Antibiofilm activity of GlmU enzyme inhibitors against catheter-associated uropathogens

Abstract

The colonization of uropathogenic bacteria on urinary catheters resulting in biofilm formation frequently leads to the infection of surrounding tissue and often requires removal of the catheter. Infections associated with biofilms are difficult to treat since they may be more than 1,000 times more resistant to antibiotics than their planktonic counterparts. We have developed an antibiofilm composition comprising an N-acetyl-D-glucosamine-1-phosphate acetyltransferase (GlmU) inhibitor and protamine sulfate, a cationic polypeptide. The antibiofilm activity of GlmU inhibitors, such as iodoacetamide (IDA), N-ethyl maleimide (NEM), and NEM analogs, including N-phenyl maleimide, N,N'-(1,2-phenylene)dimaleimide (oPDM), and N-(1-pyrenyl)maleimide (PyrM), was tested against that of catheter-associated uropathogens. Both IDA and NEM inhibited biofilm formation in Escherichia coli. All NEM analogs showed antibiofilm activity against clinical isolates of E. coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus epidermidis, and Enterococcus faecalis. The combination of oPDM with protamine sulfate (PS) enhanced its antibiofilm activity and reduced its effective concentration to as low as 12.5 microM. In addition, we found that the in vitro inhibitory activity of oPDM-plus-PS-coated silicone catheters against P. aeruginosa and S. epidermidis colonization was superior to that of catheters coated with silver hydrogel. Confocal scanning laser microscopy further confirmed that the oPDM-plus-PS-coated silicone catheters were almost free from bacterial colonization. Thus, a broad-spectrum antibiofilm composition comprising a GlmU inhibitor and protamine sulfate shows promise for use in anti-infective coatings for medical devices, including urinary catheters.

FIG. 1.

Effect of GlmU inhibitors on growth (□) and biofilm formation (▪) of E. coli P18 as determined by a microtiter plate assay. Error bars are not visible where the standard deviations are less than the area occupied by a given symbol.

FIG. 2.

Effect of GlmU inhibitors on growth (□) and biofilm formation (▪) of gram-positive catheter-associated pathogens (a) E. coli P18, (b) P. aeruginosa, and (c) K. pneumoniae as determined by a microtiter plate assay. Error bars are not visible where the standard deviations are less than the area occupied by a given symbol.

FIG. 3.

Effect of GlmU inhibitors on growth (□) and biofilm formation (▪) of gram-positive catheter-associated pathogens a) S. epidermidis and b) E. faecalis as determined by a microtiter plate assay. Error bars are not visible where the standard deviations are less than the area occupied by a given symbol.

FIG. 4.

Effect of oPDM and PS alone and in combination on growth (□) and biofilm formation (░⃞) of P. mirabilis, S. epidermidis, and K. pneumoniae as determined by a microtiter plate assay. Error bars are not visible where the standard deviations are less than the area occupied by a given symbol. Asterisks indicate a significant difference (P < 0.001) between biofilm formation in the presence of the oPDM-plus-PS combination and that in the presence of the control and oPDM and PS alone.

FIG. 5.

Confocal images of S. epidermidis biofilm formation on urinary catheters. a) Uncoated. b) Coated with oPDM plus PS.