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With mutations constantly accumulating in bacterial genomes, it is unclear whether the previously identified bacterial strains are really present in an extant sample. To address this question, we did a case study on the known strains of the bacterial speciesS.aureusandS.epidermisin 68 atopic dermatitis shotgun metagenomic samples. We evaluated the likelihood of the presence of all sixteen known strains predicted in the original study and by two popular tools in this study. We found that even with the same tool, only two known strains were predicted by the original study and this study. Moreover, none of the sixteen known strains was likely present in these 68 samples. Our study thus indicates the limitation of the known-strain-based studies, especially those on rapidly evolving bacterial species. It implies the unlikely presence of the previously identified known strains in a current environmental sample. It also called for de novo bacterial strain identification directly from shotgun metagenomic reads.