Extensive Genomic Diversity among Bovine-Adapted Staphylococcus aureus: Evidence for a Genomic Rearrangement within CC97

Abstract

Staphylococcus aureus is an important pathogen associated with both human and veterinary disease and is a common cause of bovine mastitis. Genomic heterogeneity exists between S. aureus strains and has been implicated in the adaptation of specific strains to colonise particular mammalian hosts. Knowledge of the factors required for host specificity and virulence is important for understanding the pathogenesis and management of S. aureus mastitis. In this study, a panel of mastitis-associated S. aureus isolates (n = 126) was tested for resistance to antibiotics commonly used to treat mastitis. Over half of the isolates (52%) demonstrated resistance to penicillin and ampicillin but all were susceptible to the other antibiotics tested. S. aureus isolates were further examined for their clonal diversity by Multi-Locus Sequence Typing (MLST). In total, 18 different sequence types (STs) were identified and eBURST analysis demonstrated that the majority of isolates grouped into clonal complexes CC97, CC151 or sequence type (ST) 136. Analysis of the role of recombination events in determining S. aureus population structure determined that ST diversification through nucleotide substitutions were more likely to be due to recombination compared to point mutation, with regions of the genome possibly acting as recombination hotspots. DNA microarray analysis revealed a large number of differences amongst S. aureus STs in their variable genome content, including genes associated with capsule and biofilm formation and adhesion factors. Finally, evidence for a genomic arrangement was observed within isolates from CC97 with the ST71-like subgroup showing evidence of an IS431 insertion element having replaced approximately 30 kb of DNA including the ica operon and histidine biosynthesis genes, resulting in histidine auxotrophy. This genomic rearrangement may be responsible for the diversification of ST71 into an emerging bovine adapted subgroup.

Fig 1. eBURST analysis of mastitis-associated S. aureus.

Sequence types (STs) from this study were compared to all STs of bovine origin in the MLST database. Circles represent different STs; blue represents a primary founder, yellow a subgroup founder, pink denotes STs found in both datasets, green STs found in present study dataset only and black STs in the MLST database dataset only. All lines represent a single locus difference and the size of the circles is relative to the number of isolates.

Fig 2. Relationship among the S. aureus isolates.

Hierarchical cluster dendrogram of all isolates based on their hybridization profile from the genotyping array. Bootstrap support values indicated for the major internal branches that separate the clonal complexes. Subgroups within the ST71-like group and CC151 are noted.

Fig 3. Gene differences for ST97 and ST71.

Schematic diagram of S. aureus genome showing gene differences for the closely related isolates ST97 and ST71.