Recombinant Escherichia coli as a model to study a potential pathogenic agent of Burkholderia pseudomallei-phospholipase C
Abstract number: p1179
Sadowska B., Wieckowska-Szakiel M., Rudnicka W., Korbsrisate S., Rozalska B.
Burkholderia pseudomallei is the etiological agent of melioidosis endemic disease, whose acute form is manifested as pulmonary or septicemic infection with abscesses formation. The pathogenic factors of these Gram-negative bacilli are not known well enough. However, it is worth noting, that they produce many enzymes including phospholipase C (PLC). We hypothesised that PLC can be important for B. pseudomallei interaction with host immune cells.
A parental E. coli DH5alpha strain (PLC-) and recombinant mutant E. coli pSN-1a producing phospholipase C from B. pseudomallei (PLC +) were used for the study. The mouse peritoneal macrophages were infected with FITC-labelled and "normal" bacteria to determine their adherence to the cells by a fluorimetric assay and ingestion / intracellular killing by a colony forming units-assay. The supernatants were collected to estimate the production of IL-12 and TNF-alpha by infected phagocytes using DuoSet ELISA (RandD).
The adherence of E. coli DH5alpha to the macrophages was stronger that of E. coli pSN-1a mutant, which also corresponded with better ingestion and intracellular killing of this PLC- strain by used phagocytes. Interestingly, the phagocytic activity of the cells infected with the mixed population of both E. coli (PLC + and PLC -) strains was diminished as compared with their activity against the homogenous populations of these bacteria. Two hours incubation of the macrophages with those E. coli strains led to the production of TNF-alpha (the level of IL-12 was below detection limit). However, the stimulatory effects of parental and mutant strain as well as their homo- and heterogeneous cultures were similar.
The production of phospholipase C seems to disturb phagocytic but not secretory activity of the macrophages. Thus, our data suggest that this enzyme can play an important role in the survival of B. pseudomallei inside professional phagocytes. Some investigators even suggest that phospholipase C of B. pseudomallei can play a similar role in the pathogenesis of melioidosis as phospholipase for Listeria monocytogenes, which takes place during the "escape" of these bacteria from the phagolisosomes.
Supported by Grant No. 3 PO4C 081 24 from State Committee for Scientific Research.
|Session name:||XXIst ISTH Congress|
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