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ABSTRACTWhole-genome sequencing is becoming a leading technology in the typing and epidemiology of microbial pathogens, but the increase in genomic information necessitates significant investment in bioinformatic resources and expertise, and currently used methodologies struggle with genetically heterogeneous bacteria such as the human gastric pathogen Helicobacter pylori. Here we demonstrate that the alignment-free analysis method feature frequency profiling (FFP) can be used to rapidly construct phylogenetic trees of draft bacterial genome sequences on a standard desktop computer and that coupling with in silicogenotyping methods gives useful information for comparative and clinical genomic and molecular epidemiology applications. FFP-based phylogenetic trees of seven gastric Helicobacterspecies matched those obtained by analysis of 16S rRNA genes and ribosomal proteins, and FFP- and core genome single nucleotide polymorphism-based analysis of 63 H. pylorigenomes again showed comparable phylogenetic clustering, consistent with genomotypes assigned by using multilocus sequence typing (MLST). Analysis of 377 H. pylorigenomes highlighted the conservation of genomotypes and linkage with phylogeographic characteristics and predicted the presence of an incomplete or nonfunctional cagpathogenicity island in 18/276 genomes. In silicoanalysis of antibiotic susceptibility markers suggests that most H. pylorihspAmerind and hspEAsia isolates are predicted to carry the T2812C mutation potentially conferring low-level clarithromycin resistance, while levels of metronidazole resistance were similar in all multilocus sequence types. In conclusion, the use of FFP phylogenetic clustering and in silicogenotyping allows determination of genome evolution and phylogeographic clustering and can contribute to clinical microbiology by genomotyping for outbreak management and the prediction of pathogenic potential and antibiotic susceptibility.

ABSTRACTIdentification of Neisseria gonorrhoeaeby the Bruker matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system may be affected by “B consistency categorization.” A supplementary database of 17 N. gonorrhoeaemain spectra was constructed. Twelve of 64 N. gonorrhoeaeidentifications were categorized with B consistency, which disappeared using the supplementary database. Database extension did not result in misidentification of Neisseria meningitidis.