Trends and prediction of antimicrobial susceptibility in urinary bacteria isolated in European emergency departments: the EuroUTI 2010-2016 Study

OBJECTIVES: To assess recent trends in susceptibility to antibiotics among urinary isolates isolated in European emergency departments (EDs) and to identify isolates with a high (90% or more) predicted probability of susceptibility to fluoroquinolones or third-generation cephalosporins (3GCs). METHODS: In this cross-sectional study, we included urine cultures obtained from adult patients between 2010 and 2016 in 24 European EDs. Temporal trends were assessed using time-series analysis and multivariate logistic models. Multivariate logistic models were also used to predict susceptibility to fluoroquinolones or 3GCs from patient age and sex, year, month and ED. RESULTS: We included 88 242 isolates. Time-series analysis found a significant increase in susceptibility to fluoroquinolones and no significant trend for susceptibility to 3GCs. Adjusting for patient age and sex, ED and organism, multivariate models showed that susceptibility to 3GCs decreased from 2014 to 2016, while susceptibility to fluoroquinolones increased in 2015 and 2016. Among isolates from 2016, multivariate models predicted high probability of susceptibility to fluoroquinolones in 11% of isolates (positive predictive value 91%) and a high probability of susceptibility to 3GCs in 35% of isolates (positive predictive value 94%). CONCLUSIONS: Susceptibility of ED urinary isolates to fluoroquinolones increased from 2014, while susceptibility to 3GCs decreased from 2015. Predictive models identified isolates with a high probability of susceptibility to fluoroquinolones or 3GCs. The ability of such models to guide the empirical treatment of pyelonephritis in the ED remains to be determined

Single-cell time-lapse imaging of intracellular O2 in response to metabolic inhibition and mitochondrial cytochrome-c release

The detection of intracellular molecular oxygen (O2) levels is important for understanding cell physiology, cell death, and drug effects, and has recently been improved with the development of oxygen-sensitive probes that are compatible with live cell time-lapse microscopy. We here provide a protocol for the use of the nanoparticle probe MitoImage-MM2 to monitor intracellular oxygen levels by confocal microscopy under baseline conditions, in response to mitochondrial toxins, and following mitochondrial cytochrome-c release. We demonstrate that the MitoImage-MM2 probe, which embeds Pt(II)-5,10,15,20-tetrakis-(2,3,4,5,6-pentafluorophenyl)-porphyrin as oxygen sensor and poly(9,9-dioctylfluorene) as an O2-independent component, enables quantitative, ratiometric time-lapse imaging of intracellular O2. Multiplexing with tetra-methyl-rhodamine-methyl ester in HeLa cervical cancer cells showed significant increases in intracellular O2 accompanied by strong mitochondrial depolarization when respiratory chain complexes III or IV were inhibited by Antimycin A or sodium azide, respectively, and when cells were maintained at 'physiological' tissue O2 levels (5% O2). Multiplexing also allowed us to monitor intracellular O2 during the apoptotic signaling process of mitochondrial outer membrane permeabilization in HeLa expressing cytochrome-c-eGFP, and demonstrated that mitochondria post cytochrome-c release are able to retain their capacity to respire at physiological O2 despite a decrease in mitochondrial membrane potential.</p