Masi Muriel, Dumont Estelle, Vergalli Julia, Pajovic Jelena, Réfrégiers Matthieu, Pagès Jean-Marie
UMR MD1, Aix-Marseille University, IRBA, TMCD2, Facultés de Médecine et de Pharmacie, 27 bd, Jean Moulin, 13385 Marseille cedex 05, France.
DISCO Beamline, Synchrotron Soleil, L'Orme des Merisiers, Saint-Aubin BP 48, 91192 Gif-sur-Yvette cedex, France; University of Belgrade, Faculty of Physics, Studentski trg 12, 11001 Belgrade, Serbia.
Res Microbiol. 2018 Sep-Oct;169(7-8):432-441. doi: 10.1016/j.resmic.2017.11.005. Epub 2017 Dec 5.
To understand antibiotic resistance in Gram-negative bacteria, a key point is to investigate antibiotic accumulation, which is defined by influx and efflux. Several methods exist to evaluate membrane permeability and efflux pump activity, but they present disadvantages and limitations. An optimized spectrofluorimetric method using intrinsic tryptophan fluorescence as an internal standard, as well as a complementary microfluorimetric assay following time-course accumulation in intact individual cells, have been developed. Comparing the latter population and single cell approaches can lead to an understanding of phenotypic heterogeneity within a population. The two methodologies lead to determination of parameters, concentration, accumulation rates and localization that contribute to emerging concepts (RTC2T, SICAR) with the aim of identifying and detailing antibiotic chemotypes involved in influx/efflux.
为了解革兰氏阴性菌中的抗生素耐药性，关键在于研究抗生素积累情况，抗生素积累由内流和外流定义。现有多种方法可评估膜通透性和外排泵活性，但这些方法都存在缺点和局限性。已开发出一种优化的光谱荧光法，该方法使用内源性色氨酸荧光作为内标，以及一种在完整的单个细胞中进行时间进程积累后的补充性显微荧光测定法。比较后一种群体方法和单细胞方法有助于理解群体中的表型异质性。这两种方法可用于确定参数、浓度、积累速率和定位，这些参数有助于形成新的概念（RTC2T、SICAR），目的是识别和详细描述参与内流/外流的抗生素化学类型。
