Assessment of Intestinal Transcytosis of Neonatal Escherichia coli Bacteremia Isolates.
Newborns ingest maternal E. coli strains that colonize their intestinal tract around the time of delivery. E. coli strains with the ability to translocate across the gut invade the newborn's bloodstream, causing life-threatening bacteremia. The methodology presented here utilizes polarized intestinal epithelial cells grown on semipermeable inserts to assess the transcytosis of neonatal E. coli bacteremia isolates in vitro. This method uses the established T84 intestinal cell line that has the ability to grow to confluence and form tight junctions and desmosomes. After reaching confluence, mature T84 monolayers develop transepithelial resistance (TEER), which can be quantified using a voltmeter. The TEER values are inversely correlated with the paracellular permeability of extracellular components, including bacteria, across the intestinal monolayer. The transcellular passage of bacteria (transcytosis), on the other hand, does not necessarily alter the TEER measurements. In this model, bacterial passage across the intestinal monolayer is quantified for up to 6 h post-infection, and repeated measurements of TEER are made to monitor the paracellular permeability. In addition, this method facilitates the use of techniques such as immunostaining to study the structural changes in tight junctions and other cell-to-cell adhesion proteins during bacterial transcytosis across the polarized epithelium. The use of this model contributes to the characterization of the mechanisms by which neonatal E. coli transcytose across the intestinal epithelium to produce bacteremia.
J Vis Exp
Infant, Newborn; Humans; Escherichia coli; Bacteremia; Cell Line; Epithelium; Transcytosis
Escherichia coli; Bacteremia; Cell Line; Epithelium; Transcytosis
Islam A, Wheatley JL, Chavez-Bueno S. Assessment of Intestinal Transcytosis of Neonatal Escherichia coli Bacteremia Isolates. J Vis Exp. 2023;(192):10.3791/64241. Published 2023 Feb 17. doi:10.3791/64241