Antibiofilm activity of Serratia marcescens purified gelatinase against Pseudomonas

Most bacteria can produce biofilms on avariety of surfaces in nature and it represents an important virulence factors such as the biofilm based infections by Pseudomonas aeruginosa can be life -threatening for people pationsand they can also lead to a long term infection. About 22 of Serratia marcescens clinical isolates were collected and characterized by morphological and biochemical tests even thogh PCR method . All the 22 isolates were screened for gelatinase production by culturing them on gelatine agar medium and the results showed that the S. m. no. 16 was the highest gelatinase producer which gave a lysis area of (28)mm in diameter as a primary and qualitative detection of this enzyme, also, crude extraction and quantitative detection were carried out by preparing of cell-free culture (CFS ) and detection of protein concentration of this crude extract to be equal to 12.144 by Bradford method. Purification of gelatinase was done by three steps: precipitation (80%) saturation of ammonium sulfate, followed by dialysis, ion exchange chromatography utilizing DEAE -Cellulose and gel filtration chromatography throughout Sephaeryl S-200 column. The results showed that S. marcescens 16 (S 16) gelatinase was obtained with specific activity of 3.75U\mg of protein with a fold of purification of about 49.23. Also, purified S16 gelatinase was characterized and the results showed that its molecular weight was 68 kDa by electrophoresis method (SDS-PAGE). S16 gelatinase and gentamicin's minimum inhibitory concentrations (MIC) were calculated using broth microdilution. Purifie gelatinase was tested for antibacterial activity at 8-16-32-64 µg/ml in vitro using the agar well diffusion method against five isolates of P. aeruginosa that were multidrug resistant. Antibiofilm activity of purified gelatinase at sub-MIC was more effective than the antibiofilm activity of gentamicin (P < 0.01), and the gelatinase was high active than the gentamicin in a preventing the production of biofilm. Gentamicin and purified gelatinase both possess significant antibacterial activity against P. aeruginosa isolates as compared with control