Rapid sequencing of MRSA direct from clinical plates in a routine microbiology laboratory

Background: Routine sequencing of MRSA could bring about significant improvements to outbreak detection and investigation. Sequencing is commonly performed using DNA extracted from a pure culture, but overcoming the delay associated with this step could reduce the time to infection control interventions. Objectives: To develop and evaluate rapid sequencing of MRSA using primary clinical cultures. Methods: Patients with samples submitted to the clinical laboratory at the Cambridge University Hospitals NHS Foundation Trust from which MRSA was isolated were identified, the routine laboratory culture plates obtained and DNA extraction and sequencing performed. Results: An evaluation of routine MRSA cultures from 30 patients demonstrated that direct sequencing from bacterial colonies picked from four different culture media was feasible. The 30 clinical MRSA isolates were sequenced on the day of plate retrieval over five runs and passed quality control metrics for sequencing depth and coverage. The maximum contamination detected using Kraken was 1.09% fragments, which were identified as Prevotella dentalis. The most common contaminants were other staphylococcal species (25 isolate sequences) and Burkholderia dolosa (11 isolate sequences). Core genome pairwise SNP analysis to identify clusters based on isolates that were ≤50 SNPs different was used to triage cases for further investigation. This identified three clusters, but more detailed genomic and epidemiological evaluation excluded an acute outbreak. Conclusions: Rapid sequencing of MRSA from clinical culture plates is feasible and reduces the delay associated with purity culture prior to DNA extraction.