The third and fourth sets of cells were for PMA-treated live cell dilutions and untreated live cell dilutions. Combination of qPCR with PMA treatment PMA treatment was performed as described earlier [21]. Briefly, separate live cells, heat-killed cells, and live/dead

cell mixtures were aliquoted 100 μl in three 1.5-ml microtubes. Two microliters of 10 mM PMA was added to each aliquot to a final concentration of 50 μM. The samples were first incubated at room temperature in the dark for 5 min, with gentle shaking. Then the samples were exposed to a 650-W halogen light source, followed by DNA preparation, and qPCR Citarinostat molecular weight analysis. Detection of live salmonella cells in spiked spinach and beef samples using PMA-qPCR Fresh Emricasan concentration spinach and ground beef purchased from a local retail source, which were confirmed to be free of Salmonella by standard FDA BAM methods [45], was used for the spiking studies. The studies consisted of two parts. In part 1, three Selleck LY2090314 spinach samples (25 g) and three beef samples (25 g) were inoculated with 3 × 101, 3 × 102 and 3 × 103 CFU/g Salmonella strain SARB16. In part 2, three samples three beef samples (25 g) were each inoculated with 3 × 107/g dead cells and with 3 × 101, 3 × 102, and

3 × 103 CFU/g of live cells, respectively. Each spinach or beef sample was mixed with 225 ml of LB medium and homogenized for 2 min using a stomacher (Seward, England). Five milliliters of the enriched cultures was collected at 0, 4, 8, 12 and 24 h after incubation at 37°C with shaking at 180 rpm. The collected samples were centrifuged at 600 × g for 1 min to collect leaf or fat tissues. The supernatants were transferred to 2.0-ml microtubes and centrifuged at 3000 × g for 5 min to collect cells. The cell pellets were suspended in 1.5 ml of LB medium and treated with PMA before DNA extraction and qPCR analysis. Acknowledgments The authors are in debt to Christopher A. Elkins and Ben Tall for critically reviewing this manuscript and providing insightful comments and suggestions.

We thank Huanli Liu for reading this manuscript and giving useful suggestions and Mark Mammel for help in getting Dolichyl-phosphate-mannose-protein mannosyltransferase the background information on bacterial collections in DMB. Additionally, we want to thank the three reviewers who critically reviewed the manuscript and provided useful suggestions for revising the manuscript. Electronic supplementary material Additional file 1: Table S1: Salmonella enterica strains of the SARA and SARB reference collections used in this study. (DOC 59 KB) Additional file 2: Table S2: Selective detecion of live Salmonella cells spiked in beef by PMA-qPCR. (XLS 36 KB) References 1. Alali WQ, Thakur S, Berghaus RD, Martin MP, Gebreyes WA: Prevalence and distribution of Salmonella in organic and conventional broiler poultry farms. Foodborne Pathog Dis 2010, 7:1363–1371.PubMedCrossRef 2.