Regulation of Expression of the Nonhemolytic Phospholipase C of Burkholderia cepacia (original) (raw)

1999, Current Microbiology

Burkholderia cepacia is an opportunistic pathogen that causes serious pulmonary infections in cystic fibrosis patients. We have purified and partially characterized one potential virulence factor for the organism-a nonhemolytic phospholipase C-and we studied the effect of iron restriction and choline and phosphate concentrations on the expression of phospholipase C. Iron limitation did not affect expression, the effect of choline was variable, and high phosphate concentrations repressed expression. Experiments with heat-treated spent culture supernatants suggested that autoinducers affected the expression of the phospholipase and two other potential virulence factors, a protease and a lipase. We screened 26 B. cepacia isolates for autoinducer activity: 11 induced violacein production in the autoinducer-deficient mutant Chromobacterium violaceum CV026. Spent supernatants from two strains, one that was positive in the C. violaceum assay and one that was negative, were tested for inducing early expression of phospholipase C, protease, and lipase in homologous and heterologous cultures. Expression of all three enzymes was increased or induced at an earlier stage in the growth curve in every case, suggesting not only that autoinducers were involved in the regulation of the expression of these enzymes, but also that the autoinducers were of two different classes. Burkholderia cepacia has several potential virulence factors-a protease, a lipase, and two phospholipases: one hemolytic (PLC-H) and the other, non-hemolytic (PLC-N) [9, 10, 13, 14, 18, 19]. We have purified and partially characterized the PLC-N [19]. The protein is approximately 54 kDa, hydrophobic, and its N-terminus shows significant homology with PLCs from Mycobacterium tuberculosis and Pseudomonas aeruginosa. Low phosphate concentrations enhance PLC-N expression [9], but we were interested in the effect of iron and choline concentrations on the expression of the enzyme. We were also interested in determining whether quorum sensing might play a role in PLC regulation. We studied the regulation of the B. cepacia Pc224c PLC-N and, in addition to describing the effect (or lack of effect) of iron concentration and choline on expression, we also discovered that B. cepacia produces autoinducers that do affect the expression of not only PLC, but also the protease and the lipase. Materials and Methods Bacteria and growth media. Burkholderia cepacia strains (Table 1), originally obtained from D.E. Woods