4.6. Pulsed-Field Gel Electrophoresis–PFGE

Pulsed-field gel electrophoresis (PFGE) is a method of separating large fragments of DNA and is particularly useful for characterizing and typing bacteria for epidemiological studies. In PFGE, pure bacterial strains in agarose plugs are treated with enzymes and detergents (proteases and SDS) that release chromosomal DNA. The agarose plugs are then incubated with restriction enzymes, which cut at specific sites to generate a limited number of DNA fragments. The plugs are then subjected to electric current and alternate rotations in a magnetic field (which enhances the movement of large DNA fragments), leading to the size separation of DNA fragments and emergence of a banding pattern [200]. In an outbreak investigation of cholera over seven years, fifty isolates of Vibrio cholerae were subject to molecular typing by PFGE [201]. Analysis revealed that over the years, the outbreak involved 15 different pulsotypes of V. cholerae, four pulsotypes matched published pulsotypes and there were 11 new types. Notably, PFGE typing revealed the chronological emergence of new types, which subsequently replaced the earlier pulsotype.