Standardized PFGE Protocol for K. pneumoniae

We used the 1-day, standardized PFGE protocol for K. pneumoniae [27 (link)]. Cell suspensions were placed in polystyrene tubes (Falcon; 12 × 75 mm), and their optical density was adjusted to 3.8–4.0 using a Densimat photometer (BioMérieux, Marcy l’Etoile, France). K. pneumoniae slices were digested using 50 U per slice of XbaI (Takara, Dalian, China) for 4 h at 37 °C, and electrophoresis was performed using a CHEF-DRIII system (Bio-Rad Laboratories, Hercules, CA, USA). Electrophoresis was run with a switch time of 6 s to 36 s for 18.5 h, and images were captured using a Gel Doc 2000 system (Bio-Rad) and converted to TIFF files. The TIFF files were analyzed using the BioNumerics version 5.1 software (Applied Maths, Kortrijk, Belgium).