Role of autolysins in the EDTA-induced lysis of Pseudomonas aeruginosa.

Abstract

Treatment of Pseudomonas aeruginosa with metal ion chelators, especially ethylenediaminetetraacetic acid (EDTA), causes both release of protein-lipopolysaccharide complexes and cell death. We have examined the effect of EDTA on P. aeruginosa and found that EDTA does not induce the rapid solubilization of the peptidoglycan sacculus and complete lysis as previously thought; the decrease in optical density of cultures incubated with EDTA is primarily due to the loss of the outer membrane. Of the other potential solubilizers examined, only ethylene-bis(oxyethylenenitrilo)tetraacetic acid (EGTA) resulted in some decrease in optical density. The lytic effect of EDTA on 12 strains of P. aeruginosa was examined and was found to vary greatly between strains; the sensitivity to EDTA varies from between 96% and 10% of the decrease in optical density resulting from incubation of cells with both EDTA and lysozyme. Sensitivity to EDTA is not constant during the growth of P. aeruginosa; in the early exponential phase of growth, cells treated with EDTA exhibit a 82% decrease in optical density after 30 min while in the stationary phase the optical density decreases by only 40%. Nucleic acids were observed to leak from cells following treatment with EDTA and this was greatly facilitated by DNase and RNase. The release of genetic material was much reduced when cells were incubated at 4 degrees C, supporting an enzymatic role in cell wall solubilization. We propose that only small areas of the sacculus become hydrolysed via specific peptidoglycan hydrolases, or autolysin(s), which are activated or de-regulated by EDTA.