Cronobacter sakazakii is an emerging opportunistic pathogen contaminating powdered infant formulas causing lethal threats to neonates and immune-deficient infants. It causes life-threatening infections, septicemia, neonatal meningitis, and necrotizing enterocolitis. The aim of this study was to test the commercially available formulas that are intended for consumption by 0-6 months old infants (neonates and immune-compromised infants), for the presence of Cronobacter spp., and to determine the presence of C.sakazakii in the stool of these infants who consumed these formulas through conventional methods as culturing, biochemical tests and PCR. Fifty PIF samples (different brands) retailed in upper Egypt were collected from Assiut University Children Hospital at the Gastroenterology and Hepatology and Preterm Units, and we checked the presence of C.sakazakii in them. Fifty Stool samples were also collected from the infants who were fed the studied PIF samples, to study the presence of C.sakazakii in the stool of these infants. The samples underwent three steps of pre-enrichment, enrichment procedures, and subculture onto chromogenic Enterobacter sakazakii agar plates. Biochemical tests were afterwards carried out. Finally, molecular characterization using specific PCR was done to detect Cronobacter sakazakii, targeting the ESA_02797 gene which is found in all C.sakazakii strains. The results of this study shed light on the immense need for applying effective prevention and control measures and taking all the precautions needed during the production and preparation of PIF to hinder its contamination with C.sakazakii and to prevent the spreading of such fatal infections to infants with low immunity and neonates.