title: Multilocus sequence typing of Streptococcus pneumoniae by use of mass spectrometry.
abstract: Multilocus sequence typing (MLST) is an important tool for the global surveillance of bacterial pathogens that is performed by comparing the sequences of designated housekeeping genes. We developed and tested a novel mass spectrometry-based method for MLST of Streptococcus pneumoniae. PCR amplicons were subjected to in vitro transcription and base-specific cleavage, followed by analysis of the resultant fragments by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Comparison of the cleavage fragment peak patterns to a reference sequence set permitted automated identification of alleles. Validation experiments using 29 isolates of S. pneumoniae revealed that the results of MALDI-TOF MS MLST matched those obtained by traditional sequence-based MLST for 99% of alleles and that the MALDI-TOF MS method accurately identified two single-nucleotide variations. The MADLI-TOF MS method was then used for MLST analysis of 43 S. pneumoniae isolates from Papua New Guinean children. The majority of the isolates present in this population were not clonal and contained seven new alleles and 30 previously unreported sequence types.
authors: Centre for International Child Health, Murdoch Childrens Research Institute, Parkville, Victoria 3052, Australia. eileen.dunne@mcri.edu.au
authors: Eileen M
authors: EM
authors: Dunne
authors: Eng Kok
authors: EK
authors: Ong
authors: Ralf J
authors: RJ
authors: Moser
authors: Peter M
authors: PM
authors: Siba
authors: Suparat
authors: S
authors: Phuanukoonnon
authors: Andrew R
authors: AR
authors: Greenhill
authors: Roy M
authors: RM
authors: Robins-Browne
authors: E Kim
authors: EK
authors: Mulholland
authors: Catherine
authors: C
authors: Satzke
mesh_terms: D000483:Alleles; D001331:Automation / Q000379:methods; D002675:Child, Preschool; D006801:Humans; D007223:Infant; D058885:Multilocus Sequence Typing / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013296:Streptococcus pneumoniae / Q000737:chemistry / Q000145:classification* / Q000235:genetics