Consumption of food contaminated with Escherichia coli O157:H7 and Salmonella can cause enteric disease in consumers. If not properly sanitized, knives used during animal harvest can spread these and other pathogens. This study evaluated the reduction of E. coli O157:H7 and Salmonella on harvesting knives after nonthermal sanitation. Knives were inoculated in cocktails of E. coli O157:H7 or Salmonella and treated by 30-s immersions in ambient-temperature solutions (unless temperature was specified) of 1.1% sodium metasilicate (SMS), 200 ppm of quaternary ammonium compounds (QAC), 200 ppm of chlorine (Cl 2), 5% lactic acid (LA), 82.2°C water, and 21°C water. Initial and treated counts were determined by plating onto MacConkey and xylose lysine desoxycholate for E. coli O157:H7 and Salmonella, respectively. Initial counts were determined by sampling one side of the knife blade, while treated counts were sampled from the opposite side. Plates were incubated for 24 to 48 h at 37°C. Mean attachment of E. coli O157:H7 and Salmonella was 4.51 and 5.09 log CFU/cm2, respectively. Mean log reductions on knives inoculated with E. coli O157:H7 were 1.16, 3.51, 3.38, 1.38, 3.82, and -0.41 CFU/cm2 after treatment in SMS, QAC, Cl2, LA, 82.2°C water, and 21°C water, respectively (P ≤ 0.05). Knives inoculated with Salmonella showed reductions of 0.78, 3.42, 3.40, 2.91, 4.12, and 0.36 log CFU/cm2 after treatment in SMS, QAC, Cl2, LA, 82.2°C water, and 21°C water, respectively (P ≤ 0.05). Results indicate that some ambient-temperature sanitizing agents have the potential to significantly reduce E. coli O157:H7 and Salmonella populations on knives used during animal harvest.