INTRACELLULAR SURVIVAL OF PSEUDOMONAS AERUGINOSA PAO1 IN A549 CELLS
TSVETELINA PAUNOVA-KRASTEVA1*, VASILKA VELKOVA1,2, DAYANA BORISOVA1,
STOYANKA STOITSOVA1
1- Department of General Microbiology, Institute of Microbiology, BAS, Sofia, Bulgaria;
2 -Faculty of Biology, Sofia University “St. Kliment Ohridski”, Sofia, Bulgaria
*Corresponding author: pauny@abv.bg
Keywords: Pseudomonas aeruginosa, human lung carcinoma A549 cells, GFP
Abstract: In their interaction with host epithelia, different strains of Pseudomonas aeruginosa manifest different phenotypes: they may remain adherent to the surface of the eukaryotic cells (adherent phenotype), or penetrate and survive intracellularly (invasive phenotype). The aim of the study is to characterize the interaction of strain P. aeruginosa PAO1 with human lung carcinoma A549 cells. Initially, we performed a co-cultivation of the cells with 1x104 bacteria for 2 hours followed by incubations for 1 or 24 hours of the infected cells in the absence of bacteria. We determined the number of viable intracellular bacteria by plating on agar. The result indicated that the bacterium is capable for intracellular replication. Using a three-parental labelling system (including E. coli MT 102 pBAH 8 as donor, and E. coli HB 101 pR K600 as helper) the strain was labelled with green fluorescent protein. Fluorescent microscopy of the GFP-carrying PAO1-GFP strain after co-cultivation with A549 cells confirmed the intracellular survival of the bacteria, as well as progressive disruption of the actin cytoskeleton.