Clinical and Molecular Epidemiology of an Emerging Panton-Valentine Leukocidin-Positive ST5 Methicillin-Resistant Staphylococcus aureus Clone in Northern Australia

Abstract

Recently, we identified a Staphylococcus aureus sequence type 5 (ST5) clone in northern Australia with discrepant trimethoprim-sulfamethoxazole (SXT) susceptibility results. We aimed to identify isolates of this clone using Vitek 2 SXT resistance as a proxy and to compare its epidemiology with those of other circulating S. aureus strains. We collated Vitek 2 susceptibility data for S. aureus isolates collected through our laboratory and conducted a prospective, case-control study comparing clinical, microbiological, epidemiological, and genomic data for subsets of isolates reported as SXT resistant (cases) and SXT susceptible (controls) by Vitek 2. While overall SXT resistance rates remained relatively stable from 2011 to 2018 among 27,721 S. aureus isolates, non-multidrug-resistant methicillin-resistant S. aureus (MRSA) strains almost completely replaced multidrug-resistant MRSA strains as the predominant SXT-resistant MRSA phenotype. Demographic and clinical features of 51 case-control pairs were similar, but genotyping revealed stark differences: clonal complex 5 (CC5) MRSA predominated among SXT-resistant cases (34/51 [67%]), while CC93 MRSA predominated among susceptible controls (26/51 [51%]). All CC5 isolates were an ST5 clonal lineage that possessed the trimethoprim resistance gene dfrG within SCCmec IVo; all were SXT susceptible by Etest. The replacement of Vitek 2 reported SXT-resistant multidrug-resistant MRSA by non-multidrug-resistant MRSA appears related to the emergence of an ST5-MRSA-SCCmec IVo clone that is SXT susceptible by Etest and causes clinical disease similar to that caused by ST93-MRSA-SCCmec IVa. Reliance on Vitek 2 SXT reporting may lead to unnecessary restriction of effective oral treatment options for S. aureus infections. Whether the presence of dfrG within SCCmec IVo provides a selective advantage at the population level is currently unclear.IMPORTANCEStaphylococcus aureus is an important human pathogen that causes a wide range of clinical infections. In the past 2 decades, an epidemic of community-associated skin and soft tissue infections has been driven by S. aureus strains with specific virulence factors and resistance to beta-lactam antibiotics. Recently, an S. aureus strain with discrepant antimicrobial susceptibility testing results has emerged in northern Australia. This ST5-MRSA-SCCmec IVo clone is reported as resistant to trimethoprim-sulfamethoxazole by Vitek 2 but susceptible by phenotypic methods. ST5-MRSA-SCCmec IVo is now the second most common community-associated MRSA clone in parts of Australia and causes a spectrum of clinical disease similar to that caused by the virulent ST93-MRSA lineage. Whole-genome sequence analysis demonstrates that ST5-MRSA-SCCmecIVo is causing a clonal outbreak across a large geographical region. Although phenotypic testing suggests in vitro susceptibility to trimethoprim-sulfamethoxazole, it is unclear at this stage whether the presence of dfrG within SCCmec IVo provides a selective advantage at the population level.

Keywords: Staphylococcus aureus; epidemiology; genomics; methicillin resistance; susceptibility testing; trimethoprim.

FIG 1

Proportion of 27,721 S. aureus isolates reported as SXT resistant by Vitek 2 by year (2011 to 2018), stratified by type (A) and proportion of 27,721 S. aureus isolates reported as MSSA, nmMRSA, and mMRSA by year (2011 to 2018) (B). SXT, trimethoprim-sulfamethoxazole; nmMRSA, non-multidrug-resistant MRSA; mMRSA, multidrug-resistant MRSA; MSSA, methicillin-susceptible S. aureus.

FIG 2

Genotyping results for S. aureus isolates among cases and controls. CC, clonal complex; ST, sequence type. Full genotyping results are available in Data Set S1.

FIG 3

Map of distribution of ST5 and CC93 MRSA isolates. The greater Darwin area includes Darwin City, Palmerston City, and Litchfield Council areas, Belyuen and Coomalie Community Council areas, and the Tiwi Islands.

FIG 4

Structure of SCCmec IVo. SCCmec IVo is defined as SCCmec IVc harboring a putative mobile genetic element containing dfrG and two additional open reading frames (ORFU1 and ORFU2) encoding hypothetical proteins (18), inserted upstream of IS431. The direct repeats (DR) and left and right inverted repeats (IRL and IRR, respectively) of the putative dfrG mobile genetic element are indicated.

FIG 5

Phylogenomic analysis of ST5 isolates. Shown is a maximum-parsimony tree rooted on Mu50 of ST5 genome sequences based on 2,148 biallelic single nucleotide polymorphisms located in regions orthologous to all genomes (consistency index = 0.9581). Genome sequences included were those of 34 ST5 isolates from this study (indicated with the prefix “CASE_”), 24 ST5 isolates from a recent study of trimethoprim resistance in the same region (BioProject no. PRJNA312422) (16), and 5 publicly available ST5 isolates (ED98 [GenBank accession no. [NC_013450], ECT-R 2 [NC_017343], N315 [NC_002745], Mu3 [NC_009782], and Mu50 (NCBI reference sequence NC_002758.2; GenBank accession no. [NC_002758]). Synthetic short-read data (designated Mu50) generated from the Mu50 reference genome assembly (designated Mu50_R) were also included as an internal control for the alignment. The tree is available in Newick format in Text S1. Gray blocks to the right indicate the presence of mecA, lukF/S (PVL), dfrG, and SCCmec. Vitek 2 SXT MICs are reported as the sum of the trimethoprim and sulfamethoxazole MICs which are present in a ratio of 1:19, with a resistance breakpoint of ≥80 mg/liter (29). Etest SXT MICs are reported as the trimethoprim MIC, with a resistance breakpoint of >4 μg/ml (38). Scale represents 50 SNPs.