Summation of peaks and L34 ribosomal protein in the presence and absence of antibiotics enables susceptibility testing using MALDI-TOF mass spectrometry in 2h from Escherichia coli-positive blood cultures / El sumatorio de picos y la proteína ribosómica L34, en presencia y ausencia de antimicrobianos, permiten conocer la sensibilidad a antimicrobianos mediante espectrometría de masas MALDI-TOF en 2h, en hemocultivos positivos para Escherichia coli

INTRODUCTION: We have developed a MALDI-TOF-mediated phenotypic method, which determines antibiotic susceptibility (AS) from positive blood cultures (BCs) in 2h. We developed a software for process automation. We report results on Escherichia coli-positive BCs with cefotaxime (CTX) and ciprofloxacin (CIP). METHODS: We studied CIP and CTX activity in 18 and 17 real E. coli-positive BCs, and in 56 and 45 spiked BCs, respectively. Positive BCs were incubated for 2 h without any antibiotics, and with 2 mg/l and 4 mg/l of CIP and CTX. The extraction was performed using ethanol/formic acid. Spectra were processed with specifically developed software which compares the peaks' intensity and the size of specific peaks. RESULTS: The set cut-off point was a 3-fold decrease in the summation of all peaks and/or the 5382 m/z peak value (ribosomal protein L34). In simulated BCs, the correlation of CIP 2mg/l and 4mg/l with Etest(R) was 94.6% and 98.2%, respectively; for CTX 2 mg/l and 4 mg/l, this correlation was 95.6%. In real BCs, the correlations were 100% for CIP (2mg/l and 4mg/l) and 88.2% and 94.1% for CTX 2 mg/l and 4 mg/l, respectively. Resistant isolates were always correctly classified. CONCLUSION: This method provides accurate, fast and inexpensive AS information. The method can be automated, making it easier to implement in a microbiology laboratory routine

INTRODUCCIÓN: Se ha desarrollado un método fenotípico basado en MALDI-TOF, que determina la sensibilidad a antibióticos en hemocultivos (HC) positivos en 2h. Se ha desarrollado un software que automatiza el proceso. Se presentan los resultados en HC positivos para Escherichia coli, con cefotaxima (CTX) y ciprofloxacino (CIP). MÉTODOS: Se estudió la actividad de CIP y CTX en 18 y 17HC positivos reales con E. coli, y en 56 y 45 HC simulados. Los HC positivos se incubaron durante 2h sin antibiótico, y con 2 y 4 mg/l de CIP y de CTX. La extracción se realizó con etanol/ácido fórmico. Los espectros se procesaron con un software específico, que compara la intensidad de los picos y el tamaño de los picos específicos. RESULTADOS: El punto de corte establecido fue una disminución de 3 veces en la suma de picos, y/o en el valor del pico de 5.382 m/z (proteína ribosómica L34). En hemocultivos simulados la correlación con Etest(R) para las concentraciones de CIP de 2 y 4 mg/l fueron 94,6 y 98,2%, respectivamente, y 95,6% para CTX (2 y 4 mg/l). En HC reales, la correlación con Etest(R) fue del 100% para CIP (2 y 4 mg/l), y del 88,2 y 94,1% para CTX 2 y 4 mg/l, respectivamente. Los aislados resistentes siempre se clasificaron correctamente. CONCLUSIÓN: Este método proporciona información sobre sensibilidad a antimicrobianos de manera precisa, rápida y barata. El método se puede automatizar e incluir en la rutina del laboratorio de microbiología

Summation of peaks and L34 ribosomal protein in the presence and absence of antibiotics enables susceptibility testing using MALDI-TOF mass spectrometry in 2h from Escherichia coli-positive blood cultures.

INTRODUCTION: We have developed a MALDI-TOF-mediated phenotypic method, which determines antibiotic susceptibility (AS) from positive blood cultures (BCs) in 2h. We developed a software for process automation. We report results on Escherichia coli-positive BCs with cefotaxime (CTX) and ciprofloxacin (CIP). METHODS: We studied CIP and CTX activity in 18 and 17 real E. coli-positive BCs, and in 56 and 45 spiked BCs, respectively. Positive BCs were incubated for 2h without any antibiotics, and with 2mg/l and 4mg/l of CIP and CTX. The extraction was performed using ethanol/formic acid. Spectra were processed with specifically developed software which compares the peaks' intensity and the size of specific peaks. RESULTS: The set cut-off point was a 3-fold decrease in the summation of all peaks and/or the 5382m/z peak value (ribosomal protein L34). In simulated BCs, the correlation of CIP 2mg/l and 4mg/l with Etest® was 94.6% and 98.2%, respectively; for CTX 2mg/l and 4mg/l, this correlation was 95.6%. In real BCs, the correlations were 100% for CIP (2mg/l and 4mg/l) and 88.2% and 94.1% for CTX 2mg/l and 4mg/l, respectively. Resistant isolates were always correctly classified. CONCLUSION: This method provides accurate, fast and inexpensive AS information. The method can be automated, making it easier to implement in a microbiology laboratory routine.