RNA Sequencing of Sepsis Patients Informs Tests to Quickly Diagnose Pathogens and Resistance

Purpose: Diagnosis of infection in patients with sepsis takes days via culture and appropriate treatment of pathogens are delayed awaiting results. We hypothesize that we can use RNA sequencing from patients with sepsis to identify novel targets for faster nucleic acid-based tests. Methods: Cohort study of sepsis patients admitted to the ICU with RNA sequencing done after consented. RNA sequencing data that did not map to the human genome was then aligned to resistance genes and pathogen genomes and used to design novel PCR tests. These tests were correlated with blood culture diagnosis and clinical outcomes. Results: Forty-six patients were enrolled and samples from 87 time points were collected. These samples resulted in 8.6 billion RNA sequencing reads to identify pathogen RNA. PCR target discovery focused on positive blood cultures (n=40 total) due to Escherichia coli (5 samples), Staphylococcus aureus (6 samples), and Pseudomonas aeruginosa (3 samples) as well as identification of resistance genes. From RNA sequencing reads, 40 targets were defined and tested by quantitative PCR. In a cohort of patients (9 out of 46) with available samples, some of the proposed PCRs identified all cases of positive blood cultures (Pseudomonas aeruginosa and Staphylococcus aureus); Escherichia coli had no positive blood cultures in this cohort. Conclusions: RNA sequencing from patients with sepsis can identify RNA from pathogens causing the infection. This is used to design PCR primers that identify patients with positive blood cultures. Translation of these primers to clinical microbiology machines will allow the diagnosis faster than blood culture.