Evolution of hypervirulence during bloodstream adaptation of CA-MRSA strain USA300

The epidemic community-acquired methicillin-resistant S. aureus (CA-MRSA) clone USA300 has recently become a leading cause of hospital-associated bloodstream infections (BSI). Leveraging this recent introduction into hospitals and the limited genetic variation across the USA300 strains, we combined microbial comparative genomics with phenotypic analyses to discover adaptive mutations. USA300 isolates from BSI were found to have independently evolved single nucleotide variants in the transcriptional regulator sarZ. sarZ inactivation lead to altered expression of virulence factors, resulting in increased lethality in a murine model of BSI. Thus, USA300 strains can optimize their fitness in hospitals through evolution of higher virulence. To establish the SarZ regulon in the USA300 background we performed RNA-Seq experiments in sarZ wild-type, mutant and complement strains during logarithmic or stationary growth. The laboratory-constructed strains AH-LAC + vector, AH-LAC sarZ::bursa + vector, and AH-LAC sarZ::bursa + sarZWT were used. In addition we selected two clinical USA300 bloodstream infection (BSI) isolates with a wild-type (ER00573) and mutant sarZ locus (ER00594), and constructed sarZ deletion (sarZ::tet) and complement strains. Each strain was profiled, in triplicate, after culturing for 3h (logarithmic growth) and 5h (stationary phase) in 5mL tryptic soy broth (TSB).