Whole-genome sequencing for high-resolution investigation of methicillin-resistant Staphylococcus aureus epidemiology and genome plasticity.
Methicillin-resistant Staphylococcus aureus (MRSA) infections pose a major challenge in health care, yet the limited heterogeneity within this group hinders molecular investigations of related outbreaks. Pulsed-field gel electrophoresis (PFGE) has been the gold standard approach but is impractical for many clinical laboratories and is often replaced with PCR-based methods. Regardless, both approaches can prove problematic for identifying subclonal outbreaks. Here, we explore the use of whole-genome sequencing for clinical laboratory investigations of MRSA molecular epidemiology. We examine the relationships of 44 MRSA isolates collected over a period of 3 years by using whole-genome sequencing and two PCR-based methods, multilocus variable-number tandem-repeat analysis (MLVA) and spa typing. We find that MLVA offers higher resolution than spa typing, as it resolved 17 versus 12 discrete isolate groups, respectively. In contrast, whole-genome sequencing reproducibly cataloged genomic variants (131,424 different single nucleotide polymorphisms and indels across the strain collection) that uniquely identified each MRSA clone, recapitulating those groups but enabling higher-resolution phylogenetic inferences of the epidemiological relationships. Importantly, whole-genome sequencing detected a significant number of variants, thereby distinguishing between groups that were considered identical by both spa typing (minimum, 1,124 polymorphisms) and MLVA (minimum, 193 polymorphisms); this suggests that these more conventional approaches can lead to false-positive identification of outbreaks due to inappropriate grouping of genetically distinct strains. An analysis of the distribution of variants across the MRSA genome reveals 47 mutational hot spots (comprising ∼ 2.5% of the genome) that account for 23.5% of the observed polymorphisms, and the use of this selected data set successfully recapitulates most epidemiological relationships in this pathogen group.
Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
Dhruba J
DJ
SenGupta
Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
Lisa A
LA
Cummings
Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
Daniel R
DR
Hoogestraat
Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.
Susan M
SM
Butler-Wu
Department of Genome Sciences, University of Washington, Seattle, Washington, USA.
Jay
J
Shendure
Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA Department of Microbiology, University of Washington, Seattle, Washington, USA.
Brad T
BT
Cookson
Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA stevesal@uw.edu.
Stephen J
SJ
Salipante
D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics*; D014644:Genetic Variation; D016680:Genome, Bacterial; D006801:Humans; D055624:Methicillin-Resistant Staphylococcus aureus / Q000235:genetics* / Q000302:isolation & purification; D017720:Molecular Epidemiology / Q000379:methods*; D008969:Molecular Sequence Data; D058889:Molecular Typing / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods*; D013203:Staphylococcal Infections / Q000382:microbiology*
D004269:DNA, Bacterial