Flow cytometry parameters for predicting the successful identification of gram-negative bacilli in urine by MALDI-TOF...

Direct matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) enables rapid identification of microorganisms in urinary tract infections (UTIs), accelerating diagnosis and guiding early antimicrobial therapy alongside conventional culture. However, there is no universally established threshold for significant bacteriuria determined by flow cytometry in clinical decision-making. This study analyzed 652 urine samples from suspected UTI patients at Hospital Clínico Universitario de Valencia (Jan 2023-Nov 2024). Flow cytometry was used to determine bacterial load (bacteria/µL) and Gram staining before MALDI-TOF MS and culture processing. The overall concordance between MALDI-TOF MS and culture was 96.5%. Samples successfully identified by MALDI-TOF MS had a significantly higher median bacterial load than those not identified. Receiver operating characteristic (ROC) analysis established 15,606 bacteria/µL as the optimal cut-off for MALDI-TOF MS processing (sensitivity: 83.3%, specificity: 50%, PPV: 70.9%, NPV: 67.2%). This differs from the conventional ≥ 10⁵ CFU/mL threshold, suggesting MALDI-TOF MS remains effective at lower bacterial loads, thus improving resource efficiency. Additionally, lateral flow immunochromatographic (LFIC) assays facilitated rapid detection of resistance mechanisms, aiding empirical antimicrobial therapy. Our findings support the utility of flow cytometry in efficiently selecting candidate samples for MALDI-TOF MS, enhancing clinical decision-making in UTI management.