ANTI-PHAGOCYTIC MECHANISMS IN ATYPICAL ENTEROPATHOGENIC Escherichia coli (aEPEC)

AC01

Melo, KCM; Ruiz, RC 

Laboratório de Bacteriologia, Instituto Butantan, São Paulo, Brasil

Introduction: EPEC has been associated with diarrhea in several countries. At present it is the most common bacterial agent in children endemic diarrhea in Brazil. The atypical EPEC (aEPEC) differs from the typical (tEPEC) one by not presenting the EAF plasmidium and is considered an emergent pathogen. The increase in diarrhea cases caused by aEPEC, as tEPEC diarrhea cases drop, evokes the capacity of adaptation and pathogenicity that this EPEC subtype is developing. Several virulent bacteria have evolved mechanisms to prevent phagocytosis. The interaction of tEPEC with phagocytic cells inhibits its uptake. This requires a functional type III secretion system and occurs via PI 3 kinase inhibition required for the re-assemblage of the cytoskeleton and actin polymerization, both important in phagocytosis. Developing an anti-phagocytic mechanism seems to improve the colonization of the epithelial cells by enteric bacteria, by delaying the activation of the immune response. Objective: Investigation of the existence of an anti-phagocytic mechanism in aEPEC. Methodology: Macrophages interaction assays were performed with aEPEC isolates: 7(O55:H7); 320(O55:H7) and tEPEC control (E2348/69) for 10, 30, 60 min infection pulses with different bacteria concentrations. The macrophages were treated with gentamicin and the number of intracellular bacteria was determined. Results: Isolate 7 (DO 0,6) was less phagocyted after 10 and 30 min pulses with 20,3% and 49,9% of infection, respectively, while iIsolate 320 infected 76,3% and 81,9% and   isolate E2348/69 infected 91,2I% and 94,2% at the same times. Furthermore, in isolate 7, the number of bacteria adhered or phagocyted per macrophage is significantly smaller than with isolates 320 and E2348/69. The kinetics of interaction with different bacteria concentrations (OD 1,2; 0,6; 0,06 and 0,006) showed that the anti-phagocytic effect of isolate 7 still occurs at much smaller concentrations, such as OD 0,006. In the 10 min infection pulse, both the bacterial interaction and the number of bacteria/cell were smaller than observed with the control isolate. In the 30 min pulse, only the number of bacteria/cell was smaller than with the control. Isolate E2348/69, with no anti-phagocytic effect under these conditions, showed less invasive after pre-incubation with isolate 7 supernatant in the 10 min infection pulse. Discussion: Isolate 7 may present a different anti-phagocytosis mechanism from tEPEC, witch triggers the anti-phagocytosis effect after 2h of interaction, and not presented by the other aEPEC isolate studied. These results suggest that aEPEC isolate 7 seems to induce a different anti-phagocytic effect than that presented by the tEPEC, witch depends on the adhesion of the bacterium to the macrophage. The existence of an anti-phagocytic mechanism may be of great advantage to the pathogen, since prevention or delay of the immune response may contribute to intestinal colonization.