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1.  Development of Pooled Suppression Subtractive Hybridization to analyze the Pangenome of Staphylococcus aureus 
We describe the development and application of a pooled Suppression Subtractive Hybridization (PSSH) method to describe differences between the genomic content of a pool of clinical Staphylococcus aureus isolates and a sequenced reference strain. In comparative bacterial genomics, Suppression Subtractive Hybridization (SSH) is normally utilized to compare genomic features or expression profiles of one strain versus another, which limits its ability to analyze communities of isolates. However, a PSSH approach theoretically enables the user to characterize the entirety of gene content unique to a related group of isolates in a single reaction. These unique fragments may then be linked to individual isolates through standard PCR. This method was applied to examine the genomic diversity found in pools of Staphylococcus aureus isolates associated with complicated bacteremia infections leading to endocarditis and osteomyelitis. Across four pools of 10 isolates each, four hundred and twenty nine fragments not found in or significantly divergent from the S. aureus NCTC 8325 reference genome were detected. These fragments could be linked to individual strains within its pool by PCR. This is the first use of PSSH to examine the S. aureus pangenome. We propose that PSSH is a powerful tool for researchers interested in rapidly comparing the genomic content of multiple unstudied isolates.
doi:10.1016/j.mimet.2010.01.022
PMCID: PMC2843094  PMID: 20138093
Staphylococcus aureus; pangenome; subtractive hybridization
2.  Potential Associations between Severity of Infection and the Presence of Virulence-Associated Genes in Clinical Strains of Staphylococcus aureus 
PLoS ONE  2011;6(4):e18673.
Background
The clinical spectrum of Staphylococcus aureus infection ranges from asymptomatic nasal carriage to osteomyelitis, infective endocarditis (IE) and death. In this study, we evaluate potential association between the presence of specific genes in a collection of prospectively characterized S. aureus clinical isolates and clinical outcome.
Methodology/Principal Findings
Two hundred thirty-nine S. aureus isolates (121 methicillin-resistant S. aureus [MRSA] and 118 methicillin-susceptible S. aureus [MSSA]) were screened by array comparative genomic hybridization (aCGH) to identify genes implicated in complicated infections. After adjustment for multiple tests, 226 genes were significantly associated with severity of infection. Of these 226 genes, 185 were not in the SCCmec element. Within the 185 non-SCCmec genes, 171 were less common and 14 more common in the complicated infection group. Among the 41 genes in the SCCmec element, 37 were more common and 4 were less common in the complicated group. A total of 51 of the 2014 sequences evaluated, 14 non-SCCmec and 37 SCCmec, were identified as genes of interest.
Conclusions/Significance
Of the 171 genes less common in complicated infections, 152 are of unknown function and may contribute to attenuation of virulence. The 14 non-SCCmec genes more common in complicated infections include bacteriophage-encoded genes such as regulatory factors and autolysins with potential roles in tissue adhesion or biofilm formation.
doi:10.1371/journal.pone.0018673
PMCID: PMC3082525  PMID: 21541311

Results 1-2 (2)