Metagenomic sequencing as a diagnostic tool for urine culture negative febrile urinary tract infection

OBJECTIVES: The diagnosis of febrile urinary tract infection (fUTI) by urine culture is hampered by antibiotic pre-treatment. We investigated urine metagenomics to diagnose fUTI in patients with positive blood but negative urine cultures. METHODS: We performed shotgun metagenomic sequencing on 41 culture-positive and 19 culture-negative urine samples from fUTI patients, comparing urine metagenomics to blood and urine culture including antimicrobial susceptibility testing (AST). mOTUs3.1 performed metagenomic pathogen detection and ResFinder2.0 antimicrobial drug resistance (AMR) gene detection (standard settings). Whole genome sequencing (WGS) was performed on blood culture isolates from culture-negative urine samples. BWA-MEM and sylph aligned metagenomic pathogen reads to their respective WGS assemblies. RESULTS: Metagenomics detected the blood culture isolate in 39/41 culture-positive and 17/19 culture-negative urine samples. 11/19 urine culture-negative patients were pre-treated with antibiotics, versus 8/41 urine culture-positives. The blood culture isolate was the most abundant pathogen in 33/41 culture-positive and 15/19 culture-negative urine samples. A median of 93.2% of pathogen-specific metagenomic reads mapped to their WGS assemblies with a median ANI of 98.7% (n=11). Genotypic AMR detection and phenotypic AST matched in 38-96% of cases. CONCLUSIONS: Urine metagenomics successfully detected the causative pathogen in urine culture-negative fUTI patients. Genotypic AMR prediction requires further investigation.