Thirty-three antimicrobial agents, including antimicrobial peptides (nisin, lacticin 3147, isracidin), organic acids, emulsifiers (organic acid esters), glycine, lysozyme, tocopherol, EDTA, milk fat globule membrane, and the lactoperoxidase system (LPOS) were screened for anti-Cronobacter sakazakii activity. The compounds were initially screened individually in parallel in synthetic media. Those showing antimicrobial activity were then tested in reconstituted whole milk and finally in reconstituted powdered infant formula (PIF), using mild temperatures of reconstitution and prolonged storage at room temperature. Propionic acid and monocaprylin (as POEM M-100) in combination showed inhibitory activity at sufficiently low concentrations (0.1 to 0.2%) in milk to be considered as potential antimicrobial additives for the inhibition of C. sakazakii in reconstituted PIF. More interestingly, LPOS, when combined with the broad-spectrum bacteriocins nisin or lacticin 3147, inhibited outgrowth of C. sakazakii at 37 degrees C for 8 h. The combined effects of POEM M-100 and either acetate or propionate and LPOS with lacticin 3147 or nisin were evaluated under the Food and Agriculture Organization of the United Nations-World Health Organization high-risk scenario for PIF, i.e., low temperature of reconstitution and long storage or feeding times at ambient temperature. In the presence of LPOS and lacticin 3147, growth of Cronobacter spp. was inhibited for up to 12 h when the PIF was rehydrated at 40 or 50 degrees C. These results highlight the potential of combinatory approaches to improving the safety of infant milk formula.