2476. Universal detection of healthcare-related bacterial transmissions using de novo assembly with short-read whole genome sequencing

BACKGROUND: Whole genome sequencing (WGS) is the gold standard for molecular evidence of pathogen transmission. To date, most WGS investigations within healthcare settings have used low cost short-read technologies (e.g., Illumina), aligning reads to a high quality reference genome. However, this relies on either public repositories having closely-related high quality references, which is not guaranteed for arbitrary clinical isolates, or additional long-read sequencing. De novo assembly avoids reliance on references, but the utility of fragmented assemblies of short reads is not well-established in hospital outbreak investigations. METHODS: 24 clusters of clinical isolates were identified by automated and routine methods at 17 U.S. hospitals and underwent paired-end WGS on Illumina platforms, followed by two parallel analyses. For a reference-free analysis, de novo assembly was performed with Shovill, assemblies were clustered with Mash, core-genome alignments were created with Harvest, and finally single nucleotide polymorphism (SNP) distances were visualized using the PathoSPOT phylogenomics toolkit to highlight likely transmission events. For a reference-guided analysis, references were selected for each isolate using StrainGST, and SNPs called by read alignment were used to define phylogenomic distances. Finally, conclusions from both analyses were compared. RESULTS: 92 bacterial isolates underwent WGS, of which 86 (93%) completed de novo assembly and passed quality control, comprising 10 isolates of Acinetobacter baumanii, 4 of Klebsiella oxytoca, 46 of Klebsiella pneumoniae, and 26 of Staphylococcus aureus. Reference-free analysis with PathoSPOT identified close relatedness consistent with transmission in 13 of 24 clusters (54%), while the reference-guided analysis identified the same in 12 clusters (50%). Overall, 21 clusters (88%) showed concordance between reference-free and reference-guided methods. Cohen’s kappa indicated substantial agreement (0.75; 95% CI, 0.49–1.0). CONCLUSION: De novo assembly of short reads is tenable for WGS investigation of hospital outbreaks, elides the need for reference genomes or long-read sequencing, and compares favorably with SNP calling via read alignment for detecting likely healthcare-related transmissions. DISCLOSURES: Laura E. McLean, M.Ed., HCA Healthcare: Stocks/Bonds Susan S. Huang, MD MPH, Medline Industries, Inc: Conducted studies whereby participating nursing homes and hospital patients received cleaning & antiseptic products|Xttrium Laboratories: Conducted studies where participating nursing homes and hospital patients received antiseptic products Yonatan H. Grad, MD, PhD, Day Zero Diagnostics: Board Member|GSK: Advisor/Consultant