual peak comparison, statistical analysis and proteomics database searching we were able to detect several biomarkers that can differentiate all species and isolates studied, along with common biomarkers for all N. fowleri isolates. Naegleria fowleri could be easily separated from other species within the genus Naegleria. A number of peaks detected were tentatively identified. MALDI-TOF MS fingerprinting is a rapid, reproducible, high-throughput alternative method for identifying Naegleria isolates. This method has potential for studying eukaryotic agents.
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, M.S. F-50, 4770 Buford Hwy NE, Atlanta, Georgia, 30341, USA.
authors.forename: Hercules
authors.initials: H
authors.lastname: Moura
authors.forename: Fernando
authors.initials: F
authors.lastname: Izquierdo
authors.forename: Adrian R
authors.initials: AR
authors.lastname: Woolfitt
authors.forename: Glauber
authors.initials: G
authors.lastname: Wagner
authors.forename: Tatiana
authors.initials: T
authors.lastname: Pinto
authors.forename: Carmen
authors.initials: C
authors.lastname: del Aguila
authors.forename: John R
authors.initials: JR
authors.lastname: Barr
mesh_terms: D000293:Adolescent; D000328:Adult; D000562:Amebiasis / Q000134:cerebrospinal fluid* / Q000469:parasitology; D000818:Animals; D061245:Axenic Culture; D015415:Biomarkers / Q000134:cerebrospinal fluid; D001921:Brain / Q000469:parasitology; D002417:Cattle; D020808:Central Nervous System Protozoal Infections / Q000134:cerebrospinal fluid* / Q000469:parasitology; D002648:Child; D005260:Female; D005618:Fresh Water / Q000469:parasitology; D005838:Genotype; D006801:Humans; D008297:Male; D016848:Naegleria fowleri / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification; D010449:Peptide Mapping / Q000379:methods*; D010802:Phylogeny; D040901:Proteomics / Q000379:methods; D015800:Protozoan Proteins / Q000737:chemistry / Q000145:classification / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D053058:Trophozoites / Q000737:chemistry*
chemical_list: D015415:Biomarkers; D015800:Protozoan Proteins
keywords: Free-living amoebae; Naegleria; matrix-assisted laser-desorption-ionization-time-of-flight mass spectrometry; protein fingerprints