Pulsed-field Gel Electrophoresis for Salmonella Typing

PFGE was performed according to PulseNet protocol developed by the CDC [24 ]. S. enterica serovar Braenderup (ATCC BAA 664) was used as the control strain. Briefly, agarose plugs containing genomic DNA treated with Proteinase K Buffer were incubated with 20 units of XbaI enzyme at 37 °C for 2 h (New England BioLabs, Ipswich, MA, USA). DNA separation was performed with 1% SeaKem Gold agarose gels in 0.5 M Tris borate–EDTA buffer at 14 °C for 18 h with pulse times between 2.16 and 63.8 s using a CHEF DR III apparatus (Bio-Rad, Hercules, CA, USA). Gels were stained with ethidium bromide, visualized under UV light, and photographed. DNA fingerprints were analyzed using Bionumerics 7.1 software (Applied Maths, Austin, TX, USA). A phylogenetic tree was constructed using the Unweighted Pair Group Method, with Arithmetic Mean method with Dice coefficient at an optimization setting of 1% and a position tolerance setting of 1.5%, as recommended previously [25 (link)].