The Role of Glucose in the Pathology of EHEC O157: H7

DA SILVA, WANDERSON MARQUES; LARZABAL, MARIANO; TAIBO, CATALINA; CATALDI, ÁNGEL; GARCÍA, JULIA SABIO

MICROSCOPY & MICROANALYSIS

CAMBRIDGE UNIV PRESS

Lugar: Cambridge; Año: 2020 vol. 26 p. 181 - 182

1431-9276

The pathogen enterohemorrhagic Escherichia coli (EHEC) O157: H7 is responsible for hemorrhagic colitis and hemolytic uremic syndrome in humans [1]. During the colonization process in the gastrointestinal tract, EHEC needs to adapt to changes in nutrient availability [2]. The objective of this study was to evaluate the influence of glucose on physiology and processes involved in the pathogenesis of EHEC O157: H7 in order to improve our understanding of the mechanisms controlling EHEC growth and survival in the bovine gut. In this study we first analyzed the growth rate of EHEC O157: H7 Rafaela II clade 8, a strain isolated from a bovine in Argentina, grown in the medium DMEM supplemented with either 4.5% glucose (High-glucose - DHG) or 1% glucose (Low-glucose - DLG). In addition, we assessed the bacterial adhesion capacity and actin pedestal formation induced by EHEC [3] by performing infection assays. For this purpose, Caco-2 epithelial cells were exposed for 5 h with Rafaela II grown with the different concentrations of glucose. Subsequently, the samples were fixed (paraformaldehyde 4%) and permeabilized (triton); actin and nucleic acids (DNA) were stained with rhodamine-phalloidin and TO-PRO-3, respectively. Bacterial adhesion capacity and pedestal formation of cells were evaluated using a Leica TCS SP5 laser scanning confocal microscope (MC). Each Image was acquired by monitoring a single focal plane over time (xyt scanning mode) using a 40X/1.25 oil objective lens and 543nm HeNe and 633 nm HeNe lasers. The frequency and resolution for acquiring images were set at 200 Hz and 1,024 x 1,024 pixels, while maintaining the same settings for laser powers, gain, and offset. The growth rate of Rafaela II was similar under either condition (DHG and DLG). According to the MC observations of the infection assays, however, Rafaela II grown in DLG displayed smaller cell size as well as greater ability to adhere to Caco-2. Furthermore, the cells infected with the strain grown in DLG presented higher actin accumulation. This actin rearrangement is consistent with the formation of pedestals and is characteristic of the ?attaching and effacing? lesions that contribute to the diarrheal manifestations caused by EHEC infection [4]. These preliminary assays demonstrate that glucose plays a role in processes related to EHEC physiology and pathogenesis of EHEC O157: H7 Rafaela II clade 8.