Stressed Salmonella are exposed to reactive oxygen species from two independent sources during recovery in conventional culture media

Previously, Stephens et al. [J. Appl. Microbiol. 83 (1997) 445-455] develop ed a sensitive technique for studying the resuscitation of low levels of st ressed Salmonella. Using this technique the influence on recovery performan ce of the peptone component of buffered peptone water was investigated. Wit hin 12 different peptone types as much as 3.5 log(10) cells/ml difference w as observed between the best and worst performing formulations. Poor recove ry performance was linked to reactive oxygen species (ROS) generation throu gh auto-oxidation of reducing sugars and photo-sensitisation of sensitive c omponents such as riboflavin. Supplementary recovery agents were explored w ith only Oxyrase(R), which has both enzymes to degrade ROS and the ability to rapidly turn a medium anaerobic, having any significant effect. It impro ved the speed of recovery and increased, by up to 100-fold, the number of s tressed cells recovered. Stressed cells were further studied by flow cytome try with cell sorting, based on the staining pattern from a novel fluorochr ome combination, into good and poor recovery media. It was identified that within a stressed population the removal of all oxygen protected actively r espiring cells the most by forcing them to generate energy from anaerobic m etabolism thus avoiding any risk from accidental endogenous ROS generation. The recognition of two independent sources of oxidative stress in the rout ine use of conventional culture media is discussed in relation to pathogen detection and other areas of food microbiology. (C) 2000 Elsevier Science B N. All rights reserved.