Molecular Fingerprinting of Bacterial Isolates

Genotypes and transmission patterns were determined by performing PFGE according to the method described in a previous study [19 (link)]. S. flexneri isolates were digested with the restriction enzyme NotI (TaKaRa, Japan) at 37 °C for 3 h to generate a DNA fingerprinting profile. Salmonella enterica serotype Braenderup strain H9812 was digested with XbaI (TaKaRa, Japan) and used as a molecular size standard. Electrophoresis was performed on the CHEF Mapper XA system (Bio-Rad) with a 1% agarose SeaKem Gold gel (Lonza, USA). Electrophoretic parameters were determined by performing multiple screening runs and included switching times of 2.16 to 54.17 s, a voltage of 6 v/cm, a 120° angle and a run time of 21 h. PFGE images were obtained using a Universal Hood II (Bio-RAD, USA) and analyzed using BioNumerics software version 7.1 (Applied Maths, Sint-Martens-Latem, Belgium). A clustering tree that indicated relative genetic similarity was constructed using UPGMA (Unweighted Pair Group Method with Arithmetic Mean) and the Dice-predicted similarity value with a 1.0% pattern optimization and 1.5% band position tolerance.