This study evaluated the effectiveness of warm solutions of acidic calcium sulfate (ACS), lactic acid (LA), epsilon-polylysine (EPL), ACS plus EPL and sterile distilled water (W) applied to the surface of fresh, pre-rigor beef rounds for reducing Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes applied as a cocktail at a level of 6.4 log cfu/ 50 cm(2). All treatments were applied for 15 to 20 s using a stainless steel spray cabinet at 50-55C under a constant pressure. Sequential application of warm ACS, followed by EPL significantly reduced inoculated levels of S. Typhimurium, E. coli O157: H7 and L. monocytogenes, with an extended effect over seven storage days. This combination was more effective than single treatments of ACS, LA, EPL or W alone. From these results, it appears that a sequential application of ACS and EPL could be a better strategy for pathogen reduction in meat plants than a single decontamination treatment.