Computational protocol: Whole genome sequencing to investigate a putative outbreak of the virulent community-associated methicillin-resistant Staphylococcus aureus ST93 clone in a remote Indigenous community

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Protocol publication

[…] The purpose of the contact tracing exercise was explained to community members with the aid of a flip chart and a local interpreter where required. Written informed consent was obtained from each study participant or their guardian. The investigation was initially registered as a quality assurance audit (QAAR 2013-2107), and subsequently a complete research ethics application was approved (HREC 2015-2368) by the Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research.During a 2-week period in September 2013, we visited the households of the two index cases. All members of the two households had nasal, throat and groin swabs collected, and skin lesions swabbed if present. Additionally, we collected nasal swabs from a sample of patients who had presented to the community health clinic, and we also obtained S. aureus isolates from patients with SSTI at this time.The swabs were collected and transported as previously described (). We extracted DNA from S. aureus isolates using a QiaAMP DNA Mini Kit (Qiagen) according to the manufacturer’s instructions. Genotyping to the level of clonal complex was by a high-resolution melting analysis method previously described (). In addition, PCR assays were used to detect the nuc (), mecA () and lukSF-PV () genes. Whole genome sequencing of ST93 isolates was carried out on an Illumina HiSeq platform by Macrogen or through a collaborative agreement with the Wellcome Trust Sanger Institute. The ST93 genomes already published provided a phylogenetic framework for the analysis () and we sequenced an additional 20 ST93 isolates obtained between 2005 and 2010 from Northern Territory (NT) hospital-based studies to provide local genomic context (; ).We aligned sequence data from existing isolates () and newly generated sequences against the JKD6159 reference genome (ST93, GenBank: CP002114), using spandx v2.6, with default parameters to produce a core alignment (). Orthologous SNPs were used for phylogenetic reconstruction using RAxML (). Mobile genetic elements were not excluded. […]

Pipeline specifications

Software tools SPANDx, RAxML
Applications Phylogenetics, WGS analysis, Nucleotide sequence alignment
Organisms Staphylococcus aureus
Diseases Pneumonia, Skin Diseases, Staphylococcal Infections