Subspecies Typing of Streptococcus agalactiae Based on Ribosomal Subunit Protein Mass Variation by MALDI-TOF MS

Abstract

<b>Background:</b> A ribosomal subunit protein (rsp)-based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) method was developed for fast subspecies-level typing of <i>Streptococcus agalactiae</i> (Group B <i>Streptococcus</i>, GBS), a major cause of neonatal sepsis and meningitis. <b>Methods:</b> A total of 796 GBS whole genome sequences, covering the genetic diversity of the global GBS population, were used to <i>in silico</i> predict molecular mass variability of 28 rsp and to identify unique rsp mass combinations, termed "rsp-profiles". The <i>in silico</i> established GBS typing scheme was validated by MALDI-TOF MS analysis of GBS isolates at two independent research sites in Europe and South East Asia. <b>Results:</b> We identified <i>in silico</i> 62 rsp-profiles, with the majority (>80%) of the 796 GBS isolates displaying one of the six rsp-profiles 1-6. These dominant rsp-profiles classify GBS strains in high concordance with the core-genome based phylogenetic clustering. Validation of our approach by in-house MALDI-TOF MS analysis of 248 GBS isolates and external analysis of 8 GBS isolates showed that across different laboratories and MALDI-TOF MS platforms, the 28 rsp were detected reliably in the mass spectra, allowing assignment of clinical isolates to rsp-profiles at high sensitivity (99%) and specificity (97%). Our approach distinguishes the major phylogenetic GBS genotypes, identifies hyper-virulent strains, predicts the probable capsular serotype and surface protein variants and distinguishes between GBS genotypes of human and animal origin. <b>Conclusion:</b> We combine the information depth of whole genome sequences with the highly cost efficient, rapid and robust MALDI-TOF MS approach facilitating high-throughput, inter-laboratory, large-scale GBS epidemiological and clinical studies based on pre-defined rsp-profiles.