Metagenomic Characterization of Nitrite-Negative Urinary Tract Infections and Identification of an optrA-fexA Co-Localization in a Primary Healthcare Setting in China.

INTRODUCTION: Urinary tract infections (UTIs) showing negative nitrite results create a diagnostic blind spot for Gram-positive pathogens. The plasmid-mediated resistance genes optrA (linezolid) and tet(X) (tigecycline) pose severe therapeutic challenges, but their genomic context in primary healthcare settings remains poorly characterized. METHODS: Deep metagenomic sequencing was performed on urine samples from 20 patients with complex nitrite-negative UTIs and pyuria in China. Antibiotic resistance genes (ARGs) were profiled using a read-mapping threshold (>50 reads for tet(X)) and de novo assembly. RESULTS: optrA was ubiquitously detected via read mapping. Based on our pre-established cutoff, tet(X) reads were identified in 55% (11/20) of samples. Crucially, genomic assembly captured the physical co-localization of optrA and the phenicol exporter fexA on a single contig in one sample (JX-NY001). Virulence factor profiles varied across samples. DISCUSSION: This study provides genomic evidence of high-risk resistance determinants in community nitrite-negative UTIs. However, results are limited by the DNA-level metagenomic approach without culture confirmation. Molecular surveillance is needed to monitor these cryptic multidrug-resistance clusters.