Pulsed-Field Gel Electrophoresis (PFGE) standardization protocol for Proteus mirabilis

Abstract

Objective: For the detection of outbreaks caused by Proteus mirabilis, strains clonal relations are determined methods as “pulsed-field gel electrophoresis (PFGE)”. The aim of this study was optimization of a pulsed-field gel electrophoresis for molecular typing of P. mirabilis. Methods: In this study, PFGE\' protocol is optimized for use in molecular typing of P. mirabilis. Phylogenetic analyzes of strains were evaluated with Bionumerics software system (version 6.01; Applied Maths, Sint-Martens-Latem, Belgium). Results: This protocol compared with Gram-negative bacteria PFGE protocols, NotI enzyme is suitable for this bacterium. Electrophoresis conditions should be revealed as; - block 1: initial pulse duration 1 sec, ending pulse duration 30 sec, striking angle 120d, the current 6 V/cm2, temperature 14dC, time 8 hours; - block 2: initial pulse duration 30 sec, ending pulse duration 70 sec, striking angle 120d, the current 6 V/cm2, temperature 14dC, time 16 hours; - TBE, pH=8.4. Conclusion: P. mirabilis strains were typed by PFGE and Bionumerics analysis program were determined clonal relationships. The procedure was simple, reproducible and suitable for these bacteria. Also it was evaluated, because of reducing time, the solution volumes and enzymes can be economically. Outbreaks of nosocomial infections due to bacteria studied assessment and the potential to provide useful information about the degree of prevalence. This optimized protocol is allowed different centers\' PFGE results to compare with other laboratories results. J Clin Exp Invest 2013; 4 (3): 306-312