Journal of Pharmaceutical Research International

Aims: To detect the prevalence of biofilm producers among Gram negative bacilli and Gram positive cocci bacterial pathogens along with their antimicrobial susceptibility pattern. Growth and adherence on catheter eluates and in the presence of antibiotics.
Methodology: From laboratory of microbiology, one hundred samples (100 urinary catheters and 100 urine samples from the attached drainage bags) of bladder cancer patients collected in National Cancer Institute in Cairo, Egypt, were identified to species level. Slime production was investigated by the quantitative and qualitative methods. Qualitative method was carried out by tube method. Adherence assay and quantitation of biofilm was performed by spectrophotometric method by measuring the optical densities of stained bacterial films adherent to plastic tissue culture plates. Hydrophobicity was evaluated by adhesion to P-xylene. Identification and minimum inhibitory concentration (MICs) of 26 antimicrobial agents against gram negative and 24 against gram positive bacterial isolates were determined using microscan walk away 96 SI system. Plasmid profile analysis was carried out by plasmid isolation kit. Scanning electron microscopy studies for growth, adherence and biofilm formation. Impact of gamma irradiation at a dose level of 24.41Gy was studied.
Results: From the processing of 100 samples, 98 cases were positive. Out of them 110 isolates of gram negative bacilli and 13 of gram positive cocci. They were belonging to 15 and 6 species respectively. Among them, 117 isolates showed positive results for adherence assay and biofilm/slime production. They were identified as; Escherichia coli, Klebsiella spp., Enterobacter spp., Acinetobacter, Proteus spp., Citrobacter, Alcaligenes, Empedobacter (104 strains) Staphylococcus spp. and Enterococcus (13 stains). The results obtained by different methods correlated well with strain to strain variation. Gamma irradiation resulted in changes in slime production and adherence ability for all the tested strains. Cell surface hydrophobicity (CSH) showed a hydrophobic reaction and these with increase in its value after irradiation in case of Escherichia coli. On the other hand, Staphylococcus epidermidis was moderate hydrophobic before irradiation changed to strictly hydrophilic after irradiation. All the slime producers showed reduced susceptibility to majority of antibiotics. They exhibited highest percentage susceptibility before and after in vitro gamma irradiation at a dose level 24.41Gy for both Amikacin and Imipenem. Scanning electron microscopy (SEM) confirmed growth and biofilm formation in the presence of catheter eluates only with halos surrounding the cells and visible erosion zones on catheter surfaces. The antimicrobial and adherence activity of Amikacin and Imipenem at the MIC level showed marked abnormalities in cells shape and size with significant reduction in adherence ability. Plasmid profile analysis of irradiated strains showed more extra-plasmid bands and / or difference in molecular weight.
Conclusion: The biofilm assay strategy applied in this study may constitute a tool in biomaterial related infection and antimicrobial resistant research for further studies for biomaterial modification. Early detection of biofilm forming organisms can help in appropriate antibiotic choice.