Application of thin agar layer method for recovery of injured Salmonella typhimurium

Xylose lysine decarboxylase (XLD) medium, a selective plating medium, can inhibit heat-injured Salmonella typhimurium from growing, whereas tryptic soy agar (TSA), a nonselective medium, does not. To facilitate recovery of heat-injured S. typhimurium cells while providing selectivity of isolation of S. typhimurium from other bacteria in the sample, a thin agar layer (TAL) procedure was developed by overlaying 14 ml of nonselective medium (TSA) onto prepoured and solidified XLD medium in a 8.5 cm diameter Petri dish. During the first few hours of incubating the plate, the injured S. typhimurium repaired and started to grow in the TSA. During the resuscitation of injured cells, the selective agents from XLD were diffused to the TSA top layer part. Once the selective agents diffused to the top part of the TAL, the resuscitated S. typhimurium started to produce a typical reaction (black color) and other microorganisms were inhibited by the selective agents. The recovery rate for heat-injured (55°C for 15 min) S. typhimurium with the TAL method was compared with TSA, XLD, and the traditional overlay method (OV; pouring selective agar on top of resuscitated cells on TSA agar 3–4 h after incubation). No significant difference occurred among TSA, OV, and TAL (P>0.05) for enumeration of heat-injured S. typhimurium, but they recovered significantly higher numbers than from XLD agar (P<0.05).