Effects of dam and seqA genes on biofilm and pellicle formation in Salmonella.

In this study, the effects of dam and seqA genes on the formation of pellicle and biofilm was determined using five different Salmonella serovars S. Group C1 (DMC2 encoded), S. Typhimurium (DMC4 encoded), S. Virchow (DMC11 encoded), S. Enteritidis (DMC22 encoded), and S. Montevideo (DMC89 encoded). dam and seqA mutants in Salmonella serovars were performed by the single step lambda red recombination method. The mutants obtained were examined according to the properties of biofilm on the polystyrene surfaces and the pellicle formation on the liquid medium. As a result of these investigations, it was determined that the biofilm formation properties on polystyrene surfaces decreased significantly (p < 0.05) in all tested dam and seqA mutants, while the pellicle formation properties were lost in the liquid medium. When pBAD24 vector, containing the dam and seqA genes cloned behind the inducible arabinose promoter, transduced into dam and seqA mutant strains, it was determined that the biofilm formation properties on the polystyrene surfaces reached to the natural strains' level in all mutant strains. Also, the pellicle formation ability was regained in the liquid media. All these data demonstrate that dam and seqA genes play an important role in the formation of biofilm and pellicle structures in Salmonella serovars.

Inhibition and eradication of Salmonella Typhimurium biofilm using P22 bacteriophage, EDTA and nisin.

P22 phage >105 PFU ml-1 could be used to inhibit Salmonella Typhimurium biofilm formation by 55-80%. Concentrations of EDTA >1.25 mM and concentrations of nisin >1,200 µg ml-1 were also highly effective in reducing S. Typhimurium biofilm formation (≥96% and ≥95% reductions were observed, respectively). A synergistic effect was observed when EDTA and nisin were combined whereas P22 phage in combination with nisin had no synergistic impact on biofilm formation. Triple combination of P22 phage, EDTA and nisin could be also used to inhibit biofilm formation (≥93.2%) at a low phage titer (102 PFU ml-1), and low EDTA (1.25 mM) and nisin (9.375 µg ml-1) concentrations. A reduction of 70% in the mature biofilm was possible when 107 PFU ml-1 of P22 phage, 20 mM of EDTA and 150 µg ml-1 of nisin were used in combination. This study revealed that it could be possible to reduce biofilm formation by S. Typhimurium by the use of P22 phage, EDTA and nisin, either alone or in combination. Although, removal of the mature biofilm was more difficult, the triple combination could be successfully used for mature biofilm of S. Typhimurium.

Does subinhibitory concentrations of clinically important antibiotic induce biofilm production of Enterococcus faecium strains?

Biofilm structures are the most resistant form of active microorganisms against sanitation, disinfection, and sterilization processes. One of the specific properties of biofilm is the development of antibiotic resistance that can be up to 1,000-fold greater than planktonic cells. Enterococcus faecium is a human pathogen that causes nosocomial bacteremia and at the present time, it is well known that most of the chronic infections are biofilm-based. Recent evidence suggested that subinhibitory concentrations (sub-MICs) of antibiotics have an important role in the evolution of antibiotic resistance and induction on biofilm formation. Based on this information, we aimed to determine the effect of subinhibitory antibiotic concentrations on biofilm formation and the role of the antibiotic concentrations on the enterococcal surface protein gene (esp). To determine the impact of clinically important antibiotics on biofilm production, crystal violet assay was used. Then, the effect of sub-MICs of antibiotics on the expression of the esp gene was investigated by quantitative real-time PCR. Biofilm production assays show that MIC/2 of erythromycin (ERT; 512 µg/ml), MIC/32 of vancomycin (VAN; 16 µg/ml), MIC/64 of streptomycin (STR; 32 µg/ml), and MIC/128 of kanamycin (KAN; 4 µg/ml) values induce maximum biofilm production compared with the control. According to q-PCR results, sub-MIC values of ERT, VAN, and STR antibiotics were found to enhance esp gene expression. In addition, despite the increasing biofilm production after KAN treatment, the antibiotic was not effective on the esp expression.